Mo.

wflfiffi: ‘6? 1.2-);

   
       
       
  
      

5§¢fi.§

   

 
  

{:fi‘tfi a}:
‘ $31 $42» if"
. 4.1{Qfs

«ML r‘il' fer-q
{9% :(QShg‘h‘

  

   

0 .-
.w +1.
I0 I 0
$3M.

    

4 vu)‘
'uf . .“

 

  

a‘ ‘ ' 'x'
‘ ,1‘.:.| L“
9'? (‘VL j"
.“I .I'I:\‘\. lJ.“:‘.‘ll H: ‘:'.IIII.‘I‘

        
     

..
.

"

u.}-.-

Wm.

.~
1.31

A LJ’-.\
w“

,-

:'. i it‘d-ta. o

‘5 *"t' :1 4thh-
1..

.fi
U,” n 3.73;.

-‘-

V-_._.
v, —

 

 
    

   

3 1293 10337 zealM LIBRARY

Michigan State
University

 

This is to certify that the

thesis entitled

Postpartum Reproductive and Lactational
Performance of Cows Relative to Energy,
Phosphorus and Hormonal Status

presented by
Jane Allison Carstairs

has been accepted towards fulfillment
of the requirements for

Ph.D. degree in Dairy Science

 

Major professor

/ /
Date%//@ 7/; /? 7?

0'7 639

 

POSTPARTUM REPRODUCTIVE AND LACTATIONAL PERFORMANCE OF

COWS RELATIVE TO ENERGY, PHOSPHORUS AND HORMONAL STATUS

BY

Jane Allison Carstairs

A DISSERTATION

Submitted to
Michigan State University
in partial fulfillment of the requirements
for the degree of

DOCTOR OF PHILOSOPHY

Department of Dairy Science

1978

-ABSTRACT
POSTPARTUM REPRODUCTIVE AND LACTATIONAL PERFORMANCE OF
COWS RELATIVE TO ENERGY, PHOSPHORUS AND HORMONAL STATUS
. By

Jane Allison Carstairs

To determine if energy and phosphorus status would affect postpartum
health, lactational and reproductive performance, dairy cows were assigned
at parturition, six per group, to: (1) high energy, high phosphorus; (2)
high energy, low phosphorus; (3) low energy, high phosphorus; (4) low
energy, low phosphorus. Energy status (MCal/day) fbr highland low energy
groups was 8.4 and ~3.7, (P = .001), representing 135 and 85% of require-
ments. Phosphorus status (g/day) fbr high and low phosphorus groups was
20.6 and -1.9, (P = .001) or 138 and 98% of requirements. Treatment ex-
tended through 84 days postpartum after which standard rations were fed.
Heifers were bled twice weekly. Reproductive status was monitored via
serum progesterone palpation per rectum. The entire experiment was repli—
cated. Milk yields were not statistically different until week 5 of lac-
tation. Prom weeks 5—12, low phosphorus groups yielded 1.8 kg/day more
milk than high phosphorus groups (P §_.03). Energy did not affect milk
yield until week 9 when persistency of low energy groups declined. Over-
all means for milk yield (kg/day) for high and low energy were 23.0 and
22.0, for high and low phosphorus 21.7 and 23.2. Low phosphorus groups

produced 816 kg more milk (P = .01) in 305 days than high phosphorus

Jane Allison Carstairs

groups while energy groups did not differ in total yield. High energy
groups had almost twice as much (P = .15) disease and higher (P = .006)
rectal temperatures than low energy groups. Means were 38.9 and 38.7 C.
All heifers had temperatures elevated above normal during the first month
of lactation.

Body weight, serum glucose, insulin and propionate increased with
energy status. Weight changes (kg/week) for high and low energy groups
were 0.54 and -1.49, (P = .03). In trial 1 high energy groups had more
(P = .06) serum glucose (mg/d1) than low energy groups, 75.5 versus 71.6.
In trial 2, glucose values were approximately 14 mg/dl lower than in trial
1, 60.4 and 58.8 (P = .025) for high and low energy groups. Glucose for
high and low phosphorus groups were also 60.4 and 58.8 (mg/d1) (P = .026).
In triallq insulin concentrations were 3-4 fold more (P < .0005) in high
energy than low energy groups, 16.5 versus 6.0 (uU/ml). In trial 2, values
were lower but still different, 7.3 versus 5.7 (P = .001). Serum propio—
nate increased (P = .001) with energy, 23.9 versus 15.3 uM for high and
low energy groups. By 9 weeks of lactation serum growth hormone (ng/ml)
was higher (P §_.02) in low energy than high energy groups, 4.8 versus
3.5 for weeks 9-12. Growth hormone in high energy groups declined as lac-
tation advanced, in low energy groups it did not change.

Serum acetate (over mean = 736.4 uM) was not different among groups.
Serum butyrate and isobutyrate varied inconsistently with energy and phos-
phorus status.

Phosphatemia in low phosphorus groups indicated marginal phOSphorus
deficiency. In trial 1, high phosphorus groups had more (P = .02) serum
phosphorus than low phosphorus groups, 7.1 versus 6.0 mg/dl. In trial 2,

high phosphorus groups again had higher (P = .0006) serum phosphorus, 5.6

Jane Allison Carstairs

versus 5.2, but high energy groups exceeded low energy groups in serum
phosphorus, 5.5 versus 5.3 (P < .0005). Serum calcium was inversely re-
lated to serum phosphorus, means for high and low phosphorus groups for
serum calcium were 9.5 and 9.7 mg/dl (P = .001).

Regression analysis indicated that physiological concentrations of
growth hormone, propionate, butyrate and isobutyrate varied with serum
insulin (p < .005, R2 = 0.26).

Reproductive function was almost identical among energy and phosphorus
groups. No heifer with peak progesterone prior to insemination below 2.7
ng/ml conceived to that insemination. Days to reach 3 ng/ml progesterone
were correlated with serum phosphorus, glucose (r = -0.34, -0.33, P = .05)

and growth hormone (r = 0.51, P = .05).

ACKNOWLEDGMENTS

I would like to acknowledge the following people for their help
throughout my graduate studies:

My Major Professor, Dr. Roy Emery, for his enthusiasm, discussion,
inspiration and for always being ready to help. The members of my com-
mittee, Dr. Allen Tucker for his constructive guidance, especially in
writing, Dr. Loran Bieber for his ever positive attitude, and Dr. David
Morrow for his help and advice with the reproductive aspect of this study.

Dr. Charles Lassiter and Dr. Harold Hafs for making the facilities
of the Department of Dairy Science available for the execution of this
research.

Dr. John Gill and Roger Neitzel for statistical advice and computer
programming.

James Liesman for his help in sample collection.

Thanks to all the graduate students in the Department, especially
those who ever 'lived' in 218 Anthony Hall.

Finally, special thanks to Wilson Cunningham, for helping me retain

my sanity and sense of humour.

ii

TABLE OF CONTENTS

LIST OF TABLES .
LIST OF FIGURES
INTRODUCTION .
LITERATURE REVIEW

Temporal and Non-Dietary Changes in Certain
Hormones and Metabolites .

Insulin .

Growth Hormone

Glucocorticoids .

Prolactin .

Glucagon

Glucose . . .

Non-Esterified Fatty Acids

Calcium .

Phosphorus . . . . . . . . .
Magnesium . . . . . . . . . .
Volatile Fatty Acids . . . .

Reproduction as Affected by Energy and Phosphorus Intake

Energy and Reproduction .'. . . . . . .
Phosphorus and Reproductive Function
The Interrelationship Between Hormones and Metabolites

MATERIALS AND METHODS

Experimental Design . .
Statistical Analysis

Analysis of Serum Hormones
Analysis of Serum Metabolites .

RESULTS

Feed Intake . . . .

Summary of Data Analysis . .

Energy and Phosphorus Status

Milk Production . . . . . . . .

Health . . . . . . . . . . . . . . . . . . . . .
Energy Status Relationship . . . . . . . . . . . . . .
Indicators of Phosphorus Status .

iii

Page

.viii

45

45
48
50
51

55

55
S6
S6
59
68
72
87

Table of Contents (continued)

Regulation of Serum Insulin .

Reproduction Data .
Simple Correlations .

DISCUSSION .
CONCLUSIONS
APPENDIX .

BIBLIOGRAPHY .

iv

Page
90
93
98

99

. 118

. 121

. 125

Table

10.

11.

12.

13.

LIST OF TABLES

Mean dry matter intake (kg/day) of corn silage, hay and
grain for the first 12 weeks of lactation in trials 1
and 2 . . . . . . . . . . . . . . .

Mean energy status (MCal/day) of high and low energy
groups as a function of weeks of lactation for trials
1 and 2 .

Mean phosphorus status (g/day) as a function of weeks
of lactation f0r both trials . . . . . .

Mean daily milk yield (kg/day) fOr both trials as a
function of weeks of lactation . . . . . . .

Mean daily milk yield (kg/day) for both trials for high
and low energy and high and low phosphorus groups .

Regression of energy and phosphorus status with milk
yield, milk fat % and milk protein % for both trials

Mean milk fat percent for the first 12 weeks of lactation
for both trials . . . . . . . . . . . . . . . . . . .

Mean milk protein percent for the first 12 weeks of
lactation for both trials .

Cummulative 305 day milk yield (kg) for heifers for
both trials . . . . . . . . . . . . . . . . .

Mean incidence of disease in both trials for the first
12 weeks of lactation . . . . . . . . . . .

Types of health problems in high and low energy groups
f0r the first 12 weeks of lactation . . . . . .

Mean rectal temperature (C) for the first 12 weeks of
lactation fer the second trial . . . . . . . . . .

Change in body weight (kg/week) for the first 12 weeks
of lactation f0r both trials . . . . . . . . . . . . .

Page

55

57

59

61

61

64

64

65

66

68

69

70

73

Table Page

14. Regression of energy and phosphorus status with body
weight change . . . . . . . . . . . . . . . . . . . . . . . . 74

15. Mean serum glucose concentrations (mg/d1) for the first
12 weeks of lactation for the first trial . . . . . . . . . . 75

16. Mean serum glucose concentrations (mg/d1) for the first
12 weeks of lactation for the second trial . . . . . . . . . 75

17. Regression of energy and phosphorus status with serum
glucose concentrations . . . . . . . . . . . . . . . . . . . 76

18. Mean serum insulin concentrations (uU/ml) for the first
trial as a function of weeks of lactation . . . . . . . . . . 77

19. Regression of energy and phosphorus status with serum
insulin concentrations . . . . . . . . . . . . . . . . . . . 77

20. Mean serum insulin concentrations (uU/ml) for the first
12 weeks of lactation in the second trial . . . . . . . . . . 79

21. Mean serum acetate concentrations (uM) for the first 12
weeks of lactation in the second trial . . . . . . . . . . . 81

22. Regression of energy and phosphorus status with serum
volatile fatty acids . . . . . . . . . . . . . . . . . . . . 81

23. Mean serum propionate concentrations (uM) for the first
12 weeks of lactation in the second trial . . . . . . . . . . 82

24. Mean serum butyrate concentration (uM) for the first 12
weeks of lactation in the second trial . . . . . . . . . . . 83

25. Mean serum isobutyrate concentration (uM) fOr the first
12 weeks of lactation in the second trial . . . . . . . . . . 84

26. Simple correlations between serum VFA . . . . . . . . . . . . 86

27. Mean serum growth hormone concentrations (ng/ml) as a
function of weeks of lactation . . . . . . . . . . . . . . . 87

28. Mean serum phosphorus concentrations (mg/d1) for the first
12 weeks of lactation in the first trial . . . . . . . . . . 88

29. Mean serum phosphorus concentrations (mg/d1) for the first
12 weeks of lactation in the second trial . . . . . . . . . . 88

30. Regression of energy and phosphorus status with serum
phosphorus . . . . . . . . . . . . . . . . . . . . . . . . . 89

vi

Table Page

31. Mean serum calcium concentrations (mg/d1) for the
first 12 weeks of lactation for both trials . . . . . . . . . 89

32. Regression of energy and phosphorus status with
serum calcium . . . . . . . . . . . . . . . . . . . . . . . . 90

33. Simple correlations with insulin in trials 1 and 2 . . . . . 91

34. Deletion regression analysis with serum insulin as
dependent variable: standard coefficients . . . . . . . . . 92

35. Number of days postpartum for serum progesterone to

reach 3 ng/ml in both trials . . . . . . . . . . . . . . . . 94
36. Number of days postpartum to first ovulation . . . . . . . . 95
37. Mean number of days non-pregnant for both trials . . . . . . 96

38. Mean number of inseminations per conception for both
trials . . . . . . . . . . . . . . . . . . . . . . . . . . . 97

39. Mean number and percent undetected estrus for both trials
for the first 15 weeks of lactation . . . . . . . . . . . . . 97

A1. Composition of grain mixes (%) . . . . . . . . . . . . . . . 121

A2. Dry matter, net energy and phosphorus values used to
calculate NE intake and phosphorus intake for trial 1 . . . . 122

A3. Dry matter, net energy and phosphorus values used to
calculate HE intake and phosphorus intake for trial 2 . . . . 123

A4. Simple correlations from both trials . . . . . . . . . . . . 124

vii

Figure

LIST OF FIGURES

Mean energy status (N.R.C. %) for the first 12 weeks
of lactation . . . . . . . . . . . . . . . . . .

Mean phosphorus status (N.R.C. %) for the first 12
weeks of lactation . . . . . . . . . . . . .

Mean milk yield (kg/day) for the first 12 weeks of
lactation . . . . . . . . . . . . . . . . . . . .

Mean rectal temperature (C) for the first 12 weeks of
lactation . . . . . . . .

Mean serum insulin (uU/ml) for the first 12 weeks of
lactation in trial 1 . . . . . . . . . . .

Mean serum insulin (uU/ml) for the first 12 weeks of
lactation in trial 2 . . . . . . . . .

Mean serum growth hormone (ng/ml) for the first 12 weeks
of lactation in trial 2 . . . . . . . . . . . . . . .

Deletion regression analysis with serum insulin as
dependent variable . . . . . . . . . . . .

viii

Page

58

6O

63

71

78

80

86

92

INTRODUCTION

This study was conducted to determine if energy or phosphorus
status would affect postpartum performance of dairy cows. Performance
was considered not only as milk production but health and reproduction
also since all of these parameters are part of a successful lactation.

In order to accomplish this, performance was recorded for the first
3 months postpartum in Holstein heifers subjected to widely different
intakes of energy and phosphorus. This early postpartum period is
critical since lactational and reproductive performance during this time
affects the total and subsequent lactations. Energy and phosphorus in-
take for each animal was calculated relative to published recommendations
and performance was related to this calculated status. A number of
metabolic parameters were measured to determine if they were correlated
to energy or phosphorus status and attempts were also made to correlate
these parameters with differences in performance of cows.

Efficiency of production is a primary goal of the livestock industry.
If excess nutrients have detrimental or no beneficial effect on perform-
ance this has practical implications. Feed wastage is not economically
or conservationally sound. Marginal deficiencies should also be evalu—
ated for effects on dairy cow performance. The nutritional requirements
of the postpartum cow are controversial. The aim of this research was
to define requirements for early lactation more accurately and to relate
these requirements to the physiology and endocrinology of the cow.

1

LITERATURE REVIEW

In order to assess the metabolic needs of the dairy cow in the post-
partum period it is important to understand the changes that occur at
this time. This literature review will first describe the changes in
certain hormones that occur after calving and through lactation. Then,
changes in certain metabolites will be discussed.

Next, the review will describe the changes in reproductive status
which occur after calving and how energy or phosphorus intake might affect
and be related to this.

Finally, the interactions between hormones and metabolites will be
discussed within the context of how nutrient utilization might be control-
led on a day to day basis by the lactating cow.

Hopefully, this review will form the basis for detailed discussion

of the experimental data to follow.

Temporal and Non-Dietary Changes in Certain Hormones and Metabolites

 

This section will review the changes which occur in insulin, gluca-
gon, growth hormone, glucocorticoids and prolactin in periparturient and
postpartum cows. These hormones are closely involved with control of
metabolism and utilization of nutrients. If we can understand how, and
perhaps why, these hormones change in postpartum cows we can manipulate
these hormones (perhaps by dietary means) so that nutrients might be

more efficiently used at peak lactation. An understanding of these

3

hormonal signals might also enable us to control metabolic disorders
where nutrient utilization is impaired.

Next, the changes in glucose, non-esterified fatty acids, calcium,
phosphorus, magnesium and volatile fatty acids will be described. It
is important to know whether serum values are indicative of dietary in-
take of a substance, what we can predict from examination of concentra—
tions of serum metabolites and whether these have direct application
to nutritional problems or lactational performance. Only when the normal
is established can deviations from this be discussed and solutions

arrived at.

Insulin

Plasma insulin in ruminants, as measured by radioimmunoassay, varies
between 5 and 10 uU/ml, with basal concentrations of 10-59, 15-29 and 40
uU/ml for sheep, cattle and goats, respectively (Trenkle, 1972). Doubt-
less these ranges reflect no only inter-laboratory and time of sampling
differences but also physiological and nutritional factors.

In normal cows there is no marked elevation in insulin concentra-
tions at parturition (Schwalm and Schultz, 1976). Insulin concentrations
do, however, increase with weeks after parturition (Koprowski and Tucker,
1973a; Smith et al., 1976; Gorewit, et al., 1977). Reported increases
vary from 30% in the first 56 days of lactation (Smith et al., 1976),

30% increase in the first four months postpartum (Gorewit et al., 1977)
and a more dramatic 2-3 fold increase between the fOurth and twelfth

weeks of lactation (Koprowski and Tucker, 1973a). Thereafter, insulin
remains relatively stable. Differences in reported absolute values of

insulin may, in part, be due to sampling time relative to feeding.

4
Eating causes release of insulin in sheep, cattle and other ruminants,
consequently, time of sampling can influence values. Postprandial insu-
lin response will be discussed later in this review.

Thus, insulin concentrations increase in early lactation, along
with milk production and feed intake. Indeed, in early lactation, serum
insulin was positively correlated with nutrient intake (either dry matter
intake or estimated net energy intake) (Smith et al., 1976) and parallelled
increasing serum glucose (Gorewit et al., 1977). In support of these
observations that insulin reflects changing nutritional (energy) status,
insulin concentrations were elevated almost 20% by an ad libitum grain-
restricted roughage diet (Walker and Elliot, 1973), insulin concentrations
doubled when straw was reduced from 47.5 to 17.5% of the diet and re-
placed by grain (Blair et al., 1974) and there was almost a three-fold
difference in insulin concentrations between heifers fed more than 100%
versus heifers fed 75% of their N.R.C. energy requirements (Carstairs,
1975). It should be noted that none of the above studies involved
isocaloric diets. Not only does insulin seem to reflect energy level of
the diet, however, it also may indicate the amount of roughage in iso-
caloric diets (Evans et al., 1975). Insulin concentrations were 20%
higher in the low roughage group cows compared with the high roughage
group. The insulin may have increased in response to increased propionate
type fermentation or to more glucose reaching the lower gut.

It is not clear whether serum insulin concentrations are the same
in lactating and non-lactating cows (Schwalm and Schultz, 1976; Chang
et al., 1977) or higher in the latter (Hove, 1974). Multiparous cows
have greater (33%) concentrations of insulin than primiparous cows

(Koprowski and Tucker, 1973a). It is not clear at this time why this is

5
so. In addition, insulin decreases as pregnancy progresses but is not
affected by sex of fetus or season of the year (Koprowski and Tucker,
1973a). Insulin is responsive to other metabolic stimuli, these will be

discussed in the last section of this review.

Growth Hormone (GH)

 

It has been reported that serum GH concentrations gradually increased
up to, and then doubled at, the time of parturition. This peak was of
about 12 h duration, concentrations decreased to prepartum concentrations
by about 4 days postpartum (Ingalls et al., 1973). Another study does
not support these findings, reporting, on the average, no changes, al-
though some cows did show peaks at parturition (Reynaert et al., 1976).

In contrast to insulin, GH decreases with time after parturition,
and this is well documented (Koprowski and Tucker, 1973a; Smith et al.,
1976; Halse et al., 1976). Smith et al. (1976) reported a 30% decrease
by 41 days postpartum, Halse et a1. (1976) reported a 70% decrease by 90
days. In longer term studies it has been shown that GH concentrations,
if adjusted for stage of pregnancy and season of the year, linearly
decrease by 50% from one to eleven months postpartum (Koprowski and
Tucker, 1973a). As this suggests, GH decreases as lactation advances
and increases as gestation progresses. GH is not affected by season or
sex of fetus but in contrast to insulin it is not affected by cow parity
either. The milking stimulus does not effect GH release (Koprowski and
Tucker, 1973a).

Finally, it has been reported that GH concentrations in early lacta-
tion were negatively correlated with dry matter intake and estimated net

energy intake (in contrast to insulin which was positively correlated)

6
(Smith et al., 1976). Further support for this idea that GH may reflect
nutritional status (in the opposite way to which insulin might) is not
available at the present time. Other regulators of GH release will be

discussed in the last section of this review.

Glucocorticoidsg(GCD)

 

There is little doubt that GCD concentrations increase very rapidly
at parturition. Concentrations may increase as much as 3, 5 or 8 times
over prepartum values (Edgerton and Hafs, 1973; Convey et al., 1974;
Arije et al., 1974; Paterson and Linzell, 1974).

During lactation, GCD concentrations are extremely variable. Values
varied between 11 and 62 ng/ml from day 15 postpartum in a study with
beef cows (Arije et al., 1974). One the average, however, it seems that
GCD concentrations change little as lactation advances (Koprowski and
Tucker, 1973a; Edgerton and Hafs, 1973; Convey, 1974). Not all studies
support these findings, however, because Blom and Halse (1975) found the
lowest GCD concentrations in cows within the first month of lactation and
the highest at 2.5-3 months. Paterson and Linzell (1974) found GCD dur-
ing lactation 3 times higher than in late pregnancy.

Neither gestation, season, fetal sex nor parity affect GCD concen-
trations but GCD is released by the milking stimulus and this release is
not diminished as lactation proceeds (Koprowski and Tucker, 1973a).

At this time there is no evidence available to suggest that serum
GCD concentration in any way reflects nutritional status. GCD may, how-
ever, be involved in the regulation of metabolism and this will be dis-

cussed later in this review.

Prolacting(PRL)

 

Serum PRL concentrations begin to increase a few days prepartum, in-
creases 5-6 times at or just before parturition and remain elevated for
some days or weeks after calving (Edgerton and Hafs, 1973; Arije et al.,
1974; Convey, 1974). Beef cows perhaps have higher and more variable serum
PRL concentrations in the peripaturient period (Arije et al., 1974). This
may be due to the more freguent suckling stimulus applied by the calf.

PRL is dramatically affected by season (more Specifically temperature
and daylength) (Koprowski and Tucker, 1973b; Bourne and Tucker, 1975;
Peters and Tucker, 1978). During April to September in Michigan (warmer
temperatures and longer days) mean serum PRL concentrations are twice
as high as from October to March (Koprowski and Tucker, 1973b). It is
well documented that PRL is released in response to the milking (or
suckling) stimulus. This response increases up to, and is highest at,
about 8 weeks of lactation (corresponding to peak milk yield) and gradu-
ally declines as lactation advances. By 32 weeks of lactation (when milk
yield was about 20 kg/day in this experiment) PRL was no longer released
in response to milking (Koprowski and Tucker, 1973b).

Multiparous cows in the above experiment produced more milk and had
greater PRL concentrations than primiparous cows. No pregnancy effect was
noted but lactating non-pregnant cows did have greater PRL concentrations
than lactating pregnant cows. There was no influence of fetal sex or stage
of the estrous cycle on PRL concentrations (Koprowski and Tucker, 1973b).

Evidence for a direct effect of diet on PRL concentrations is not
available at this time but its possible involvement with metabolic regu-

lation will be discussed later.

Glucagpn (GLG)

 

There is, unfortunately, not much information on serum GLG concen-
trations in cows because of the unavailability up to this time of a
bovine GLG radioimmunoassay. The cow has both pancreatic and gut GLG as
do the non-ruminants which have been studied (Manns, 1972). Separating
these two GLG species from plasma, Manns (1972) showed that GLG like
immunoreactivity (GL1) increases 3-5 fold in the initial days of
lactation. This increase followed a similar course to milk production
and was found to be primarily of gut rather than pancreatic origin.
Concentrations remained elevated until 12 days postpartum when the experi-
ment was terminated. It is possible, the author suggests, that the two-
fold increase in grain intake with lactation might have resulted in glu-
cose entering the small intestine thus elevating gut GLI secretion.

The role of gut GLG is not clear at this time but possibilities
will be discussed in a later section of this review.

Serum GLG concentrations in monthly serum samples from dairy cows
increased from 579 pg/ml at 1 month to a maximum of 693 pg/ml at 2
months, falling to 633 pg/ml at 4 months of lactation, (Gorewit et al.,
1977). These data are limited and probably include both species of GLG
but they are of the same order of magnitude as those reported by Manns
(1972).

The role of GLG in metabolism will be discussed further. As yet

there is not enough data to know whether GLG would reflect energy status.

Glucose
Serum glucose concentrations increase 56-66% at parturition and

return to prepartum concentrations by 24 h postpartum (Schwalm and

9
Schultz, 1976; Bolenbaugh, 1977). For the first 3 or 4 months of lacta-
tion glucose concentrations increase gradually, after this time there is
a decline (Blom and Halse, 1975; Smith et al., 1976; Gorewit et al.,
1977). It should be noted that some studies have shown no variance of
plasma glucose during pregnancy and lactation (Paterson and Linzell,
1974) and also wide variation (43-88 mg/dl) at different stages of lacta-
tion (Halse et al., 1976). No doubt these differences are due to factors
such as experimental design, time and frequency of sampling and nutri-
tional regimes.

Certainly, nutritional and metabolic status seems to influence glu-
cose concentrations. Ketotic cows, immediately postpartum, have lower
serum glucose concentrations than their non-ketotic contemporaries
(Schwalm and Schultz, 1976). It has been reported that glucose concen-
trations were positively correlated with either dry matter intake or esti-
mated net energy intake during the first 2 months of lactation (Smith et
al., 1976), that glucose concentrations tended to increase as energy in
the diet increased (Blair et al., 1974). In addition, mid-lactation
cows, (150 days postpartum), showed an increase of 18 mg/dl glucose over
initial values in response to grain feeding (Jenny et al., 1974) and cows
fed 75 or 50% of their previous voluntary feed intake had significantly
lower serum glucose concentrations than those fed 100% (Fisher et al.,
1975).

Finally, there is some evidence that glucose concentrations may
respond not only to energy intake in the diet but also to roughage con-
tent. It has been reported that glucose concentrations were significant-
ly higher in cows fed a low, compared with a high, roughage diet even

though these diets were isocaloric (Evans et al., 1975). Whether this

10
is due to increased propionate type fermentation or more carbohydrate

reaching the lower gut in the low roughage fed cows is not clear.

Non-esterified Fatty Acids (NEFA)

 

Kronfeld (1965) reported mean jugular plasma NEFA concentrations of
286 qu/l (14 mg/dl). The range was 46-864 qu/l and distribution was
skewed so that 88% of values were between 50 and 450 and another 11%
between 450 and 659 qu/l. This illustrates the variability of NEFA con-
centrations in blood.

NEFA concentrations either double (Reynaert et al., 1976) or in-
crease as much as six times at parturition (Schwalm and Schultz, 1976).
Concentrations remain elevated for 6 or 7 weeks postpartum and then
begin to decline to prepartum values (Schwalm and Schultz, 1976; Chang
et al., 1977). The largest NEFA concentrations usually occur at peak
milk production (Bowden, 1971). The obvious conclusion from these data
is that NEFA are a reflection of greater or lesser energy status (Belyea
et al., 1975a) but there are limitations to this generalization. Indeed,
some studies have shown that NEFA concentrations were closely associated
with energy status (Erfle et al., 1974; Fisher et al., 1975; Carstairs,
1975), some have failed to show NEFA responding to dietary changes
(Blair et al., 1974) possibly because the variability was too large to
enable differences to be detected. In conclusion, it seems that using
NEFA concentrations as a criterion of energy status during lactation is
of limited usefulness.

The reader is referred to Bowden (1971) and Belyea et al. (1975a)
for tables summarizing NEFA concentrations in cattle in various physiolo-

gical and nutritional states from a number of additional literature sources.

ll

Calcium4(Ca)

 

Serum Ca concentrations in dairy cows are relatively stable. Analy-
ses of serum from 4000 cows of various breeds showed a mean of 9.55 mg/
d1, with a coefficient of variation of only 7.8% (Sansom, 1973). In
another study utilizing samples from 236 Guernseys mean TCA soluble
blood Ca was 7.4 mg/dl (range 3.7-12.0 mg/dl) (Lane et al., 1968).

Concentrations increase 1-2 mg/dl at parturition then begin to in-
crease further postpartum (Belyea et al., Bolenbaugh, 1977). In weeks
1 to 10 of lactation serum Ca gradually increases (Belyea et al., 1975a;
Carstairs, 1975). Month of lactation, however, seems negatively corre-
lated with serum Ca (Lane et al., 1968).

Pregnancy does not influence Ca concentrations, season apparently
does, although this may in part be due to nutritional programs which are
seasonal (Lane et al., 1968). It has been shown by an extensive study
(involving hundreds of cows) that there is a significant decrease in
mean serum Ca concentrations with increasing age (Tumbleson et al., 1973).
Mean serum Ca for Holsteins declined from 11.6 mg/dl at less than 0.5 hr
of age to a mean of 10.5 mg/dl at 5 yr of age. From 3 through 10 yr of
age, mean Ca was significantly lower in Holsteins than Guernseys. It
should also be noted that plasma Ca concentrations are inversely corre-
lated with plasma phosphorus concentrations (for plasma phosphorus be-
tween 3.7 and 26.0 mg/dl) (Symonds and Manston, 1974).

In addition, the reader is referred to a summary of Ca concentra-

tions from various literature sources (Belyea et al., 1975a).

12

Phosphorus (P)

 

When 236 Guernseys were sampled a mean phosphorus concentration of
6.1 mg/dl (range 3.7-10.1 mg/dl) (Lane et al., 1968) was found. Neither
season nor milk production affected serum concentrations. It is not
clear at this time whether stage of pregnancy affects serum P levels.
Serum P concentrations decrease at parturition and have increased almost
to prepartum values 24 h after calving (Bolenbaugh, 1977).

Serum P significantly decreases as age of the animal increases
(Lane et al., 1968; Tumbleson et al., 1973). At less than 0.5 yr of age
mean serum P was 10 mg/dl and this decreased to 5.8 mg/dl by 4-6 yr of
age (Tumbleson et al., 1973). The mean for 2-3 yr olds was approximately
6.6 mg/dl.

Except in cases of severe deficiency, it is not clear that serum
P concentrations reflect intake variations. Certainly, when cows were
fed differing amounts of P (although all diets provided all nutrients
equal to or greater than N.R.C. requirements) serum P was not different
among groups (Belyea et al., 1975a). Even when 75% of N.R.C. require-
ments were fed to heifers for the first 12 weeks of lactation there was
only a one mg/dl serum P difference (significant nonetheless) between
this group and heifers consuming greater than 100% of their requirements
(Carstairs, 1975). In a practical situation a 15% difference such as
this would be difficult to detect in individual animals. Only in extreme
(rather than marginal) circumstances, in the field or in controlled ex-
periments will P concentrations decrease to half of normal (2.5-3.0 mg/

d1) (Eckles et al., 1935; Palmer et al., 1941; Morrow, 1969).

13

Magnesium (Mg)

Extensive studies with dairy cows have shown serum Mg concentrations
to be about 2.4 mg/dl (with a coefficient of variation of about 17%)
(Lane et al., 1968; Sansom, 1973). In ruminants Mg absorption rate
varies considerably, low absorption is caused by high potassium content
and/or low fibre content in feed and high pH in rumen fluid (Meyer, 1976).

Blood Mg concentrations are primarily controlled by renal excretion,
if blood levels drop too low, excretion ceases. The following classifi-
cation of Mg concentrations for sheep and cattle was described by

Meyer (1976):

Normal 1.8-3.2 mg/dl
Slight hypomagnesemia 1.2-1.8 mg/dl
Severe hypomagnesemia §_ 1.2 mg/dl

Mg reserves of the body are quite low, mainly in the skeleton, this
makes compensating for Mg deficiency quite difficult.

Pregnancy does not influence Mg concentrations but Mg is positively
correlated with milk production (Lane et al., 1968). There is no change
in serum Mg concentrations at parturition (Bolenbaugh, 1977). When in-
takes of Mg are adequate (relative to requirements) serum Mg does not
seem to reflect intake variations (Belyea et al., 1975a). In addition,
a summary of some typical Mg values are presented in the paper by Belyea

et al., (1975a).

Volatile Fatty Acids (VFA)

 

- There are not many data available where serum VFA have been examined

in the periparturient cow. Schwalm and Schultz (1976) measured serum

l4
acetate (C2) from 20 days prior to calving to 48 days after. Prior to
calving, C2 remained fairly constant at about 1.3 mM, very little change
was seen at calving, but after calving, in normal cows, C2 increased to
l.67-2.0 mM. Cows that were sub-clinically ketotic however, had consi-
derably higher serum C2 concentrations of 2.5-3.33 mM and this differ-
ence persisted for at least 45 days. This two-fold difference between
normal and ketotic cows was also noted in data summarized by Annison and
Armstrong (1970), although in one experiment there was no difference
between cows.

It is not clear whether lactating cows have different blood concen-
trations of VFA than non-lactating cows. If there are differences they
are probably of dietary origin. Baird et al., (1975) reported that C2
was higher in dry cows than in lactating cows (3.3 vs 2.5 mM), that
serum propionate (C3) was the same and that butyrate (C4) in lactating
cows was higher (0.042 vs 0.022 mM). In contrast, Annison and Armstrong
reported data in which there was no difference in serum C2 levels.
Values of C2 concentrations for lactating cows fed normal diets range
from 1-2.5 mM (Huber et al., 1969; Yousef et al., 1969; Annison and
Armstrong, 1970; Bickerstaffe et al., 1974; Baird et al., 1975). Only
limited data is available for other serum VFA. Reported values f0r
arterial concentrations, in lactating cows, for C3 range from 0.04-0.13
mM (Bickerstaffe et al., 1974; Baird et al., 1975), for C4, 0.042 mM
was reported (Bickerstaffe et al., 1974).

In fasted goats or steers C2 concentrations decrease about 50%
(Annison and Armstrong, 1970). Serum VFA do seem to be sensitive to
dietary changes, presumably this is due to the induction of different

fermentation patterns with different feeds. It has shown that when

15

lactating cows were fed high grain, restricted roughage diets that plasma
CZ concentration decreased about 20% (Huber et al., 1969; Evans et al.,
1975). C4 levels did not change, both C2 and isobutyrate (isoC4) in-
creased (45 and 56%, respectively) on the low roughage diet, reflecting
the probable changes in fermentation (Evans et al., 1975). Neither the
addition of whey or minerals (NaHCO3 and MgO) had any further effect on
plasma C2 concentrations in cows fed high grain (Huber et al., 1969).
In contrast, Walker and Elliot (1973) found no differences between cows
fed high grain diets in either C2 or C3 concentrations, means were 2.5
and 0.115 mM for C2 and C3, respectively. The authors suggested that
perhaps their assay was not sensitive enough to show differences.

In conclusion, data are limited, but there do seem to be dietary
effects on serum VFA concentrations. Other environmental effects remain
to be described. Involvement of VFA in regulation of metabolism will

be discussed in the last section of the review.

Reproduction as Affectedppy Energy and Phosphorus Intake

 

Many theories have been p05tulated as to the cause of infertility,
anestrus, cessation of ovulation and other reproductive failures.
Nutrition has been one of the factors implicated in infertility. Rank
deficiencies of certain nutrients are known to impair reproductive func-
tion. However, the effects of excesses, marginal levels of nutrients or
nutritional imbalance have not been fully elucidated. Reproductive
efficiency is of prime importance to the producer. The characterization
and application of the relationship between nutrition and infertility
will greatly aid this efficiency.

In this part of the review the influence of energy and phosphorus

16
on postpartum estrus and fertility will be discussed. Possible mechanisms
for the action of these nutrients on reproductive function also will be

examined.

Energy and Reproduction

 

It has been suggested that liveweight change is a measure of plane
of nutrition and evidence has been reported that there is a positive re-
lationship between plane of nutrition and fertility (Broster, 1973).

The idea that low planes of nutrition can lead to anestrus and reduced
conception rates is widely accepted. The evidence, however, is not so
clear.

A ten percent decrease in liveweight from calving to the time of
mating has been associated with infertility (McClure, 1970). A one per-
cent change in conception to first service per one percent change in
liveweight has been noted by King (1968). In that study, the cows that
were gaining weight had a much higher conception rate at first service
than those losing weight. Similarly, Schilling and England (1968) re-
ported that with each kg of bodyweight lost from fall to spring there
was a 0.123% reduction in calving rate and with each kg of bodyweight
gained during the summer an increase of 0.203% in calving rate resulted.
Youdan and King (1977) concluded that longterm weight changes were
associated with fertility and that managing cows so that they were
gaining weight at the time of service resulted in improved fertility.

No evidence was found to suggest that short term bodyweight gain was bene-
ficial. Similarly, in Rhodesian beef cattle it was found that in cows
fed either to gain 12-14% or lose 12-14% of their peak liveweight from

early pregnancy to mid-breeding season (12-16 months), that estrus

17
occurred 9 days earlier in the high than the low plane cows but again,
short term fluctuations in nutrient supply had no apparent effect on the
incidence of estrus (Holness et al., 1978).

Using a condition scoring system where 0 represents 'emaciated' and
5, 'very fat' it has been shown in 15-20 suckler herds that optimum score
for mating was 2-2.S with extended calving intervals recorded for very
lean and very fat cows (Lowman and Scott, 1976). Heifers with good/
very good body condition score, fed at a high level of nutrition, showed
evidence of ovarian abnormalities. The data suggest greater irregulari-
ties in estrus cycle length or that behavioral signs of estrus were mani-
fested to a lesser extent (Leaver, 1977) than in those heifers in moder-
ate condition. Similarly, in dairy cows, conception rate at first ser-
vice was 52-60% when the condition score was 2 or less but 72-79% when
the score was greater than two (Mulvany, 1977). After many studies with
beef cows Wiltbank (1974, 1977) showed that when cows were fed low levels
of feed following calving there was an increase in the number of cows
that did not show heat and a decrease in the number that conceived to a
single service. Wiltbank concluded that to successfully rebreed cows
they should calve in moderate body condition and receive sufficient feed
following calving to maintain their weight.

Not all studies, however, have found a significant relationship be-
tween liveweight change and fertility. Although Boyd (1972) found evi-
dence for increased conception rates when cows were gaining weight,
fertility differences were not statistically different between weight
change groups and he concluded that weight loss had no adverse effect on
fertility. Similarly, Broster (1973) quotes work by Munro (1970, un-

published) in which no conclusive relationship between fertility and

18
liveweight change was found. Downie and Gelman (1976) found that ferti-
lity seemed more related to serum glucose concentrations than to whether
cows were gaining or losing weight. Finally, bodyweight changes were
not found to be related to time of first ovulation in dairy cows
(Stevenson, 1977). Liveweight change may indeed be related to fertility
but it is probably only one of many factors involved.

The question of the greater vulnerability of the cow with a higher
yield potential to the adverse effects of undernutrition compared with
the lower yielding cow is not fully answered. King (1968), reviewing
work in this area found no evidence to suggest that cows yielding more
milk showed reduced fertility compared with lower yielding cows.
Similarly, returns to service were equally distributed among cows of
differing yield potential in New Zealand herds (Simpson, 1972). Dawson
(1972) suggested that Simpson analyzed his data in a way that was biased
towards finding no differences. In the United States a positive correla-
tion between milk production and the interval from calving to conception
as well as services per conception, especially for animals producing more
than 7272 kg of milk per lactation, has been reported (Morrow et al.,
1966). Further reports, with some limitations, essentially support these
data. Hewett (1968) showed that repeat breeders averaged 86.4 kg more
milk during the first 120 days of lactation when compared to contemporary
herd controls. This study may have some limitations due to the fact that
two-thirds of the herds examined had less than 15 cows and in some cases
a control could not be found and thus some repeat breeders must have
been excluded from the analysis. In another more extensive study of
Swedish herds, Hewett (1974) failed to show that milk yield potential

affected fertility.

19

Also in a recent study, 393 calving intervals were used to compare
the effects of breeding at first postpartum estrus after 74 days as
modified by two different levels of nutrition and two different genetic
levels of production (Whitmore et al., 1974). The interval to first
postpartum estrus was longer in cows with superior genetic potential
for milk production than for those which were genetically inferior pro-
ducers. After an eleven year study of Czech dairy cattle it was con-
cluded that when yields were above 3000-3500 kg conception rates signi-
ficantly declined, the corresponding yield limit for older cows was 4000
kg (Brauner, 1975). Using Holstein dairy cows it was reported that in-
terval to first ovulation was extended due to increased milk yield
(Stevenson, 1977). Cows which ovulated after day 20 postpartum produced
significantly more milk than those which ovulated earlier, cows which
returned to estrus before day 15 produced significantly less milk.

Studies examining the effects of diets with below normal recommenda-
tions for energy have suggested that fertility was impaired. Dunn et al.,
(1969) studied these effects in two year old heifers from 140 days pre-
partum to 120 days postpartum. Two levels of digestible energy, low
and high, were fed prepartum. At parturition, the groups were further
divided to assess the effects of low, moderate and high energy levels.
It should be noted that these designations were for convenience only,
all were below the levels recommended by the National Research Council.
Pregnancy rate 120 days after calving was directly related to postpartum
energy level. Pregnancy rates at 120 days were 87, 72, and 64% for
cows fed high, moderate and low levels of energy postpartum, respectively.
Estrus was delayed in the heifers receiving the low level of energy pre-

partum. Detrimental effects of feeding low levels of energy just prior

20
to parturition were partially overcome by feeding higher levels post-
partum. Similarly, Brahman X Hereford heifers fed high energy pre and
postpartum returned to estrus before those fed below recommendations
(Randel and Welker, 1977). When heifers were fed high or low energy
rations for 90 days prepartum and adequate energy postpartum, a higher
percent of the high group were pregnant 60 days postbreeding compared
with the low group (Bellows et al., 1972). Other studies have shown
that low energy rations postpartum impair fertility (Macfarlane et al.,
1977) and that energy supplementation for 6 weeks prior to, and for 6
weeks after mating improves conception (Drew et al., 1976). Nevertheless,
not all work has established an effect of energy intake on fertility.
In a study of 64 dairy cows throughout a lactation with high and low
energy levels neither level of energy intake nor level of production
before or after calving or both, had a significant effect on fertility
(Gardner, 1969b).

Most evidence would suggest that adequate energy intake is more
important or critical than protein intake for maintaining reproductive
function (Wiltbank et al., 1962; Wiltbank et al., 1964; and McClure,
1968a). Protein intake, however, cannot be ignored or discounted. In
a recent study of dairy herds in Israel involving 2725 total lactations
it was shown that digestible protein intake, energy level and total dry
matter fed were all significant factors influencing good conception
rates. The nutrition of herds with good (345%) and low (32%) conception
rates was examined very closely and the authors concluded that proper
amounts of these constituents in the ration during the entire lactation
or inter-calving period was essential for providing appropriate fertility

levels in high producing dairy herds (Francos et al., 1977).

21

It was also found in Israeli dairy herds, that incidence of many
types of fertility disorders was much higher in herds fed a low plane of
nutrition when compared to herds fed according to recommendations.
Francos (1974) concluded that repeat breeder syndrome is associated with
feeding a ration deficient in energy during all stages of lactation and
the final stages of pregnancy.

The practice of overfeeding has also been implicated in fertility
problems. Excessive feeding of energy and protein to medium, low yield
and dry cows in the second half of pregnancy has been related to an in-
creased incidence of mastitis after normal parturition. It was suggested
that this practice predisposed the cow to faulty involution of the uterus
and to metritis (Francos, 1970). Once the diet was corrected, the inci-
dence of metritis declined. After an extensive survey of Swedish dairy
herds Hewett (1974) also suggested that the incidence of infertility due
to excess feeding was increasing.

Another aspect of feeding which has been associated with fertility
problems is that of reduced roughage in the diet. It has been reported
that there is a significant relationship between milk fat percent and
conception rate (Francos, 1968, 1969). This Israeli data suggested that
to maintain adequate fertility, the ration should be composed so as to
ensure 3.3% milk fat, even at high milk yields. The observed decrease
in fat percent was related to decreased acetate production in the rumen
due to shortage of roughage, feeding pelleted concentrates, or both.
Francos suggested that decreased acetate in the blood, as reflected by
low milk fat %, somehow resulted in lower fertility. It could be that
acetate insufficiency interfered with cholesterol and steroid synthesis,

resulting perhaps in estrogen and progesterone deficiencies. This appears

22
to be a long term dietary effect since Francos noted that it took 4-5
months before 10w roughage diets induced a decline in fertility. A rise
in conception rates were noted 1-2 months after fiber content of the
ration was raised. Francos (1969) recommended that dry matter from
roughage should make up at least 30% of the total dry matter in the
ration.

Assuming that reproductive function is also impaired with decreased
energy intake, several hypotheses have been suggested as to how this
effect is mediated. Some years ago it was suggested that restricted
energy intake caused hypoglycemia and that this in turn caused reproduc-
tive hypofunction. This observation was made after fertility was signi-
ficantly improved in an Australian herd when randomly selected cows were
fed an additional 5.6-6.4 kg of hay per day from calving to 3 weeks after
mating (McClure, 1965). The blood concentrations of glucose of the fertile
cows at the time of mating were increasing, and averaged 28 mg/dl, where-
as concentrations in the infertile cows were decreasing, and averaged 22
mg/dl. Body weight followed the same pattern. This hypothesis was
reiterated a few years later (McClure, 1968a) when low fertility syndrome
herds in Australia and New Zealand were characterized. Affected herds
lost 5-10% of their body weight between parturition and mating and had
low blood glucose concentrations of between 20 and 30 mg/dl. Supplementa-
tion of the rations of these herds with energy rich concentrations signi-
ficantly increased both blood glucose and fertility (McClure, 1972).

Further support was given by a study in which cows were made hypo-
glycemic with insulin. Cows mated 0-2 days after 3-4 days of daily
insulin treatment or those treated with insulin daily for the first 4

days after mating were significantly less fertile than cows treated with

23
insulin at other times, or control untreated cows (McClure, 1968b).
Similar results were observed in mice (McClure, 1967). McClure (1967)
suggested that the hypothalamus was failing to control the anterior
pitultary as a direct failure of supply and utilization of glucose.

It should be noted that the nature of the malnutrition described by
McClure (1968a) was apparently not a simple energy deficiency but was
complicated by deficiencies of other nutrients such as protein, phos-
phorus, and carotene. Also, the nutritional and hypoglycemic conditions
he describes are extreme when compared with a concentrate fed dairy cow
in North America. McClures' hypothesis, however, was supported by
Howland et al., (1966) who also suggested that hypoglycemic effects were
mediated through the hypothalamus and resulted in a loss of ovarian
activity. Additionally, a significant negative correlation was found
between plasma glucose concentrations and postpartum interval to occur-
ence of a 10 mm follicle and ovulation in a study of primiparous cows
(Oxenreider and Wagner, 1971). Both energy and lactation had a signifi-
cant effect on plasma glucose levels during the first 8 weeks postpartum.
Lactation and low energy significantly delayed postpartum follicular
growth and ovulation. More recently, further support for this idea has
been published. Downie and Gelman (1976) reported that increasing glu-
cose concentrations during the 20 days preceding estrus was characteris-
tic of fertile services in suckling beef cows regardless of body weight
change. Parker and Blowey (1976) also reported that plasma glucose con-
centrations within + or - 3 days of first service of cows which conceived
was higher than that of cows which did not conceive. This difference
only approached significance at the 5% level but supports the above data.

Additionally, in a large study of ovarian function and carbohydrate

24
status in dairy cows in Norway it was found that in periods before 24
days or after 40 days postpartum the cows without corpora lutea had sig-
nificantly lower plasma glucose concentrations than cows with a corpus
luteum (Benjaminsen, 1977).

On the negative side of this question neither Blowey et a1. (1973)
or Hewett (1974) were able to show any relationship between glucose con-
centrations and fertility in extensive surveys of herds in England and
Sweden. The reasons for these discrepancies are not clear at this time.
Throughout these studies, however, it is difficult to say that energy
status or intake, lactation, and glucose are independent of each other.
This leads to a problem of interpreting which is the main cause and which
is effect.

Numerous endocrinologists have studied the effects of energy intake
on postpartum ovarian activity and changes in serum hormone concentrations
pre and postpartum have been studied. Unfortunately, metabolite concen-
trations were not concurrently measured with alterations in reproductive
function. Normal ovarian and hormonal changes have been reported by
numerous workers such as Echternkamp and Hansel (1973).

The effects of different energy intakes prepartum on postpartum
progesterone and estradiol concentrations in beef heifers have been
examined (Corah et al., 1974). Prepartum energy intake had no signifi-
cant effect on either progesterone or estradiol, either prior to, or
following, parturition. Reducing energy intake for 100 days prior to
calving markedly reduced body weight and fat cover but the interval to
first estrus was not influenced by the reduced energy intake. Of
particular interest in this study was the marked elevation of proges—

terone in those cows which conceived, suggesting that a period of

25

elevated progesterone may be necessary for conception at first estrus.
This effect has also been noted in diary cows maintained on high and low
levels of nutrition (Folman et al., 1973). Cows that conceived after
one insemination had significantly higher progesterone concentrations
during the estrous cycle preceding insemination than did cows that did
not conceive. The concentration of progesterone required appeared to be
around 3 ng/ml in this case. Also, during the luteal phase preceding in-
semination, cows that conceived after first insemination gained weight
whereas cows that did not were losing weight, the difference approached
significance. In a more recent study it was found that the timing of
this peak may also be important. In fertile cows, peak progesterone
occurred within 4-7 days prior to estrus rather than 8-11 days prior to
estrus. This was found to be significant, however, only in the cows
cycling in the summer and not in the winter (Rosenberg et al., 1977).

Other studies have shown that level of nutrition can affect peri-
pheral reproductive hormone levels. Plasma luteinizing hormone (LH) and
progesterone concentrations were examined for 3 estrous cycles in groups
of Holstein heifers fed 100 or 62% of energy requirements (Gombe and
Hansel, 1973). Plasma LH increased progressively from the first to the
third estrous cycle in heifers fed the low energy ration. This increase
was first seen in the maximum LH of the cycle but by the third cycle LH
was also higher throughout the cycle. During the first cycle progesterone
was slightly higher in the cows fed 62% compared to 100% of their energy
requirements but became progressively lower in the subsequent cycles.
Total progesterone and progesterone concentration in the corpora lutea
of heifers fed 62% of their energy requirements was lower than in their

normally fed counterparts.

26

Evidently, ovarian hypofunction in cases of energy deficiency is not
due to reduced circulating levels of LH as was previously thought by such
workers as Wiltbank et al., (1962). The first effect may be a reduced
ability of the ovarian tissue to respond to LH. Decreasing progesterone
has been noted before in heifers receiving 25% of the total feed consumed
by the controls (Donaldson et al., 1970). Declines in plasma progesterone
have occurred within even 5 days of reduction in feed intake (Hill et al.,
1970). This undernutrition, which was about 85% of maintenance, also
temporarily reduced the number of medium sized follicles, altered the
length of the estrous cycle and reduced the proportion of heifers with
normal fertilized ova. Corpora lutea weights in undernourished heifers
were about 70% of control values. It should be noted that Donaldson et
al. (1970) and Hill et a1. (1970) varied both energy and protein content
of the ration whereas Gombe and Hansel (1973) reduced only energy intake.

Evidence suggests that the observed reduction in plasma progesterone
is a reflection of a smaller corpus luteum, containing less total and a
lesser concentration of progesterone (Gombe and Hansel, 1973). The
simplest suggestion is that restricted energy intake affects some step
in steroidogenesis within the corpus luteum, resulting ultimately in re-
duced plasma progesterone concentrations. Several additional suggestions
have been made as to how this effect is mediated (Gombe and Hansel, 1973).

Recently, using multiple regression analysis, factors affecting post-
partum return to estrus were examined in dairy cows. The interaction be-
tween factors involved in calculated energy status and serum glucocorti-
coids accounted for more than 35% of the variation in days to first ovula-
tion (Stevenson, 1977). Estradiol and the interaction between LH and estra-
diol also significantly contributed to the model. LH was negatively

correlated to days to first ovulation. In a study with

27
Hereford Heifers fed either 115% or 60% of N.R.C. requirements pre and
postpartum it was shown that level of energy did not affect estrogen
secretion but, the pattern of LH release after GNRH was altered by
feeding. The peak of LH after GNRH injection was 30 min later in the
cows fed low energy (Lishman et al., 1977). Staigmiller et al., (1977)
concluded that level of nutrition could influence follicular development
in response to exogenous FSH but does not significantly affect or alter
ovulation rate.

Accumulating evidence suggests there is an interrelationship between
energy intake and reproductive function in the cow. The characteristics
of this relationship are still not clearly defined and the mechanism of
action is not fully elucidated. The evidence does suggest that re-
stricted energy, in certain circumstances, can cause reproductive fail-
ures. This relationship is not simple and must be considered concurrently
with body weight, stage of lactation, specific type of diet and other

physiological factors.

Phosphorus and Reproductive FunCtion

 

A relationship between phosphorus and fertility was postulated many
years ago and appears to be widely accepted. Evidence suggests that the
phosphorus deficiency required to impair fertility is extremely severe.
It has been more difficult to show effects of marginal deficiency on
reproductive function. However, a sub-clinical or marginal deficiency
may be of more importance since it may indeed cause production losses
but not be overly detectable. Clinical signs of severe deficiencies may
not be easily diagnosed but are easily noticeable.

The widespread occurrence of a natural deficiency of phosphorus

28
affecting cattle has been described by Tuff (1923), Theiler et a1. (1924),
Eckles et a1. (1926), Hart and Guilbert (1928) and numerous others. As
early as 1906, in a study of the effect of phosphorus compounds in the
diet of milking cows it was noted that phosphorus deficiency was accom-
panied at times by the cessation of estrus (Jordan et al., 1966).

South African workers were among the first to report reproductive
hypofunction in cattle maintained on veld deficient in phosphorus. By
supplementing the naturally deficient animals with bonemeal, calf crops
were increased from 51 to 80% (Theiler et al., 1928). Bonemeal supple-
mentation also increased blood phosphorus concentrations from 2.2 mg/dl
to normal levels in cows grazing phosphorus deficient pasture (Malan et
al., 1928). Hyp0phosphatemia was reported in animals fed a hay-oats
phosphorus deficient diet but effects on reproduction were not mentioned
(Palmer and Eckles, 1927). Years later in Ireland both clinical and
sub-clinical aphosphorosis of cattle grazing phosphorus deficient pasture
was noted. The cattle developed all the symptoms of phosphorus deficiency
plus 'temporary sterility' (Sheechy, 1946) or 'anestrus or estrus with
repeated failures to conceive after service' (O'Moore, 1950). Hypo-
phosphatemia was noted in all these cases and phosphorus supplementation
seemed to solve the problem.

In the United States, Eckles et a1. (1935) followed the example of
the South African workers such as du Toit et a1. (1934), and tried to
obtain experimental data regarding reproductive function in cattle on
controlled uncomplicated phosphorus deficient rations. They recognized
that much of the reported field data was confounded, knowingly or not,
by definiencies of other nutrients. Eckles et al. (1935) fed phosphorus

so as to maintain plasma phosphorus at approximately 2.5 mg/dl, about

29

one half normal. This concentration was selected as being representative
of cattle in a severely deficient area. The daily milk yields of these
cows were low by today's standards, between 3.2 and 11.1 kg, and probably
did not stress the animal excessively. After 3 years of experimentation
no evidence was obtained to show that phosphorus deficiency influenced
the estrous cycles of the cows. It did appear, however, that breeding
efficiency was reduced. The authors concluded that the disturbances in
estrus and reproductive function seen in naturally deficient areas were
probably due to the complication of other nutrient deficiencies.

More recently, heifer infertility problems in a herd were attributed
to phosphorus deficiency (resulting from depleted phosphorus in the soil
and consequently in the crops), (Morrow, 1969). Calculations of intake
and requirements showed that phOSphorus intake was deficient. Clinical
signs consisted of rough coat, depraved appetite and infertility. Low
blood phosphorus was observed. Treatment of the condition with dicalcium
phosphate supplements returned blood levels to normal and fertility was
restored. The fertility problem was not due to length of estrous cycles
or frequency of silent estrus but rather of conception failure.

In order to more accurately simulate a deficiency in the field the
effects of a combined phosphorus and protein deficiency have been
examined. Cows were observed for at least 24 months and up to 59 months
on a protein and phosphorus deficient ration (Palmer et al., 1941). Cows
showed delayed sexual maturity and silent heat but normal, regular
ovulation and conception was not impaired. Breeding efficiency was not,
reduced. In the Northern territory of Australia, supplementation of
range cows with 8 g P/head/day improved body weight and increased the

pregnancy rate of lactating cows 20% over their controls (Hart and

30
Mitchell, 1965). The authors did comment, however, that the provision
of protein in their opinion was of equal, if not more importance, than
phosphorus.

In support of the latter comment, although bonemeal supplementation
to native Rhodesian cattle grazing on veld pasture throughout the year,
increased the weight of calves weaned by 21%, it did not appear to im-
prove conception rates or have much effect on the duration of postpartum
anestrus (Ward, 1968). Supplementation with groundnut cake in the winter,
on the other hand, doubled conception rates over that of control cows
indicating that protein was more limiting than phosphorus. Similarly,
supplementation of Australian cows fed native hay with phosphorus had
no significant effect on feed intake, liveweight change, milk production
or reproductive activity. In cows which received adequate protein,
however, estrus activity was markedly increased whether phosphorus was
supplemented or not (Teleni et al., 1977).

Long term supplementation to dairy heifers of phosphorus above
N.R.C. minimum daily requirements has also failed to show any reproduc-
tive advantages. N0 effects were noted in estrus exhibition, services
per conception and pregnancies (Hecht et al., 1977) or on weight gains,
feed conversion efficiency, animal health or conception rate (Noller et
al., 1977). In contrast, studies with beef cows nursing calves under
drylot conditions have shown that increasing P intake to 150% of N.R.C.
requirements from 30 days postpartum to weaning improved rebreeding
efficiency and apparently improved milk production. Eighty-nine percent
of cows on high phosphorus treatment conceived to first insemination as
compared to 59% of those fed the 100% N.R.C. diet (Taylor et al., 1976).

Another approach to the Australian problem of low fertility rates

31
in cattle grazing native pasture has been to supplement the pasture,
rather than the cows, with superphosphate fertilizer. Application of
a high level (377 kg/ha/yr) of this fertilizer significantly increased
total pasture dry matter available and improved pregnancy rates and
growth patterns of breeding cattle. Statistical analyses strongly sug-
gest that improved conception was a direct effect of superphosphate and
not attributable to liveweight differences (Williams et al., 1971;
Siebert, 1973). At lower rates of fertilization, phosphorus content
of the pasture was less than 0.15% for much of the year, apparently only
at higher fertilization rates did the pasture approach an adequate phos-
phorus content to support pregnant and lactating cows (Ritson et al.,
1971). It is not clear at this time whether further supplementation or
protein additions would have improved fertility further.

Except in cases of severe deficiency it has been difficult to de-
monstrate a clear correlation between serum phosphorus levels and ferti-
lity in dairy cows. Attempts have been made to do this to aid in manage-
ment of commercial dairy herds._ Rowland et a1. (1977) found no difference
between these parameters in dairy cows between 40 and 100 days postpartum.
Similarly, when serum phosphorus concentrations were examined in cows
from 15 dairy herds there were no differences between those cows which
conceived and those that did not (Parker and Blowey, 1976). It was
suggested that when serum phosphorus concentrations were within the
normal range it was unrealistic to expect to be able to detect differ-
ences.

Some years ago it was suggested that breeding efficiency of cattle
might be related to the Caz? ratio of the feed consumed, (assuming that

the intake of each element alone was sufficient and adequate) (Hignett,

32
1950). It was suggested that a 1:1 ratio was ideal for fertility, where-
as herd fertility could be impaired when the ratio was 2:1 or more,
especially when the phosphorus intake was only slightly in excess of
minimum requirements. After work was completed on another 802 cows,
it was concluded that the generally accepted recommendations for phos-
phorus in England at that time were not high enough for high fertility
in dairy cows (Hignett and Hignett, 1951). Decreased fertility was ob-
served in rapidly growing heifers fed rations deficient in manganese and
unbalanced in calcium and phosphorus (Hignett, 1959). It should be noted
that even Hignett and Hignett (1951) regarded their results with caution.
Most of their work was of the field survey type and only feed analyses
and breeding data were used. No blood parameters were measured. Their
caution appeared to have been justified since a large scale controlled
experiment at Weybridge in 1958 and 1959 failed to demonstrate any
significant relationship between the Ca:P ratio of the diet and fertility
in dairy heifers (Littlejohn and Lewis, 1960). A high Ca:P ratio did,
however, depress growth rate. In addition, in a study of 15 commercial
dairy herds, where Ca:P ratios ranged from l.19-3.55:l, no effect on
fertility was noted (Parker and Blowey, 1976). Hignett (1950) originally,
had also warned against the feeding of too much phosphorus, associating
this with infertility. More recently this speculation was raised again
when an extensive survey of Swedish dairy cattle showed that low herd
fertility was associated with greater than normal serum concentrations
of phosphorus, potassium, total protein and urea nitrogen (Hewett, 1974).
Hewett interprets his results to indicate that excessive intake of cer-
tain nutrients occurs as a result of trying to provide the high yielding

cow with sufficient energy, feeding proportionately more concentrates,

33

so that the ration is more likely to be in excess than that of low yield-
ing cows. He suggests that sterility due to heavy feeding is steadily
increasing and that 'starvation sterility' has little relevance today.
In another study of dairy herds in British Columbia, however, it was
found that the Ca and P content of the silage was important for fertility.
As the intake of Ca, P and Cu/Mo was increased from silage as opposed to
other sources, non-return rates decreased (Peterson and Waldern, 1977).

If phosphorus affects reproductive function, and evidence suggests
that it may, the mechanism is not clear. It has been suggested that
phosphorus deficiency acts at the anterior pituitary level causing
'cessation of estrus, lack of sexual libido, testicular and accessory
organ atrophy' (Guilbert, 1942). Also the 'ovary in phosphorus defici-
ency becomes quiescent and infantile' (Guilbert and Hart, 1930; Kleiber
et al., 1936). None of these reports are enlightening as to how de-
ficiency causes these effects. Kleiber et a1. (1936) reported that
phosphorus deficiency lowers the total efficiency of energy utilization
partly by depressing appetite and partly by lowering the utilization of
the food eaten. Fasting catabolism did not appear to be altered. New
evidence suggests that phosphorus intake could have its action directly
in regulating cellular metabolism. It is postulated that cellular
respiratory rate and NADH generation rate are regulated by the [ATP] /
[ADP] - [PI] ratio. This means that cellular processes could be regu-
lated by intracellular phosphorus rather than energy charge (Erecinska
et al., 1977). It is possible then that phosphorus may directly affect

metabolism of the reproductive tissues.

34

The Interrelationship Between Hormones and Metabolites

 

For many years attempts have been made to correlate peripheral con-
centrations of hormones and metabolites in ruminants. The idea that
substrates and hormones might interact to regulate metabolism to the
advantage of the animal is logical. The fact that the nutritional
physiology of the ruminant is peculiar to that species suggested that
the emphasis of regulation of metabolism should be different from that
of the non-ruminant animal.

The most important features of ruminant metabolism and digestion
which need to be considered in this context are: (l) the production
from dietary carbohydrates by microbial fermentation in the rumen of
large quantities of short chain fatty acids (principally C2 (70%), C3
(20%), and C4 (10%)), and absorption of these acids from the rumen into
portal blood either direCtly or after conversion of some C3 to lactate
and C4 to BOHB in the rumen wall; (2) the virtual absence of hexose from
the lower parts of the tract except on certain high grain starch con-
taining diets; and (3) the degradation of much dietary protein and the
synthesis of microbial protein in the rumen (Bassett, 1975). As a con-
sequence of this, ruminants are almost entirely dependent on gluconeo-
genesis for provision of glucose in the fed and fasted state and utilize
acetate (C2) instead of glucose as a major substrate for energy storage
and oxidation in the fed state.

As a consequence, the rate of gluconeogenesis is highest and the
availability of gluconeogenic precursors (C3, amino acids) are greatest
in the immediate postprandial period. Bassett (1975) suggested that the
hormonal responses to nutrient intake in ruminants, therefore, should

favour Co oxidation or incorporation into fat and the maintenance of

35
high rates of glucose synthesis to preserve metabolic homeostasis.
Bassett suggests that the responses seen may differ little from other
species consuming a low carbohydrate or predominantly protein diet.
However, ruminants differ from other species eating low carbohydrate
diets, in that C3 is a major precursor of glucose and may account for
about 50% of the glucose produced.

Although the processes of digestion and absorption in adult rumi-
nants are prolonged when compared with those in monogastrics, some
diurnal variations of hormone and metabolite patterns have been observed
that are associated with feed ingestion. The examination of postprandial
changes has been useful. The alternative approach has been to observe
changes after administration of hormones or metabolites to the animal.

In both sheep and cows, plasma insulin concentrations increase
significantly after feeding (Trenkle, 1971a; McAtee and Trenkle, 1971;
Ross and Kitts, 1973; Bassett, 1974a, 1974b, 1975; Jenny and Polan, 1975;
Evans et al., 1975). The increase which occurs postprandially may peak
anywhere from 2-6 h postfeeding (McAtee and Trenkle, 1971a; Bassett,
1974a; Jenny and Polan, 1975; Evans et al., 1975). The act of ingestion
itself causes vagal reflexes which cause insulin release. This can be
seen in suckling lambs at 5 min after the initiation of suckling, there
is a second peak at 45-90 min after feeding which is not a neural effect
but due to absorbed glucose (Bassett, 1974b). In adult sheep, an extra
feeding, given 12 h after the first or usual feeding, resulted in a repe-
tition of the changes in plasma insulin concentrations seen after the
normal feed (Bassett, 1974a). With sheep fed once daily on 3 different
quantities of a mixed hay/grain diet Bassett (1974a) showed that the time

that insulin was high increased as the amount of food increased. Bassett

36
(1975) concluded that the amount of food ingested by sheep clearly in-
fluenced the magnitude of the insulin response. It has been reported
that insulin levels increase less after feeding hay than concentrates
(Bassett, 1975). Also, lactating cows fed a high grain ration had signi-
ficantly higher insulin concentrations at all hours postfeeding when com-
pared to control cows (Jenny and Polan, 1975). Bassett suggests that
these differences are probably due to slower digestion of forage type
diets and a smaller amount of digestible organic matter. The flow of
protein digesta into the abomasum and small intestine may alsolxaimpor-
tant in determining insulin levels in sheep (Bassett et al., 1971;
Bassett, 1975). This was suggested after it was found that insulin
levels were higher in sheep fed a diet of protected protein.

As would be expected from the above data, fasting decreases insulin
concentrations (Trenkle, 1970a; McAtee and Trenkle, 1971a; Trenkle, 1976).
It seems that dietary effects are greater than those of age or sex. No
differences in plasma insulin could be related to age (18-371 days old)
or breed (Angus, Hereford, Shorthorn and crossbred cattle) in a study
conducted by Irvin and Trenkle (1971). Steers had higher insulin concen-
trations than heifers at 371 days of age, but, the steers were fed a
finishing ration, the heifers a lower energy ration and thus, the dif-
ference was judged to be dietary. Certainly, in another study yearling
steers fed finishing rations increased insulin concentrations during the
feeding period (Trenkle, 1970b).

There is also evidence that serum GH concentration in both young
and adult ruminants are influenced by feeding and dietary factors. The
most striking features of plasma GH in adult ruminants is their great

variability, suggestive or episodic secretion of the hormone (Bassett,

37

1975). Fasting does not cause any consistent increase in plasma GH
(Bassett, 1975; Trenkle, 1971, 1976), however, it appears that fasting
increased the halflife of injected GH from 9.2 to 13.4 min in sheep and
from 20.3 to 31.9 min in calves (Trenkle, 1976).

Oscillations in CH concentrations seem greatest in animals not fed
and it appears that one affect of feeding may be to decrease the size
and frequency of these secretory'bursts(Bassett, 1975). It was recently
suggested that this suppression may in part be due to somatostatin re-
lease from the intestine and/or the pancreas (Bassett, 1975; Trenkle,
1978). In sheep fed maintenance diets once daily, feeding causes a
marked decrease in plasma GH and suppresses the oscillations for 2-4 h
(Bassett, 1974a, 1975). After this time concentrations return, with
marked oscillations, towards prefeeding values. In sheep fed larger
quantities of food GH levels tend to be generally lower and an effect
of feeding is less clearly seen (Bassett, 1974a). In sheep fed every
3 h mean plasma GH concentrations were negatively correlated with digesti-
ble organic matter intake and with plasma insulin concentrations (Bassett
et al., 1971). The response of plasma GH levels in lambs to milk feeding
differs substantially from the changes following feeding in adult sheep
(Bassett, 1974b). Suckling initially causes a rapid decrease in plasma
GH but this period is quickly followed by increases in plasma GH levels
to high values 30-90 min after milk ingestion. The functional signifi-
cance of these differences is not clear at this time, but is possibly
partly due to rumen bypass and difference in composition of the diet.
Since GH appears to be episodically secreted, unless samples are taken
fairly frequently, interpretation becomes difficult.

The logical question is, by what mechanisms are hormone concentration

38
influenced by diet and how might this control metabolism? It cannot be
assumed, for example, that insulin and glucagon are regulated by the
same mechanisms as in a non-ruminant fed a low carbohydrate diet because
of the special features of ruminant digestion.

In view of the extensive metabolism of dietary hexose to VFA in the
rumen, attention has naturally been directed towards the VFA as possible
regulators of insulin (and glucagon) release in ruminants. It is now
well established that intravascularly administered acids C3-C8, but not
C2, stimulate insulin secretion in sheep and cattle (Manns and Boda,

1967; Manns et al., 1967; Horino et al., 1968; Hertelendy et al., 1969;
McAtee and Trenkle, 1971a; Trenkle, 1971a). Usually, C4 is more insulino-
genic than C3. In some experiments where glucose was tested at the same
time, glucose was much less effective than C3 and C4 in stimulating in-
sulin secretion (Manns and Boda, 1967; Trenkle, 1971a; McAtee and Trenkle,
1971a). In addition to C2, neither acetocetate nor B-hydrobutyrate (BOHB)
is insulinogenic, either in vivo (Manns and Boda, 1967; Horino et al.,
1968) or in vitro (Jordan and Phillips, 1978). While C3 has no effect

on insulin secretion in rats, rabbits or pigs (Horino et al., 1968) it

is interesting to note that VFA do stimulate insulin secretion in newborn
lambs (Hertelendy et al., 1969), suggesting that the capacity of the
ruminant to respond to short chain fatty acids is a constitutional rather
than an acquired characteristic. Due to these reports it is tempting to
conclude that the VFA were regulators of insulin secretion in response

to food ingestion in ruminants.

Despite these studies there is still considerably controversy about
the physiological importance of VFA in insulin release. Bassett (1975)

summarizes these problems: (1) C3 and C are largely removed from the

4

39
blood by a single passage, yet C2, the only VFA occurring in peripheral
plasma in substantial quantities, has no significant effect on insulin
secretion and probably none on glucagon secretion either since it does
not cause hyperglycemia; (2) the hyperglycemic and insulinogenic actions
of VFA have been observed following injections and infusions of probably
unphysiologically high concentrations, hence, perhaps reflecting a
pharmacological effect; and (3) when VFA were administered via the gastro-
intestinal tract in physiological quantities they were ineffective
(Stern et al., 1970). On the other hand, as Bassett (1975) points out,
in their experiments, increases in insulin and glucagon in sheep fed
once daily occur at the same time as increases in plasma VFA levels.
Ross and Kitts (1973) found that plasma C4 and isoC4 were most related to
postprandial insulin concentrations in sheep and Trenkle (1978) reports
work in which intraruminal infusion of physiological concentrations of
C2, C3 and C4 in sheep fasted for 36 h increased insulin concentrations.
It is suggested that Stern et a1. (1970) failed to find a response be-
cause the animals in that study had been allowed free access to feed
during the experiments and that the higher concentrations of insulin
in ad libitum fed goats may have masked any additional response to
VFA (Trenkle, 1978).

Certainly, VFA do increase in the circulation after feeding, despite
the high extraction of C3 and C4 by liver and metabolism of C4 to BOHB
in the rumen wall (Ross and Kitts, 1973; Bassett, 1975; Evans et al.,
1975). The timing of these increases relative to changes in insulin
is variable. At any rate it appears that maximum concentrations occur-
ring do not approach 200 uM and may be lO-fold lower (Bassett, 1975).

Bassett (1975) suggests that it would be surprising if such low

40
concentrations were to have any marked influence on insulin secretion
although this question remains to be definitively answered. Certainly,
it is not likely that VFA are the only way in which insulin and glucagon
secretion are regulated in ruminants.

Due to the absence of hexose from the digesta in ruminants, changes
in plasma glucose have not been generally considered important in regu-
lating insulin release. Bassett (1975) suggests that plasma glucose con-
centrations are the end result of hormonal regulation. Generally,
evidence is weak that increases in insulin after feeding are the result
of increases in glucose. Correlations between postprandial glucose and
insulin concentrations are usually low (Bassett et al., 1971; McAtee
and Trenkle, 1971a, Ross and Kitts, 1973; Evans et al., 1975; Bassett,
1975; Trenkle, 1978). Insulin concentrations may be more related to
glucose entry rates rather than to concentrations per se (Bassett et al.,
1971).

The next obvious candidate for a regulator of insulin secretion
would be protein or amino acids. In fact, it has been shown that in-
travascular administration of leucine, phenylalanine, arginine or hydro-
lysed casein will significantly increase insulin concentrations in rumi-
nants (Hertelendy et al., 1969; McAtee and Trenkle, 1971a; Davis, 1972;
Trenkle, 1978). This increase was apparently 'much more than could be
accounted for by the glucogenic effects of amino acids' (McAtee and
Trenkle, 1971a). The amounts of amino acids infused, however, are
generally considered to be unphysiological and in fact, after once daily
feeding, plasma amino nitrogen levels decrease rather than increase and
so do not suggest a significant role for regulating plasma insulin

(Bassett, 1975; Trenkle, 1978). One study has shown that plasma

41
of tyrosine, valine, isoleucine, and phenylalanine were correlated with
plasma insulin concentrations in sheep, the highest correlation, however,
was with the amount of protein digested in the intestine (Bassett et al.,
1971).

The effects of protein on insulin and glucagon release may be
mediated by the secretion of gastrointestinal hormones (Bassett, 1975).
In non-ruminants gastrin, secretin and pancreozymin can stimulate insulin
secretion and gastrin and pancreozymin can also stimulate glucagon re-
lease (Bassett, 1975). These hormones also seem able to potentiate the
insulin secretory responses to glucose and amino acids although there is,
as yet, no evidence that any of them are effective at physiological con-
centrations. The GI hormones secretin, and cholecystokinin-pancreozymin
stimulate insulin secretion in ruminants (Baile et al., 1969; Trenkle,
1972), and it has been pr0posed that these hormones may be involved in
the initial increase in plasma insulin after feeding (Bassett, 1975;
Trenkle, 1978). Perhaps gut glucagon as well as other GI hormones are
involved in this mechanism too, although the function and regulation of
these hormones in ruminants is poorly defined. In view of the long time-
course of digestion in ruminants and the apparently well utilized by-pass
protein and fat in some ruminant diets however, one might speculate that
these hormones may well be involved in the potentiation of responses to
other stimuli of insulin and glucagon secretion.

Several mechanisms seem to be involved in regulation of serum insu-
lin concentrations in re5ponse to ingestion and digestion of feed. The
first response is probably a release of GI hormones resulting from pas-
sage of feed into the intestine which causes the first release of insulin.

The second increase in plasma insulin is probably caused by a combination

42
of shortchain fatty acids from the rumen as well as absorption of gluco-
genic precursors from the digestive tract, (Trenkle, 1978).

Relationships between GH and metabolites at physiological concen-
trations are not well established. A number of studies have shown that
GH is released in a number of situations. Decreasing plasma glucose
(from induced hyperglycemic levels) seem to stimulate GH release, while
hypoglycemia per se does not (Hertelendy et al., 1969; Trenkle, 1971b;
McAtee and Trenkle, 1971b). Short chain fatty acids (C2, C3 and C4) do
not seem to affect GH levels (McAtee and Trenkle, 1971b). Although one
study reported that very high doses of C4 released GH (Reynaert et al.,
1975). Glucose and VFA do not seem to be important in the regulation
of GH concentrations in normal animals. Amino acids have been shown to
stimulate GH secretion many times. Arginine and also leucine signifi-
cantly increase plasma GH concentrations when administered intra-vascular-
1y into cows, heifers, steers, lambs and sheep (Hertelendy et al., 1969;
McAtee, 1970; Hertelendy et al., 1970; McAtee and Trenkle, 1971b, Davis,
1972).

There is some evidence that fasting alters the sensitivity of the
pituitary to factors which cause secretion of GH since there are greater
responses to arginine in fasted than fed heifers (McAtee and Trenkle,
1971b).

Trenkle (1978) also reports that GH can be released by injections
of unphysiological amounts of glucose, insulin and glucagon. These in-
creases are not due apparently to altering blood glucose concentrations
since increases in GH occurred both duringzidecrease (during administra-
tion of insulin and C4) and an increase (during administration of glucose

and amino acids) of blood glucose. Furthermore, inhibition of glucose

43
utilization with 2-deoxy-g1ucose did not change plasma GH (McAtee and
Trenkle, 1971b). Even though intra-vascular infusion of amino acids in-
creases plasma GH, plasma concentrations of amino acids do not seem to
play a significant role in secretion of GH, because concentrations of
GH in sheep plasma are correlated negatively with free amino acids in
plasma and the amount of protein digested in the intestine after feeding
(Bassett et al., 1971). A decrease in free fatty acids in plasma, which
is common with the injection of hormones and metabolites that result in
increases in CH, has been suggested to induce the rise in plasma GH
(Reynaert et al., 1975).

In general, it is suggested that GH tends to increase when avail-
ability of nutrients becomes limiting. Especially when there are in-
creased metabolic demands during pregnancy and lactation. The increase
in GH would mobilize energy from adipose tissue to satisfy the needs for
metabolism (Bassett, 1975; Trenkle, 1978). This hypothesis is supported
by the observation that GH concentrations were lower in ketotic than
non-ketotic cows (Reynaert et al., 1977).

GH and insulin are involved in homeostasis of nutrient metabolism.
The pattern of postprandial secretion is such that higher insulin with
adequate GH favors uptake of glucose and amino acids by non-hepatic
tissues for tissue synthesis or storage. Increasing glucagon favors
hepatic removal of glucogenic precursors and favors synthesis of glucose.
These hormonal changes after feeding favor efficient use of dietary nutri-
ents, which usually are consumed periodically rather than continuously
(Trenkle, 1978).

In times of greater metabolic demand GH and GCD usually increase,

insulin and glucagon decrease, insulin relatively more than glucagon.

44
Higher GH alone with low insulin favors transfer of calories from
adipose tissue to other tissues. Decreased uptake of amino acids by
non-hepatic tissues combined with GCD action on muscle to result in loss
of amino acids eventually results in increased gluconeogensis. This is
facilitated by higher GCD and a higher glucagon: insulin ratio (Trenkle,
1978).

Although data are limited at this time, PRL and GCD are perhaps
also involved in metabolism and utilization of nutrients. PRL appears
to follow the pattern of insulin, increasing at feeding, decreasing with
fasting and increased metabolic demands (Trenkle, 1978). GCD although
not apparently changing with feeding may increase with fasting and
greater metabolic demand (Trenkle, 1978). The role of these hormones

in the integration of metabolism remains to be clearly defined.

MATERIALS AND METHODS

Experimental Desigp_

 

Two levels of energy and phosphorus (>100 and 75% of N.R.C., 1971)
requirements were fed to 24 primiparous Holstein heifers from the
Michigan State University herd using a 2 x 2 factorial design. This
trial was then replicated so that in total 48 heifers completed the ex-
periment. At parturition these heifers weighed between 437 and 625 kg
in the first trial and between 405 and 633 kg in the second trial. At
parturition heifers were assigned to one of four treatment groups, in
groups of six to: (1) high energy, high phosphorus; (2) high energy,
low phosphorus; (3) low energy, high phosphorus; (4) low energy, 10w
phosphorus. Heifers were assigned so as to represent equivalent genetic
potential within each treatment group. Treatment began on the day of
parturition and extended to 84 days postpartum. At that time, heifers
were returned to the herd ration and some measurements were taken for a
further 21 days (the experiment ending at 105 days of lactation).

In the first trial the high energy rations were corn silage treated
with NH4OH fed until feed refusals were about 10% of intake and grain
mixture fed to the maximum which the heifers would consume. Grain was
fed daily at 0800 and 1300 h. Grain intake was restricted to 2.3 kg
at 0800 h for the low energy rations. .All cows were fed 2.3 kg of alfal-
fa-brome hay once daily. The corn silage was fed once daily in the
morning. Phosphorus was restricted by replacing the dicalcium phosphate

45

46

in the grain mixture with corn and CaCO Composition of the concentrate

3.
mixtures may be seen in Appendix Table 1.
In the second trial the dietary regime was essentially the same,
the only important difference was the time of feeding. In this case, the

grain was fed daily at 1800 and 0800 h. For the low energy rations,
grain intake was again restricted to 2.3 kg at 1800 h. Corn silage was
fed once in the evening and hay was fed once during the day. It is also
important to note that in the second trial a mistake was made in mixing
the high energy, low phosphorus grain at the feed mill. This grain was
not low in phosphorus and was fed for approximately 3 or 4 weeks before
it was noticed and corrected. Consequently, some of the heifers in this
group were not in negative phosphorus balance for some time. The levels
of intake will be clarified further in the results.

For both trials daily records of disease, feed intake and milk pro-
duction were kept. Composite morning and evening milk samples were taken
every two weeks and analyzed for fat and protein. Milk fat in both trials
was analyzed by standard Dairy Herd Improvement Association (DHIA) proce-
dures by DHIA personnel (E. Lansing, MI). In the first trial, protein
content was measured by routine microneldahl analysis. In the second
trial, the first few samples were analyzed by the microneldahl method
and the rest were measured using a dye method, (Udy Analyser Co., Boulder,
CO). The dye binding method used acid orange 12 dye, a measured amount
of this was added to 2.240 ml of thoroughly mixed milk sample. After
shaking vigorously and filtering, the percent transmission of the fil-
trate was measured with a spectrophotometer (Udy Color Analyser) set to
a reference point. Unknowns were calculated from a conversion table

supplied by the manufacturer. For several weeks both Kjeldahls and dye

47
binding methods were perfbrmed. Since values were in close agreement,
following this, only the dye binding method was used.

Body weights were measured at parturition and once a week thereafter.
This was done at approximately the same time each week. Once weekly,
throughout the second trial only, rectal temperatures were recorded.
Blood was collected at approximately 1000 h twice weekly throughout the
experiment via the tail vein using a 20 guage 25 mm vacutainer needle
and 20 ml vacutainer tubes (Becton Dickenson Inc., Rutherford, NJ). The
blood was allowed to stand at room temperature for 1 h after collection
and was then placed in a 5C cooler for 5 h. It was then centrifuged for
20 min at 1000 x g and the serum was removed. Serum was stored at -15C
until assayed for hormones and metabolites. Reproductive status was
determined by the concurrent examination of serum progesterone concentra-
tions and weekly rectal palpation data.

Feed samples were taken and analyzed for dry matter, protein and
phosphorus (analysis was done in the analytical laboratory of the Depart-
ment of Biochemistry, M.S.U. or in the analytical laboratory at Wooster,
OH).

From these composition measurements of the feed and N.R.C. feed com-
position tables a value for Net Energy (NE) and phosphorus was calculated
for each feed used in the experiment. (Feed assay data and NE and other
values used are shown in Appendix Tables 2 and 3.) From these values and
the feed intake data, milk production and composition measurements, a
calculated energy status was determined for every heifer once every two
weeks over the first 12 weeks of lactation.

First, energy requirements (MCal NE lactation/day) were calculated

for maintenance and lactation from N.R.C. (1971) tables using the two

48
week average f0r body weight, milk production and milk fat percent.
N.R.C. recommendations to add 20 percent to the maintenance allowance
for growth during first lactation and an increase of 3 percent feed for
each 10 kg of milk produced over 20 kg/day were not included in these
calculations. Then energy intake was calculated (MCal NE lactation/day)
using feed intake data and the energy values of the feeds. Thus, energy
status was calculated from the difference between the requirements and
actual intake. Phosphorus status was calculated in the same way. Energy
status or balance indicates the difference between input and output of
energy. If the energy consumed is not enough to meet these requirements
then the heifer would have to supply this energy difference from endo-
genous sources or decrease production. The heifer would in these cir-
cumstances be in negative energy balance or status. If the amount con-
sumed is equivalent to the amount required then the animal is at zero
balance (or 100% N.R.C.). Thus, status is an expression of intake of

energy and phosphorus relative to requirements.

Statistical Analysis

 

Because of the variation in the energy and phosphorus status of the
cows in both trials, data were analyzed not only by standard analysis
of variance procedures but also be regression analysis. In this way,
since absolute status values were used in the analysis, inaccurate group
designations did not affect subsequent conclusions. Absolute values of
energy and phosphorus status and their interaction were used as the
independent variables in a multiple regression analysis. The regression

equation is described below:

49

Y = a + 8X1 + 8X2 + Bxlxz
where:
Y = dependent variable
a = constant
X1 = energy status
X2 = phosphorus status
XIX2 = interaction of X1 and X2

Each heifer had a calculated X1 and X2 for each two week period of
the experiment. Thus, there were six values for each heifer and 144
observations were used in the regression analysis in each trial. In
addition to regression analysis all data were analyzed by two way analy-
sis of variance to determine if means were different. Also, because the
experiments were repeat measure designs the possibility of interactions
between the main effects and time period could not be ignored. Conse-
quently, all data were analyzed as shown below to test for time con-

fbunding factors:

U
’11

Source of Variance M.S.

I'm

 

Energy status (E)
Phosphorus status (P)
E x P

Error A (Anim./EP)
Period

E x Period

P x Period

E x P x Period

Error B (Anim. x time) 100 (220)*
Total 143 (287)*

N
UlU'IMMOt-‘HH

(44)*

 

*Degrees of Freedom (D.F.) for when both experiments were included in
the analysis.

50

Error 8 was used to test for significance of the period terms in
the analysis. If E x Period, P x Period or E x P x Period were signifi-
cant the data were shown to be confounded with time and reanalyzed by
considering each period separately. In this way, means from each period
were compared and tested and the repeat measure confounding factor was
removed. If these terms were not significant then no further analysis
of variance was necessary. Results in all cases were prepared by
analyzing each trial independently as well as together where possible
(i.e. where parameters were measured in both trials and were comparable).

All variables were analyzed to see if they were correlated with
time, whether the parameter increased, decreased or did not change with
time (or stage of lactation).

Other specific analyses were carried out to test specific hypotheses

and these will be discussed further in the results.

Analysis of Serum Hormones

 

Progesterone
Progesterone was measured by radiommunoassay as described by
Louis et a1. (1975). Progesterone was measured twice weekly for all
heifers throughout 15 weeks of lactation in both trials.
Insulin
Insulin was measured by radiommunoassay courtesy of Dr. E.
Beehuizen of Eli Lilly and Co., Indianapolis, IN. Insulin was measured
once every two weeks in the first trial and once weekly in the second
trial.
Growth Hormone

Growth hormone was measured weekly, in the second trial only,

51

by radioimmunoassay, as described by Purchas et al., (1970).

Analysis of Serum Metabolites

 

In the first trial serum glucose, calcium and phosphorus were
measured using a Hycel Mark X clinical autoanalyzer, (Hycel Inc.,
Houston, TX). The test principles used by this analyzer are described
under individual metabolite headings. In the second trial, since the
autoanalyzer was no longer available, alternative methods were used.
These are described below also.

Glucose

In the first trial, using the autoanalyzer, the test principle
used was of O-toluidine reacting with glucose in hot acetic acid to form
glucosylamine and a Schiff base. The end product was blue-green and was
quantified spectrophotometrically.

In the second trial, glucose was measured by a coupled reaction
enzyme system, using glucose oxidase and peroxidase, of a Glucostat
Reagent set (Worthington Biochemical Corporation, Freehold, NJ). Again,
the glucose is quantified spectrophotometrically.

In both trials glucose was measured weekly in all heifer throughout
the experimental period.

Calcium and Phosphorus

In the autoanalyzer method serum albumin bound calcium was
first released by acid. The total serum calcium was then complexed by
cresopthalein complexone. This was measured colorimetrically after the
complex was turned purple with base. Serum phosphorus was measured by
reacting serum inorganic phosphate with ammonium molybdate, this forms

ammonium-phosphomolybdate which was then measured colorimetrically.

52

In the second trial serum calcium and phosphorus were measured by
alternative methods using glassware washed with a 2:1, water:hydrochloric
acid solution. All solutions used in the assays were made with deionized
distilled water. Both assays required deproteinization of serum. Serum
was deproteinized in duplicate with 12.5% trichloracetic acid (TCA).
The serum was diluted 1:5 (0.5 m1 serum plus 2 ml TCA). The solution
was vortexed, allowed to stand for 5-10 min and centrifuged at 1000xg for
15 min. The supernatant was decanted into another set of acid washed
tubes for calcium and phosphorus analysis. Composite standards were
used in all assays and were run through all procedures beginning with
deproteinization. In addition, a blank deionized distilled water sample
was run with all assays to use in the phosphorus analysis.

Calcium

0.5 ml of supernatant was diluted with 3 ml of 10000 ppm

strontium solution, (strontium as strontium chloride, SrC12-6H20). The
solution was vortexed and read on an atomic absorption spectrophotometer
(Model IL 453, Instrumentation Lab. Inc., Lexington, MA).

Phosphorus

The Gomorri modification of the Fiske-SubbaRow inorganic phos-

phorus determination was used (Fiske and SubbaRow, 1925). To 0.5 m1 of
supernatant, 2.5 m1 of molybdate-sulphuric acid solution and 0.25 ml of
p-methyl-amino-phenolsulphate solution were added. The samples were
vortexed and incubated at room temperature for 45 min at which time they
were read at 700 my on a Gilford 24005 spectrophotometer, (Gilford

Instrument Laboratories Inc., Oberlin, OH).

S3
Reagents
10000 ppm Sr solution: 60.86 g SrC12-6H20 and 10.0 g NaCl were
diluted to 1 liter with deionized distilled water. This was diluted 1:1
with deionized distilled water to give 10000 ppm Sr solution.

Molybdate-sulphuric acid solution: 5 g of sodium molybdate (Na2M04
-2H20) in 500 m1 deionized distilled water, 14 m1 H2S04 (conc.) diluted
in approximately 200 m1 of deionized distilled water was added to the
molybdate solution. This solution was then made up to 1 liter with
deionized water.

P-methyl-amino-phenolsulphate (elon) solution: 1 g of elon was dis-
solved in 100 ml of 3% sodium bisulphite (NaHSOs). It was protected from
the light while mixing. The solution was stored in a brown bottle.

Serum Volatile Fatty Acids

Serum VFA determinations were done using the ethanolic extrac-
tion method described by Remesy and Demigne (1974). The concentrated
extract was quantified with a Model No. 5730A gas liquid chromatograph
(Hewlett Packard, Avondale, PA).

Two hundred ul of serum were placed in a 16 x 100 m1 glass tube and
1 m1 of pure ethanol was added. The solution was then vortexed and centri-
fuged for 15 min at 1000 X g. The supernatant was then transferred into
a 12 x 75 mm glass culture tube and made alkaline with 20 ul of 0.1
M NaOH. It was then evaporated in a slight air current at 20 C. The
dry residue was redissolved in 15 ul of water a few minutes before
analysis. Just before injection on the column, 5 ul of 25% (v/v) ortho-
phosphoric acid was added. Acidification at the last moment avoided
the risk of volatilization of the VFA and of micellar separation of

isobutyric acid, which is poorly soluble in acid conditions. Thus, a

54
solution with a concentration 10 times that of the original sample was
obtained. Using a microsyringe (Hamilton Co., Reno, Nev.) 3 ul of the

20 ul was injected on the column.

Conditions

 

Column: glass, 2.7m long, 2mm i.d.

Packing: 3% carbowax 20M, 0.5% H3P04 on 60/80 mesh carbopack B
(Supelco Inc., Bellefonte, PA).

Oven temperature: 160 C

Port and Detector temperature: 200 C

Detector: Hydrogen flame ionization.

Carrier gas: Nitrogen, flow rate 25 ml/min.

Peak area: Measured by an electronic integrator. Unknowns were calcu-
lated by comparing peak areas of a standard mixture of VFA with peak
areas of unknowns. Standards were about the same concentration as VFA
in serum. They were subjected to the same treatment as the unknowns.
The standard solution contained acetate, propionate, butyrate and

insobutyrate.

RESULTS

Feed Intake

 

Dry matter intakes of corn silage, hay and grain for trial 1 and
2 are shown in Table 1. In the first trial high energy (HE) groups con-
sumed 7.1 kg more grain but 2.8 kg less corn silage daily than low energy
(LE) groups. Hay intake was approximately constant, each row was offered
2.3 kg/day and consumed on average 1.7 kg/day.

Table 1.--Mean dry matter intake (kg/day) of corn silage, hay and grain
for the first 12 weeks of lactation in trials 1 and 2.

 

 

 

High Energy Low Energy

Feed Trial High Phos. Low Phos. ‘High Phos. Low Phos.
Corn silage 1 7.83 7.50 10.74 10.19
(with NPN) 2 6 46 6 58 7.28 7.22
(kg/day)

Hay 1 1.65 1.64 1.78 1.77
(kg/day) 2 2.08 2.08 2.08 2.09
Grain* 1 8.92 9.20 2.00 1.98
(kg/day) 2 8.71 8.91 2.02 2.03

 

*Dicalcium phosphate replaced by CaCO3 and corn in low P grain.

In the second trial, heifers consumed less corn silage, more hay
and approximately the same amount of grain as in the first trial. De-
pressed silage intake was partly due to the higher moisture content of
the silage and also due to deliberate restriction of silage fed, to
control energy intake. Treatment means are shown in Table 1. The HE

SS

56
groups consumed 6.8 kg more grain but 0.7 kg less corn silage. Hay in-
takes were not different among groups. There was no difference in silage
intake between high and low phosphorus groups.

Consequently, in trials 1 and 2 the HE groups consumed 49% of their
dry matter intake as grain mix, 51% of their intake as corn silage and
hay. In trial 1, the LE group consumed 14% and 86% of their dry matter
intake as grain and corn silage/hay respectively. In trial 2 the cor-

responding figures were 18 and 82% for the LE group.

Summary of Data Analysis

 

Data for both trials have been combined throughout this presentation
except where trial affected results.

Because trials were repeat measure experiments all data were tested
for period-nutrient status interaction. Where data were confounded by
period, data were analyzed within periods, otherwise data are presented

analyzed over all periods.

Energy and Phosphorus Status

 

Where status is an expression of intake of energy on phosphorus
relative to requirements.

Energy Status

HE groups were in positive energy status throughout the experi-

ment, and thus were receiving more than N.R.C. (1971) requirements, while
LE groups were receiving less than requirements for maintenance and milk
production. Energy status data from both trials are presented in Table
2.

Energy status for treatment groups is represented as a function of

stage of lactation versus energy expressed as a percent of N.R.C.

57
requirements in Figure 1. This enables a slightly different perspective
to be taken of the data than absolute values. In all groups energy
status increases with time, particularly in the HE groups, due to in-
creasing dry matter intake.

Table 2.--Mean energy status (MCal/day) of high and low energy groups
as a function of weeks of lactation for trials 1 and 2.

 

Weeks of Lactation

 

 

1-2 3-4 5-6 7-8 9-10 ll-12
High energy* 3.72 5.94 8.44 9.71 11.01 11.78
Low energy -5.03 -4.27 -3.76 -3.09 - 3.37 - 2.46

 

*HE > LE (P = .001) in all weeks. Differences between HP and LP were
N.S. There was no significant interaction of E and P status.

Phosphorus Status

Phosphorus status was not as easily controlled as energy status.

In both trials it was difficult to maintain negative phosphorus status
when combined with a high energy ration. Further complications were
added when in trial 2 the high energy, low phosphorus grain was incor-
rectly mixed and the resulting grain was not restricted in phosphorus con-
tent. Before this was discovered and corrected, some of the heifers
were not in negative phosphorus status.

Phosphorus status data for both trials are presented in Table 3.
It can be seen from this study only the low energy, low phosphorus group
was in negative phosphorus status throughout the experiment. When the
two trials were combined, the high energy, low phosphorus group was not
in negative status (for the previously mentioned reasons) but generally,

they were much less positive than either of the high phosphorus groups.

S8

.8330: no 9.00: «H yawn 0:».u0u an away 25ch amuse 50: .H. 0.23?"
8:55 ho SENS

2 . S . a. e e

D P b P D

’N

 

 

 

 

‘33
E
I044

T .\\..

om

cad

OHH

. 82

and

 

\\ I OmH
I

smwsmmma

XOXN

59

Table 3.--Mean phosphorus status (g/day) as a function of weeks of
lactation for both trials.

 

 

 

Weeks of High Energy* Low Ener
lactation High P** L_ow_P W _I_.p_w_li
1-2 16.90 4.69 11.64 - 6.55
3-4 22.93 3.55 8.72 - 9.14
5-6 27.62 6.02 9.84 - 9.76
7-8 35.19 4.50 12.18 -10.35
9-10 38.42 5.77 10.17 - 9.00
11-12 40.66 5.82 13.40 - 8.70
* In weeks 1-12, HE > LE groups in P status (P §_.002).
**In weeks l-12, HP > LP groups in P status (P = .001).

In weeks 1-6, interaction EXP, NS.

In weeks 5-12, interaction EXP, (P §_.04)
Thus, only the low energy, low phosphorus were marginally deficient in
phosphorus status. Only the high energy, high phosphorus group tended
to increase in phosphorus status as lactation advanced, whereas the other
groups were quite stable. Phosphorus status expressed as a percent of
N.R.C. requirements and as a function of stage of lactation is shown in
Figure 2. When high phosphorus groups were averaged they represented
138% of requirements. When low phosphorus groups were averaged they

represented 98% of requirements (average of 110% and 86%).

Milk Production

 

(a) Average daily milk yield
Milk yield for both trials combined is presented in Tables 4

and 5.

 

6O

.coaunuuoa mo axon: «a umuwm onu now an umzv magnum mauonauona coo:

.~ unawam

 

 

 

 

 

 

 

acquauuuq mo axed:
NH OH w o c N
P D l b p b m
on
4 tom
OOH
\ ‘ ‘
‘
r 2.4
I_
\ I OcH
\-
\I v 0nd
. I
\ mum..— ‘
I 3m: 0 _. o:
him: .
r 63

Z DUN anneas snioquoqa

61

Table 4.--Mean daily milk yield (kg/day) for both trials as a function

of weeks of lactation.

 

 

 

 

 

 

Weeks of High Energy Low Energy
lactation H_1_gh__P_ M H_i_g_h_I: L_ow__P
1-2 17.02 18.71 19.21 19.19
3-4 21.12 23.33 22.74 23.00
5-6 22.74 26.21 23.43 23.81
7-8 22.83 26.31 22.33 23.76
9-10 23.17 25.86 22.27 22.87
11-12 22.33 26.26 21.13 22.00
Mean 21.54 24.45 21.85 22.44

 

Table 5.--Mean daily milk yield (kg/day) for both trials for high and

low energy and high and low phosphorus groups.

 

 

 

 

 

 

Weeks of
lactation High Energy* Low Energy High P** Low P***
1-2 17.60 19.20 18.12 18.95
3-4 22.27 22.87 21.97 23.16
5-6 24.48 23.62 23.09 23.81
7-8 24.57 23.05 22.58 25.04
9-10 24.52 22.57 22.72 24.37
11-12 24.30 21.56 21.73 24.13
")1 22.96 22.15 21.70 23.24
* Weeks 1 , HE and LE NS.
Weeks 9 2, HE > LE (P §_.05).
** Weeks 1 HP and LP NS.
Weeks 5 11-12, LP > HP (P < .03).
Weeks 9 0, LP > HP (P = .10).
***Low P = 98% of NRC P requirements.

(Mean of 110 and 86%).

62

These data show there was no significant difference in mean daily
milk yield among groups for the first 4 weeks of lactation. From the
third through the twelfth week the daily yield of the low phosphorus
(LP) group exceeded that of the high phosphorus (HP) group. This dif-
ference ranged from 0.72 kg/day in weeks 5-6 to 2.46 kg/day in the weeks
7-8. Only in weeks 8-12 of lactation did the HE group yield significantly
more milk (P f_.05) than the LE group. This difference was 1.95 kg/day
in weeks 9-10 and 2.75 kg/day in the weeks ll-12. There were no signi-
ficant differences in average daily milk yield between heifers designated
best, worst and control genetic groups, (means were: 23.4, 22.0 and
21.9 kg/day, respectively).

Milk yields for HE and LE and HP and LP groups are graphically re-
presented in Figure 3. This graph shows that HE groups did not begin to
yield more milk until approximately 5 weeks of lactation, and after this
were more persistent than the LE groups. The second graph shows that LP
groups (which averaged 98% of N.R.C. requirements) consistently produced
more milk than HP groups throughout the experiment. When groups are

ranked by average milk yield the order is as follows:

1. HELP
2. LELP
3. LEHP
4. HEHP

When ranked by energy and phosphorus groups, the order is:

1. LP
2. HE
3. LE
4. HP

Regression analysis support the above data by showing that milk
yield increased as energy status increased and declined as phosphorus

status increased. Regression coefficients show that, relatively,

MILK YIELD IKgI DAYI

MILK YIELD (Kg! DAYI

26

24

22

20

18

16

14

26

24

22

20

'63

 

 

 

 

 

 

 

_—. +
p-
00..¢.......DIIIOIII I...
«on-«.90. ...
00.....O
I High EnerQY
. a Low EMMY
I L . l l 1
2 4 6 8 10 12
WEEKS on: LAcTAT'ON ‘
p-
b
..,-" I Low PhOsI’hmus
.- U
a High Phospmms
l l ‘ l L I
4 a B 10 12

WEEKS OF LACTATION

Figure 3. Mean milk yield (kg/day) for the first 12 weeks of
lactation.

64

phosphorus status was a more important component of the regression equa—

tion than energy status (Table 6).

Table 6.--Regression of energy and phosphorus status with milk yield,
milk fat % and milk protein % for both trials.

 

Standard Regression Coefficient*

 

 

 

 

 

Dependent Variable Enepgy Status Phos. Status EXP
Milk yield 0.30 -0.52 NS**
Milk fat % -0.57 NS** 0.31
Milk Prot. % 0.17 0.35 -0.19

 

* Standardized coefficients so that comparison may be made within
variables without regard to units.
**NS = P > .05.
(b) Milk fat percent
Milk fat percent combined data can be seen in Table 7. LE
groups had more (P = .02) fat percent than thelfligroups. This was probably
due to the LE group yielding slightly less milk than the HE groups.
Phosphorus groups did not significantly differ. This was a significant

Table 7.--Mean milk fat percent for the first 12 weeks of lactation for
both trials.

 

 

 

 

Phosphorus

Energy Hig Low Mean
High 3.32 3.13 3.22
Low 3.34 3.52 3.43
Mean 3.33 3.32

E P = .02

P NS

EXP P = .04

 

65
interaction between energy and phosphorus status. This was due mostly
to the low energy, low phosphorus group which had the highest milk fat
percent of 3.52%. As expected milk yield and milk fat % were negatively
correlated, r = -0.15, (P < .05). These data are supported by regres-
sion analysis which indicated that fat percent decreases as energy
status increases (P < .0005). The interaction between energy and phos-
phorus status was also significant (Table 6).
(c) Milk protein percent

HE groups had greater (P < .0005) milk protein percent than LE
groups and HP groups had greater (P = .01) protein percent than LP groups
(Table 8).

Table 8.--Mean milk protein percent for the first 12 weeks of lactation
for both trials.

 

 

 

 

Phosphorus

Energy High Low Mean
High 3.31 3.27 3.29
Low 3.18 3.02 3.10
Mean 3.24 3.14

E P < .0005

P P = .01

EXP NS

 

As expected milk yield and protein percent were negatively corre-
lated, r = -0.44, (P < .05). Regression analysis supports the conclusions
of the analysis of variance. As energy and ph05phorus status increased,
protein percent increased. Coefficients indicate that phosphorus status

had more effect on protein percent than energy status. The negative

66
interaction coefficient shows that energy and phosphorus effects were
less than additive (Table 6).
(d) 305 day actual milk yields

Since the first part of the lactation curve, including peak
milk yield, determines to a large extent the total of the lactation, it
was considered important to examine the total first lactation yields of
heifers involved in both trials. It seemed important to determine if
the treatments imposed in the first 3 months of lactation had any ad-
verse or beneficial effects on the whole lactation relative to their per-
formance in the first 3 months. Of course this assumes that even though
heifers were assigned to various other experiments after 3 months that it
is probable that treatment differences in subsequent experiments were
balanced and cancelled out. Table 9 shows the 305 day milk yields for

heifers from both trials.

Table 9.--Cumulative 305 day milk yield (kg) for heifers from both trails.

 

 

 

 

Phosphorus

Energy High Low Mean
High 5357 6515 5936
Low 5642' 6117 5880
Mean 5500 6316

E NS

P P = .01

EXP NS

 

There were no significant differences between energy groups in yield.
Consistent with the results in the first 3 months however, heifers fed at

98% requirements in early lactation yielded 816 kg more (P = .01) milk

67
in 305 days than heifers fed excessive phosphorus in early lactation.
Ranking in milk production was HELP, LELP, LEHP and HEHP and LP, HE, LE,
HP. These are the same order as they were after 3 months of lactation.
Thus, it could be that differences which occurred in the first 3 months
of lactation may have affected the total lactation. It is not clear
whether these differences were due to nutritional treatment or whether
the 305 day yields, or 3 month yields, reflect differences in genetic
ability among cows.

When these differences are expressed over the total 305 day lacta-
tion heifers which were fed excessive phosphorus for 84 days postpartum
gave 2.68 kg of milk less per day throughout lactation than those fed
at about N.R.C. requirements (i.e., the LP groups). HE and LE groups
were approximately the same, means were 19.46 and 19.28 kg/day, respec-
tively.

Further examination of the data showed that in the first 84 days
of lactation heifers had produced on the average just over 30% of their
total 305 day yield. This is shown below by group:

HEHP HELP LEHP LELP
33.8% 31.5% 32.5% 30.8%

If the 3 month treatment period is excluded and yield for the last
221 days of lactation was examined the groups ranked in yield as before:
HELP, LELP, LEHP and HEHP. When the average daily yield of the first
84 days is compared with the average daily yield of the last 221 days
it was found that for the HEHP group yield per day in the last 221 days
was 74.5% of the first 84 days, the corresponding value for the HELP group
was 82.6%. For the LEHP and LELP groups values were 78.8 and 85.3%, re-

spectively.

68

Health

(a) Health scores

Records were kept of the incidence of postpartum health problems

such as retained placenta, metritis, mastitis, displaced abomasum, respira-
tory and foot and leg problems. A scoring system of one point was used
for each recorded incidence of disease. In the case of mastitis another
point was scored for each incidence more than 3 days from the last record
of mastitis. Thus, a score was established for each group. Prior to
the second trial it was decided to add another more objective indicator
of health, and rectal temperatures were taken weekly on all heifers through-
out the second experiment. Also, culling data were collected from all
heifers which were culled after the first lactation, reasons for leaving
the herd were noted.

Mean health scores for each group with combined trials are shown in
Table 10.

Table 10.--Mean incidence of disease in both trials for the first 12
weeks of lactation.

 

 

 

 

Phosphorus
Energy High Low Mean
High 2.0 2.6 2.3
Low 1.1 1.6 1.3
Mean 1.5 2.1
E P = .15
P NS

EXP NS

 

69

Although means were not highly statistically different, the trend
was the same in both trials, HE groups tended to have more health problems
than LE groups, means were 2.3 and 1.3, respectively. There was little
difference in disease incidence for HP and LP groups. Generally, it is
difficult to show disease differences in small experiments but the bio-
logical significance of these data should not be ignored. Twice as many
cows would be needed to show significance at P = .05 level given similar
variance.

Health problems involved in these scores are shown for HE and LE
groups in Table 11. These data show that incidence of mastitis and

Table ll.--Types of health problems in high and low energy groups for
the first 12 weeks of lactation.

 

 

 

 

 

High Energy Low Energy

Health Number of Number of
Problems Incidences Percent Incidences Percent
Mastitis 17 32 16 49
Metritis 12 22 10 30
Respiratory/pneumonia 8 15 3 9
Foot and leg problems* 5 9 0 0
Displaced abomasum and

digestive upsets* 6 11 0 0
General infections* 4 7 0 0
Edema 2 4 1 3
Retained placenta** _Q_ 0 _§. 9
Total 54 100 33 100
* HE > LE (P = .05).
**LE > HE (P = .05).

70
metritis are similar in HE and LE groups. HE groups, however, had more
respiratory, foot and leg, digestive, general infection and edema problems.
Total number of disease incidences for HE groups was 54, almost twice as
high as for the LE groups with a total score of 33.
(b) Rectal temperatures

First of all, intra-animal correlations with time showed that
in each group rectal temperatures significant (P < .05) declined over
the first 12 weeks of lactation, r = -0.44 to -0.55. During this time
the HE groups had higher (P = .006) temperatures than LE groups. Means
were 38.87 and 38.74 C, respectively (38.61 E 101.5F E normal temperature).
There were no differences between phosphorus groups and no interaction
between the main effects. Average temperatures for high and low phos-
phorus groups were 38.78 and 38.83 C (Table 12).

Table 12.--Mean rectal temperature (C) for the first 12 weeks of lacta-
tion for the second trial.

 

 

 

Phosphorus
Energy High Low
High 38.88 38.85
Low 38.69 38.80
E P = .006
P NS
EXP NS

 

Regression analysis did not show that rectal temperature was dependent
upon energy or phosphorus status. The reason for this contradiction in
results is not clear.

When temperature was examined as a function of time it was clear it

declined as lactation advanced (Figure 4). Temperatures for HE and LE

71

£0quou no 3.003 Nu any: on» new 3v cyanogen-Du noun: 50:. .c was»:
:3ququ we 9.003

 

 

 

 

  

.2 2 2 m . m A m m . e m N _.
a q . u a _ , q u q q u — q “.mm
.9...
0...... ocean I O. m .m m
4 C m.—.o: 0m.mm
5mm
uw.mm
>mcocw 26.. o .m.mm
>mcocm £91 I
.o.mm
Adm
i

 

(3) BUHLVHBleBI. “11.338

72
groups in the first week of lactation were 39.0 and 38.9 C. Temperatures
did not decline to normal levels (38.6 C) until approximately 8 or 9 weeks
of lactation in LE groups, whereas by week 12, HE group temperatures were
still elevated. It was interesting that both milk yield and corn silage
intake were negatively correlated to rectal temperature I = -0.30 and
-0.43, respectively, (P < .05).
(c) Culling after first lactation

Examination of culling records showed that 42% (or 10/24) of
heifers fed excessive energy in the first 3 months of lactation were
culled compared with only 12.5% (or 3/24) of those fed low energy rations,
(P = .05). Culling percentages were 3.33 (8/24) and 20.8% (5/24) for HP
and LP groups, respectively. It should also be noted that no heifer was
culled after first lactation in the low energy, low phosphorus group in
either trial. Of 10 heifers culled in the HE group, 5 were because of
fertility problems, three were for poor condition and health and 2 were
because of udder problems. All three of the heifers culled in LE groups
were because of fertility problems. These culling data are somewhat
biased, however, since no cows were culled for production due to an on-

going genetics experiment.

Energnytatus Relationship
(a) Body weight change
Change in body weight expressed as kg/week of the treatment
period generally followed energy status of cows in both trials. When
data from both trials were combined HE groups gained on average 0.54 kg/
week whereas the LE groups lost 1.49 kg/week (P = .034). There was no

difference in weight loss between phosphorus groups and no interaction

73

of energy and phosphorus status (Table 13).

Table l3.--Change in body weight (kg/week) for the first 12 weeks of
lactation for both trials.

 

 

 

Phosphorus
Energy High Low
High 0.475 0.575
Low -1.18 -1.75
E P = .034
P NS
EXP NS

 

Variation within each group was considerable. For example, the range of
weight change in the HEHP groups was -5.15 to +5.73 kg/week, and range
for the LELP group was -6.33 to +1.85 kg/week. This shows why, when
averaged, body weight changes seem small. Of the four groups: HEHP,
HELP, LEHP and LELP: 6, 7, 3 and 4 out of 12 heifers were gaining weight
over the 12 week period, whereas the rest, on average, lost weight.
Regression analysis shows that body weight change in dependent upon
energy status but not in a highly predictive manner (r = 0.51, P < .01)
(Table 14). The groups that were gaining and losing the most weight,
HELP and LELP, were ranked first and second in milk production. Body
weight change and milk production were negatively correlated, r = -0.54,

(9 < .01).

74

Table 14.--Regression of energy_and phosphorus status with body weight

 

 

 

 

 

change.
Variable Energy Status Phos. Status EXP
Body weight Trial 1 0.54* NS NS
change

Trial 2 0.44** NS NS

Combined trials 0.51*** NS NS
* P = .07.
** P = .15.
***P = .007.

(b) Serum glucose
Response of serum glucose to energy and phosphorus status dif-
fered slightly in each trial. Since, also, serum glucose should be
examined by trial when considering insulin responses later in these re-
sults, glucose data will be presented separately.

In the first trial serum glucose concentrations of HE groups were
greater (P = .06) than those of LB groups by approximately 4 mg/dl (Table
15). Neither phosphorus status nor the interation of the main effects
had any influence on glucose concentrations.

In the second trial glucose concentrations were, on average, 13-14
mg/dl less than in the first trial. It is not clear if this is due to
assay differences, real differences in cows due perhaps to feeding
schedule differences (since cows in trial 1 were sampled nearer to feed-
ing) or due to some factor the author is not aware of. However, serum
glucose concentrations were higher (P = .025) in cows fed excess energy
than in those fed low energy. Glucose also increased, however, (P = .026),
in those cows fed HP when compared to those fed LP (Table 16). There was

no significant interaction between energy and phosphorus status and

75

Table 15.--Means serum glucose concentrations (mg/d1) for the first 12
weeks of lactation for the first trial.

 

 

 

 

Phosphorus
Energy High Low Mean
High 75.9 75.0 75.5
Low 70.4 72.8 71.6
E P = .06
P NS
EXP NS

 

Table l6.--Mean serum glucose concentrations (mg/d1) for the first 12
weeks of lactation for the second trial.

 

 

 

 

Phosphorus

Energy High Low Mean
High 61.1 59.8 60.4
Low 59.8 57.8 58.8
Mean 60.4 58.8

E P = .025

P P = .026

EXP NS

 

glucose levels. Serum glucose increased 1.6 mg/dl when energy or phos-
phorus was increased. Thus, these glucose data do not reflect energy
status quite as well as data from the first trial.

Regression analyses support analysis of variance results. Coeffi-
cients from the first trial show that as energy status increases, glucose
concentrations increase (Table 17). However, energy and phosphorus were
not entirely independent of each other since the interaction term was

significant. In the second trial only the phosphorus status term was

76

Table l7.--Regression of energy and phosphorus status with serum glucose

 

 

 

 

 

 

concentrations.
Standard Regression Coefficients
Dependent Variable Energy Status Phos. Status EXP
Serum Trial 1 0.42 NS 0.20
glucose
Trial 2 NS 0.45 NS

 

significant (Table 17). This appears to be due to a wider range of phos-
phorus status values compared to the range of energy status values and
this, coupled with glucose increasing equally when energy or phosphorus
was increased meant phosphorus status expressed glucose concentrations
better than energy status in the regression equation.

Finally, analysis showed that in no group did serum glucose increase
or decrease significantly as lactation advanced. Serum glucose was posi-
tively correlated with energy status, r = 0.30, (P <.01). Serum glu-
cose was negatively correlated to serum acetate, r = -.19, (P < .05) but
was not significantly correlated to other serum VFA's measured.

(c) Serum insulin

Since insulin data from each trial were different, each will
be discussed separately. In the first trial serum insulin concentrations
responded markedly and positively to energy status increases (Table 18).
The difference between energy groups, except, in weeks 9-10, was quite
dramatic. On the average HE groups had four-fold higher insulin than
LE groups. It is important to remember that these differences were seen
in samples taken approximately 3 h postfeeding of the main feed of the
day. In no week was there any difference between phosphorus groups or

a significant interaction between energy and phosphorus status.

77
Regression analysis strongly support the analysis of variance re-
sults, showing that as energy status increased, insulin concentrations
increased (P < .0005), (Table 19). Figure 5 shows the relationship be-
tween HE and LE groups graphically as a function of stage of lactation.

Table 18.--Mean serum insulin concentrations (uU/ml) for the first trial
as a function of weeks of lactation.

 

 

Weeks of High Energy* Low Energy
lactation High_§_ ppy_g_ High_£_ Lgy_£_
1-2 15.10 12.62 4.03 3.23
3-4 15.68 10.83 5.03 3.98
5-6 27.00 18.57 3.50 4.42
7-8 23.73 13.48 5.77 6.32
9-10 14.83 13.48 11.23 15.70
ll-12 18.70 14.37 2.02 5.38

 

*In weeks 1-8, and ll-lZ, HE > LE (P §_.05).
In weeks 9-10, HE and LE NS.

Table l9.--Regression of energy and phosphorus status with serum insulin

 

 

 

 

 

concentrations.
Dependent Variable Energy Status Phos. Status EXP
Serum Trial 1 0.50 NS NS
insulin

Trial 2 0.51 NS NS

 

In the second trial the response in serum insulin concentrations to
changing energy status was not as marked. This may be in part due to
difference in sampling times relative to feeding. In the second trial

the LE group were sampled approximately 15 h postfeeding, the HE group

.N Houuu cw :ofiuouuoa mo mxoo: NH umuum onu new Aa6\=:v :«aamcu Eamon coo: .m ouswwh

cowuouuoq mo mxooz
NH 3 . m o e

 

 

 

N
n a _ lb P p
0’ \O I G
. .lIlAu
.uo
.um
.Ioa
. :3
.0
O /
\\\\\\\\\\\\\\\\\ .uwH
I
. .ImH
C
xmuocu 30A 0 IS
km N
Saw a :1 I INN
.0
rs

(Iw/nn) utlnsul mnias

79

was sampled approximately 3 h after half of the grain ration was fed.

Insulin concentrations in the HE group were 28% higher (P = .001)
than in the LE group (Table 20). There were no differences between
phosphorus groups in insulin concentrations. There was a significant
energy and phosphorus status interaction due to the HELP group which had
the highest mean insulin concentration. These results were supported
by the regression analysis which again showed that insulin concentrations
were highly dependent (P < .0005) on energy status (Table 19). The
interaction term in the regression equation was near significance (P =
.063) showing that, as in the analysis of variance, energy and phosphorus

Table 20.--Mean serum insulin concentrations (uU/ml) for the first 12
weeks of lactation in the second trial.

 

 

 

Phosphorus
Energy High Low Mean
High 6.46 8.03 7.25
Low 5.96 5.41 5.68
E P = .001
P NS
EXP P = .02

 

status were not entirely independent in their influence on serum insulin.
Figure 6 shows serum insulin in the second trial for HE nad LE groups as
a function of stage of lactation. From this figure it can be seen that
after only 2 1/2 weeks of lactation the separation between HE and LE

groups is clear.

80

«A

ad

.N Hugh» 6% acqumuoma uo mxooz NH umuau on» new AHE\D:V awazmcw asuoo coo: .e ouawwm

 

83303 L6 8.3:

S a w .a o n
D b b D

hwuoem 36.— D

.885 .3: I

L...
row
Inn
.9...
1......
no;
rm;
8.»
a.»

no.m

 

(IN/fin) uttnsul mnias

81
(d) Serum volatile fatty acids
(1) Serum acetate
Acetate (C2) was measured in the second trial only. There were
no significant differences between groups in serum acetate concentrations
overall mean was 736.4 uM (Table 21).

Table 21.--Mean serum acetate concentrations (uM) for the first 12 weeks
of lactation in the second trial.

 

 

 

Phosphorus
Energy High Low
High 713.5 823.7
Low 761.5 737.0
E NS
P NS
EXP NS

 

Regression analysis supports these conclusions showing that energy and
phosphorus status had no significant influence on serum C2 concentrations
(Table 22).

Table 22.--Regression of energy and phosphorus status with serum vola-
tile fatty acids.

 

 

 

 

 

Dependent Standard Partial Regression Coefficients
Variable Energy Status Phos. Status EXP
Serum acetate NS NS NS
Serum propionate 0.37 NS NS
Serum butyrate 0.57 NS NS

Serum isobutyrate NS NS NS

 

82

Serum C2 was negatively correlated with serum glucose, r = -0.19, ( P <
.05), but was positively correlated with milk yield and corn silage in-
take, r = 0.24 and 0.22, respectively (P < .05).

(2) Serum propionate

Serum propionate (C3) was greater (P = .001) in the HE than in
the LE groups, means were 23.9 and 15.3 uM, respectively. C3 was also
different between phosphorus groups, the LP groups had greater (P = .022)
C3 concentrations than the HP groups, means were 22.5 and 16.6 uM (Table
23). The significant interaction between energy and phosphorus status

Table 23.--Mean serum propionate concentrations (uM) for the first 12
weeks of lactation in the second trial.

 

 

 

Phosphorus
Energy High Low
High 17.8 29.9
Low 15.4 15.2
E P = .001
P P = .022
EXP P = .02

 

(P = .02) was due mainly to the high energy low phosphorus group which
had the highest C3 level. Serum C3 and grain intake were positively
correlated, r = 0.29, (P < .05) but C3 was not significantly correlated
to serum glucose. Regression analysis showed that serum C3 increased as
energy status increased (P = .013) but not that phosphorus status or the

interaction of energy and phosphorus status had any effect (Table 22).

(3)

In general, where means were different (Table 24), HE groups

Serum butyrate

83

had higher (P f .044) butyrate (C4) concentrations than LE groups and

LP groups had higher (P §_.09) serum C4 than HP groups.

5-6 was this relationship not consistent.

Only in weeks

It is not clear why this is so.

Regression analysis showed that as energy status increased, serum C4 in-

creased.

Regression analysis did not, however, show that phosphorus

status or the interaction between energy and phosphorus status had any

influence on serum (C4) (Table 22).

Serum C4 was not correlated to serum

glucose but was positively correlated to grain intake, r = 0.37 (P < .05).

Table 24.--Mean serum butyrate concentration (uM) for the first 12 weeks
of lactation in the second trial.

 

 

 

 

 

 

 

Weeks

only, EXP (9:

.003).

Weeks of
lactation HE* LE HP** LP
1-2 12.52 5.15 4.55 13.13
3-4 17.32 7.45 8.42 16.35
5-6 7.43 20.14 19.94 7.63
7-8 14.52 5.00 8.23 11.29
9-10 11.23 3.01 4.62 9.62
11-12 10.88 6.34 7.74 9.47
* Weeks 1- 2,5- -6, NS.
Weeks 3- 4,7- -12, HE > LE (P < .044).
**Weeks 5- 8,11- -12, NS.
Weeks 1-4,9-10, LP > HP (P < .09).
3-40

84

(4) Serum isobutyrate

Analysis of variance showed no differences between energy groups
in serum isobutyrate. The LP group had higher (P = .005) serum isobuty-
rate (isoC4) than the HP group, means were 2.50 and 1.73 uM. The signi-
ficant interaction of energy and phosphorus status was due to the high
energy low phosphorus group and the high energy high phosphorus group
which had the highest and lowest means, respectively (Table 25).

Table 25.--Mean serum isobutyrate concentration (uM) for the first 12
weeks of lactation in the second trial.

 

 

 

 

Phosphorus
Energy High Low
High 1.40 2.84
Low 1.92 2.17
E NS
P P = .005
EXP P = .05

 

Regression analysis showed that only energy status was near significance
(P = .1) in influencing isoC4 concentrations (Table 22). It is not clear
why this is contradictory to analysis of variance results.

Finally, all serum VFA were positively correlated with one another,

(P < .05), values are shown:h1(Table 26).

85

Table 26.--Simple correlations between serum VFA.

 

 

 

VFA Simple Correlation
C2 x C4 0.65
C2 x isoC4 0.46
C3 x C4 0.77
GS X isoC4 0.46
C4 x isoC4 0.46

 

(e) Growth hormone
Serum growth hormone results are presented by weeks in Table 27.
In weeks 1-2 HE groups had greater (P = .08) serum GH than LE groups. In
weeks 3-8 there was no significant difference between HE and LE groups
but by week 9 it was apparent that HE groups had decreased whereas LE
groups had not changed in CH concentrations. In weeks 9-12 LE groups
had greater (P §_.02) GH concentrations than HE groups. GH as a function
of stage of lactation for HE and LE groups is presented in Figure 7.
Regression analysis indicated that GH was not significantly influenced
by energy or phosphorus status.

Analysis showed that GH in the HEHP and HELP groups declined as
lactation advanced, r = -0.39 and -0.43, respectively (P < .05). There
was no significant increase or decline in the two low energy groups.

GH was negatively correlated with milk yield, r = -0.23, (P < .05), and
corn silage intake, r = -0.22, (P < .05). It was positively correlated

with serum C4 concentrations, r = 0.23 (P < .05).

86

.N away» a“ sawumuuoa mo mxooa Na umuqm osu new AHE\ch ocosuo; canon» enuom .s ouawah

caguouooa mo wxooz

 

 

NH HA OH a m h o M Q n N H
Pll - . p p p p p p . — .
O. 1.
\.\ /. .\O
.

/..\ / \./.

\ z<x /\/\/

hwuocm swam O

 

o.¢

o.m

o.o

o.n

(Im/Hu) auomaon q2n019 WHJOS

87

Table 27.--Mean serum growth hormone concentrations (ng/ml) as a func-
tion of weeks of lactation.

 

 

 

 

 

 

 

Weeks of
lactation HE* LE HP** LP
1-2 5.00 3.82 4.13 4.69
3-4 5.75 4.59 4.57 5.77
5-6 4.93 4.01 4.43 4.51
7-8 3.48 3.88 3.59 3.78
9-10 3.50 5.39 3.95 4.94
11-12 3.51 4.. 3.89 3.84
* Weeks 1-2, HE > LE, P = .08.
Weeks 3-8, HE and LE, NS.
Weeks 9-12, LE > HE, P 5 .02.
**Weeks 1-12, HP, LP, NS.‘

Indicators of Phosphorus Status

(a) Serum phosphorus

Because analysis of variance results were not the same in each

trial results are presented separately. In the first trial serum P
reflected P status since the HP groups had greater (P = .02) serum P
concentrations than the LP group. Neither energy status nor the inter-
action of energy and phosphorus status affected serum P levels (Table 28).
Regression analysis showed that serum P increased as phosphorus status
increased, decreased as energy status increased and that these two effects
were not additive (Table 30).

In the second trial HP groups had greater serum P than LP groups
(P < .005) but the HE groups also had greater (P = .0006) serum P than

the LE groups (Table 29).

88

Table 28.--Mean serum phosphorus concentrations (mg/d1) for the first 12
weeks of lactation in the first trial.

 

 

 

Phosphorus

Energy High Low
High 6.67 6.22
Low 7.52 5.68
Mean 7.1 6.0

E NS

P P = .02

EXP NS

 

Table 29.--Mean serum phosphorus concentrations (mg/d1) for the first 12
weeks of lactation in the second trial.

 

 

 

 

Phosphorus

Energy . High Low Mean
High 5.72 5.33 5.53
Low 5.54 4.97 5.25
Mean 5.63 5.15

E P = .0006

P P < .0005

EXP NS

 

Regression analysis confirmed that as phosphorus status increased, serum
P increased but did not show that energy status influenced serum P con-

centrations (Table 30).

89

Table 30.--Regression of energy and phosphorus status with serum phos-

 

 

 

 

 

 

phorus.
Standard Regression Coefficients
Dependent Variable Energy Status Phos. Status EXP
Serum Trial 1 -0.33 0.59 -0.25
Phos.
Trial 2 NS 0.44 NS

 

(b) Serum calcium

Since there were no important differences between trials data
are presented for both experiments combined. Serum Ca was greater (P =
.001) in the LP groups than the HP groups, means were 9.69 and 9.45 mg/
d1 for LP and HP. Thus, there is an inverse relationship between serum
Ca and P. There were no differences in serum Ca between energy groups
but the interaction of energy and phosphorus status was significant
(P = .04) (Table 31).

Table 31.--Mean serum calcium concentrations (mg/d1) for the first 12
weeks of lactation for both trials.

 

 

 

Phosphorus
Energy High Low
High 9.56 9.65
Low 9.35 9.73
E NS
P P = .001
EXP P = .04

 

Regression analysis supports the conclusion that serum Ca decreases as

phosphorus status increases and the interaction of energy and phosphorus

90
status increases and the interaction of energy and phosphorus status
(Table 32). This interaction is due to increasing energy blocking the
effect of phosphorus. This may be the result of confounding of energy
and phosphorus particularly in trial 2. In all but the HEHP group serum

Ca increased as lactation advanced, r = 0.41 to 0.51, (P < .05).

Table 32.--Regression of energy and phosphorus status with serum calcium.

 

Standard Regression Coefficients
Dependent Variable Energy Status Phos. Status EXP

 

 

 

 

 

Serum Ca combined trials NS -0.31 0.32

 

Regulation of Serum Insulin

 

These experiments presented an opportunity to examine serum insulin
as it responded to differences in energy intake and to attempt to relate
these changes to other serum hormones or metabolites.

Because more data were available in trial 2 a number of variables
were selected from the second trial for a regression analysis involving
serum insulin as the dependent variable. Variables for analysis were
chosen because they were significantly correlated with serum insulin.
Grain intake was significantly correlated with insulin but since it was
so highly related to energy status (r = 0.95) it was not included.

By analysis of variance and regression analysis insulin was shown
to be related to energy and phosphorus status. Table 33 shows variables
that insulin was correlated with in both trials 1 and 2. Serum glucose
and insulin concentrations were significantly correlated, as were energy

status and insulin. However, in trial 2, serum C2, C3 and C4 were more

91
correlated with insulin concentrations than glucose. Serum C2, C3,
C4 and isoC4 were not measured in trial 1 and so comparable information
is not available. Although milk yield was not well related to insulin

in either trial it was chosen as one of the variables for analysis.

Table 33.--Simple correlations with insulin in trials 1 and 2.

 

Simple Correlations

 

 

 

Trial 1 Trial 2
glucose x insulin 0.35* 0.22*
C2 x insulin -- 0.26**
C3 x insulin -- 0.42**
isoC4 x insulin -- -0.048
C4 x insulin -- 0.40**
GH x insulin -- -0.27*
E status x insulin 0.54* 0.32*
P status x insulin 0.33* 0.10
milk yield x insulin -0.l7* 0.13
* P < .05, r = 0.159, n = 144.
**P < .05, r = 0.205, n = 92.

Figure 8 shows the stepwise deletion regression analysis used. In
dependent variables that significantly contributed to the equation (F <
.0005, R2 = 0.26) were serum GH, 03, c4 and 15004. Variables with 9 > .1
were deleted (that is, energy status was deleted first, glucose last).
Coefficients in Figure 8 are absolute values and are not standardized.
Table 34 shows standard regression coefficients of the previous equation
(Figure 8) so that independent variables may be compared with each

other.

92

y = 6.4 + 177.6x1 - 1092.9x2 - 0.23x3 + 143.6x4

where y insulin
x1 = butyrate
x2 = isobutyrate
x3 = growth hormone
x4 = propionate

(9 < .0005, R2 = 0.26)

Deleted: (1) E status (2) P status (3) milk
yield (4) acetate (5) glucose

Figure 8. Deletion regression analysis with serum insulin

as dependent variable.

Table 34.--Deletion regression analysis with serum insulin as dependent
variable: standard coefficients.

 

Stand.

 

 

 

Variable Regression Coeff. R2 Deletes* Sig.
Growth hormone -0.338 0.16 .001
Propionate 0.310 0.22 .040
Butyrate 0.265 0.24 .088
Isobutyrate -0.261 0.21 .016

 

*Original Rigs 0.26.

Examination of these coefficients show GH and C3 concentrations to
be the most important variables since they have the highest coefficients.
GH and insulin were negatively related and C3 and insulin were positively
related. C4 and isoC4 fellow with approximately equal coefficients. The
R2 deletes column indicates the R2 or proportion of total variance that

would result if that single variable were dropped from the equation. R2

93
decreases the most when GH is eliminated from the equation, (from R2 =
0.26 to 0.16). If either C3, C4 or isoC4 are excluded the R2 also de-
clines (Table 34). C4 was positively related to insulin, isoC4 was
negatively related. This analysis suggests that serum GH and serum VFA
(excluding C2) are more important in regulation of serum insulin concen-

trations than serum glucose concentrations or other parameters tested.

Reproduction Data

 

Serum progesterone concentrations were used in conjunction with
rectal palpation data to establish occurrence of estrus, ovulation and
normal cycles. Number of days to first ovulation (it was not in all
cases detected as an overt estrus), and number of days postpartum to
reach a progesterone concentration of 3 ng/ml were calculated. Time
taken to reach 3 ng/ml was measured because this level is considered to
be the peak needed in the estrous cycle preceding insemination in order
to permit conception (Folman et al., 1973; Corah et al., 1974; Rosenberg
et al., 1977). It was hoped that these two parameters would provide
an independent measure of reproduction, however, days to each 3 ng/ml
progesterone and days to first ovulation were well correlated, r = 0.74,
(P < .01).

Days to reach 3 ng/ml progesterone and days to first ovulation were
regressed with only energy and phosphorus status data from periods pre-
ceding these events for each heifer. Of course, energy or phosphorus
status after an event does not affect the event. Throughout these re-

productive data, where P > .25 the effect is designated non-significant.

94
(a) Days postpartum to reach 3 ng/ml progesterone
Although there were no significant differences between groups,
LP groups took longer (P = .19) to reach 3 ng/ml than HP groups (by
approximately 9 days) (Table 35). Calculation shows that given the
same variation in data approximately n = 100 would be needed to show a
difference between P groups (at P = .05 level).

Table 35.--Number of days postpartum for serum progesterone to reach
3 ng/ml in both trials.

 

 

 

 

Phosphorus

Energy High Low Mean
High 43.8 52.8 48.3
Low 36.7 46.5 41.6
Mean 40.3 49.7

E NS

P NS (P = .19)

EXP NS

 

When these data were analyzed by regression with energy and phosphorus
status there was little indication that energy or phosphorus status in-
fluenced progesterone concentrations reaching 3 ng/ml. Correlation data
showed that as serum phosphorus or glucose increased, days to reach 3
ng/nu.progesterone decreased (occurred sooner), r = -0.34, -0.33 (P < .05),
respectively. Days to 3 ng/ml progesterone was positively correlated
with serum GH concentrations (r = 0.51, P < .05).

(b) Days to first ovulation

HE groups, on average, ovulated for the first time approximately

4 days prior to LB groups. HP and LP groups both ovulated at 27 days

95
postpartum. There were no significant differences between groups by

analysis of variance (Table 36).

Table 36.--Number of days postpartum to first ovulation.

 

 

 

 

Phosphorus
Energy High Low Mean
High 27.0 22.0 24.5
Low 26.8 31.2 29.0
Mean 26.9 26.6

E, P, EXP, NS

 

Regression analysis supports the hypothesis of increasing energy
status decreasing time to first ovulation. The regression coefficient
for energy status was -0.38 (P = .07), phosphorus status and the inter-
action of the two main effects had no effect on first ovulation. Serum
insulin was negatively correlated to days to first ovulation, r = -0.55,
(P < .01) in trial 2, but these parameters were not significantly corre-
lated when both trials were combined. Heifers apparently ovulated bef0re
progesterone concentrations reached 3 ng/ml, on average this difference
was 18 days.

(c) Days non-pregnant

Heifers could leave the herd only if one of the following condi-
tions was met: (1) they were diagnosed pregnant; (2) they had been in-
seminated 3 times; (3) they were more than 150 days in lactation. One
hundred and fifty days was selected as a realistic cut off point beyond

which heifers would normally be culled if apparently infertile.

96
Actual pregnancy dates were used for heifers which became pregnant
prior to 150 days postpartum. All other heifers were arbitrarily assigned
150 days non-pregnancy. When this was done all groups had similar days

non-pregnant (Table 37).

Table 37.--Mean number of days non-pregnant for both trials.

 

 

 

 

Phosphorus
Energy High Low Mean
High 117.8 123.8 120.8
Low 110.1 121.4 115.8
Mean 114.0 122.6

E, P, EXP, NS

 

Given the same variation calculations indicate a sample size of at
least 250 would be needed to show differences between phosphorus groups
and over 700 to show differences in energy groups in pregnancy (at P =
.05 level).

(d) Inseminations per conception

There are no significant differences between groups in insemina-
tion per conception (Table 38).

Given the same variation a sample size of about 280 would be needed
to show differences between energy groups (at P = .05 level).

There were some differences however, in number of cows pregnant.

For HE and LE groups, 8 (or 33%) and 3 (or 12.5%) heifers, respectively,
did not conceive six (25%) of HP and 5 (21%) of LP groups did not conceive.

These differences cannot be attributed to treatments, however.

97

Table 38.--Mean number of inseminations per conception for both trials.

 

 

 

 

Phosphorus

Energy High Low Mean
High 2.56 1.86 2.2

(9)* (7) (16)
Low 2.44 2.67 2.6

(9) (12) (21)
Mean 2.5 2.3

(18) (19)

E, P, EXP, NS

 

*Number cows pregnant.

(e) Absolute number and percent undetected estrus
There were no differences in either number or percent of un-
detected estrus (Table 39). Overall means were 1.45 (out of 3.3 ovula—
tions) or 43.8% undetected estrus during the first 15 weeks of lactation.

Table 39.--Mean number and percent undetected estrus for both trials for
the first 15 weeks of lactation.

 

 

 

 

Phosphorus

Energy High Low Mean
High 1.3 1.6 1.5

(41.7)* (48.6) (45.2)
Low 1.2 1.7 1.4

(35.4) (49.3) (42.4)
Mean 1.3 1.6

(38.6) (49.0)

E NS

P NS (P < .25)

EXP NS

 

*Percent undetected estrus.

98
An estimated sample size of 116 would be needed to detect differences be-
tween phosphorus groups at the P = .05 level.

(f) Interval to second and third ovulations

There were no significant differences in interval to second or
third ovulation. Forty-four of 48 heifers had a second ovulation, 38
of 48 had a third. Overall means were 20.6 days and 21.2 days for second
and third interestrual intervals, respectively. All first, second and
third ovulations used in these analyses occurred within 13 weeks post-
partum, during and just after the dietary treatment period.

There were no differences between phosphorus groups in mean number
of ovulations that occurred within the 15 week experimental period, mean
was 3.3 ovulations per heifer. HE groups had less ovulations (P < .25)
than LE groups, means were 3.1 and 3.5, however, it was estimated that
a sample size of 116 would be needed to detect differences at P = .05

level.

Simple Correlations

 

Other correlations which were statistically significant but are not
reported in the text and which may be interest are listed in Appendix

Table 4. Non-significant correlations are not presented.

DISCUSSION

Results of these experiments show postpartum energy and phosphorus
status affects milk production and possibly health. Certain serum meta-
bolites and hormones responded to changes in energy or phosphorus status.
Relationships between energy and phosphorus status and reproductive
function were not definitive. Status, is an expression of intake of
energy or phosphorus relative to requirements for maintenance and milk
production. This must be distinguished from intake per se. Analysis of
variance really tested differences in intake rather than status. Whereas
multiple regression analysis tested for influence of status on variables
measured, not of intake. This explains why results of these two analyses
are not always the same.

Milk production responses of heifers to energy and phosphorus status
were of practical significance. Heifers fed excessive phosphorus (138%
N.R.C.) in the first 3 months of their lactation yielded 6.5% less milk
per day for 84 days postpartum than those heifers which were fed almost
to requirements (98% N.R.C.). When 305 day yields were compared, heifers
fed excessive phosphorus early in lactation yielded 13% less milk (816
kg) than heifers fed to recommendations. If excess phosphorus suppressed
milk yield it is not immediately clear how this occurred. The excess
phosphorus may have affected metabolism at the tissue level. The possi-
bility that differences between phosphorus groups were due to differ-
ences in genetic potential, post-treatment or unknown factors cannot be

99

100
excluded. There was no apparent difference in health of phosphorus
groups so this at least was probably not the cause of differences in
yield.

Because of fertilization and natural soil content, phosphorus levels
in feeds may well meet requirements of cows without additional phosphorus
supplementation. It cannot be assumed that phosphorus 30-50% in excess
of requirements is without effect. In fact, data from these experiments
suggest it is better to feed just below requirements rather than over
them. Energy status also affected milk yield. HE groups did not begin
to yield more milk than LE groups until week 5 of lactation and this
difference did not become significant until week 8 when LE groups were
less persistent.

These responses could in part be due to postpartum disease since
heifers fed in excess of requirements had nearly twice as many incidences
of disease when compared with heifers fed below requirements. Also,
whereas the majority of disease incidence in LE and HE groups occurred
within the first 4 weeks of lactation, HE groups also had more disease
problems from week 5 onwards, (17 versus 5 incidences for weeks 5-12 for
HE and LE groups). Prolonged reoccurrences of health problems may have
suppressed milk yield potential in HE groups. By 9 weeks incidence of
problems was low in all groups. Incidences of mastitis and metritis
were approximately equal between HE and LE groups. HE groups, however,
had more respiratory, pneumonia and edema problems. LE heifers had no
displaced abomasums or digestive problems, no general infections or foot
and leg problems. Previous studies have linked liberal pre- and post-
partum grain feeding to increased disease but not always with statistical

significance. The type of disease varies from trial to trial suggesting

101

a nonspecific mechanism, (Armstrong et al., 1966); Emery et al., 1969;
Trimberger et al., 1972; Belyea et al., 1975a). It is possible that
feeding excess energy lowers resistance to disease in some way.

Rectal temperatures in all groups declined as lactation advanced
and reached 'normal' levels at approximately 9 weeks. It is tempting
to suggest that HE groups had higher temperatures because of higher
incidence of disease, especially since after 9 weeks, disease incidence
was practically zero and temperatures were 'normal'. Both HE and LE
groups had elevated temperatures at the beginning of lactation which is
consistent because both groups had a fairly high incidence of disease
at this time. However, if one examines heifers with a zero disease
score, (no clinical disease problems), which also have rectal temperature
measurements then HE group heifers still had higher temperatures than
LE heifers for the first 4 weeks of lactation and in both groups tempera-
tures were elevated above normal. These limited data suggest that energy
status rather than disease per se affected temperature. Also, tempera-
tures at beginning of lactation seemed to be above normal regardless of
nutritional regime or health status. This elevation is probably not due
to increased feed since feed intake was increasing as temperatures were
declining. It could be that temperature effects observed were due in
part to health and in part to energy status and attempts of cows to
dissipate extra heat from feed. As far as the author is aware, tempera-
ture elevation at the beginning of lactation has not been previously
examined.

Subsequent culling data support excess energy status-health problem
associations. Three times more heifers were culled from the original

HE groups than LE groups and most of these heifers culled in HE

102
groups were for reasons of poor condition and health or udder problems.
Heifers culled in LB groups were all for fertility problems.

In addition to health problems, heifers fed energy in excess of
requirements were apparently not as efficient in utilization of energy
in feeds offered as heifers deficient in energy. This is probably
another reason why heifers fed excess energy did not yield more milk.
Some of the extra energy was partitioned into body tissue since HE
groups on average gained weight. Failure, however, to find a perfect
correlation between weight change and energy status must be due to
changing body composition and to variation in efficiencies of nutrient
utilization. Calculations indicated that the average HE group heifer
partitioned less than 10% of the excess energy fed to body tissue.

The rest was presumably wasted since it was obviously not partitioned
to the mammary gland. Weight losses in this study were quite similar
to those reported by Gordon (1977) for cows fed low and high amounts of
concentrates and to those reported by Belyea et al. (1975b) for cows in
early lactation fed to meet requirements.

The beneficial effects of feeding limited rations early in lacta-
tion have previously been reported. Cows fed limited grain for the first
45 days postpartum and fed grain ad libitum from day 46 to 180 gave the
greatest amount of milk, fat corrected milk, fat and solids-not-fat
when compared with cows fed ad libitum from parturition (Thomas et al.,
1974). These data however, disagree radically with work by Broster et
a1. (1975) who fbund a marked response over the full lactation to extra
food given in weeks 1 to 4 of lactation in first calf heifers. Broster
et a1. (1975) concluded that level of intake in the first few weeks of

lactation appeared more critical in increasing yield than supplementary

103
feeding fOr the first several weeks of lactation. Heifers in the above
study were fed three times daily and this may have helped utilization
of fOOd offered in early lactation. In addition, Broster had no group
that was restricted in feed and then switched to high energy feeding
to compare with data in the present study.

These milk production and health data strongly suggest that heifers
should be fed rations moderate in energy immediately after calving,
gradually building up their energy intake to a high level by 4-5 weeks
of lactation. Phosphorus should be fed according to recommendations
during this time. These data suggest there can be no justification for
feeding in excess of energy or phosphorus requirements in early lacta-
tion. If rations are fed to these recommendations milk yield should be
optimized and health problems minimized. Management systems are present-
ly available that would enable these recommendations to be followed.

Now, responses of various serum metabolites to energy or phosphorus
status will be discussed. Within the range of phosphorus intake or status
in this study serum P did not reflect either very well. In extreme condi-
tions serum P may decrease by 50%, but in the present study serum P only
decreased by 11% in LP groups. No symptoms of phosphorus deficiency
were noticed. LELP groups in both trials (which were approximately 85%
N.R.C. for P) had the lowest mean serum P concentrations. According to
data of Tumbleson et al. (1973) LELP group values are rather low for 3
year old Holsteins. These authors quoted mean serum P of around 7.0 mg/
d1 fOr heifers for this age. Previous studies which involved experimental
or accidental severe deficiencies reported much lower values. Eckles et
a1. (1935) reported 2.5 mg/dl in phosphorus deficient cows and no diffi-

culty was experienced in producing a state of phosphorus deficiency in

104
any of the animals. This was accomplished by using a ration of prairie
hay (grown on phosphorus deficient soil), and a grain mix low in phos-
phorus. It was calculated that cows producing 9.1 kg of milk per day
were consuming about 19 g or less of phosphorus per day. Palmer et a1.
(1941) reported serum phosphorus values of 2.5-3.0 mg/dl with an average
intake of phosphorus of about 6 g per day, these animals, however, had
very low milk yields. In the field, serum values of 4.0 mg/dl were
reported for yearling heifers fed phosphorus deficient rations (Morrow,
1969). In the present study cows fed phosphorus deficient rations were
consuming 45-65 g/day, considerably more than in the previous studies
cited. They were, however, producing 2 or 4 times as much milk as cows
studied by Eckles et al., (1935) or Palmer et al., (1941). In a recent
study it was concluded that cows fed at or greater than N.R.C. for
phosphorus did not reflect differences in intake very well (Belyea et al.,
1975a). Cows in the present study were at most 15% below N.R.C. require-
ments and serum P concentrations did not indicate that there was a defi-
ciency problem of any proportions. This means that serum P will not
indicate cases of anything but severely inadequate P intake in a practical
situation. Feed analysis and actual calculation of intake and require-
ments would be needed.

Serum Ca was inversely related to serum P. Serum Ca concentrations
generally decrease as phosphorus status and serum P increase. The nega-
tive correlation between Ca and P has been previously reported (Symonds
and Manston, 1974). Serum Ca concentrations were quite stable and
differences between groups were small. In all but the HEHP group in
trial 2, serum Ca concentrations gradually increased as lactation progres-

sed as previously shown by Ward, Blosser and Adams (1952), Belyea et al.,

105
(1975c) and Belyea et al., (1975a). Concentrations in the present study
were comparable to those in other reports cited. In the short term, any-
way, both serum P and Ca did not appear to be detrimentally affected by
the range of P intake in this study. Body weight change, serum glucose,
certain VFA's, insulin and GH all responded to changes in energy status.
If these parameters were measured in a herd with apparent nutritional
problems however, it would be difficult to accurately predict that energy
was limiting or in excess in the rations.

Serum glucose concentrations were a better indicator of energy
status in the first trial than the second. In general, cows fed excess
energy had significantly higher serum glucose levels than those defici-
ent in energy. In the second trial, however, serum glucose increased
also with excess phosphorus. Glucose concentrations have previously
been associated with dry matter intake and estimated net energy intake
in the first 2 months of lactation (Smith et al., 1976), increasing
amounts<xfconcentrates (and therefbre energy) in diets (Blair et al.,
1974; Jenny et al., 1974). In the present studies glucose was correlated
with grain intake (r = 0.21, P < .05) but was not correlated with hay or
corn silage intake. Glucose concentrations, however, did not signifi-
cantly increase or decrease during the firSt 3 months of lactation con-
firming reports of Paterson and Linzell (1974) and Halse et al., (1976)
but contradicting Blom and Halse (1975), Smith et al., (1967) and Gorewit
et a1. (1977) who reported increased glucose concentrations in the first
3-4 months of lactation. These differences are probably due to experi-
mental design, time and frequency of sampling and nutrient regimes. LE
groups were not hypoglycemic and the slightly lower serum glucose concen-

trations of the LE groups did not appear to limit milk production.

106

Not all serum VFA responded to changes in energy status. Serum C2
was not different among groups, neither did regression analysis indicate
a significant relation between C2 and energy status. This agrees with
a previous report that cows fed high or normal levels of grain did not
differ in serum C2 concentrations (Walker and Elliot, 1973). In contrast,
however, other reports (Huber et al., 1969; Evans et al., 1975) found a
20% decrease in C2 concentrations in cows fed high grain, restricted
roughage diets. It is possible that since heifers in the present study
were not roughage restricted that sufficient C2 fermentation occurred
in the rumen in both groups so that C2 levels did not change. C2 values
fer cows in this study are similar to those previously reported for
lactating cows fed normal diets (Huber et al., 1969; Emery et al., 1969;
Annison and Armstrong, 1970; Annison and Linzell, 1974; Baird et al.,
1975; Evans et al., 1975; Schwalm and Schultz, 1976).

Serum C3 increased as energy status increased, however, by analysis
of variance phosphorus groups were also significantly different. Serum
C3 was also positively correlated with grain intake (r = 0.29, P = .05).
This confirms previous work where serum C3 increased 45% with a low
roughage diet (Evans et al., 1975) but does not agree with another study
(Walker and Elliot, 1973). Means in the present study were lower than
in the studies just cited (also Bickerstaffe et al., 1974 and Baird et
al., 1975). Differences between heifers fed high and lower amounts of
concentrates were presumably due to greater C3 production ruminally and
subsequent greater absorption into the blood. Also, since more glucose
would probably be absorbed from the intestines with concentrate by-pass,
gluconeogenesis may have decreased somewhat and C3 may have been spared,

hence increasing in serum.

107

Serum isoC4 did not reflect energy status but was increased in LP
groups compared with HP groups. Serum isoC4 has previously been shown
to increase 56% when roughage was restricted and replaced with grain
(Evans et al., 1975). This increase was possibly due to increased valine
degradation in the rumen. Values for isoC4 in the present study were
about one tenth as high as those reported by Evans et a1. (1975) and its
relationship to energy or phosphorus status is not clear. It appears
that when HE was combined with excess phosphorus, isoC4 levels were
depressed, but when HE was combined with phosphorus below or around re-
quirements isoC4 was increased (over either of LB groups). This accounts
for the significant interaction term in the analysis of variance.

It is not apparent that serum VFA in these studies affected per—
formance of different treatment groups in early lactation. However,
their relationship to serum insulin was of interest and will be discus-
sed later in this section.

In the first trial insulin was a very sensitive indicator of energy
status. Differences in energy groups were easily noticeable. Cows fed
excess energy had serum insulin concentrations three times as high as
cows fed deficient energy. These differences were in part due to excess
energy but were also accentuated by sampling close to feeding time.
Response to high energy or grain feeding and increases in serum insulin
have been previously reported (Walker and Elliot, 1973; Blair et al.,
1974; Jenny et al., 1974 and Evans et al., 1975). Increases in insulin
postprandially are also well documented (Trenkle, 1971a; McAtee and
Trenkle, 1971a; Ross and Kitts, 1973; Bassett, 1974a, 1974b, 1975; Jenny

and Polan, 1975; Evans et al., 1975).

108

In the second trial, although serum insulin increased with energy
status and differences between HE and LE groups were significant, they
were not as great as in the first trial (increases of 28% compared to
300-400%). This may partly be due to feeding schedule changes between
first and second trials resulting in insulin samples being taken many
hours postfeeding. This may also account for why values in the second
trial are low compared with other reports, even for primiparous cows
(Koprowski and Tucker, 1973a; Trenkle, 1972; Jenny and Polan, 1975;
Walker and Elliot, 1973; Evans et al., 1975). Values were comparable,
however, to those reported by others (Smith et al., 1976; Schwalm and
Schultz, 1976; Blair et al., 1974; Gorewit et al., 1977).

Insulin has been reported to increase with time after parturition
(Koprowski and Tucker, 1973a; Smith et al., 1976; Gorewit et al., 1977),
in the present studies only the LELP group in the first trial and the
HEHP group in the second trial significantly increased with time, the
rest did not significantly increase or decrease. This would suggest
that insulin may not follow intake or milk yield as well as previously
reported.

In the first trial insulin was negatively correlated with milk
yield (r = -0.l7, P = .05), in the second trial the correlation was not
significantly different from zero (r = 0.13, NS). Other studies which
have shown or implied a negative relationship between insulin and milk
yield are numerous (Kronfeld et al., 1963; Schmidt, 1966; Koprowski and
Tucker, 1973a; Jenny et al., 1974). In contrast, a positive relationship
is implied by data from Baldwin et a1. (1973) in which cows treated with
insulin did not experience a fall in production whereas milk yields of

untreated controls fell 18% in the same period. Apparent contradictions

109
among these trials may be due to variation in energy status and glucose
concentrations of the animals and other unknown factors. At any rate,
in the present study, insulin was not well correlated with milk yield
in either experiment. This suggests that insulin concentrations do not
directly influence milk yield in lactating cows.

In the first trial insulin was negatively correlated with milk fat
percent, (r = —O.33, P = .05) confirming reports by others (Walker and
Elliot, 1973; Blair et al., 1974; Jenny et al., 1974). In the second
trial, insulin and milk fat were not significantly correlated. Insulin,
as expected, was related to other variables, which will be discussed
further later in this section.

Serum GH, although taking some weeks, did seem to respond to in-
creasing energy status by decreasing in concentration in an opposite
manner to the response of insulin. These data suggest that in response
to excess energy supply metabolic demands of HE cows declined and GH
decreased in response to this. In LE heifers on the other hand, meta-
bolic demands of lactation in the face of deficient energy supplies
were considerable and GH remained at normal postpartum levels possibly
to facilitate endogenous energy mobilization.

GH has previously been reported to decline as lactation advanced,
presumably due to decreasing metabolic demands as feed intake increased
and milk yield declined (K0prowski and Tucker, 1973a; Halse et al., 1976;
Smith et al., 1976). None of the cows in the above studies were fed
restricted diets. LE groups in the present study did not significantly
increase or decrease, suggesting that GH only declines in response to
increasing and sufficient energy intake. GH was negatively correlated

with dry matter, estimated net energy intake and digestible organic

110

matter intake (Bassett et al., 1971; Smith et al., 1976). Increasing
GH has been previously associated with increasing metabolic demands
(Bassett, 1975; Trenkle, 1978). Data from the present studies support
the hypothesis that GH increases and insulin decreases in time of
greater metabolic demand. It is possible that GH and insulin responded
as they did to maintain endogenous energy supplies. In HE groups, higher
insulin and lower GH would favor substrates moving into tissue for syn-
thesis and storage, whereas in LB groups, low insulin and higher GH
would favor endogenous mobilization of substrates necessary to maintain
milk production. The fact that insulin responds so quickly and GH much
more slowly to dietary changes suggests that insulin responds to energy
intake whereas GH responds to changes in energy status. Undoubtedly,
g1ucagon,cnrticoidsand other hormones are involved in this regulation
of nutrient supply. Knowledge of optimum hormonal status for milk
production presents opportunities to improve milk yield in future work.

GH was not correlated with serum glucose concentrations (r = -0.075),
Halse et al. (1976) did not find a high correlation between these vari-
ables either (r = -0.15, P = .05). GH was negatively correlated with
milk yield (r = -0.23, P = .05), this does not agree with results from
Koprowski and Tucker (1973a) who reported no significant correlation
between GH and milk yield early in lactation, (up to 12 weeks) although
correlations were largely negatively, especially after 24 weeks post-
partum. GH concentrations in this study were comparable to those reported
by other workers for early lactation cows, (Koprowski and Tucker, 1973a;
Convey, 1974; Reynaert et al., 1976; Halse et al., 1976) but much lower

than those reported by Smith et a1. (1976).

111

Results from this investigation help explain regulation of serum
insulin. Physiological concentrations of serum C3, C4 and isoC4 (but
not C2), may be involved in insulin regulation. These data are supported
by the studies of Ross and Kitts (1973) who concluded that circulating
C4 and isoC4 were the most important VFA with respect to postprandial
insulin concentrations in sheep. VFA concentrations from that study
were physiological and very similar to levels in the present study.

Why GH was retained and energy status, phosphorus status, milk yield,
acetate and glucose were deleted from the equation is not clear. There
is a possibility that GH and insulin may directly influence each other,
perhaps at the receptor level. The fact that GH responds so slowly,
however, and insulin responds so quickly to energy or metabolic status
changes weakens the suggestion of a direct effect. If GH and insulin
indirectly affect each other, the mediator in this indirect effect is
not obvious. It cannot be glucose since GH and glucose were not corre-
lated and glucose was deleted from the equation anyway. Neither is it
likely to be glucose entry rate since this is highly related to milk
yield and GH was not strongly correlated with milk yield and milk yield
was also deleted from the equation. Alternative suggestions are that
GH is reflecting the amount of protein digested. Bassett et a1. (1971)
found a very high correlation between insulin concentrations and protein
digested in the small intestine, additionally, insulin and GH were highly
negatively correlated. It is however, not likely that in the present
study that there was enough difference in protein reaching the lower gut
to account for these changes in insulin and GH concentrations. Finally,
it may be that GH varies with insulin merely because GH is decreasing

and insulin increasing with advancing lactation.

112

Evidence from these experiments suggest that energy and phosphorus
status did not affect postpartum reproductive function in any dramatic
way. Heifers fed HE rations ovulated for the first time only 4.5 days
prior to heifers fed LE rations and only by regression analysis did this
relationship approach significance (P = .07). There were no significant
differences in time postpartum to reach 3 ng/ml progesterone, LP groups,
however, took 9 days longer than HP groups. Heifers usually ovulated
once before reaching this level of progesterone, average time between
these events were 18 days. Using the criteria of Folmann et a1. (1973),
Corah et a1. (1974) and Rosenberg et a1. (1977) this would mean that
only the second ovulation in these heifers would be viable for conception.
In order to test the hypothesis that serum progesterone concentrations
should be at least 3 ng/ml for conception to occur, data from this study
were examined. Progesterone peaks preceding inseminations resulting in
pregnancy or non-pregnancy were tabulated. As a result 41 and 16 peaks
occurring prior to non-pregnancy and pregnancy were recorded. Means were
similar, 5.74 and 5.07 ng/ml, respectively. However, no heifer with a
serum progesterone peak below 2.7 ng/ml became pregnant to the subsequent
insemination. Peak progesterone before nonconceiving inseminations
ranged from 0.7-10.71 ng/ml progesterone. ‘Thus, although there appeared
to be no guarantee that 3 ng/ml would result in pregnancy, there was a
suggestion that at least that much was needed to permit conception, con-
firming reports by previously cited authors that a critical amount of
progesterone is needed for conception. Heifers in the study of
Rosenberg et a1. (1977) ovulated fbr the first time at 27 days postpartum
(some time after multiparous cows) and this compares well with data from

the present study. Also, heifers in Rosenber'g study reach 3.4 ng/ml of

113
progesterone at a time comparable to those in the present study (40-50
days). When best, worst and control genetic groups were examined, days
to first ovulation were 27.0, 27.5 and 25.9 days, respectively. Thus,
as far as designated genetic potential there was very little difference.

There were no differences in days to conception which could be
attributed to energy or phosphorus status or intake differences. This
confirms one other study with dairy cows (Gardner, 1969b) but contra-
dicts several other reports where deficient energy rations did lengthen
days nonpregnant (Wiltbank et al., 1962, 1964; McClure, 1968a; Dunn et
al., 1969; Bellows et al., 1972; Drew et al., 1976; Randel and Welker,
1977; Macfarlane et al., 1977). These studies, however, were either
conducted with beef cows or dairy cows under marginal nutritional condi-
tions where energy status was at a more critical level.

No evidence was found that lower phosphorus intake delayed preg-
nancy which confirms previous work in which experimental phosphorus defi-
ciency, (where blood values of P were half normal), did not impair ovula—
tion or conception (Palmer et al., 1941). Severe phosphorus deficiency
has been implicated with fertility problems (Eckles et al., 1935;

Morrow, 1969) but heifers in the present experiment were on average, at
N.R.C. recommendations for P. Serum P concentrations within a wide range
have not been shown to influence fertility (Parker and Blowey, 1976;
Rowlands et al., 1977). Experiments with beef cows where phosphorus
supplementation improved fertility (Williams et al., 1971; Taylor et al.,
1976; Ritson et al., 1971) suggest that published P requirements for
beef cows are too low and also that cows were supplied with greatly
insufficient phosphorus to start with. Other studies with similarly

poor nutritional conditions have not shown improvements in fertility

114
after phosphorus supplementation (Ward, 1968; Teleni et al., 1977).
There was no evidence in the present study that excessive phosphorus
was associated with infertility as suggested by Hewett (1974). In all
cases, if anything, phosphorus at recommended levels caused problems.
In this study serum P concentrations were negatively correlated with
days to 3 ng/ml serum progesterone (r = -0.34, P = .05). Although
serum P was not correlated to days to ovulation, ovulation and time to
reach 3 ng/ml progesterone were highly related.

There were no significant differences among treatment groups in
inseminations per conception, although, three times as many HE heifers
did not become pregnant compared with LE heifers. These differences
in pregnancies may have been the result of management rather than nutri—
tional problems, since some heifers became pregnant after treatment
ended.

There were no dramatic differences in number or percent undetected
estrus among treatment groups. The trend was that groups fed excess
phosphorus had slightly less undetected estrus than heifers fed to re-
quirements. Phosphorus deficiency has previously been associated with
silent estrus (Palmer et al., 1941), temporary sterility (Sheehy, 1946)
and 'anestrus or estrus with repeated failures to conceive after service',
(O'Moore, 1950). It must be remembered, however, that these were appar-
ently cases of severe deficiency where serum P concentrations were de-
pressed as much as half. Morrow (1969) did not observe silent estrus
in heifers in his study.

There was no evidence that treatments affected subsequent ovulations
once cyclic behavior was established. Intervals to second and third

ovulations were normal (20.6 and 21.2 days). Undernutrition has been

115

quite severe (85% of maintenance) in cases where estrous cycle length
was altered (Hill et al., 1970). HE groups had 0.4 fewer ovulations
that LE groups in the 15 week experimental period. This difference
approached significance at the P = 0.1 level. At any rate decreases
in ovulations in HE groups could not be attributed to pregnancies since
more HE group heifers were open at this time than LE groups (16 and 13
open, respectively).

As previously summarized in the literature review several hypotheses
exist as to how nutrition or other factors influence reproduction.
Several workers have suggested that increases in body weight were not
important for ovulation and conception (King, 1968; Schilling and England,
1968; McClure, 1970; Folman et al., 1973; Wiltbank, 1974; Youdan and King,
1977; Holness et al., 1978). In the present study there was little
evidence for differences between groups in time to first ovulation. Thus,
although on average HE groups gained weight and ovulated a few days sooner
than LE groups, which on average lost weight, there was much variability
and evidence is certainly not conclusive. This confirms reports by
several workers (Gardner, 1969b; Boyd, 1972; Broster, 1973; Downie and
Gelman, 1976 and Stevenson, 1977) that body weight change does not play
an important role in postpartum reproductiOn under normal circumstances.

High milk production has often been cited as a cause of delayed first
estrus and ovulation in postpartum cows (Morrow et al., 1966; Hewett,
1968; Whitmore et al., 1974; Brauner, 1975; Stevenson, 1977). Since
there was no significant difference in milk production between energy
groups for the first 8 weeks of lactation this cannot be used as a reason
for HE and LE groups differing in ovulation times (which they did not,

significantly). There were no differences among phosphorus groups in

116
time to first ovulation even though LP groups gave significantly more
milk from weeks 5-12 than HP groups. This confirms conclusions of King
(1968) and Simpson (1972). Heifers in the present trials, however, were
not excessively high producers (average 305 day yield was 5908 kg) and
this may be one reason for lack of a correlation between these variables.

Francos (1968, 1969) suggested that 3.3% milk fat was required for
optimum fertility. Since all groups, on average, in this study met this
criterion, low milk fat and not enough roughage in the ration cannot be
cited as a reason for fertility problems.

Hypoglycemia has received much attention in association with repro-
ductive hypofunction (McClure, 1965, 1967, 1968a, 1968b, 1972; Howland
et al., 1966). In neither trial in this study was there very much dif-
ference in glucose concentration between groups and neither would they
be considered hypoglycemic. In this study the only data suggesting that
glucose might be involved was that serum glucose was negatively corre-
lated (r = -0.33, P = .05) with days to reach 3 ng/ml progesterone which,
as already discussed, may be involved in conception. Similarly,
Oxenreider and Wagner (1971) found a negative correlation between glucose
and postpartum interval to occurrence of a 10 mm follicle and ovulation
in primiparous dairy cows. McClure and Payne (1978) found that non-
return rates increased as blood glucose concentrations increased. In
contrast, Blowey and Davis (1973) and Hewett (1974) fbund no relationship
between glucose concentrations and fertility after extensive study of
many herds.

Most reports that energy status might alter levels of reproductive
hormones can neither be confirmed nor denied since only progesterone was

measured in these experiments. Since there were no dramatic differences

117

in other reproductive functions measured it seems unlikely that proges-
terone was altered significantly by either energy or phosphorus status.
Previous reports would suggest that undernutrition or energy deprivation
had to be quite severe to effect changes in progesterone concentrations
or corpus luteum size and function (Hill et al., 1970; Donaldson et al.,
1970; Gombe and Hansel, 1973). In fact recent evidence, where beef
heifers were fed one-third of recommended energy, showed no differences
in blood levels of progesterone or LH, number of follicles nor folli-
cular or luteal volume. However, the ovary containing the corpus luteum
was 57% larger in heifers fed adequate energy than in those restricted
in energy intake (Spitzer et al., 1978). Reduced pregnancies in heifers
restricted in energy appeared not to be a fertilization failure, but due
to some later causative factor.

Finally, there was no evidence that heifers fed LE in this present
study had a higher incidence of fertility disorders compared with HE

groups as suggested by Francos (1974).

CONCLUSIONS

Cows supplied with excess phosphorus yielded less milk throughout
the first 3 months of lactation than those fed just below recommendations.
Phosphorus in excess of recommendations may have had negative effects
on milk production. Cows fed excess energy did not begin to yield more
milk than cows fed deficient energy until the fifth week of lactation.
By eight weeks cows fed excess energy had greater persistency than those
deficient in energy. It is not clear why excess energy did not improve
milk production further. Health may have contributed to this since cows
fed high energy rations had almost twice the incidence of disease of
those fed deficient energy. Incidences of mastitis and metritis were
similar among energy groups. Heifers fed excess energy, however, had
more other types of disease and also higher rectal temperatures than
those fed deficient energy rations. All heifers, however, had elevated
temperatures at the beginning of lactation. In addition, three times
more heifers were culled after first lactation from groups fed excess
energy fbr three months postpartum compared with those fed deficient
energy. Differences between phosphorus groups in milk production persisted
throughout the 305 day lactation. Whether these differences in culling
and total production were a result of treatments imposed in early lacta-
tion is not know.

It is suggested that heifers should be fed rations moderate in
energy immediately after calving, gradually building up their energy

118

119
intake to a high level by 4-5 weeks of lactation. During this time
phosphorus should be fed at recommended levels. These observations con-
cerning negative effects of excess phosphorus and energy on production
and health warrent further investigation.

Body weight, serum glucose, insulin and propionate increased with
increases in energy status. None of these alone, except postprandial
insulin concentrations, would be suitable for diagnostic purposes, how
ever, since differences would be too small to detect by sampling just a
few animals. Serum growth hormone, after some weeks, seemed to respond
to increasing status by significantly decreasing. Growth hormone
decreased in cows fed excess energy, possibly due to declining metabolic
demands. By eight weeks postpartum growth hormone was significantly
lower in cows fed excess energy. In cows fed deficient energy growth
hormone did not significantly change. Serum acetate was not different
among groups. Serum butyrate and isobutyrate were not consistently re-
lated with energy or phosphorus status.

Serum phosphorus was the only indicator of phosphorus status and
it showed that a marginal phosphorus deficiency was accomplished at
least in the low energy, low phosphorus groups. Serum calcium was in-
versely related to serum phosphorus.

Results from this investigation help explain regulation of serum
insulin. Physiological concentrations of serum prOpionate, butyrate,
isobutyrate and growth hormone may be involved in insulin regulation.
Whether growth hormone has a direct or indirect effect on insulin is
not known. These parameters only accounted fbr 26% of variance in serum
insulin concentrations and thus were not the only variables involved but

these data add to evidence implicating serum VFA in insulin regulation.

120
Energy and phosphorus status did not affect postpartum reproductive
function significantly. Progesterone data indicated that no heifer with
peak progesterone prior to insemination below 2.7 ng/ml conceived to that
insemination. Days to reach 3 ng/ml serum progesterone was negatively
correlated with serum phosphorus and glucose. Examination of estrous
cycles and pregnancy data indicated that milk yield, body weight change

and hypoglycemia did not affect postpartum reproduction.

APPENDIX

121

Table Al.——Composition of grain mixes (%).

 

 

 

High Energy Low Energy
Component High P ng_g_ High P £2!_E
Shelled Corn 73.9 74.3 -- 1.8
Soybean Meal 18.8 18.8 86.1 86.1
Molasses 5.0 5.0 5.0 5.0
Dicalcium Phosphate 1.2 -- 6.0 --
Limestone 0.5 1.3 0.2 4.4
Trace Mineral Salt ' 0.6 0.6 2.7 2.7

 

 

 

 

Total 100.0 100.0 100.0 100.0

 

.mwmmn App m :o pommoumxo mozam> w no paw a .mz H~<

122

 

 

 

 

 

 

mam.o -- ao.~ oo.om .axa cea-VA\N~\m omaaam aaou
omm.o -- ac.a oo.mm a5\am\m-VA\eH\m amaaam :aou
omm.o H.NH mo.a oo.ma ak\ma\m-en\km\~ amaaam :aou
oom.o o.~H mo.a oo.aa ek\oN\~-v5\HM\H amaaam :aou
wm~.o o.mH mo.a oo.oe ek\~m\a-mn\m\oH amaaam auou
oom.o m.m~ ma.a oa.wm .axa ega-VA\a~\m am:
Ham.o o.a~ ma.a om.om «A\om\~-mA\m\oH Ra:
oom.o m.ma 4H.H co.oo pacemaoace omaaxaz
ome.H H.ma mm.a oo.wm pacemaoase aaaao who:
mwo.o a.ma am.~ oo.am pacemaoage aaaao mama
mAA.H n.54 mo.~ oo.ww uaonwaoasp aaaau arms
kmm.o a.wa om.a oo.ow pacemaoaae guano aam:
omo.o v.8H om.~ oo.km pacemaoane guano azm:

flag flaw. .Idmmwmmmmq may com: aaaa> aouan eoaa

msuonmmonm :fiououm ovsuu Smuocm poz uouum: xun

 

.H Huang you
oxmucfi monogamocm paw oxmucw mz mamasofimo ou pom: monam> msuonmmozm paw Amnoco no: .uouuma anon-.~< canny

123

.mflmmp xuw m :o commoumxo mosdm> x no van a .mz qq<

 

 

 

 

 

 

am.o o.o~ ao.~ m.m~ .axa eaa-oA\oH\~ awafiam anon
aa.o «.ma mo.a m.e~ oA\m\~-mA\w\HH amaqam :aoo
w~.o a.mH mH.H m.mm .axo e=0-65\ma\v am:
om.o m.a~ mH.H v.Hm ck\aa\e-m5\w\aa Ra:
mo.o a.om AO.N m.om pacemaoaee aaaao aqma
mm.a G.Hm No.~ ~.om “nonmaoane :aaao amma
mm.o o.AH mm.~ w.aw .axo eaa-oA\wH\H eaaau mam:
Ne.o m.oH o~.~ m.ww GA\AH\H-mA\w\HH caaau aamm
wo.o m.ma mm.~ w.aw .axo eaa-cA\wH\fi aaaau arm:
mv.o c.- 6H.~ m.mw GA\AH\H-mA\m\H~ aaaau azmz

flag “we .Iflwmwmmmmq flag cam: aaaa> maaaa eaaa

monogamonm :flouonm opsuu Amuocm uoz Houumz xun

 

.N Hmfiuu How
oxmucfi monogamonm use oxmucfl mz oumfisofimo ou pom: mo=Hm> msuonmmonm one xwuoco no: .Houuws knaun.m< oHQMH

124

Table 4.--Simple correlations from both trials.

 

 

 

Variables r

E Status x P Status 0.64
E Status x Glucose 0.30
E Status x Insulin 0.38
E Status x Fat % -0.36
E Status x Serum P -0.27
P Status x Serum P 0.17
P Status x Insulin 0.20
P Status x Milk -0.23
P Status x Protein % 0.33
Serum Ca x Serum P -0.43
Serum Ca x Glucose -0.4l
Serum Ca x Fat % 0.17
Serum P x Glucose 0.46
Serum P x Fat % 0.40
Glucose x Fat % -0.35
Glucose x Wt. change 0.33
Glucose x Protein % 0.18
Insulin x Fat % -0.32
Insulin x Protein % 0.26
Milk x Wt. change -0.42
Fat % x Wt. Change -0.41
Temperature x GH* 0.22
Temperature x Fat %* 0.28
Temperature x C2* -0.28
GH x Serum Ca* -0.18
E Status x C3* 0.28
E Status x C4* 0.35
Serum P x Temperature* -0.25
Fat % x isoC4* -0.20

 

*These variables from trial 2 only, (P f_.05), (r = 90-144). All other
variables reported from both trials combined (P §_.05), (r = 288, except
where wt. change used, r = 48).

BIBLIOGRAPHY

BIBLIOGRAPHY

Annison, E.F. and D.G. Armstrong. 1970. Volatile fatty acid metabolism.
Physiology of Digestion and Metabolism in the Ruminant. A.T.
Phillipson (Ed.) Oriel Press Ltd. Newcastle upon Tyne, England.

P. 422.

 

Arije, G.R., J.N. Wiltbank and M.L. Hopwood. 1974. Hormone levels in
pre- and postparturient beef cows. J. Anim. Sci. 39:338.

Armstrong, D.V., L.D. Brown, J.W. Thomas and S.M. Getty. 1966. High
level grain feeding and herd health. J. Dairy Sci. 49:730. (Abstr.)

Baile, C.A., Z. Glick and J. Mayer. 1969. Effects of secretin and chole-
cystokinin-pancreozymin in pancreatic juice and insulin secretion
of goats. J. Dairy Sci. 52:513.

Baird, G.D., H.W. Symonds and R. Ash. 1975. Some observations on meta-
bolite production and utilization in vivo by the gut and liver of
adult dairy cows. J. Agric. Sci., Camb. 95:281.

Baldwin, R.L., J.R. Reichl, 8. Louis, N.E. Smith, Y.T. Yang and E. Osborn.
1971. Effects of age, pregnancy and lactation on rat, guinea pig
and cow adipose enzyme activities and cow adipose metabolism. J.
Dairy Sci. 56:340.

Bassett, J.M. 1974a. Diurnal pa-terns of plasma insulin, growth hormone,
corticosteroid and metabolite concentrations in fed and fasted
sheep. Australian J. Biol. Sci. 27:167.

Bassett, J.M. 1974b. Early changes in plasma insulin and growth hormone
levels after feeding in lambs and adult sheep. Austrialian J.
Biol. Sci. 27:157.

Bassett, J.M. 1975. Dietary and gastro-intestinal control of hormones
regulating carbohydrate metabolism in ruminants. Digestion and
Metabolism in the Ruminant. I.W. McDonald and A.C.I. Warner (Eds.)
Univ. of New England Publishing Unit, Armidale, N.S.W., Australia.
P. 383.

 

 

Bassett, J.M., R.H. Weston and J.P. Hogan. 1971. Dietary regulation of
plasma insulin and growth hormone concentrations in sheep.
Australian J. Biol. Sci. 24:321.

125

126

Bellows, R.A., L.W. Varner, R.E. Short and O.F. Pahnish. 1972. Gesta-
tion feed level, calf birth weight and calving difficulty. J.
Anim. Sci. 35:185. (Abstr.)

Belyea, R.L., G.B. Coppock and G.B. Lake. 1975a. Effects of silage
diets on health, reproduction, and blood metabolites of dairy
cattle. J. Dairy Sci. 58:1336.

Belyea, R.L., G.B. Coppock, W.G. Merrill and S.T. Slack. 1975b. Effects
of silage based diets on feed intake, milk production and body
weight of dairy cows. J. Dairy Sci. 58:1328.

Belyea, R.L., G.A. Martz and R. Ricketts. 1975a. Plasma metabolites in
early postpartum cows. J. Anim. Sci. 41:343. (Abstr.)

Benjaminsen, E. 1977. Plasma progesterone and ovarian findings related
to plasma glucose and acetoacetate in the postpartum period in
the cow. Nordisk Verterinaer Medecin. 29:337.

Bickerstaffe, R., E.F. Annison and J.L. Linzell. 1974. The metabolism
of glucose, acetate, lipids and amino acids in lactating dairy
cows. J. Agric. Sci. (Camb.) 82:71.

Blair, T., D.A. Christensen and J.G. Manns. 1974. Performance of lacta-
ting cows fed complete pelleted diets based on wheat straw, barley
and wheat. Can. J. Anim. Sci. 54:347.

Blom, A.K. and K. Halse. 1975. Corticosteriods in nocturnal blood plasma
of cows in the field related to stage of lactation and plasma ace-
toacetate. Acta Endocrin. 78:306.

Blowey, R.W., D.W. Wood and J.R. Davis. 1973. A nutritional monitoring
system for dairy herds based on blood glucose, urea and albumin
levels. The Veterinary Record. 82:691.

Bolenbaugh, D.L. 1977. The effects of various dietary levels of potas-
sium on subsequent parturition and lactation in the dairy cow.
M.S. Thesis, Michigan State University, E. Lansing.

Bourne, R.A. and H.A. Tucker. 1975. Serum prolactin and LH responses
to phot0period in bull calves. Endocrin. 97:473.

Bowden, D.M. 1971. Non-esterified fatty acids and ketone bodies in
blood as indicators of nutritional status in ruminants: A review.
Can. J. Anim. Sci. 51:1.

Boyd, H. 1972. Weight change and fertility in one herd of dairy cattle.
The Veterinary Record. 91:193.

Brauner, P. 1975. Relationships between the milk performance and the
conception capacity in Czech red-spotted cattle. Acta Vet. Brno.
44:23.

127

Broster, W.H. 1972. Effect of milk yield of the cow on the level of
feeding during lactation. Dairy Sci. Abstr. 34:265.

Broster, W.H. 1973. Liveweight change and fertility in the lactating
dairy cow: A review. The Veterinary Record. 93:417.

Broster, W.H., V.J. Broster, T. Smith and J.W. Siviter. 1975. Experi~
ments on the nutrition of the dairy heifer IX. Food utilization
in lactation. J. Agric. Sci. (Camb.) 84:173.

Carstairs, J.A. 1975. Postpartum reproduction and metabolites in the
cow as affected by energy and phosphorus intake. M.S. Thesis,
Michigan State University, E. Lansing.

Chang, C.H., D.M. Henricks, T. Gimenez, A.R. Ellicott and K. Cruse.
1977. Plasma glucose, total free fatty acids and insulin in
lactating and non-lactating beef cows. Am. Soc. Anim. Sci.
Meeting Abstracts, P. 144.

Convey, E.M. 1974. Serum hormone concentrations in ruminants during
mammary growth, lactogenesis and lactation: A review. J. Dairy
Sci. 57:905.

Corah, L.R., A.P. Quealy, T.G. Dunn and C.C. Kaltenbach. 1974. Prepartum
and postpartum levels of progesterone and estradiol in beef heifers
fed on two levels of energy. J. Anim. Sci. 39:380.

Davis, S.L. 1972. Plasma levels of prolactin, growth hormone, and insu-
lin in sheep following the infusion of arginine, leucine and
phenylalanine. Endocrin. 91:549.

Dawson, F.L.M. 1972. Fertility and production in dairy cows. New
Zealand Vet. J. 20:168.

Donaldson, L.E., J.M. Bassett and G.D. Thorburn. 1970. Peripheral plasma
progesterone concentrations of cows during puberty, oestrus cycles,
pregnancy and lactation and the effects of undernutrition or exo-
genous oxytocin on progesterone concentration. J. Endocrinol. 48:
599.

Downie, J.G. and A.L. Gelman. 1976. The relationship between changes in
bodyweight, plasma glucose and fertility in beef cows. The
Veterinary Record. 99:210.

Drew, 8., D.F. Wishart and D. Thomson. 1976. The effect of nutrition on
the fertility of dairy heifers. Anim. Prod. 22:145 (Abstr.)

Dunn, T.G., J.E. Ingalls, D.R. Zimmerman and J.N. Wiltbank. 1969. Repro-
ductive performance of 2-year-old Hereford and Angus heifers as

influenced by pre- and post-calving energy intake. J. Anim. Sci.
29:719.

128

du Toit, P.J., A.I. Malan and J.W. Groenewald. 1934. Minimum mineral
requirements of cattle. Onderstepoort J. Vet. Sci. and Anim.
Ind. 2:565.

Echternkamp, S.E. and W. Hansel. 1973. Concurrent changes in bovine
plasma hormone levels prior to and during the first postpartum
estrous cycle. J. Anim. Sci. 37:1362.

Eckles, C.H., R.B. Becker and L.S. Palmer. 1926. A mineral deficiency
in the rations of cattle. Univ. of Minn. Agr. Exp. Station Bull.
229:1.

Eckles, C.H., L.S. Palmer, T.W. Gullickson, C.P. Ritch, W.L. Boyd, L.
Bishop and J.W. Nelson. 1935. Effects of uncomplicated phos-
phorus deficiency on estrous cycle, reproduction and composition
of tissues of mature dairy cows. Cornell Veterinarian 25:22.

Edgerton, L.A. and H.D. Hafs. 1973. Serum luteinizing hormone, prolac-
tin, glucocorticoid and progestin in dairy cows from calving to
gestation. J. Dairy Sci. 56:451.

Emery, R.S., H.D. Hafs, D. Armstrong and W.W. Snyder. 1969. Prepartum
grain feeding on milk production, mammary edema and incidence of
diseases. J. Dairy Sci. 52:345.

Erecinska, M., M. Stubbs, Y. Miyata, C.M. Ditre and D.F. Wilson. 1977.
Regulation of cellular metabolism by intracellular phosphate.
Biochem. Biophys. Acta. 462:20.

Erfle, J.D., L.J. Fisher and F.D. Sauer. 1974. Interrelationships be-
tween blood metabolites and an evaluation of their use as criteria

of energy status of cows in early lactation. Can. J. Anim. Sci.
54:293.

Evans, E., J.G. Buchana - Smith and G.K. Macleod. 1975. Postprandial
patterns of plasma glucose, insulin and volatile fatty acids in
ruminants fed low- and high-roughage diets. J. Anim. Sci. 41:1474.

Fisher, L.J., P.E. Donnelly, J.B. Hutton and D.M. Duganzich. 1975. Rela-
tionships between levels of feeding and certain blood metabolites
in dairy cows in mid-lactation. J. Agric. Sci. 84:29.

Fiske, C.H. and Y. SubbaRow. 1925. The colorimetric determination of
phosphorus. J. Biol. Chem. 66:375.

Folman, Y., M. Rosenberg, Z. Herz and M. Davidson. 1973. The relation-
ship between plasma progesterone concentration and conception in
postpartum dairy cows maintained on 2 levels of nutrition. J.
Reprod. Fertil. 34:267.

Francos, G. 1968. The relationship between the milk fat percentage and
fertility in dairy herds. Quart. Israel Vet. Med. Ass. 25:32.

129

Francos, G. 1969. Observations on the relationship between fertility
and butter fat percentage in three dairy herds. Quart. Israel
Vet. Med. Ass. 26:121.

Francos, G. 1970. Observations on the relationship between overfeeding
and the incidence of metritis in cows after normal parturition.
Quart. Israel Vet. Med. Ass. 27:148.

Francos, G. 1974. Possible effect of different nutritional planes on
the incidence of reproductive disorders is Israeli dairy herds.
Quart. Israel Vet. Med. Ass. 31:27.

Francos, G., M. Davidson and E. Mayer. 1977. The influence of some nu-
tritional factors on the incidence of the repeat breeder syndrome
in high producing dairy herds. Theriogenology 7:105.

Gardner, R.W. 1969a. Interactions of energy levels offered to Holstein
cows prepartum and postpartum. I. Production responses and blood
composition changes. J. Dairy Sci. 52:1973.

Gardner, R.W. 1969b. Interactions of energy levels offered to Holstein
cows prepartum and postpartum. II. Reproductive performance.
J. Dairy Sci. 52:1985.

Gombe, S. and W. Hansel. 1973. Plasma luteinizing hormone (LH) and
progesterone levels in heifers on restricted energy intakes. J.
Anim. Sci. 37:728.

Gordon, F.J. 1977. The effect of protein content on the response of
lactating cows to level of concentrate feeding. Anim. Prod. 25:
181.

Gorewit, R.C., P.A. Oltenacu and G.B. Lake. 1977. Serum glucagon, glu-
cose and insulin in Holstein cows over four months of lactation.
Amer. Soc. Anim. Sci. Meeting Abstracts. P. 164.

Guilbert, H.R. 1942. Some endocrine relationships in nutritional repro-
ductive failure (A review). J. Anim. Sci. 1:3.

Guilbert, H.R. and G.H. Hart. 1930. Some effects of varying calcium and
phosphorus intake on the estrous cycle and reproduction in the
rat. Hilgardia 5:101.

Halse, K., A.K. Blom and K. Hove. 1976. Growth hormone related to insu—
lin and sugar in nocturnal blood plasma of lactating cows. Acta
Endocrin. 82:767.

Hart, C.H. and H.R. Guilbert. 1928. Factors influencing percentage of
calf crop in range herds. U. of California, College of Agr. Bull.
458:1.

130

Hart, B. and G.L. Mitchell. 1965. Effect of phosphorus supplementation
on the fertility of an open range beef cattle herd on the Barkly
Tableland. Australian Vet. J. 41:305.

Hecht, D., M.E. Wells, L.J. Bush and G.D. Adams. 1977. Effect of dietary
phosphorus levels on reproductive efficiency in dairy heifers.
Okla. Ag. Exp. Sta. Anim. Sci. Res. Rep. P. 126.

Hertelendy, F., L.J. Machlin and D.M. Kipnis. 1969. Further studies on
the regulation of insulin and growth hormone secretion in the
sheep. Endocrin. 84:192.

Hertelendy, F., K. Takahashi, L.J. Machlin and D.M. Kipnis. 1970. Growth
hormone and insulin secretory responses to arginine in the sheep,
pig and cow. Gen. Comp. Endocrinol. 14:172.

Hewett, G.C. 1968. A survey of the incidence of the repeat breeder cow
in Sweden with reference to herd size, season, age and milk yield.
Brit. Vet. J. 124:342.

Hewett, C. 1974. On the causes and effects of variations in the blood
profile of Swedish dairy cattle. Acta Vet. Scand. Suppl. 50.

Hignett, S.L. 1950. Factors influencing herd fertility in cattle. The
Veterinary Record. 62:652.

Hignett, S.L. 1959. Some nutritional and other inter-acting factors
which may influence the fertility of cattle. The Veterinary
Record. 71:247.

Hignett, S.L. and P.G. Hignett. 1951. The influence of nutrition on re-
productive efficiency in cattle. I. The effect of calcium and
phosphorus intake on the fertility of cows and heifers. The
Veterinary Record. 63:603.

Hill, Jr., J.R., D.R. Lamond, D.M. Henricks, J.P. Dickey and G.D. Niswender.
1970. The effects of undernutrition on ovarian function and ferti-
lity in beef heifers. Biol. Reprod. 2:78.

Holness, D.H., J.D.H. Hopley and D.H. Hale. 1978. The effects of plane
of nutrition, live weight, temporary weaning and breed on the
occurrence of oestrus in beef cows during the postpartum period.
Anim. Prod. 26:47.

Horino, M., L.J. Machlin, F. Hertelendy and D.M. Kipnis. 1968. Effect
of short chain fatty acids on plasma insulin in ruminants and non-
ruminant species. Endocrin. 83:118.

Hove, K. 1974. Nocturnal plasma insulin levels in cows with varying
levels of plasma ketone bodies; relations to plasma sugar and
acetoacetate. Acta Endocrin. 76:513.

131

Howland, B.E., R.L. Kirkpatrick, A.L. Pope and L.E. Casida. 1966. Pitui-
tary and ovarian function in ewes fed on two nutritional levels.
J. Anim. Sci. 94:716.

Huber, J.T., R.S. Emery, J.W. Thomas and I.M. Yousef. 1969. Milk fat
synthesis on restricted roughage rations containing whey, sodium
bicarbonate and magnesium oxide. J. Dairy Sci. 52:54.

Ingalls, W.G., E.M. Convey and H.D. Hafs. 1973. Bovine serum LH, GH and
prolactin during late pregnancy. Proc. Soc. Exp. Biol. Med. 143:
161.

Irvin, R. and A. Trenkle. 1971. Influences of age, breed and sex on plasma
hormones in cattle. J. Anim. Sci. 32:292.

Jenny, B.F. and C.E. Polan. 1975. Postprandial blood glucose and insulin
in cows fed high grain. J. Dairy Sci. 58:512.

Jenny, B.F., C.E. Polan and F.W. Thye. 1974. Effects of high grain
feeding and stage of lactation on serum insulin, glucose and milk
fat percentage in lactating cows. J. Nutrition 104:379.

Jordan, H.N. and R.W. Phillips. 1978. Effect of fatty acids on isolated
ovine pancreatic islets. Am. J. Physiol. 234:El62.

Jordon, W.H., E.B. Hart and A.J. Patten. 1906. A study of the metabo-
lism and physiological effects of certain phosphorus compounds
with milch cows. Amer. J. Physiol. 16:268.

King, J.O.L. 1968. The relationship between the conception rate and
changes in body weight, yield and SNF content of milk in dairy
cows. The Veterinary Record. 83:492.

Kleiber, M., H. G055 and H.R. Guilbert. 1936. Phosphorus deficiency,
metabolism and food utilization in beef heifers. J. Nutrition 12:
121.

Koprowski, J.A. and H.A. Tucker. 1973a. Bovine growth hormone, corti-
coids and insulin during lactation.‘ Endocrin. 93:645.

K0prowski, J.A. and H.A. Tucker. 1973b. Serum prolactin during various
physiological states and its relationship to milk production in
the bovine. Endocrin. 92:1480.

Kronfeld, D.S. 1965. Plasma non-esterified fatty acid concentrations in
the dairy cow: responses to nutritional and hormonal stimuli, and
significances in ketosis. The Veterinary Record. 77-30.

Kronfeld, D.S., G.P. Mayer, J. McD. Robertson and F. Raggi. 1963. De-
pression of milk secretion during insulin administration. J.
Dairy Sci. 46:559.

132

Lane, A.G., J.R. Campbell and G.F. Krause. 1968. Blood mineral composi-
tion in ruminants. J. Anim. Sci. 27:766.

Leaver, J.D. 1977. Rearing of dairy cattle. 7. Effect of level of
nutrition and body condition on the fertility of heifers. Anim.
Prod. 25:219.

Lishman, A.W., S.M.J. Allison, R.L. Butcher, and E.K. Inskeep. 1977.
Release of LH, ovulation and CL function after GNRH in underfed
lactating beef cows primed with FSH. Amer. Soc. Anim. Sci.
Meeting Abstracts. P. 184.

Littlejohn, A.I. and G. Lewis. 1960. Experimental studies of the rela-
tionship between the calcium—phosphorus ratio of the diet and
fertility in heifers: A preliminary report. The Veterinary
Record. 72:1137.

Louis, T.M., H.D. Hafs and B.E. Seguin. 1973. Progesterone, LH, estrus
and ovulation after prostaglandin F20 in heifers. Proc. Soc. Exp.
Biol. Med. 143:152.

Lowman, B.G. and N.A. Scott. 1976. An evaluation of factors influencing
fertility in commercial suckler beef herds. Anim. Prod. 22:158.
(Abstr.)

Macfarlane, J.S., S.H. Somerville, B.G. Lowman and D.W. Deas. 1977.
Effect of nutrition and other factors on the reproductive perfor-
mance of beef cows. Anim. Prod. 24:131. (Abstr.)

Malan, A.I., H.H. Green and J.P. du Toit. 1928. Composition of bovine
blood on phosphorus deficient pasture. J. Agr. Sci. 18:376.

Manns, J.G. 1972. Separation of pancreatic and gut glucagon-like
immunoreactivity (G.L.I.) with observations on plasma concentra-
tions of these hormones during lactation. Can. J. Phys. Pharm.
50:554.

Manns, J.G. and J.M. Boda. 1967. Insulin release by acetate, propionate
and butyrate and glucose in lambs and adult sheep. Amer. J.
Physiol. 212:747.

Manns, J.G., J.M. Boda and R.F. Willes. 1967. Probable role of propionate
and butyrate in control of insulin secretion in sheep. Am. J.
Physiol. 212:756.

McAtee, J.W. 1970. Metabolic regulation of somatotropin and prolactin
secretion in ruminants. Ph.D. Thesis, Iowa State Univ., Ames.

McAtee, J.W. and A. Trenkle. 1971a. Metabolic regulation of plasma in-
sulin levels in cattle. J. Anim. Sci. 33:438.

133

McAtee, J.W. and A. Trenkle. 1971b. Effect of feeding, fasting and in-
fusion of energy substrates on plasma growth hormone levels in
cattle. J. Anim. Aci. 33:612.

McClure, T.J. 1965. A nutritional cause of low non-return rates in
dairy herds. Australian Vet. J. 41:119.

McClure, T.J. 1967. Infertility in mice caused by fasting at about the
time of mating. J. Reprod. Fertil. 13:393.

McClure, T.J. 1968a. Malnutrition and infertility of cattle in Australia
and New Zealand. Australian Vet. J. 44:134.

McClure, T.J. 1968b. Hypoglycemia, an apparent cause of interfility of
lactating cows. Brit. Vet. J. 124:126.

McClure, T.J. 1970. A review of deve10pments in nutrition as it is re-
lated to fertility in cattle: 1964-9. New Zealand Vet. J. 18:61.

McClure, T.J. 1972. Blood glucose and female fertility. The Veterinary
Record. 91:193.

McClure, T.J. and J.M. Payne. 1978. Observations on the first service
non-return rates of some hypoglycemic, concentrate-fed dairy herds.
Australian Vet. J. 54:7.

Meyer, H. 1976. The physiology of magnesium metabolism in ruminants.
In: Magnesium in Ruminant Nutrition. A Symposium. Israel
Chemicals Ltd., Tel-Avid, Israel. P. 35.

Morrow, D.A. 1969. Phosphorus deficiency and infertility in dairy
heifers. J.A.V.M.A. 154:761.

Morrow, D.A., S.J. Roberts, K. McEntee and H.G. Gray. 1966. Postpartum
ovarian activity and uterine involution in dairy dattle. J.A.V.M.A.
149:1596.

Mulvany, P.M. 1977. A body condition scoring technique for use with
British Fresian cows. Anim. Prod. 24:157 (Abstr.)

Noller, C.H., A.G. Castro, W.E. Wheeler, D.L. Hill and N.J. Moeller.
1977. Effect of phosphorus supplementation on growth rate, blood
minerals and conception rate of dairy heifers. J. Dairy Sci. 60:
1932.

N.R.C. 1971. Nutrient requirements of domestic animals. No. 3. Nutrient
requirements of dairy cattle. National Research Council, Washington,
DOC.

O'Moore, L.B. 1950. Aphosphorosis in Ireland. Nature, London 165:192.

134

Oxenreider, S.L. and W.C. Wagner. 1971. Effect of lactation and energy
intake on postpartum ovarian activity in the cow. J. Anim. Sci.
33:1026.

Parker, B.N.J. and R.W. Blowey. 1976. Investigations into the relation-
ship of selected blood components to nutrition and fertility of
the dairy cow under commercial farm conditions. The Veterinary
Record. 98:394.

Palmer, L.S. and C.H. Eckles. 1927. Effect of phosphorus deficient ra-
tions on blood composition in cattle. Proc. Soc. Exp. Biol. Med.
24:307.

Palmer, L.S., T.W. Gullickson, W.L. Boyd, C.P. Fitch and J.W. Nelson.
1941. The effect of rations deficient in phosphorus and protein

on ovulation, estrous and reproduction of dairy heifers. J. Dairy
Sci. 24:199.

Paterson, J.Y.F. and J.L. Linzell. 1974. Cortisol secretion rate, glu-
cose entry rate and the mammary uptake of cortisol and glucose
during pregnancy and lactation in dairy cows. J. Endocrinol. 62:
371.

Peters, R.R. and H.A. Tucker. 1978. Prolactin and growth hormone responses
to photoperiod in heifers. Endocrin. 103:229.

Peterson, R.G. and D.E. Waldern. 1977. A survey of dairy heifers in the
Fraser Valley of British Columbia to determine possible causes of
unsatisfactory reproductive performance. Can. J. Anim. Sci. 57:
395.

Purchas, R.W., K.L. Macmillan and H.D. Hafs. 1970. Pituitary and plasma
growth hormone levels in bulls to one year of age. J. Anim. Sci.
31:358.

Randel, R.D. and G.A. Welker. 1977. Effect of energy intake and once
daily suckling on postpartum interval in Brahman X Hereford
Heifers. Amer. Soc. Anim. Sci. Meeting Abstracts. P. 198.

Remesy, C. and C. Demigne. 1974. Determination of volatile fatty acids
in plasma after ethanolic extraction. Biochem. J. 141:85.

Reynaert, R., W. DeGroot, 5. Marcus and G. Peeters. 1976. Serum growth
hormone and free fatty acid levels in heifers and cows before,
during and after parturition. Annales d'Endoc. (Paris) 37:75.

Reynaert, R., M. DePaepe, S. Marcus and G. Peeters. 1975. Influence of
serum free fatty acid levels on growth hormone secretion in lacta-
ting cows. J. Endocrinol. 66:213.

Reynaert, R., W. Top and G. Peeters. 1977. Bovine serum growth hormone
levels in clinical ketosis. The Veterinary Record. 101:36.

135

Ritson, J.B., L.A. Edye and P.J. Robinson. 1971. Botanical and chemical
composition of a townsville stylo-spear grass pasture in relation
to conception rate of cows. Australian J. Agric. Res. 22:993.

Rosenberg, M., Z. Herz, M. Davidson and Y. Folman. 1977. Seasonal varia-
tions in postpartum plasma progesterone levels and conception in
primiparous and multiparous dairy cows. J. Reprod. Fert. 51:363.

Ross, J.P. and W.D. Kitts. 1973. Relationship between postprandial plasma
volatile fatty acids, glucose and insulin levels in sheep fed dif-
ferent feeds. J. Nutrition. 103:488.

Rowlands, G.J., W. Little and B.A. Kitchenham. 1977. Relationships be-
tween blood composition and fertility in dairy cows - a field
study. J. Dairy Res. 44:1.

Sansom, B.F. 1973. Clinical problems in preventive medicine. Mineral
nutrition and production disease in dairy cows. Brit. Vet. J.
129:207.

Schilling, P.E. and N.C. England. 1968. Some factors affecting repro-
duction in beef cattle. J. Anim. Sci. 27:1363.

Schmidt, G.H. 1966. Effects of insulin on yield and composition of milk
of dairy cows. J. Dairy Sci. 49:381.

Schwalm and Schultz. 1976. Relationship of insulin concentration to
blood metabolites in the dairy cow. J. Dairy Sci. 59:255.

Sheehy, E.J. 1946. Aphosphorosis in cattle in Offaly. Nature, London
157:442.

Siebert, B.D. 1973. The effects of temporal variation in the water cli-
mate of beef cattle reproduction in the seasonally dry tropics
of Australia. Int. J. Biometeor. 17:123.

Simpson, B.H. 1972. The relationship between production and infertility
in dairy cows. New Zealand Vet. J. 20:7.

Smith, R.D., W. Hansel and C.E. COppock. 1976. Plasma growth hormone
and insulin during early lactation in cows fed silage based diets.
J. Dairy Sci. 59:248.

Spitzer, J.G., G.D. Niswender, G.B. Seidel, Jr., and J.N. Wiltbank. 1978.
Fertilization and blood levels of progesterone and LH in beef heifers
on a restricted energy diet. J. Anim. Sci. 46:1071.

Staigmiller, R.B., R.B. Short, R.A. Bellows and J.B. Carr. 1977. Effect
of nutrition on response to exogenous FSH in beef cows. Amer. Soc.
Anim. Sci. Meetings Abstracts. P. 211.

136

Stern, J.S., G.A. Baile and J. Mayer. 1970. Are propionate and butyrate
physiological regulators of plasma insulin in ruminants? Amer. J.
Physiol. 219:84.

Stevenson, J.S. 1977. Luteinizing hormone, progesterone, estradiol, glu-
cocorticoids and changes in energy balance before first ovulation
in postpartum dairy cows. M.S. Thesis, Michigan State University,
E. Lansing.

Symonds, H.W. and R. Manston. 1974. The response of the bovine kidney
to increasing plasma inorganic phosphorus concentrations. Res.
Vet. Sci. 16:131.

Taylor, 8., M. Selke, S. Swingle, J. Phillips and H. Vinson. 1976. Dry-
lot cow-calf research phosphorus levels for beef cows during lacta-
tion and breeding. Technical Release, Animal health and nutrition
division. I.M.C. Corp., Libertyville, IL.

Teleni, E., B.D. Siebert, R.M. Murray and G.D. Nancarrow. 1977. Effects
of supplements of phosphorus or phosphorus and protein on the
ovarian activity of cows fed native pasture hay. Australian J.
Exp. Agric. Anim. Hus. 17:207.

Theiler, A., H.H. Green and P.J. du Toit. 1924. Phosphorus in the live-
stock industry. J. Dept. Agr. (Union of S. Afr.) 8:675.

Theiler, A., H.H. Green and P.J. du Toit. 1928. Breeding of cattle on
phosphorus deficient pasture. J. Agr. Sci. 18:369.

Thomas, J.W., R.S. Emery and L.D. Brown. 1974. Variable grain intake
during two portions of the lactation. J. Dairy Sci. 57:463.

Trenkle, A. 1970a. Effects of short-chain fatty acids, feeding, fasting
and type of diet on plasma insulin levels in sheep. J. Nutrition.
100:1323.

Trenkle, A. 1970b. Plasma levels of growth hormone, insulin and plasma
protein-bound iodine in finishing cattle. J. Anim. Sci. 31:389.

Trenkle, A. 1971a. Postprandial changes in insulin secretion rates in
sheep. J. Nutrition. 101:1099.

Trenkle, A. 1971b. Influence of blood glucose levels on growth hormone
secretion in sheep. Proc. Soc. Exp. Biol. Med. 136:51.

Trenkle. A. 1976. Estimates of the kinetic parameters of growth hormone
metabolism in fed and fasted calves and sheep. J. Anim. Sci. 43:
1035.

Trenkle, A. 1978. Relations of hormonal variations to nutritional
studies and metabolism of ruminants. J. Dairy Sci. 61:281.

137

Trimberger, G.W., H.F. Tyrrell, D.A. Morrow, J.T. Reid, M.J. Wright, W.E.
Shipe, J.T. Merrill, J.K. Loosli, C.E. Coppock, L.A. Moore and
C.H. Gordon. 1972. Effects of liberal grain feeding on health,
reproductive efficiency, economy of milk production and other re-
lated responses of the dairy cow. New York Food Life Sci. Bull.
8:Animal Sci. 1.

Tuff, P. 1923. Osteomalacia and its occurrence in cattle in Norway. J.
Comp. Path and Ther. 36:143. Proc. World's Dairy Congress 2:1494.

Tumbleson, M.E., W.E. Wingfield, H.D. Johnson, J.R. Campbell and C.C.
Middleton. 1973. Serum electolyte concentrations, as a function
of age, in female dairy cattle. Cornell Veterinarian 63:58.

Walker, G.K. and J.M. Elliot. 1973. Effect of roughage restriction on
serum insulin in the dairy cow. J. Dairy Sci. 56:375.

Ward, H.K. 1968. Supplementation of beef cows grazing on veld. Rhod.
J. Agric. Res. 6:93.

Ward, G.M., T.H. Blosser and M.F. Adams. 1952. The relation of prepartal
mineral balances to the occurrence of parturient paresis in dairy
cows. J. Dairy Sci. 35:587.

Whitmore, H.L., W.J. Tyler and L.E. Casida. 1974. Effects of early post-
partum breeding in dairy cattle. J. Anim. Sci. 38:339.

Williams, W.T., K.P. Haydock, L.A. Edye and J.B. Ritson. 1971. Analysis
of a fertility trial with droughtmaster cows. Australian J. Agric.
Res. 22:979.

Wiltbank, J.N. 1974. Management programs to increase reproductive effi-
ciency of beef herds. J. Anim. Sci. 38:58 Supplement 1.

Wiltbank, J.N. 1977. Effect of level of nutrition on growth and repro-
duction of beef female. In: Proc. Georgia Nutr. Conf. P. 139.

Wiltbank, J.N., W.W. Rowden, J.E. Ingalls, K.E. Gregory and K.M. Koch.
1962. Effect of energy level on reproductive phenomena of mature
Hereford cows. J. Anim. Sci. 21:219.

Wiltbank, J.N. W.W. Rowden, J.E. Ingalls and D.R. Zimmerman. 1964. In-
fluence of postpartum energy intake on reproductive performance of
Hereford cows restricted in energy intake prior to calving. J.
Anim. Sci. 23:1049.

Youdon, P.G. and J.O.L. King. 1977. The effects of body weight changes
on fertility during the postpartum period in dairy cows. Brit.
Vet. J. 133:635.

Yousef, I.M., R.S. Emery, E.W. Askew, J.D. Benson, J.W. Thomas and J.T.
Huber. 1969. Comparison of several enzymes and components of milk,
blood and tissues between cows and ewes. J. Dairy Sci. 52:1577.

"I11111111111111“!"1111“