‘l

[tel-V 3 ‘
d- _
I~ "
.

 

)4 a. 4- ‘
a: #¢4¢{3%,“fi° ”
;_ ‘¢."‘ 5190.3“;
- -. 23*“
4

fi. - -
“.1.

«Mo

.
u

’.‘.lf 0'.‘ h N
I
"“‘k

.- v.1. ,A.- mung-
. ' ‘ I‘c‘ V - ‘A
‘ :

‘i’::§v .“g‘z-
J" - V:.. $
1 a“??? $3"? 159;:
gamw'fig
{.‘wfifififi if..." ‘2' "(‘3‘ 1:45?
$4.953??? ? ‘
:Ivmwv“v'
'4'?" I. p
. 4 (0' ‘y. f‘MJ “"5; I?" 2i“:
.p ’ '1‘": H
I? 0’9. 9-!- ' :51 513:?th X
‘ .2 "'4'“
"if; ‘3‘“: fix I?
‘u i’ t “‘
3741 g: $3313..
”9‘1!“ ".0332?’ ‘ .
~ A k .- .‘ ‘
g: m» afiflww ~
‘ am ”3% ‘filiff‘i‘g-wfi.
"fizy‘f’x’

 

6
9“

‘ Q
hw‘fiy-fi
.

a???

g

4

.5:

I ($412,
5.1:”.
”6'1“!"
I «w
.‘ .1 ..

L137:

5?»

‘-‘4‘-‘m- , ,
». ff?-

‘ It
fit

’1‘ In". 7',’
‘ ‘
l} ‘ - ”W '3'" "3.1;? '4“).
n A“ F) V . v". .‘
bush]! 1.: 'L 1 . f; {565%- ‘1!- .I.

F 63:? I? ‘3‘“
S «12“? r:
y .
(3" - i" u)»- ‘02:!“
5,..1" Wm, :ch
.3313" 5

*‘ 1::
3%

-$
‘15:, . ‘

am

W
. r
. - “E...
u
g} - m
L' '
, .
"t"?
i‘n; -
«$13
. .

.‘s
5
ti
nu.

. 1’1

2.?

S.
«It

-'v"0\una( . I.
-'.u-'l_";€,’~, "751.3,,

(,5:
‘1‘, -;
.‘;

.

2.:

:1.
‘. «\g.
3 {a

n h
-h

on
,2,"
(1- 35;:6‘:

- .4:-
_:4;"le

)1- ,,
w

vu-

f." i
If? y f."
. 4
.. ._ ‘ , _ I ’3'” «WWI
fikuH‘r‘r‘ _; 4., . 4 _ .‘ 4 ‘1 a, "9’3 wv‘F/YSI
I T25; g 4 ,4 «2‘: I «5‘5
4 '~ ’3‘
, .

W

5!
.

 

 

 

llllllllllllllllllllllllllllllll

llllllll \lllllllllllllllllllllllll

Thtlfl-‘fi 31
LIBRARY
Michigan State
University

 

This is to certify that the

dissertation entitled

THE EFFECT OF LNG-TERM HEAT
ACCLIMTIZATIN N THE CHARACTERISTICS
OF SEHEN IN THE HHITE LEGHORN ROOSTER

presented by

MSTAFA IBRAHIH ZAKI

has been accepted towards fulfillment
of the requirements for

 

Ph.D. degree in AFN-M] SCience
MajcMofessor
Date ML

 

utu.‘.....1m._.- . - r1 u." . , . . 0-1 1

 

 

MSU

LIBRARIES
n

 

 

RETURNING MATERIALS:
Place in book drop to
remove this checkout from
your record. FINES will
be charged if book is
returned after the date
stamped below.

 

 

 

 

 

 

THE EFFECT OF LONG-TERM HEAT ACCLIMATIZATION
ON THE CHARACTERISTICS OF SEMEN
IN THE WHITE LEGHORN ROOSTER

BY

Mostafa Ibrahim Zaki

A DISSERTATION

Submitted To
Michigan State University
In Partial Fulfillment Of The Requirements
For The Degree 0f

DOCTOR OF PHILOSOPHY

Department Of Animal Science

1988

 

CO

UR

ac:

CO?

in

CO

ad

WI;

Ur.

A]

we

at

 

ABSTRACT
THE EFFECT OF LONG-TERM HEAT ACCLIMATIZATION

ON THE CHARACTERISTIC OF SEMEN
IN THE WHITE LEGHORN ROOSTER

BY

Mostafa Ibrahim Zaki

The effect of temperature on the body's functions is a
concern which is global in nature. This investigation was
undertaken to determine the effect of long-term heat
acclimatization on the characteristics of semen in single
comb White Leghorn roosters. In addition, this
investigation examinedg the effect of long-term heat on feed
consumption, body weight, rectal temperature, testes and
adrenals.

The experiment was carried out on 32 sexually mature
White Leghorn roosters for 541 days at Michigan State
University's Poultry Science Research and Teaching Center.
All roosters were housed in individual cages with feed and
water provided ad libitum. The photoperiod was maintained
at 15 hours light: 9 hours dark. Twenty-five roosters were
kept at a high temperature which ranged from 30.0 to 32.1 C
(86.0 to 89.8 F) with a relative humidity of 51 percent in

the heating Vivarium for 541 days. Seven birds formed the

control group. These birds were housed in a neutral
temperature range of 19.2 to 20.8 C (66.5 to 69.5 F) with a
relative humidity of 55 percent. The data indicated that
under heat stress body weight, feed consumption, sperm
motility, testicular weight, seminiferous tubule volume,
number of rows of spermatogenic cells within the
seminiferous tubules, surface to volume ratio of
interstitial cells, adrenal cortical cell volume, and
surface to volume ratio of medullary cells were reduced
significantly. The percentage of dead sperm, the number of
interstitial cells, interstitial cell volume, surface to
volume ratio of seminiferous tubules, adrenal weights,
adrenal medullarly cells volume and surface to volume ratio
of cortical cells were significantly increased. The
percentage of live sperm, sperm count spermatocrit, and

rectal temperature were unchanged by the heat treatment.

This dissertation is dedicated to my brother
Mahmoud Ibrahim Zaki, his wife Aida, their
daughters Manal and Iman, son Yasser, my nephew
Yehia Mohamed Zaki, my Mother—in-law Catherine
Nagel, and my dear wife Sue Nagel. Their love,
understanding, and financial support contributed
immensely to the completion of this dissertation.

iv

ACKNOWLEDGEMENTS

I wish to express appreciation to Dr. Steven Bursian,
my major professor and chairperson of my Graduate Committee
for his help, patience and advice during my Ph.D. program at
Michigan State University. I sincerely appreciate the
advice of Dr. Robert K. Ringer and his help in carrying out
the steps of the experiment. I would like to express my
deep thanks to Dr. Richard Balander for his help in milking
the roosters and collecting the blood samples and for
allowing me to use his laboratory facilities. I appreciate
cordially the help and advice of Dr. Al W. Stinson for
allowing me to use his laboratory facilities and for his
assistance with the study and orientation associated with
the electron microscopy. I would like to express my
gratitude to Dr. Robert Echt (1929-1987) for his assistance
and design of the histological studies. My gratitude is
expressed to Dr. John Gill for his consultation in the
mathematical analysis of the data. My thanks are due
Mrs. Barbara Wheaton for her kind assistance with
histological techniques. My fellow graduate student,
Michael Mckinney graciously assisted with the heating
devices and overall facilities at the Poultry Science
Research and Teaching Center.

V

TABLE OF CONTENTS

Page No.

Chapter 1-IntrOductionOOOOOOOOOOOOOOOOOOOOOOOOO 1

Chapter 2 - Review of Literature................. 4

1. Temperature and Stress................... 5

2. Temperature and Testes................... 9

3. Temperature and Spermatozoa..............l4

4. Temperature and the Adrenal Glands.......18

5. Temperature and Feed.....................28

Chapter 3 - Objectives...........................31

Chapter 4 - Materials and Methods................32

Chapter 5 - Results and Discussion................36

Appendix A Live Dead Stain Nigrosin-Eosin Stain..82
B. Procedure for the Determination of

Sperm Cell Concentration..............83

C. Procedures for Histological Studies...85

D. Model and Statistical Analysis Tests..89

BibliograthOOOOOOOOOOOOO...00.0.0000000000000000095

vi

TABLE

1.

2.

10.

LIST OF TABLES

PAGE

Average Vivarium Temperature.................37
Average Rectal Temperature of Control
and Heated Rooster.........................39
Feed Consumption and Body Weight Before
and After Heating...........................42
Characteristics of Semen in Control and
Heated Roosters..............................44
The Effect of Temperature on Absolute
and Relative Adrenal Gland Weights..........54
The Effect of Temperature on Absolute
and Relative Testes Weights.................56
Morphometric Measurements of the Testes......58
Surface to Volume Ratios of Seminiferous
Tubules and Interstitial Cells
of the Testes................................66
Morphometric Measurements of Medullary and
Cortical Cells of the Adrenal Gland..........74
Surface to Volume Ratios of Cortical and

Medullary Cells of the Adrenal...............75

vii

10.

11.

12.

LIST OF FIGURES
Schmematic Diagram Describing the Temperature
Regulation System with Multiple Sensors.......6
Spermatozoa From a Control Rooster...........45
Spermatozoa From a Heated Rooster............46
Electron Micrograph of a Section of the
Testis from a Control Rooster................48
Electron Micrograph of a Section of
the Testis from a Heated Rooster.............49
The Seminiferous Epithelium Lining a
Testicular Tubule from a Control Rooster.....60
The Seminiferous Epithelium Lining a
Testicular Tubule from a Heated Rooster......6l
Higher Magnification of the Seminiferous
Epithelium from a Control Rooster............62
Higher Magnification of the Seminiferous
Epithelium from a Heated Rooster.............63
Interstitial Cells of Leydig of a Testis
From a Control Rooster.......................67
Interstitial Cells of Leydig of a Testis
From a Heated Rooster........................68
Electron Micrograph of a Section of the

Testis from a Control Rooster................69

13. Electron Micrograph of a Section of the

viii

14.

15.

16.

17.

18.

Testis from a Heated Rooster.................70
Adrenal Parenchymal Cells from a Control
Rooster......................................77
Adrenal Parenchymal Cells From a Heated
Rooster......................................78
Electron Micrograph of the Adrenal

Parenchyma From a Control Rooster............79
Electron Micrograph of Chromaffin Cells from

a Heated Rooster.............................80

Morphometric GridOOOOOOOOOOOOOOOO0.000.000.0093

ix

CHAPTER 1
INTRODUCTION

In order to develop livestock programs which will
counteract all or a portion of the adverse effects of the
environment, there must be an understanding of the
interrelationships of environment and the physiological
function of animals. Generally, studies of the degree of
response to hot environments have been correlated with one
or more climatic variables over a relatively narrow range of
temperature. Such relationships have proven informative but
do not lend themselves to broad use because of differences
in environmental conditions or animal differences. In spite
of these known variables, a number of researchers have
attempted to derive, from laboratory and field tests,
indices which might be put to general use. The most serious
criticism of the indices proposed to date is that, at their
best, they are still rather nonspecific and serve largely as
indicators of a lack of ability to maintain heat balance or
of general adjustments the animal has made to the total
environment. Further, none of the indices has given
consideration to males, which make a much greater
contribution to future generations than individual females.
The available indices may, however, suffice for general

1

selections of females if only heat balance in the body is
concerned, but if the primary interest is productive
performance, they hold only limited promise.

Temperature has obvious significance for the poultry
industry in countries having hot seasons or a tropical
climate. In addition, environmental temperature has a
pronounced effect on semen characteristics in chickens.
Boone et a1. (1963) showed lower, but nonsignificant, values
for semen volume, sperm concentration and number of sperm
per ejaculate immediately following high temperature stress.
Heywang (1944) mentioned that fertility and hatchability
were lowered when chickens are kept under high environmental
temperature. Takeda (1982) indicated that cock spermatozoa
were reversibly inactivated by a temperature of 40 C (104 F)
in certain saline media. V0 (1980) demonstrated that semen
quality was only slightly affected by the highest ambient
temperature (35 C). Studies of the effect of environmental
temperature on deep body temperature (Kadono et a1., 1978)
indicated that no significant differences in deep body
temperature were observed until ambient temperature
reached 32 C (89.6 F.) but highly significant increases

were detected between 32 and 38 C (89.6 and 100.4F) and

diurnal differences in body temperature ranged from 0.6 to
1.1 C (33.08 to 33.98 F). The thermoneutral zone of the
adult fowl within which the performance of the fowl is not
adversely affected by temperatures is from 12.8 to 26.0 C
(55.04 to 78.8 F) (Oluyemi et a1., 1979). The primary aim
of this study was to investigate the effect of long-term
heat acclimatization on reproductive parameters in the male

chicken.

CHAPTER 2
REVIEW OF THE LITERATURE
An understanding of reproduction in the male requires

knowledge of the complexity of physiological and
environmental variables that interact to determine the
reproductive capability of the individual. The study of
animal heat involves an essential vital condition of the
"milieu interieur”, an important attribute of the blood
plasma in which all the anatomical elements are immersed.
As Claud Bernard (1878) stated, "The stability of the milieu
interieur is the primary condition for freedom and
independence of existence; the mechanism which allows of
this is that which ensures in the milieu interieur the
maintenance of all the conditions necessary to the life of
the elements."
It is obvious that the effect of high environmental
temperature on the physiological functions of the body is a
very broad subject. Thus, the subject will be covered under
the following items:

1. Temperature and stress

2. Temperature and testes

3. Temperature and spermatozoa

4. Temperature and the adrenal glands

5. Temperature and feed

1. Temperature and Stress:

 

Yousef (1985) defined stress physiology as "a study of
the animal's physiological, biochemical, and behavioral
responses to the various factors of the physical, chemical,
and biological environment." Thermal stress is defined by
Bligh et a1. (1973) as "any change in the thermal relation
between an organism and its environment, which if
uncompensated by a temperature-regulatory response, would
disturb the thermal equilibrium." Disturbance of this
thermal equilibrium will upset the physiological functions
of the body and its organs. In line with the principle of
complexity of thermoregulatory control, Hillman et a1.
(1985) presented an illustration to indicate multiple
controllers as well as multiple thermal sensors (Figure 1).
Evidence indicates that'these controllers may function both
independently and in concert with each other. Therefore,
the direct and indirect effects of temperature per se on
semen characteristics should be re-evaluated in view of
recent progress in nutrition, endocrinology, enzymology,
embryology and behavior.

Studies on birds have generally emphasized adaptation
to natural habitats and relatively few species have been

acclimated to heat or cold under laboratory conditions.

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

Stress Physiology in Livestock
,- ---------------- -. euvmowuem:
: commune: : temperature
: g All Movement
: [raEEEflEII : “‘Wwi'w‘
O F ,
' [— Lower
' : “=55; em 3' - «muons: eoov:
c m— " "' Shiveclne Cece IlMOMOflI j—u-
:'_.... ’ 5 Petition ttypothetemue -———-
. IIII ' , u Veeemetlee ——-‘ Ildtmtn --
‘ a "5% Hypothetemee H‘" PtlIeereetlen Splint Cord -- I
TM 3 ll l lehevter Stun -
‘OQ' ............ 04'
1.. Multiple fleedbeclt Sinele

 

 

 

 

Figure 1. Schematic diagram describing the temperature
regulation system with multiple sensors, multrple

controllers, and multiple effectors in poultry. (From
Hillman et a1., 1985).

Therefore, the area of temperature acclimatization presents
a vast array of studies which, unfortunately to this date,
do not form a highly organized body of information.

A report by Huston (1975) indicated that chronic
exposure to either an 8 or 30 C environment reduced
fertility in avian males when compared to males maintained
in a 19 C environment. Edens (1983) reported the effect of
environmental stresses on avian male reproduction. Based on
his work, and that of others, one has to conclude that a
generalized stress response is associated with suppression
of reproductive development in immature males and depression
of reproductive potential in the mature male.

Studying heat stress in cows indicated that even
short-term exposure to heat stress affects spermatogenesis
and fertility adversely (Skinner et a1., 1966).

Birrenkott et a1. (1983), in their experiments on the
effect of caponization and adrenal cortical manipulation on
subsequent heat stress survival of five-week old capons and
normal broiler males, suggested an important adrenal
contribution to the ability of young chickens to withstand
high environmental temperatures.

Kadono et a1. (1978) studied the body temperature of

chickens at various ambient temperatures. It was indicated

that no significant differences in deep body temperature
were observed until the ambient temperature reached 32 C
but highly significant increases were detected between 32-
and 38 C.

With an increase in heat stress, the mean rectal
temperature is higher and the skewness greater in both
acclimatized and nonacclimatized men, but at a given level
of heat stress, the mean rectal temperature is higher and
the skewness greater in the nonacclimatized men (Chaffee,
1971).

The study on the strain differences in heat resistance
to acute heat stress between the Bedowin Dessert Fowl, the
White Leghorn and their crossbreeds indicated that the Sinai
desert inhabiting breed was significantly more heat
resistant than the Leghorn. This superiority was expressed
in its longer survival time, its efficient regulation of
body temperature and its high lethal body temperature (Arad
et al., 1982a).

2. Temperature and Testes

 

Testes of the male bird are paired and, unlike those of
most mammals, are located within the body cavity, ventral and
toward the cephalic border of the kidneys. Each testis is
attached to the body wall by the mesarchium and is
encapsulated by a fibrous inner coat, the tunica albuginea,
and a thin outer layer, the tunica vaginalis (Johnson,
1986).

Johnson (1986) mentioned that environmental temperature
has been shown to modify the rate of testicular
recrudescence under experimental conditions in a seasonal

breeder such as the white crowned sparrow (Zonotrichia

 

Leucophrys Gambelii).

A modifying effect of temperature on testicular
activity of mature Coturnix quail was reported by Kato and
Konishi (1968) who found that reducing ambient temperature
from 24-25 C to 5-10 C, 8L:16D caused testicular regression.
In chickens under a short photoperiod (8L:16D), high
temperature (32 C) increased the initial growth phase of
the testes compared with a 22 to 24 C environment and short
photoperiod (Ingkasuwan and Ogasawara, 1966). With a
stimulatory photoperiod (14L:10D) and high temperature,
Ingkasuwan and Ogasawara (1966) demonstrated an initial
stimulation of the early growth phase of chicken testes. V0

9

10

et a1. (1980) reported that age of sexual maturity decreased
in males reared in continuous light, and a 35 C
environment compared to males reared in a continuous light,
21 C environment. Lamoreaux (1943) and Hoffman and Shaffner
(1950) reported significantly increased testicular size in
chickens held in warm (27 to 29 C) environments as compared
to those held in a cold (2 to 4 C) environments. Huston
(1975) reported that chronic exposure to an 8 C
environment delayed sexual maturation while exposure to a
30 C environment accelerated the rate of maturation.

Cummings and Huston (1976) found that acute exposure of
chickens to temperatures of either 8 or 30 C cause
significant alterations in semen pH, sodium, potassium and
magnesium concentrations. Kosin (1958) found depressed
metabolism in turkey semen taken during summer months (35 C)
from males exposed to natural elements compared to semen
taken from males held in a temperature and light controlled
environment (18 C, l4L:10D).

Volcani (1953), in his study on the seasonal variations
in spermatogenesis of some farm animals under the climatic
conditions of Israel, indicated that in the winter-rut
period of the camel, the testes grow considerably in size

but during the hot summer months, they recede between the

11

hind legs towards the abdomen. It seems that the seasonal
changes in spermatogenesis are largely determined in the
camel by the high ambient temperature prevailing in the
summer.

Histological examination of the testes and the
epididymis in the Arabian bull indicate conspicuous seasonal
changes in the condition of the seminiferous tubules.
There was a depression of activity and appearance of summer-
sterility symptoms (June — August). In the Awassi ram,
symptoms of summer-sterility can be detected, similar to
those described in reference to the camel and the bull
(Volcani, 1953).

Damber et a1. (1980) demonstrated that heat treatment
(43 C) of the testis resulted in a reduction of testicular
weight in the rat (1.15 +/- 0.08 9, mean +/- SD) when
compared to the control group held at 33 C (1.82 +/- 0.099.).
The tubular content of the testes from rats subjected to 43
C was highly damaged. Exposing the scrotum of the rat to
33 and 43 C indicated that there was no significant
difference in basal plasma testosterone concentration
between the two groups, but the response to LH stimulation
was significantly depressed in the group exposed to 43 C

(Damber et a1., 1980). The cause of testicular degeneration

12

on removal of the organ from the scrotum into the peritoneal
cavity is found to be due to a higher temperature of the
peritoneal cavity in guinea pigs (Moore, 1924). Steinberger
et a1. (1959) reported that a 15 minute exposure of the
scrotum of the rat to a temperature of 45 C or 43 C
produced a progressive destruction of the entire germinal
epithelium. The earliest cytologic changes were observed in
spermatids.

In other experiments by Collins et a1. (1967), the
scrotal regions of adult male Wistar rats were exposed to a
temperature of 43 C for 10, 15, 20, 25, or 30 minutes, and
the animals were killed 3 days, 7 days, 3 weeks or 6 weeks
after treatment. Animals in the lS-minute group exhibited a
pattern of mild heat damage which was due to a failure of
the early transitional spermatocytes and those in late
pachytene and beyond. With more prolonged exposure to heat,
the damage was extended to other stages of germ development
(Collins et a1., 1967).

Mammalian testes shifted from the scrotum into the
abdominal cavity, where the temperature is a few degrees
higher, cease to produce spermatozoa; degeneration of the
spermatogenic tissue sets in and spermatogenic function is

not resumed while experimental cryptorchidism prevails

13

(Mann, 1964). Artificial cryptorchidism is followed about
the 21st day after operation by a transient increase in the
numbers of interstitial and Leydig cells of the testis of
the rat, and the proportion of non-senile Leydig cells is
increased over the whole experimental period (Clegg, 1961).
Younger Leydig cells are those most active in the production
of androgen and these cells are contained within the
category of non-senile Leydig cells. In the guinea pig, a
complete cessation of spermatogenesis was brought about
experimentally by scrotal application of heat (6 C above the
normal body temperature) for a period of 10 minutes (Moore,
1924, 1951). A similar effect was produced in rams; semen
collected from such animals a week or two later contained
only a small number of spermatozoa, mostly dead or
degenerate; the seminal plasma, on the other hand, retained
its normal composition or shows a slightly elevated content
of fructose, which might be interpreted as a result of
temperature stimulated endocrine activity of the testes
(Glover, 1955).

In Hereford bulls exposed to high ambient temperature,
it was found that spermatogenesis, evaluated by semen
characteristics and histological examination of testes at
the termination of the experiment, was impaired by heat

(Rhynes et a1., 1973).

3. Temperature and Spermatozoa

 

The motility and metabolic activity of spermatozoa are
inhibited at low temperatures, resulting in an extension of
their life span and fertilizing ability in vitro (Salisbury
and Lodge, 1962: Mann, 1964).

Ashizawa et al. (1983b) mentioned that when fowl
spermatozoa were incubated at 41 C with tissue from the
infundibulum, magnum, isthmus, shell gland, uterovaginal
junction and vagina, their motility, assessed at room
temperature (20 to 25 C), was maintained for 4 to 12 d. The
survival period was much longer for spermatozoa incubated
with the tissues from the infundibulum and uterovaginal
junction (with sperm-host glands, 11 to 12 d) than for those
incubated with tissues from the other regions (lacking host
glands, 4 to 7 d). In the tissues of the infundibulum and
uterovaginal junction, spermatozoa entered the sperm-host
glands and were more closely associated with the epithelial
cells than they were in tissues from other regions (Ashizawa
et a1., 1983b).

Clark et a1. (1984) studied the viability and
morphology of undiluted and diluted chicken and turkey

spermatozoa. They compared the spermatozoa when incubated

14

15

at either 41, 25, 15, or 5 C for 0 (control), 3, or 6 hr.
They mentioned that increasing the incubation time to 6 hr,
raising the incubation temperature to 41 C or both resulted
in higher numbers of dead spermatozoa in semen from both
species. At 41 C, it was found that sperm motility was
at its lowest.

Turkey semen was collected, diluted 1:1 with Beltsville
Poultry Semen Extender, and held for 0 or 18 hr at 5, 15,
25, or 35 C Changes in spermatozoa motility and sperm
numbers were monitored before and after holding. It was
found that samples held at 15 and 25 C had motility scores
of 40 and 8%, respectively. Samples held at 35 C for 18 hr
were immotile. As semen holding temperature increased from
5 to 35 C Sperm numbers decreased during the 18 hr holding
period by 11, 16, 28, and 45% of the unstored control
(Giesen et a1., 1983).

Renden et a1. (1984), in their studies on reproductive
performance of broiler breeds exposed to cycling high
temperature from 17 to 20 weeks of age, reported that the
percent of abnormal sperm was greater in heat-treated males
than controls from 28 to 31 weeks of age. Male chickens
kept at 19 C had a higher fertility capacity than males

kept in either 30 C or 8 C (Huston, 1975).

16

The fertilizing capacity of the semen of male turkeys
maintained in a "comfortable" environment was significantly
greater than that of turkey males partially protected or
wholly unprotected from temperature fluctuations (Burrows et
a1., 1953).

Ashizawa et a1. (1983) studied the morphology of fowl
oviducal cells cultured at 41 C with and without fowl
spermatozoa and observed with the scanning electron
microscope. At the beginning of incubation, abnormal
spermatozoa were comparatively few, spermatozoa were found
on the surface of the cultured cells. Morphological
alteration of spermatozoa increased with increasing
incubation interval and included head and tail coiling with
the occasional separation of heads from flagella (Ashizawa
et a1., 1983)

Egbunike et a1, (1980) studied the influence of short-
term exposure to tropical sunlight on boar seminal
characteristics. During the period of exposure, both
respiratory rate and rectal temperature increased
significantly by 276.84 and 5.13% respectively in the
exposed boars compared to unexposed boars, thus indicating a
high degree of hyperthermia. Sperm motility and

concentration deteriorated in the stressed animals. Sperm

17

output per ejaculate dropped drastically only in the week
following exposure (58 to 28 billion sperm as compared to of
54 to 47 billion by the unexposed boars. Similarly, the
frequency of sperm abnormality was higher in the stressed
boars during this period, after which the animals appeared
to have recovered.

Ingkasuwan and Ogasawara (1966) mentioned that exposure
of fully matured White Leghorn males to high temperature
(32.2 C, 8L:16D) resulted in low spermatozoal concentration
compared to birds maintained at 20 C, 8L:16D. Indeed, in
most males of other species the biochemistry of semen is
altered by hyperthermia and hypothermia (Mann, 1964;
Blackshaw, 1977). Hyperpyrexia frequently causes a
temporary azoospermia in man, and a hot climate is believed
to be the principal cause of certain forms of subfertility
among domestic animals in tropical countries (Mann, 1964).
There was evidence that cold and high temperature changed
the pH and cation concentration of semen from donor males.
This may indicate that there is an alteration in metabolic
activity of semen from both cold and hot donor birds
(Cummings and Huston, 1976). Semen pH of the 30 C males was

decreased and semen pH of 8 C males increased.

4. Temperature and the Adrenal Glands:

The adrenal glands constitute one of the major
homeostatic organs of the mammalian body. They are composed
of two separate endocrine organs that differ in
embryological orgin, type of secretion, and function. In
mammals, the two organs are arranged as an outer cortex and
an inner medulla surrounded by a common capsule. In the
other vertebrate classes, the two tissues may be completely
unassociated or intermingled to a greater or lesser degree.
In these cases, the homolog of the mammalian medulla is
called Chromaffin tissue, whereas the tissue corresponding
to the cortex of mammals is called interrenal tissue.

The proportion pf the cortex and the medulla in

 

 

the adrenal gland:

 

Harvey et a1. (1986) mentioned that the paired avian
adrenal glands are composed of intermingled Chromaffin and
cortical (interrenal) tissue. The glands are located
anterior and medial to the cephalic lobe of the kidney.
These glands are flattened and lie close together, even
fusing in some species. Avian adrenal glands are not
clearly divided into outer cortex and inner medulla, as in
mammals. Cortical and Chromaffin tissue is intermingled in

birds, with clusters or strands of Chromaffin cells

18

l9

distributed throughout the cortical tissue.

In birds chromaffin tissue constitutes about 15-25% of
adrenal tissue. The chromaffin cells are in close
association with blood spaces and appear to be abundant in
the middle of the gland, which is enriched with vascular
tissue. Two distinct types of chromaffin cells exist in the
avian adrenal, releasing epinephrine (E) and norepinephrine
(NE) respectively. There is some controversy as to the
relative proportion of cell types. Ghosh (1980) reported a
greater number of NE chromaffin cells in all birds studied
(except passerines) while Unsicker (1973) considered that E
chromaffin cells predominate.

Cortical tissue accounts for 70-80% of the female avian
adrenal. The cortical cells are arranged in numerous cords
with each being composed of a double row of parenchymous
interrenal cells. The cords radiate from the center of the
gland, branching and anastomosing frequently, and loop
against the inner surface of the connective tissue capsules.
The arrangement of Specific cell types along the cords
results in some structural zonation. In birds, adrenal
zonation is less clear than in mammals. There are two
zones, a subcapsular zone (SCZ) and an inner zone (12),

found on birds. The SCZ is about 20-40 cells thick and

20

produces aldosterone and the more extensive 12 produces
corticosterone.

Due to a variety of experimental techniques and the
physiological state of the avian, results vary as to the
percent of cortical tissue. Sturkie (1965) stated that the
cortex of the male chicken adrenal comprises 40% of the
gland, according to Kar (1947), or 44.2% according to Sauer
and Latimer (1931). The interrenal tissue of the female
gland amounts to 71% according to Kar. The mean proportion
of cortical tissue in chicken adrenal was significantly
greater in the female (57%) than in the males (50%) (Siller
et a1., 1975). In proportion to body weight, females have
30% more cortical tissue than males (Sauer and Latimer,
1931).

Oakberg (1951) measured the cortical and medullary
tissue in the adrenal of 120 day old White Leghorn chickens.
He found that percentage on the means in the males was 46
percent for the medulla and 50 percent for the cortex. The
females were 43 for the medulla, and 52 for the cortex.

Sauer and Latimer (1931) reported the weight of an
adrenal gland and the proportion by volume for each of the
cortex and medulla tissue were 0.1060 g., 44.2 percent and

31.3 percent respectively, in the male single-comb White

21

Leghorn chickens. The weight of entire gland; cortex and
medulla, as a percent of body weight of the same birds were
0.00510 +/-0.0002., 0.00211 +/- 0.00006 and 0.00164 +/-
0.00012, respectively.

Serial sections of six suprarenal glands, (from two
males and three female) single comb White Leghorns were
prepared and ratios of cortical and medullary, or chromaffin
cells were determined. Sectors of every twentieth section
were projected onto heavy drawing paper and the areas
representing the two types of cells were cut out and
weighed, thus giving the ratios of the cortical and
medullary cells. The medullary cells of the two male glands
averaged 71 % of the cortex. The individual percentages
were 65 and 79% of the cortex. In the females, the medulla
averaged only 37% of the volume of the cortex, with a range
from 28 to 71%. This seems to indicate a sex difference in
the relative amounts of cortical and medullary cells in the
chicken suprarenal (Latimer, 1925).

Social interaction plays an additive role of stress on
the already heat stressed avian male. Siegel and Siegel
(1961) studied the relationship of social competition with
endocrine weights and activity in male chickens. Fifty

strange White Leghorn cockerels, were housed in pens,

22

adrenal and body weights were compared. The right adrenal
gland of the experimental group weighed 6.04 +/- 0.87 mg
1/100 g of body weight and the left adrenal weighed 5.46 +/-
0.64 mg. In the control, the weights were 5.97 +/- 1.32 mg.
for the right adrenal gland and 4.87 +/— 0.76 mg. for the
left adrenal gland. The total combined right and left
adrenal weight in mg. /100 g body weight showed the the
experimental group to be 11.50 +/- 1.43 and 10.84 +/- 1.88
in the control. It is interesting to note the dramatic
increase in weight seen solely in the left adrenal of the
avian experimental group.

The adrenal glands (called suprarenal glands in human
beings because of their upright posture) lie
retroperitoneally near the anterior poles of the kidney and
are embedded in the perirenal adipose tissue. The combined
weight is about 8 g, the medulla is approximately 10% of the
weight of the gland (Long, 1983). Fawcett (1986) mentioned
that the weight of the two glands is 15 g, and the gland is
composed of interrenal tisue and chromaffin tissue. In
mammals, these are arranged as cortex and medulla
respectively, but in other vertebrate classes they may be
intermingled in a variety of patterns or may be entirely

dissociated.

23

The cortex, which forms the bulk of the gland, has
three distinguishable concentric zones: a thin, outer Zona
Glomerulosa adjacent to the capsule; a thick middle layer,
the Zona Fasciculata; and a moderately thick inner Zona
Reticularis contiguous with the medulla. In man these make
up respectively 15, 78 and 7% of the total cortical volume.
The transition from one zone to another in histological
sections is gradual but may appear sharper in preparations
injected to show the vascular pattern (Fawcett, 1986). Long
(1983) mentioned in his description of the histology of the
adrenal cortex in humans that the Zona Glumerulosa accounts
for approximately 15% of the total cortical volume, the Zona
Fasciculata about 78%, and the Zona Reticuloris about 7%.
These proportions were in agreement with Fawcett (1986).

Hester et a1. (1981) studied the effect of prolonged
heat stress on adrenal weight, cholesterol and
corticosterone in White Pekin ducks. They reported that the
adrenals of heat stressed ducks exposed to a constant
temperature of 29.4 C for 45 to 46 days showed an increase
in relative adrenal weight and cholesterol concentration,
but the corticosterone concentration remained at control
levels.

Freeman (1983) stated that extremes of temperature

24

stimulate a rapid increase in the concentration of
corticosterone in the plasma of the older bird (Nir et a1.,
1975, Ben Nathan et a1., 1976; Edens and Siegel, 1976;
Beuving and Vander, 1978; Nir et a1., 1975; Freeman, 1982 )
but not in the newly hatched chick (Freeman, 1982).

Deviche (1983) mentioned that the pituitary-adrenal
hormones are partly responsible for the short and longer-
term depression of the pituitary-gonadal axis which were
observed in stressful conditions. The mode action of
adrenal hormones on the reproductive system of male birds is
probably very complex and remains largely unknown. Evidence
suggests that corticosterone favors the transformation of
testosterone into biologically inactive androgens in the
pituitary gland (Deviche, 1983).

The adrenal cortex, whose secretory rate is controlled
by hormones produced in the adenohypophysis and the kidney,
produces steroid hormones that affect carbohydrate and
protein metabolism, resistance to physiological stress, and
electrolyte distribution. The medulla, under nervous
control, secretes catecholmines that affect heart rate and
smooth muscle function in blood vessels and the viscera.
Various aspects of carbohydrate and lipid metabolism are
also influenced by catecholamines. The right and left

glands have somewhat different shapes in human beings, the

25

left gland being somewhat broader. The combined weight of
the adrenals from human adults dying immediately of
accidental causes is about 8 g. It should be remembered
that both weight and size of the glands vary considerably
with age and physiological condition of the organism. In
gross section, the cortex, which constitutes the largest
part of the gland, appears yellow because of the presence of
lipids. The medulla, which represents approximately 10 % of
the weight of the adrenal gland is reddish, or brown. (Long,
1983).

Cramer (1928) stated that adrenaline released on heat
exposure would be expected to raise the body temperature
still further through its calorigenic action, aided perhaps
by some reduction of peripheral vasodilatation (Whelan et
a1., 1963). But he suggests that this is not necessarily
always the case. For an animal without sweat glands,
Cramer (1928) speculated that ”the organism will react to
exposure to heat by an inhibition of the adrenal gland" for
if this was not so, heat hyperpyrexia might ensue. Such an
inhibition would be analogous to that which the thyroid
undergoes with moderate degrees of heat exposure. In the
dog, the plasma catecholamine concentration was reported to

be unchanged during acute heat exposure (Fiorica et a1.,

26

1967).

In the bovine and the horse (with adrenaline-sensitive
sweat glands), the adrenal medulla is responsive to heat
stimulation, whereas in the rat and man (with
cholinergically innervated ecocrine glands on the pad and on
the whole body surface, respectfully) there is a lack of
response by the adrenal medulla to heating of the body
(Collins et a1., 1968).

Attention was first drawn to the involvement of the
adrenal cortex in the response to high environmental
temperatures by the morphological changes observed in the
adrenal glands of animals kept in hot conditions (Cramer,
1919; Cramer, 1928).

Histological and histochemical changes occur in the
adrenal glands of animals exposed to tropical conditions
(Bernstein, 1940 and 1941; Cramer, 1928; Flexner et a1.,
1939; Schmmidt et a1., 1938). A relative increase in actual
size has been reported in rats bred for several generations
in conditions of high temperature and humidity (Clark 1938;
Tepperman et a1., 1943).

There is evidence that prolonged exposure to a high
ambient temperature (43 C) would cause acute adrenal

cortical insufficiency (AACI) in young chickens (Edens,

27

1976). The development of AACI in heat stressed broiler
cockerels was characterized by a sharp increase in levels of
plasma corticosteroids which was followed by a rapid
decline. The decline of plasma corticosteroid levels was
associated with the significant reduction of adrenal
cortical levels of the corticosteroids. There was a
concurrent decline in plasma levels of glucose,
sodium/potassium ratio, total calcium, and inorganic
phosphate. These findings conformed to the description of
AACI in humans suffering from Addison's disease (Edens,

1978).

5. Temperature and Feed:

The hypothalamus exercises an important controlling
influence on the level of feeding which has been emphasized
by a number of research workers. Bilateral lesions in the
lateral border of the ventromedial nuclei produce obesity
with or without hyperphagia, while lesions in the lateral
hypothalamic region cause aphagia. Whether the aphagia is
intrinsically due to motor failure or to lack of motivation
is still under discussion (cited by Collins and Weiner,
1968). An elevated caloric intake is required to maintain
heat production in low temperature conditions. Conversely,
at high temperatures suppression of food intake may become a
crucial factor in counteracting hyperthermia, particularly
in the nonacclimated state.

Hot ambient temperatures characteristically reduce feed
intake activity and growth rate (Whittow, 1965). At very
high temperatures, feed efficiency can be reduced (Dale and
Fuller, 1980).

Taher et a1. (1985) studied the feeding pattern
responses to changes in dietary energy or environmental
temperature in the domestic fowl. They reported that the
change to high energy or low energy diets consumed low or

high amounts of feed, respectively. Those fed the high

28

29

energy diet tended to decrease meal size and meal duration
and to increase the number of meals. Roosters changed to
the low energy diet decreased meal size and meal duration
and increased the number of meals eaten. The results tend
to confirm the chemostatic mechanism in birds as food intake
was related to energy in the diet.

Roosters changed to high or low environmental
temperature responded by decreasing or increasing their feed
intake, respectively. Roosters changed to the high
environmental temperature significantly decreased meal size,
meal duration, and increased the number of meals. Those
changed to a low environmental temperature significantly
increased meal size and decreased meal frequency and meal
duration (Taher et a1., 1985).

Hurwitz et a1. (1983), in their studies on male turkeys
at different environmental temperatures, reported that
weight gain was severly depressed by a constant 35 C but
was lower at 27 C than at 10 C and 20 C. Feed efficiency
hardly changed between 10 and 20 C but was depressed at the
higher temperatures. The calculated energy needs for
maintenance were only slightly reduced at 20 C compared
to 10 C, but they declined continuously with an increase in

temperature up to 35 C (Hurwitz et a1., 1983).

30

Wilson et a1. (1980) studied the effects of high
environmental temperature on feed passage time and
performance traits of White Pekin ducks. They mentioned
that the high environmental temperature caused an increase
in feed passage time, suppressed growth, had no effect on
feed/gain ratios, and increased the relative weight of the
right adrenal gland (Wilson et a1., 1980).

The difference in energy metabolism at different
environmental temperature is strikingly illustrated in
studies on rats. Animals acclimated to a temperature of 4
C were found to eat three times as much as those acclimated
at 36 C, yet both maintained a steady weight. However,
hyperphagia and increases in weight could be induced in both
heat and cold acclimated rats when lesions were placed in

the ventromedial nuclei of the hypothalamus (Kennedy, 1953).

CHAPTER 3
OBJECTIVES
Experiments concerning the effects of temperature on
the body's physiological functions are difficult to
interpret because so many sites in the body are influenced
such as the cardiovascular system, the respiratory system,
the nervous system, the digestive system, the excretory
system, and the endocrine system. In addition, all these
systems are acting in combination under the heat stress.
Thus, the objectives of this study were to determine
the effect of long-term heat acclimatization in White
Leghorn roosters regarding:
1. Body temperature
2. Feed consumption and body weight
3. Semen characteristics
4. Adrenal and testes weight
5. Histological features of the testes

6. Histological features of the adrenal gland.

31

CHAPTER 4

MATERIALS AND METHODS

A. Experimental Design:

This experiment was initiated on July 1, 1984 and
terminated on March 13, 1986, for a period of 541 days. The
experiment was conducted with 32 sexually mature single comb
White Leghorn roosters at Michigan State University's
Poultry Research and Teaching Center. All roosters were
housed in individual cages (42.5 cm. deep by 45 cm. high by
40 cm. wide) with feed and water provided ad libitum. The
photoperiod was 15 hours light and 9 hours dark (15L:9D),
controlled by an Intramatic timer (Intermatic Timer
Controls, Intermatic Incorporated, Spring Grove, IL. 60081)
for a continuous 31 day cycle.

The heating stage began on September 20, 1984 and
continued 541 days until March 13, 1986. Twenty-five
roosters were kept in a high temperature environment ranging
from 30 C to 32.1 C (86.0 to 89.8 F) with a relative
humidity of 51 percent. The heating apparatus consisted of a
gas brooder heater. Seven birds formed the control group
which was housed in a neutral temperature range environment
of which from 19.5 to 20.8 C (66.5 to 69.5 F) with a
relative humidity of 55 percent. Before the initial

32

33

adjustment phase to the higher temperature and relative
humidity, the birds were trained for sperm collection during
August and September 1984, for 51 days.

B. Data Collected:

 

1. Body Weight:

 

Body weights of all birds were obtained with the use of
a Toledo balance (Toledo Scale Co., Toledo, OH). Weights
were recorded to the nearest gram. Weights were determined
each 15 days from the beginning on July 1, 1984.

2. Feed Intake:

 

Individual feed consumption for all the roosters was
determined at 15 day periods. The Toledo balance was used.
Feed consumption was corrected for wastage.

3. Vivarium Temperature:

Maximum and minimum temperature of the room and the
relative humidity were recorded daily for the duration of
the experiment. The hygrometer was manufactured by the Ohio
Thermometer/Sign Co., Springfield, Ohio.

4. Rectal Temperature:

 

Rectal temperature was recorded for each bird, every 15
days at noon for the duration of the experiment. The rectal
tale-thermometer was manufactured by Yellow Springs

Instrument Co., Inc., Yellow Springs, Ohio.

34

5. Semen Characteristics:

 

Semen was collected, from each rooster, by abdominal
massage and immediately examined for the following
characteristics:

1. Spermatocrit was determined by drawing semen up in

microhematocrit tubes and centrifuging the tubes for 15

minutes.
2. Sperm count (Number of sperm/m1) was determined by
use of a hemocytometer. Values obtained from the

hemocytometer were then compared with a standard curve
(Appendix B) to obtain final sperm counts.

3. The motility of the sperm was determined by the
hanging drop method immediately after collection. Motility
was subjectively assessed on a scale from 0 to 5 with 0
representing no motility and 5 representing maximum
motility.

4. The percentage of living and dead sperm was
determined by using the live-dead stain (Appendix A). The
procedure involved making a smear of semen on a slide,
staining it with Nigrosin-Eosin, stain then allowing it to
dry. Then, a count of 100 sperm was taken, using the light
microscope with the oil immersion lens. From each one

hundred sperm on each slide, the percentages of live and

35

dead sperm were counted. The slides were made in duplicate
for every sample from each individual rooster.

6. The Relative Weights g£_Adrena1s and Testes:

 

 

The live weight of each rooster was recorded. The
percentage of the testes and adrenal glands to the brain and
to 100 g/body weight for each rooster was determined.

7. Histological Studies:

 

At the end of the experiment on March 13, 1986, 15
roosters from the heated group and seven roosters from the
control group were weighed and killed. Immediately after
killing, the testes, the adrenal gland, and the brain of
each rooster were removed and weighed. The testes and
adrenals were then prepared for histological studies using
light and electron microscopy (Appendix C).

8. Volume Estimation:

 

For the volume estimations of the interstitial cells
and seminiferous tubules of the testes, and the medullary
and cortical cells of the adrenals the method of Gaunt and
Gaunt, (1978) was used (Appendix D).

9. Statistical Analysis:

 

The statistical model is presented in Appendix D.
The procedures outlined by Gill (1981) for the t-test and

transformation of the percentage to the log was used

CHAPTER 5
RESULTS AND DISCUSSION

The effect of long-term high temperature on the semen
characteristics of the roosters was the principal parameter
examined in this investigation. In addition to semen
production, there were other systems in the body which were
affected at the same time by this long-term high
temperature. Some of the data were recorded daily and
others were collected periodically or at the end of the
experiment.

A. Body Temperature:

 

The maximum and minimum temperature and relative
humidity of the Vivarium were recorded daily throughout the
period of the experiment (541 days). The temperature and
percent relative humidity were averaged over a monthly
period. The average Vivarium temperature for the heated
roosters and control roosters are presented in Table l. The
range of temperature for the heated roosters was 30 to 32 C
(86.0 to 89.8 F) with a relative humidity of 51.4%. The
range of the Vivarium temperature for the control rooster
was 19.2 to 20.8 C (66.5 to 69.5 F) with a relative humidity
of 55.3%. This latter temperature range is in the

thermoneutral zone of the adult fowl within which

36

37

TABLE 1. AVERAGE VIVARIUM TEMPERATURE

 

 

CONTROL HEATED
MAXIMUM 20.83 +/- 0.28 32.20 +/- 1.20 a
TEMPERATURE
MINIMUM 19.20 +/- 1.20 30.00 +/- 1.10
TEMPERATURE
RELATIVE 55.30 +/- 3.35 51.40 +/- 2.20
HUMIDITY

 

a Values are expressed as mean +/- standard error.
Temperature is in degrees centigrade.
The values for the control group are averaged over 4 months,

and the values for the heated group are averaged over 19
months.

38

performance is not adversely affected by temperature.
According to Oluyemi et a1., (1979) the thermoneutral zone
for the adult fowl is 12.8 to 26.0 C.

Birds native to hot environments, like those from cold
regions, have basal body temperatures not significantly
different from those of temperate-zone birds (Dawson et a1.
1964 and Crawford et a1. 1967). To investigate this point,
the rectal temperatures of the heated and control roosters
were recorded. It was found that the average rectal
temperatures were 41.3 and 41.6 C (106.8 and 106.9 F) for
the control and heated group, respectively (Table 2).

Kadono et a1. (1978) indicated that no significant
difference in deep body temperature was observed until the
ambient temperature reached 32 C (which was the maximum
temperature in this experiment), but highly significant
increases were detected between 32 and 38 C. In the
present experiment, the difference between rectal
temperatures of the heated and control roosters was .067 C
(0.1 F). This is in agreement with Kadono et a1. (1979) even
though the period of heating was considerably longer in the
present experiment (541 days verses 8 days). Other
experiments indicated that body temperature began to

increase significantly when ambient temperature reached

39

TABLE 2. AVERAGE RECTAL TEMPERATURES
OF CONTROL AND HEATED ROOSTERS

 

 

CONTROL HEATED
(7) (15)
RECTAL TEMPERATURE 41.3 a +/- 0.21 41.6 +/- 0.03

 

a Values are expressed as mean +/- standard error.
Temperature is in degrees centigrade.
The values for the control group are averaged over 4 months,

and the values for the heated group are averaged over 19
months.

40

beyond 27.5 C. (Van Kampen et a1., 1978).

With increases in ambient temperature, the mean rectal
temperature of the human also increases and the skewness is
greater in both acclimatized and nonacclimatized men
(Chaffee, 1971). This contrasts with acclimatized chickens
in the present investigation.

Observations of the behavior of the roosters as heating
was initiated indicated that roosters became very restless,
excited, and were jumping inside the cage for approximately
the first 9 to 10 days. This was the first defense of their
nervous system against heat stress. After this period,
their endocrine system began to acclimatize under the
present situation.

B. Feed Consumption and Body Weight:

 

 

The diffence in feed consumption and body weight
before and after heating the roosters was presented in Table
3. Feed consumption was 120.0 and 86.8 grams per day for
the before and after heated roosters, respectively. The
difference between the two groups was highly significant.
The average body weight of the heated roosters was
significantly less than before heating and the difference
was significant.

As mentioned previously, the hypothalamus has a

41

controlling influence on the amount of feed consumed. At
the same time, the hypothalmus plays a regulatory role in
body temperature. Thus, the quantity of feed consumed
depends to a great extent on the dietary energy and
environemtnal temperature (Taher et a1., 1985).

Reduced feed consumption under high temperature in the
present experiment is in agreement with data for cockerels
reported by Bottje and Harrison (1985). Those
investigators stated that heat stress reduced average daily
gain, feed intake, and feed efficiency. A depression in the
body weight is in agreement with Hurwitz et a1. (1983) who
studied male turkeys. They mentioned that weight gain was
severly depressed by a constant 35 C but was also lower at
27 C than at 10 and 20 C. In the present experiment, the
average of maximum and minimum temperature was 31.1 C for
the heated roosters during 541 days (Table 1). At the
average temperature (31.1 C) the body weight was found to
decline by about 7 percent as compared with the body weight
before heating the roosters (Table 3). There are close
relationships among feed intake, hormones, and temperature.
Denbow (1983) reported that the injection of catecholamines
into the lateral ventricle of the turkey brain decreased

colonic body temperature and feed intake. Denbow's results

42

TABLE 3. FEED CONSUMPTION AND BODY WEIGHT
BEFORE AND AFTER HEATING

 

 

BEFORE HEATING AFTER HEATING
(25) (15)
FEED CONSUMED 120.0 +/- 3.30 86.8 +/— 2.51 b
GRAN/DAY
BODY WEIGHT 2.3134 +/- 0.029 2.1523 +/— 0.034 b
(KG)

 

a Values are expressed as mean +/- standard error.
Number in parentheses refers to sample size.

b Significantly different from control at P<0.05.

43

suggested that brain catecholamines were involved in the
neural control of feed intake and body temperature of
turkeys.

There was evidence that feed intake, heat production,
and maintenance energy requirements all increase linearly
with a decrease in temperature (Farrell and Swain, 1977).
Feed passage time which, increases under high temperature,
is another factor which suppressed growth (Wilson et a1.,
1980). Thus, the decrease in body weight may be caused by
decreasing feed intake plus increasing feed pasage. It was
found that animals acclimated to a temperature of 4 C ate
three times as much as those acclimated at 36 C (Kennedy,
1953). Since so many factors interact, accurate measuring
devices for separating the effect of temperature on the
hypothalamus, feed intake, diet energy and the body's
hormones are yet to be researched.

C. Semen Characteristics:

Comparisons of the characteristics of the semen between
the control and heated roosters (Table 4) indicated that in
the heated roosters motility was reduced significantly and
the percent of dead spermatozoa increased significantly
(Figures 2 and 3). The percent of live sperm and sperm

count (billion/ml) and spermatocrit were affected by heat.

44

TABLE 4. CHARACTERISTICS OF SEMEN IN CONTROL
AND HEATED ROOSTERS

 

 

PARAMETER CONTROL HEATED

(7) (15)
MOTILITY 4.57 +/- 0.133 b 3.21 +/- 0.194 c
% LIVE SPERM 96.36 +/- 0.416 83.72 +/- 3.093
% DEAD SPERM 3.64 +/- 0.416 16.28 +/- 3.093 C
NUMBER OF SPERM/ML 7.63 +/- 0.983 6.51 +/- 0.407
IN BILLION
% SPERMATOCRIT 11.75 +/- 1.550 9.98 +/- 0.640

 

a Motility was assessed on a 5 point scale with 0 representing
no motility and 5 representing maximum motility.

b Values are expressed as mean +/- standard error.
Values expressed in percent were transformed to their
respective logarithmic values for statistical analysis.

c Significantly different from control at P<0.05.

45

 

Figure 2.

SPERMATOZOA FROM A CONTROL ROOSTER.

Structures shown include (1) acrosome, (2) head (3) neck,

(4) middle-piece and (5) tail.

Nigrosin Eosine stain x 400.

46

 

Figure 3. SPERMATOZOA FROM A HEATED ROOSTER
The presence of stain in the head of the spermatozoa
indicates that the sperm is dead.

Nigrosin Eosin stain x 400

47

This is in agreement with Ingkasuwan et a1. (1966). In
fact, it was found that temperature affects all phases of
semen production. It stimulates testicular growth in the
early phase and promotes increased semen volume and
concentration.

Figures 2 and 3 show darkly stained dead spermatids
which accepted the live-dead stain and live spermatids which
are pale in contrast. The data presented in Table 4
indicates that percent of dead spermatozoa in the heated
roosters is four to five times that of the control. Electron
micrographs of the spermatids from control and heated
roosters indicated that nuclei in control spermtids were
more active than the nuclei in the heated group (Figures 4
and 5).

These changes in semen quality under high environmental
temperature are in agreement with Casady et a1. (1953).
Volcani (1953) noticed the same stages of spermatogenesis
and the state of the seminiferous tubules as a result of
heat stress. Johnston et a1. (1963) noticed spermatozoan
abnormalities. Moore (1924) and Steinberger et a1. (1959)
reported a testicular degeneration and germinal epithelium

destruction. Collins et a1. (1967) studied spermatogonia

48

 

Figure 4. ELECTRON MICROGRAPH OF A SECTION OF THE TESTIS
FROM A CONTROL ROOSTER.

The figure shows that a manchette (2) is comprised of a
wide ring of intermediate density surrounding the nucleus of
the spermatid. The cytoplasm (4) of the Sertoli cell fills
the space between the spermatids. The nuclei (1) show as an
accumulation of coarse granules. The two centrioles (3)
appear in the middle spermatid at the left. Inside the
nuclei, a honeycomb structure is seen.

Lead citrate and uranyl acetate x 19,000.

49

 

Figure 5. ELECTRON MICROGRAPH OF A SECTION OF THE TESTIS
FROM A HEATED ROOSTER.

The spermatids (l) are engulfed in the Sertoli cell (2) with

bands of ordered microtubules around the nuclei (3).

Lead citrate and uranyl acetate x 10,000.

50

development in adult male rats under high temperature (43
C). They found heat damage of the germinal epithelium and
the damage incited by the increasing the exposure period to
heat.

The motility and metabolic activity of spermatozoa are
inhibited at low temperature (Salisbury and Lodge, 1962;
Mann, 1964). Clark et a1. (1982) mentioned that sperm
motility in diluted and undiluted chicken and turkey semen
was lowest at 41 C. Raising the incubation temperature to
41 C resulted in higher numbers of dead spermatozoa in semen
from chicken and turkey (Clark et a1., 1984). These results
are in agreement with the present experiment, in that the
high temperature reduced the motility of the spermatozoa and
increased the number of dead spermatozoa (Table 4).

Thus, it appears that chronic exposure to high
environmental temperature has an adverse effect on the
reproductive potential of the avian male as indicated by
decreased motility and increased percent dead spermatozoa in
the heated roosters as compared to the control group (Table
4). It is probable that high temperature would have
suppressed reproductive capacity as indicated by decreased
semen quality and a depression of spermatogenesis processes

(Table 4).

51

In mammals, high temperature and humidity have
deleterious effects on semen quality as evidenced by lowered
initial motility, concentration, total number of
spermatozoa, and an increase in spermatozoa abnormalities
(Johnston et a1., 1963). Mann (1964) reported that dog and
rabbit spermatozoa lost their motility at 45 C. In the boar
exposed to tropical sunlight, Egbunike et a1. (1980)
observed that the sperm motility and concentration
deteriorated.

There is a combined effect of high temperature and
photoperiod on the growth phase of the testes. The high
temperature modifies the photoperiodic effect on male
reproduction (Burger, 1948; Farner and Newalt, 1952;
Kendeigh, 1941; Engels and Jenner, 1956). This combined
effect differs between sexually mature birds and immature
birds. Ingkasuwan and Ogasawara (1966) observed that after
sexual maturation in the chicken, the stimulatory
interaction of light and warm temperature declined in the
maintenance of reproductive potential and in fact depressed
semen production.

According to Cummings and Huston (1976), after the very
sensitive light-required phase high temperature suppressed

reproductive capacity and decreased semen quality and

52

quantity. Temperatures of either 8 or 30 C. caused
significant alteration in semen composition, pH, Na, K, and
Mg. This indicates that there is an alteration in metabolic
activity of semen under cold or hot condition. In the
current investigation, it was observed that feed intake
decreased significantly for the heated roosters, and this is
another factor which affects the quality and quantity of the
semen (Tables 3 and 4). Boone and Hughes (1969) showed
that semen volume decreased during starvation.

There is experimental evidence which suggests that the
vascular pattern of the testes in mammals has a
thermoregulatory function, because spermatogenesis occurs in
the testes at temperatures lower than the body temperature.
This is not true for the chicken, because the rooster testes
are located inside the body, which means that spermatogenesis
must occur at body temperature. The air sacs do not cool
the testes of the rooster and the germinal epithelium has
adapted to the high body temperature (Herin et a1., 1960;
Braganza et a1., 1978). Some investigators suggested that
the mature avian spermatozoa cannot withstand normal avian
temperature, thus, they are stored in the external cloacal
protuberance which contains the convoluted portion of the

vas deferens (Salt, 1954, cited by Hafez, 1964).

53

In mammals, the anatomical features of the testes and
scrotum help to protect these structures against
overheating. Semen quality of the bovine is usually
reduced, in the summer, as a result of environmental
temperature. Jersy and Holstein cattle, Kumauni cattle of
India, buffalo, European breed cattle in India, camel, rat,
guinea pig, and man all exhibited semen quality depression
(Volcani, 1953; Damber et a1., 1980).

D. Adrenal and Testes Weight:

1. Adrenal weight

Data presented in Table 5 indicate that the absolute
weights of the adrenal glands as well as their weight
relative to brain weight are higher in the heated roosters
when compared to the control roosters. The absolute weight
of the adrenals from the heated roosters was 1.6 times
higher than the weight of adrenal glands from the control
birds. An increase of the adrenal weights under heat stress
is in agreement with Hester et a1. (1981) who worked with
White Pekin ducks. In mammals, a relative increase in
actual size of the adrenal has been reported in rats bred
for several generations in conditions of high temperature
and humidity (Clark, 1938; Tepperman et a1., 1943).

The weight of the adrenal gland relative to body weight

54

TABLE 5. THE EFFECT OF TEMPERATURE ON ABSOLUTE
AND RELATIVE ADRENAL GLAND WEIGHTS a

 

 

CONTROL HEATED
(7) (15)

BODY WEIGHT (9) 2139 +/- 0.6 2034 +/— 0.1 b
BRAIN WEIGHT (9) 3.25 +/- 0.081 3.32 +/- 0.042
ADRENAL WEIGHT (9) 0.187 +/- 0.22 0.298 +/- 0.02 b
ADRENAL WEIGHT (mg) 8.75 +/- 0.77 14.66 +/- 0.84 b
AS % BODY WEIGHT
ADRENAL WEIGHT (9) 5.83 +/- 0.74 8.99 +/- 0.46 b

AS % BRAIN WEIGHT

 

a Mean +/- standard error. Number in parentheses refers
to sample size.

b Significantly different from control at P< 0.05

55

was also different with the weight being significantly
greater in the heated group when compared to the control
group. The increase in the adrenal weight in the heated
mature rooster is opposite to the findings of Braganza et
a1. (1978). In their research on S-week old male Japanese
quail exposed to cyclic temperature (10 C to 34 C) there
was a reduction in adrenal weights. These results indicate
that the effect of high temperature on the adrenal weights
is different between mature and immature birds. Tepperman
(1943) pointed out that both weight and size of the adrenal
glands vary considerably with age and physiological
condition of the organism.

2. Testes Weight

In the present experiment, the total weight of the
testes and their weight relative to brain weight were 7.0 gm
and 210 percent for the heated roosters, and 10.2 gm and 319
percent for the control group (Table 6). The absolute
testes weights of the heated group were significantly
smaller than control testes. However, when testes weight
were expressed as a percent of brain weight, this difference
was no longer significant. On the other hand, when testes
weights are expressed as a percent of body weight, the

heated group has significantly lower weights when compared

56

TABLE 6. THE EFFECT OF TEMPERATURE ON ABSOLUTE
AND RELATIVE TESTES WEIGHTS a

 

 

CONTROL HEATED
(7) (15)

BODY WEIGHT (9) 2139 +/- 0.6 2034 +/- 0.1 b
BRAIN WEIGHT (9) 3.25 +/- 0.081 3.32 +/- 0.042
TESTES WEIGHT (9) 10.26 +/- 2.057 7.05 +/— 1.290
TESTES WEIGHT (9) 0.479 +/— 0.0933 0.346 +/— 0.0588 b
AS % BODY WEIGHT
TESTES WEIGHT (9) 319 +/- 67.8 210 +/- 37.6

AS % BRAIN WEIGHT

 

a Mean +/- standard error. Number in parentheses refers
to sample size.

b Significantly different from control at P< 0.05

57

to the control group. This reduction in the testes weight in
the heated roosters is in agreement with Braganza et a1.
(1978) who did experiments with Japanese quail.

E. Histological Features 9; the Testes:

 

 

Testes have exocrine and endocrine functions. Both
functions are affected by ambient temperature. Temperature
affects all phases of semen production. Cummings and Huston
(1976) found that there is an alteration in metabolic
activity of semen from both cold and hot donor birds.

As demonstrated in Table 7, the long-term heating had
an effect on testes histology. It is obvious that the
number of rows of spermatogenic cells inside the
seminiferous tubules in the controls are higher than that of
the heated roosters. The number of rows of cells in the
control is 2.5 times that of the heated group and the
difference is highly significant (Tables 7, and Figures 6,
7, 8, and 9).

Figures 6 and 8 illustrate that the rows of
spermatogenic cells were complete until the lumen of the
seminiferous tubule in the control rooster, while in Figures
7 and 9, the number of rows of spermatogenic cells in the
heated rooster were incomplete with growth stopping at the

secondary spermatocye stage. Higher magnification of the

58

TABLE 7. MORPHOMETRIC MEASUREMENT OF THE TESTES

 

 

PARAMETER CONTROL HEATED

(7) (15)
SEMINIFEROUS TUBULE 82 +/— 0.023 a 67 +/- 0.008 b
VOLUME (% OF TOTAL
VOLUME)
INTERSTITIAL CELL 13 +/- 0.018 29 +/- 0.008 b
VOLUME (% OF TOTAL
VOLUME)
AVERAGE NUMBER OF 4.30 +/- 0.340 42.28 +/- 5.460 b

INTERSTITIAL CELLS

ROWS OF 11.30 +/- 0.204 4.57 +/- 0.140 b
SPERMATOGENIC CELLS

 

a Values are expressed as the mean +/- SE. Numbers in
parentheses refer to sample size.

b Significantly different from control at P< 0.05.

59

seminiferous epithelium in control and heated roosters with
stages of spermatogenic cells are shown in Figures 8 and 9.
This is in agreement with Volcani (1953) who reported that
under summer heat stress, there were signs of degeneration
of the seminiferous tubules in farm animals. Fertilizing
capacity was reduced when there was high ambient
temperature. In fact, high temperature appeared to depress
semen production of birds under both gonadal-stimulatory and
gonadal-nonstimulatory photoperiods (Funk, 1935; Heywang,
1944; Huston, 1975). If spermatogenesis is used as a
criterion for reproductive potential, high temperature has a
depressive effect in chickens (Ingkasuwan and Ogasaware,
1966; Huston, 1975) and in turkeys (Kosin and Mitchell,
1955; Law and Kosin, 1958).

Spermatogenic activity in the present experiment was
determined by counting the number of rows of the
spermatogenic cells, measuring the volume of the
seminiferous tubules, and calculating the surface to volume
ratio of the seminiferous tubules in the testes of the
heated and control roosters. In the heated roosters, there
were depressive effects on the number of spermatogenic

cells, and the volume of seminiferous tubules (Table 7, 8,

60

 

Figure 6. THE SEMINIFEROUS EPITHELIUM LINING A

TESTICULAR TUBULE FROM A CONTROL ROOSTER.

The major stages in spermatozoa formation are (1)
spermatogonia, (2) primary spermatocyte, (3) secondary
spermatocyte, (4) spermatid and (5) spermatozoa.

Numerous intermediate stages can also be seen.

H. and E. x 160.

61

 

Figure 7. THE SEMINIFEROUS EPITHELIUM LINING A TESTICULAR
TUBULE FROM A HEATED ROOSTER.

Most of the spermatozoa stages stop at the secondary

spermatocyte under high heat stress.

In this figure, (1) represents spermatogonia, (2) is a

primary spermatocyte, (3) refers to a secondary spermatocyte

and (4) refers to spermatozoa.

H. and E. x 160.

62

 

Figure 8. HIGHER MAGNIFICATION OF THE SEMINIFEROUS
EPITHELIUM FROM A CONTROL ROOSTER.

The number (1) represents spermatogonia, (2) refers to

primary spermatocyte, (3) is a secondary spermatocyte, (4)

is a spermatid and (5) refers to spermatozoa.

H. and E. x 256.

63

 

Figure 9. HIGHER MAGNIFICATION OF THE SEMINIFEROUS
EPITHELIUM FROM A HEATED ROOSTER.

In this figure, (1) represents spermatogonia, (2) primary

spermatocyte, (3) secondary spermatocyte, (4) spermatozoa.

H. and E. x 256

64

and Figures 7 and 9). This is in agreement with Kosin
and Mitchell 1955; Law and Kosin 1958; Ingkasuwan and
Ogasaware 1966; Huston 1975. The number of rows of
spermatogenic cells inside the seminiferous tubules in the
control group is 2.3 times greater than the number of rows
for the heated roosters, this difference is significant
(Table 7). The percent volume of the seminiferous tubules
from the testes of heated roosters is less than their volume
from the control roosters and the difference is highly
significant (Tables 7 and 8).

The number of interstitial cells in the heated
roosters' testes is 9.8 times the number in the control
birds (Table 7), and the difference is highly significant
Interstitial cell volume as a percent of total volume and
the average number of interstitial cells were higher in the
testes of heated roosters compared to control roosters
(Table 7). Figure 11 shows obvious hyperplasia of
interstitial cells from heated roosters when compared to the
control group (Figure 10). Electron micrographs indicated
that the nuclei of the interstitial cells are more active in
the heated than in the control roosters (Figures 12 and 13).
To confirm the characteristics of interstitial cell

activity, the surface to volume ratio was determined for the

65

control and heated roosters. Table 8 indicates that the
surface to volume ratio of interstitial cell decreased in
the heated roosters. This means that the volume of the
interstital cells in the heated rooster increased as
indicated in Table 8. Hyperplasia and hypertrophy of the
interstitial cells in the heated roosters are in agreement
with reports for the rat (Clegg, 1961) and the human (Rock
et a1., 1965).

The percent interstitial cell volume in the heated
group is significantly greater than the the control value
(Tables 7 and 8). The volume of these cells in the heated
group is 2.3 times the volume in the control group (Figures
10, 11, 12, and 13). The ratio of surface area to volume for
the interstitial cells is 31.49 for the control, and 13.79
for the heated group. This means that the volume of these
cells increased in the heated roosters (Table 8).

Table 8 shows the surface to volume ratios. It is shown
that the surface area to volume ratio of the seminiferous
tubules in the control is 4.88 while it is 5.98 in the
heated roosters. This means that the volume of the
seminiferous tubules decreased in the heated roosters and

that is confirmed in Table 7.

66

TABLE 8: SURFACE TO VOLUME RATIOS OF SEMINIFEROUS TUBULES
AND INTERSTITIAL CELLS OF THE TESTES

 

TISSUE CONTROL HEATED

 

SEMINIFEROUS TUBULES 4.88:1 5.98:1

INTERSTITIAL CELLS 31.49:1 13.79:1

 

67

 

Figure 10. INTERSTITIAL CELLS OF LEYDIG OF A TESTIS
FROM A CONTROL ROOSTER.
The interstitial cells of Leydig occur in the
interstices between three seminiferous tubules.

H. and E. x 256.

68

 

Figure 11. INTERSTITIAL CELLS OF LEYDIG OF A TESTIS

FROM A HEATED ROOSTER.
There is hyperplasia and hypertrophy of Leydig cells when
compared to the control (Figure 10).

H. and E. x 256.

69

 

Figure 12. ELECTRON MICROGRAPH OF A SECTION OF THE TESTIS
FROM A CONTROL ROOSTER.

The surface of the nucleus of the interstitial cell appears

to be wavy and is less active when compared to interstitial

cell nuclei from a heated rooster.

Lead citrate and uranyl acetate x 7,200.

70

 

Figure 13. ELECTRON MICROGRAPH OF A SECTION OF THE TESTIS
FROM A HEATED ROOSTER.

The interstitial cells (2) show hypertrophy with respect to

cellular size and an increase in quantities of Leydig cell

organelles including the mitochondria, golgi membrane and

smooth endoplasmic reticulum. The nucleus (1) appears to be

active.

Lead citrate and uranyl acetate x 7,000.

71

In other words, in the heated roosters when the
surface to the volume ratios of the seminiferous tubules
increased their volume decreased. When this ratio decreased
as it did with the interstitial cells it means the volume
increased (Tables 7, 8, Figures 10 and 11).

In mammals, the anatomical features of the testes and
the scrotum described in the review of literature help to
protect these structures against overheating but, if the
environmental heat load is too high, spermatogenesis is
adversely affected. Histological changes occur in the testes
of animals exposed to high environmental temperature and
these changes have been correlated with changes in semen
quality (Erb et a1., 1942; Mukherjee and Bhattacharya, 1952;
Patrick et a1., 1954; Patrick et a1., 1958; Brown, 1959;
Bielanski et a1., 1961; Sen Gupta et a1., 1963; Rock and
Robinson, 1965), sperm mobility and sperm concentration
(Casady et a1., 1953; Asaj and Vergles, 1961), the state of
the seminiferous tubules and spermatogenesis (Volcani,
1953), spermatozoan abnormalities (Johnston et a1., 1963),
for testicular degeneration and germinal epithelium
destruction (Moor, 1924; Steinberger et a1., 1959),
spermatagonia development (Collins et a1., 1967),

interstitial and Leydig cells (Clegg, 1961).

72

The data of the histological features of the testes
under heat stress of the present experiment features the
following, which is in agreement with the previously
mentioned investigators for avian and mammalian species. In
the testes there was a decrease in the spermatogenic cell
row number, the percent of seminiferous tubules volume, and
in sperm motility, while the percentage of dead sperm, the
percent of interstitial cell volume and the number of
interstitial cells increased.

F. Histological Features pf the Adrenal Gland:

 

 

Histological and histochemical changes occur in the
adrenal glands of animals exposed to tropical conditions
(Bernstein, 1940, 1941; Cramer, 1928; Flexner et a1., 1939;
Schmidt et a1., 1938). An increase absolute and relative
adrenal size has been reported in rats bred for several
generations in conditions of high temperature and humidity
(Clark, 1938; Tepperman et a1., 1943; Kotby et a1., 1967).

Table 9 presents the morphometric measurements of the
volume of medullary and cortical cells of the adrenal in
heated and control roosters. The percentage of the cortical
cell volume is significantly higher in the control group
than the volume in the heated group. The volume of these

cells in the control roosters is 1.1 times that of the

73

volume in the heated group. The percentage volume of the
medullary cells in the heated group is significantly higher
than volume of the control group.

To confirm the increased volume of medullary cells and
decreased volume of cortical cells in the heated roosters,
the surface to volume ratios were calculated for both kinds
of cells in heated and control roosters. It is shown for
the heated roosters that as the surface to volume ratio of
cortical cells increased there was a decrease for the
medullary cells (Table 10). The surface area to volume
ratios for the cortical cells of the adrenal in the control
is 7.33, while it is 8.29 in the heated roosters. This
indicates that the volume of these cells decreased in the
heated roosters (Table 9,and 10 Figure 15). The surface to
volume ratio of the medullary cells is 10.68 in the
control, while it is 9.60 in the heated roosters. This
means that the volume of the medullary cells increased in
the heated group (Table 9 and 10).

Figures 14 and 15 present the types of adrenal
parenchymal cells in control and heated roosters
respectively. Figure 16 illustrates that the blood sinusoid
increased in the adrenal of the heated rooster. The stain is

heavier for the medullary cells as a result of the granular

74

TABLE 9. MORPHOMETRIC MEASUREMENTS OF MEDULLARY
AND CORTICAL CELLS OF THE ADRENAL GLAND

 

PARAMETER

CONTROL
(7)

HEATED
(15)

 

CORTICAL CELL
(8 OF TOTAL VOLUME)

MEDULLARY CELLS
(% OF TOTAL VOLUME)

55 +/- 0.6 a

48 +/- 0.4 b

42 +/- 0.4 b

 

a Values are expressed as the mean +/- SE.

parentheses refer to sample size.

Numbers in

b Significantly different from control at P<0.05.

75

TABLE 10. SURFACE TO VOLUME RATIOS OF CORTICAL AND
MEDULLARY CELLS IN THE ADRENAL

 

TISSUE CONTROL HEATED

 

CORTICAL CELLS 7.33:1 8.29:1

MEDULLARY CELLS 10.69:1 9.60:1

 

76

increase and activity of these kinds of cells.

Electron micrographs (Figures 16 and 17) indicated that
the medullary cells in the heated roosters were more active
than those in the control group. Two distinct types of
medullary cells could be distinguished. One type of
medullary cell released norepinephrine (NE) and the other
type of medullary cell released epinephrine (E).

There is some controversy as to the relative proportion
of cell types in the adrenal. In the present experiment,
the percent volume of cortical and medullary cells was 55 %
and 37 % respectively, for the control roosters, while the
percent volumes were 48 % and 42 %, respectively, for the
heated group (Table 9). These findings are in disagreement
with Harvey et a1. (1986) who stated that the medullary
cells constitute about 15 to 25 % of the adrenal tissue,
while the cortical tissue accounts for 70 to 80 % of the
avian adrenal. Several authors vary in their recording of
the percent of the cortex of the adrenal gland for the male
chicken. Sturkie (1965) quoted 40% according to Kar (1947)
and Sauer and Latimar (1931) reported 44.2%. Oakberg (1951)
reported that the percentages of adrenal cell types were 46
% for the medulla and 50 % for the cortex in lZO-day old

White Leghorn male cockerels.

77

 

Figure 14. ADRENAL PARENCHYMAL CELLS FROM A CONTROL ROOSTER.
The adrenal parenchyma composed of cortical cells (1) and
medullary cells (2).

H. and E. x 256.

78

 

Figure 15. ADRENAL PARENCHYMAL CELLS FROM A HEATED ROOSTER.
Blood sinusoids (3) are interspersed among cortical cells
(1) and medullary cells (2).

H. and E. x 256.

79

 

Figure 16. ELECTRON MICROGRAPH OF THE ADRENAL PARENCHYMA

FROM A CONTROL ROOSTER.

Features include (1) norepinephrine containing cell, (2)
epinephrine storage cell (3) the nucleus, (4) sinusoid
capillary and (5) nucleolus.

Lead citrate and uranyl acetate x 7,200.

 

Figure 17. ELECTRON MICROGRAPH OF CHROMAFFIN

CELLS FROM A HEATED ROOSTER

Features include (l)norepinephrine containing cell (2)
epinephrine storage cell, (3) nucleus and (4) nucleolus.

Lead citrate and uranyl acetate x 4,500.

81

In the present experiment, the medullary volume
averaged 67 percent of the cortex volume for the control
roosters, while the medullary volume for the heated group
averaged 88% of the cortex volume. This is close to the
value reported by Latimar (1925) who mentioned that the
medullary cells averaged 71 percent of the cortex. From
this contrast in data, it could be concluded that the
percentages of the cortical and medullary cell tissue
depends to a great extent on the physiological condition of
the animal, the circumstances under which it is living
including the temperature, age, sex, and the method of
determination. All of these factors above combine together

for their effect on the cortical and medullary percentages.

APPENDIXES

APPENDIX A

LIVE-DEAD STAIN

NIGROSIN-EOSIN STAIN

82

LIVE-DEAD STAIN

NIGROSIN-EOSIN STAIN

Sodium Glutamate 1.920 g.
Potassium Citrate 0.128 g.
Sodium Acetate 0.513 9.

Magnesium Dichloride 0.068 g.

Sodium Chloride 0.167 g.
Eosin Blue 1.000 g.
Distilled Water 100.000 ml.

The stain is then filtered through
No. 2 filter paper.

APPENDIX B
PROCEDURE FOR THE DETERMINATION

OF SPERM CELL CONCENTRATION

83

The sperm cell count was obtained by using a bright-
line hemocytometer (American Optical Corporation,
Buffalo, NY 14215) and a light microscope.

Samples were prepared by filling the tip of a RBC
pipette to the 0.5 mark with semen by capillary
action.

A 2 percent formalin solution in 0.085% saline was
used to dilute the semen to the pipette's 101 mark.
This gave a dillution of 200. The formalin immobilizes
the spermatozoa, thus facilitating easy counting.

The samples counted were done in duplicate. Each
sample, consisted of 5 squares or counting chambers (l
x 0.1 mm), was located on opposite sides of the
hemocytometer. Each chamber had a depth of 0.1 mm and
consisted of 16 small squares. Thus a total of 80
squares was counted.

A cover slip was placed over each replicate.

From the pipette, a small amount of sample was
released at one side of each cover slip, from which
the sample spread, covering all the counting
chambers.

The sperm cell counts used at any point on the
standard curve was the average of the two replicates

for a particular sample.

84

The following formula was used for the calculations:

Sperm cell/cubic mm =3 pf cells counted x dilution x 4000
# of small squares counted

 

number counted

5 20 x 4000
80

 

number of cells counted x 10,000

concentration of sperm cells inseminated per volume =
number of sperm cells/cubic mm. x number of cubic

mm/cc (ml) x volume.

APPENDIX C

PROCEDURES FOR

HISTOLOGICAL STUDIES

I.

A.

3.

85

PREPARATION OF TISSUE FOR HISTOLOGICAL STUDIES BY LIGHT

MICROSCOPE:

Fixation of Tissue

The

tissue was fixed immediately after dissection

in Bouin's Solution for 24 hours.

Bouin's Solution:

Picric acid, saturated aqueous solution 750.0 ml.
37 - 40 % Formalin 250.0 m1.
Glacial Acetic Acid 50.0 ml.
Wash several times in 50 % ethanol for 4 to 6

hours, agitating constantly, to insure proper

removal of the picric acid. Then, store in 70%
ethanol. Since paraffin is not miscible with
water, the tissue must be dehydrated and then

cleared in solution miscible with paraffin before

impregnation.

The dehydration process continues by upgrading the

ethanol to absolute ethanol by the following

procedure:

a. 70 % ethanol for 2 hrs.
b. 80 % ethanol for 2 hrs.
c. 80 % ethanol for 2 hrs.
d. 95 % ethanol for 2 hrs.
e. 95 % ethanol for 2 hrs.
f. 100 % ethanol for 2 hrs.

9. 100 % ethanol for 2 hrs.

86

h. Xylene for 2 hrs.

i. Xylene for 2 hrs.

j. Paraffin for 2 hrs.

k. Paraffin for 2 hrs.
Transfer the tissue to melted wax under vacuum for
30 - 45 minutes to get rid of the air bubbles.
Embed the tissue in melted paraffin in certain size
of metal mold. When the paraffin is solidified, it
provides a firm medium for keeping intact all parts
of the tissue when sections are cut.
Cut sections from the paraffin blocks into slices 5

microns thick with a microtome and sharp knife.

Hematoxylin-Eosin Saining of Tissue

1.

Clean slides with acid alcohol (70 % alcohol and
hydrochloric acid).

Place a small drop of albumin on the slide and
spread. Place slide on slide warmer. Label slide.
Place water on slide.

A slide is made of every tenth section. This slide
is returned to the slide warmer for about 3 minutes.
The section is then carefully rolled with a damp

paper towel to remove air bubbles.

87

6. The sections are then stained by the following

steps:

a.

The slide is placed in xylene for 3
minutes, 100% ethanol for 3 minutes,100%
ethanol for 1 minute, 95% ethanol for
3 minutes, 80% ethanol for 3 minutes,
70% ethanol for 3 minutes.

Place the slide in henatoxylin for 1
minute and then transfer.

Place the slide in acid alcohol to get
the blue out.

Place the slide into water for 30 minutes
until the purple color goes to blue.
Place the slide into Eosin.

Blot the slide to avoid contamination.
Use an upgrade of alcohol by placing the
slide in 70% ethanol for 3 minutes,
95% ethanol for 3 minutes, and 100% ethanol
for 3 minutes.

Place the slide in xylene.

Immediately after xylene, put a drop of
paramount over tissue. Cover slide with
a cover slide. Then, place slide on the

hot plate to dry.

88

II. PREPARATION OF TISSUE FOR ELECTRON MICROSCOPE

HISTOLOGICAL STUDIES

1.

10.

11.

Fix the tissue immediately in 2 % Glutaraldehyde
and 2 % Osmium Tetroxide at 4 C for 3 to 4 hours.
Dehydrate in upgrade acetone. Place tissue in 75%
acetone for 15 minutes, 95% acetone for 15 minutes,
and 100% acetone for 15 minutes. Let tissue sit
overnight.

Place tissue in one part acetone to one part epoxy
resin for 2 to 3 hours (or Optional overnight).
The tissue was embedded in a mold using 100% fresh
epoxy resin making sure that the sample was at the
edge of the mold. Place tissue sample under vacuum
for 1 hour, or overnight.

Place mold into a desiccator for l to 2 hours.

Poke out any air bubbles in the resin.

Polymerize sample for 48 or more hours in oven at
65 C.

Remove mold and allow to cool 10 to 15 minutes
before popping out each block.

Store the block in a desiccator.

Use a sharp glass knife when using the microtome to
cut the section 1 micron thick.

Hold the section on a grid and stain it with lead

citrate and uranyl acetate.

APPENDIX D

MODEL AND STATISTICAL

ANALYSIS TESTS

89

I. RESPONSE: Y = U + Ti + R + P + (TP) + E
ijk (i)j k jk (ijk)
U = The mean

Ti = Fixed effect of the temperature

R
(i)j = Rooster nested in temperature
treatment
P
k = Period
(TP) = interaction between temperature & period
E

(ijk) =Residual error
II. T-TEST FOR SIGNIFICANT DIFFERENCE IN TREATMENT

t-test (table A.4, from Gill, 1978)

 

t = (Y - Y ) / [8 7/(l/r ) + 1/r )]
l 2 l 2

 

_ n V r
S _ +7/{[Zr'flf -— $3.975] {gfii -(2;3..-)2/4j} my“)
FEOE Gill, 1981. equation number 1.83
III. SEMEN CHARACTERISTICS

a. For sperm motility, the normal t-test was used.

b. For percent live and dead sperm the percentages
were transformed to the proportions by using
the following equation:

live percent / 100 = proportion.
Then, logs of these proportions were obtained by

using the following equation: Y = log [F/(l - F)]

90

F = the proportion.
Then, use normal t-test.
c. For counting sperms, the following equation was
used: Y = log (count)
Then, the t-TEST was used on the obtained number.
d. For Spermatocrit:
The same procedure as outlined in b.
IV MORPHOMETRIC ANALYSIS
Morphometry implies the use of quantitative data in the
description of structural features. Morphometric data can
be obtained by a variety of measuring procedures performed
on any type of specimen. It is a branch of applied
mathematics used for the three dimensional analysis of
organs and materials from two-dimensional measurements.
Morphometry is the application of stereologic axioms
allowing quantitative analysis of volume, surface, and
number of cells, or organelles.
1. VOLUME ESTIMATION
From a test specimen, a selection of an
appropriate number of sections is ascertained. A
regular network of points is superimposed on each
section to estimate the volume ratios. The proportion

of the volume occupied by a component C (V ) is
vc

91

estimated from the proportion of test points lying on

C (P ). This proportion is the number of test points
pc

coinciding with C (PC) divided by the total number of

test points, P. Thus,

V P = PC/P

vc Pc
In the following example, nine fields are counted
using a grid that has 36 points. Of these fields 81
points are found to lie on profiles of a component C.

P , the point fraction of C, is then:
PC

Pc/P = 81/ (9 x 36) = 0.25
Hence, the component C occupies 25 % of the volume of
the speciman. As this is a statistical estimation it
is meaningless without some indication of the
reliability of the result. This is expressed in terms

of the standard deviation:

o’= P (1-P)/P
c 7/ Pc Pc

which gives the following example:

 

 

C

o’ = / 0.25 ( 1-0.25) / (9 x 36)

 

7/ 0.1875 / 18
0.024

Thus, P = 0.25 + / - 0.024. In other words, 68.4 %
pc

 

of

Th

Cl‘

 

 

92

of the results should fall between 0.226 and 0.274.
The standard deviation serves a useful purpose in the
choice of sampling size and method (Gaunt and Gaunt,
1978). The grid which was used in this experiment has
168 points and measured, for each section, 5 fields at
random (Figure 18).

V. ESTIMATION OF SURFACE-TO-VOLUME RATIOS:

The surface to volume ratio of structures proves
to be a very useful measure. On the one hand, it is
related to geometric features of the structures, and
on the other hand, it can often serve as a direct
measure of some physiological properties of these
structures since it gives the size of their contact
area with the surrounding per mass of structure. It
is essentially a straight forward combination of
point-counting volumetry with surface estimation by
line intersection. The test system consists of short
lines of equal length Z, which again are randomly
placed on random sections. The two end-points of
these test lines are used as markers for volumetry:
end-points lying on sections of structures are counted
as hits and recorded as Pi. Simultaneously, that is

without displacement of the test system, the number of

93

 

?

F-—H r———4 h——4 h—-H t———t hr-d g
r———1 p——4 U——-4 h——« r———t h——-§
h——4 r—-« F-;4 P—-* *-* "-‘ §
p——q b——d h——q h——H t———1 h——d é
h—-—4 h——n t--1 h——4 t——-4 h——~§
tu——-g t——-t h——-| h—d i——-t l-—-i g
h——4 h——d h——4 h——n h—-4 h-—4 é
h——4 h—-4 h——4 h——d h——d h——d§
h—-4 b——4 h——4 h——4 h——q p——4 g
h———t h——d .h———t h——4 &———t h——4§

 

i («444 M1“

Figure 18. MORPHOMETRIC GRID

Minimum-w: to' .t s} stem M I“

('altlwatiom .! is‘ltf'ilt ut shun lute wvnwm
llcmlmital (mm v. with Lid

Vemwl [Lune \\ ucltlt l.‘.| .‘al

94

intersections, Nzi, of test lines and surface contour
of the structures is recorded. The surface-to-volume
ratio Si /Vi, of individual structures i, follows this

form:

Si / Vi = 4(Ni) / Z (Pi)

Si = Surface area of structure 1

Vi = Volume of structure 1

Ni = Number of structure i

Pi = Collection number of test points lying

in i.
Z = Short lines of equal length

(Weibel et a1., 1966).

BIBLI OGRAPHY

Ar.

Asl

As]

Be:

Be]

Be:

Ber

Ben

sir

Bla.

95

Arad, Z. and J. Marder, 1982. Strain difference in heat
resistance to acute heat stress, between the Bedouin
Desert Fowl, the White Leghorn, and their crossbreeds.
Comp. Biochem. Physiol. 72A 191-193.

Ashizawa, K., and H. Nishiyama, 1983. Prolonged survival
of fowl spermatozoa in the oviductal tissues in organ
culture. Br. Poult. Sci. 24:27-32.

Ashizawa, K., Y. Tokudome, O. Keizo, 1983. Scanning elcetron
microscopy of fowl oviduct cells and HeLa and BHK-21
cells incubated with or without fowl spermatozoa in
Vitro. Poultry Sci. 62:1711-1714.

Ben Nathan, D., E. D. Heller and M. Perek, 1976. The effect
of short heat stress upon leucocyte count, plasma
corticosterone level, plasma and leucocyte ascorbic
acid content. Br. Poult. Sci. 17:481-485.

Bernard, C. 1878. Lecons sur les phenomenes de la vie
communs aux animaux et aux vegetaux. J. B. Bailliere
et Fils Librairie, Paris. 32:67, 111-114, 123-124.
Cited by Fulton, J. F., and L. G. Wilson, 1966.
Selected Reading 13 The History 9: Physiology.
Charles C.Thomas Publisher. Springfield, Virginia.

 

Bernstein, J. G., 1940. Experimental variations in the
histological structure of the adrenal cortex of the
albino rat as a result of artificial fever induced by
ultra-high frequency radio emanations. Am. J. Anat.
66:177-196.

Bernstein, J. G., 1941. The effect of thermal environment
on the morphology of the thyroid and adrenal cortical
glands in the albino rat. Endocrinology) 28:985-998.

Beuving, G., and G. M. A. Vonder, 1978. Effect of stressing
factors on corticosterone levels in the plasma of
laying hens. Gen. Comp. Endocrinol. 35:153-159.

Birrenkott, G. P. Jr., and A. R. Ezzat, 1983. Response of
broilers and capons to heat stress following adrenal
manipulation. Poultry Sci. 62:702-704.

Blackshaw, A. W., 1977. Temperature and seasonal influences.
in: The Testis. Advances in Physiology, Biochemistry

and Function. 4:517-545 Eds: A. D. Johnson and W. R.
Games, Academic Press, New York.

 

 

J.

Bligh

Boone

Boone

Brag

Burg

BUIJ

Cas

Cha

C15

C1;

C1

96

Bligh, J. and K. G. Johnson, 1973. Glossary of terms for
thermal physiology. Journal of Applied Physiology.
35:941.

Boone, M. A. and T. M. Huston, 1963. Effect of high
temperature on semen production and fertility in the
domestic fowl. Poultry Sci. 42:670-676.

Boone, M. A. and B. L. Hughes, 1969. Starvation and its
effects on semen quality. Poultry Sci. 48:1723-1729.

Braganza, A. and W. O. Wilson, 1978. Effect of acute and
chronic elevated air temperature. Constant (23 C) and
cyclic (10-34 C) on brain and heart norepinephrine of
male Japanese Quail. Gen. Comp. Endocrinol. 36:233-
237.

Burger, J. W., 1948. The relation of external temperature
to spermatogenesis in the male starling. J. Exp. 2001.
109:259-266.

Burrows, W. T., and I. L. Kosin, 1953. The effects of
ambient temperature on production and fertilizing
capacity of turkey spermatozoa. Physiol. 2001. 26:131-
146.

Casady, R. B., R. M. Myers and J. E. Legates, 1953. The
effect of exposure to high temperature on
spermatogenesis in dairy bull. J. Dairy Sci. 36:14:23

Chaffee, R. R. J., and J.C. Roberts, 1971. Temperature
acclimation in birds and mammals. Ann. Rev. Physiol.,
33:155.

Clark, A., 1938. Climate, diet and toxic substances in
their association with adrenal condition. J. Trop. Med.
Hyg. 41:315-316.

Clark, R. N., T. J. Sexton, and M. A. Ottinger, 1982.
Effects of holding temperature and storage time on
respiratory rate, motility, and fertility of chicken
and turkey semen. Poultry Science 61:1912-1917.

Clark, R. N., M. R. Bakst, and M. A. Ottinger, 1984.
Morphological changes in chicken and turkey spermatozoa

incubated under various conditions. Poultry Science
63:801-805.

Cle

Col

Co]

Crz

Cra

Cu]

Da

Da

De

EC

97

Cleggr E. J., 1961. Further studies on artificial

cryptorchidism: Quantitative changes in the
interstitial cells of the rat testis. J. Endocrin.
21:433.

Collins, K. J. and J. S. Weiner, 1968. Endocrinological
aspects of exposure to high environmental temperatures.
Physiol. Rev. 48:785-839.

Collins, P. and D. Lacy, 1967. Effect of heat on the
germinal epithelium and steroid metabolism of the
seminiferous tubules and intestitial cells of the rat.
Gen. Comp. Endocrinol. 9:509.

Cramer, W., 1919. Observation on the functional activity of
the suprarenal gland in health and in disease. Imp.
Cancer Res. Fund 6th Sci. Rept. 1-19.

Cramer, W., 1928. Fever, Heat Regulation, Climate and The
Thyroid-Adrenal Apparatus. Longmans, London.

 

Cummings, V. T. and T. M. Huston, 1976. The influence of
short term exposure to two different environmental
temperatures on electrolyte concentrations of fowl
semen. Poultry Sci. 55:857-861.

Dale, N. M., and H. L. Fuller, 1980. Effects of diet
composition on feed intake and growth of chicks under
heat stress. II. Constant versus cycling temperatures.
Poultry Sci. 59:1434-1441.

Damber, J. E., A. Bergh, and P. O. Janson, 1980. Leydig cell
function and morphology in the rat testis after
exposure to heat. Andrologia 12:12-19.

Deviche, P. 1983. Interactions between adrenal function and
reproduction in male birds. In: Avian Endocrinology:
Environmental and Ecological Perspectives. Eds:Mikami,
S., K. Homma, and M. Wada. Japan Sie. Soc. Press,
Tokyo/Springer-Verlag, Berlin, pp.243-254.

 

 

Edens, F. W., and H. S. Siegel, 1975. Adrenal Responses
in high and low ACTH response lines of chickens during
acute heat stress. Gen. and Comp. Endocrinol. 25:64-73.

98

Edens, F. W., 1976. Symptoms of adrenal insufficiency in
young chickens exposed to a high ambient temperature.
Poultry Sci. 55:1594.

Edens, F. W., 1978. Adrenal cortical insufficiency in young
chickens exposed to a high ambient temperature.
Poultry Sci. 57:1746-1750.

Edens, F. W., 1983. Effect of environmental stressors on
male reproduction. Poultry Sci. 62:1676—1689.

Egbunike, G. N., and T. I. Dede, 1980. The influence of
short-term exposure to tropical sunlight on boar semen
characteristics. Int. J. Biometeor. 24:129-135.

Engels, W. L. and C. E. Jenner, 1956. The effect of
temperature on testicular recandescence in Juncos at
different photoperiods. Biol. Bull. 110:129-137.

Farner, D. S., and L. R. Newalt, 1952. The relative roles of
roles of photoperiod and temperature in gonadal
recrudescence in male Zontotrichia Leucophyrs Gambelii.
Anat. Rec. 113:612.

 

Farrell, D. J. and S. Swain, 1977. Effects of temperature
treatments on the energy and nitrogen metabolism of fed
chickens. Br. Poultry Sci. 735-748.

Fawcett, D. W., 1986. Adrenal Glands and Paraganglia. In:
A Textbook g: Histology by Fawcett, D. W. (ed.). W. B.
Saunders Co., London.

 

Fiorica, V., P. F. Iampietro, E. A. Higgins, and R. Moses,
1967. Sympathico-adrenomedullary activity in dogs
during acute heat exposure. J. Appl. Physiol. 22:16-20.

Flexner, L. B., and A. Grollman, 1939. The reduction of
osmic acid as an indicator of adrenal cortical activity
in the rat. Anat. Record 75:207-221.

Freeman, B. M., 1982. Stress non-responsiveness in the
newly-hatched fowl. Comp. Biochem. Physiol. 72 A:251-
253.

GI

G1

G1

Ha

Ha:

Hex

99

 

Freeman, B. M., 1983. Body temperature and
thermoregulation. In: Physiology and Biochemistry 9f
the Domestic Fowl. Ed: Freeman, B. M., Vol 4.

Academic Press, New York.

Funk, E. M., 1935. Factors influencing hatchability in the
domestic fowl. Missouri Agric. Exp. Sta. Bull. 314.

Gaunt, P.N. and W. A. Gaunt, 1978. Three Dimensional
Reconstruction 12 Biology. Pitman Medical Publishing
Co. Ltd. P.O. Box 7, Tunbridge Wells Kent, TNlXH,
England.

 

Gill, J. L., 1978. Design and Analysis 9; Experiments ig
the Animal and Medical Sciences. Vol 3. Iowa State
University Press, Ames, Iowa.

 

 

Gill, J. L., 1981. Design and Analysis pf Experiments la
the Animal and Medical Sciences. Vol 1, Iowa State
University Press, Ames, Iowa.

 

 

Ghosh, A., 1980. Avian adrenal medullazstructure and
function. In: Avian Endocrinology. Eds: A. Epple and
M. H. Stetson. Academic Press, New York, New York.

Giesen, A. F. and T. J. Sexton, 1983. Beltsville poultry
semen extender. 9. Effect of storage temperature

on turkey semen held eighteen hours. Poultry Sci.
62:1305-1311.

Glover, T. D., 1955. Some effects of scrotal insulation on
the semen of the Ram. Proc. Soc. Study of Fertility.
7:66—75.

Hafez, E. S., 1964. Effects of high temperature on
reproduction. Int. J. Biometeror. 7:223-230.

Harvey, S., C. G. Scanes and K. I. Brow, 1986. Adrenals.
In: Avian Physiology. Ed:Sturkie, P. D. Springer-
Verlag, New York.

Herin, Reginald A., N. H. Booth and R. M. Johnson, 1960.
Thermoregulatory effects of abdominal air sacs on
spermatogenesis in domestic fowl. Am. J. Physiol.
198:1343-1345.

100

Hester, P. Y., S. G. Smith, E. K. Wilson, and F. W. Pierson,
1981. The effect of prolonged heat stress on adrenal
weight, cholesterol, and corticosterone in White Pekin
ducks. Poultry Sci. 60:1583-1586.

Heywang, B. W., 1944. Fertility and hatchability when the
environmental temperature of chickens is high. Poultry
Sci. 23:334-339.

Hillman, P. E., N. R. Scott, and A. Van Tienhoven, 1985.
Physiology responses and adaptations to hot and cold
environments. In: Stress Physiology in_Livestock. Vol
III. Poultry. Ed: Yousef, M. K. CRC Press Inc.,
Boca Ratan, Florida.

 

 

 

Hoffmann, E., and C. S. Shaffner, 1950. Thyroid weight and
function as influenced by environmental temperature.
Poultry Sci. 29:365—376.

Hurwitz, S., E. Wax, and I. Bengal, 1983. Performance and
energy needs of 20 week old male turkeys at different
environmental temperatures. Poultry Sci. 62:1327-1329.

Huston, T. M., 1975. The effects of environmental
temperature on fertility of the domestic fowl. Poultry
Sci. 54:1180-1184.

Ingkasuwan, P., and F. X. Ogasawara, 1966. The effect of
light and temperature and their interaction on the
semen production of White Leghorn males. Poult. Sci.
45:1199.

Johnson, A. L., 1986. Reproduction in the male. In: Avian
Physiology ed: Sturkie, P. D. Springer-Verlag, New
York, New York.

 

Johnston, J. E., H. Naelapaa, and J. B. Frye Jr., 1963.
Physiological responses of Holstein Brown, Swiss and
Red Sindhi crossbred bulls exposed to high temperatures
and humidities. J. Animal Sci. 22:432-436.

Kadono, H., and E. L. Besch, 1978. Telemetry measured body
temperature of domestic fowl at various ambient
temperatures. Poultry Sci. 57:1075-1080.

101

Kar, A. B., 1947. The adrenal cortex, testicular reactions
in the fowl: The effect of castration and replacement
therapy on the adrenal cortex. Anat. Rec. 99:177-197.

Kato, M., and T. Konishi, 1968. The effect of light and
temperature on the testicular growth of the Japanese
quail. Poultry Sci. 47:1052-1056.

Kendeigh, S. C., 1941. Length of day and energy
requirements for gonad development and egg laying in
birds. Ecology 22:237-248.

Kennedy, G. C., 1953. The role of depot fat in the
hypothalamus control of feed intake in the rat. Proc.
Roy. Soc. (London) Ser. B. 140:578-598.

Kosin, I. L., M. S. Mitchell, and E. St. Pierre, 1955.
Ambient temperature as a factor in turkey reproduction.
1. The effect of preheating males and females on their
subsequent breeding pen performance. Poultry Sci. 34:
484-496. 2. The effect of artificially lowered air
temperature on the breeding activity of males in late
spring and in summer. Poultry Sci. 34:499-505.

Kosin, I. L., 1958. Metabolism of turkey semen as affected
by the environment of donor birds. Poultry Sci.
37:376-388.

Kotby, S. and H. D. Johnson, 1967. Rat adrenal cortical
activity during exposure to a high (34 C) ambient
temperature. Life Science 6:1121-1132.

Lamoreux, W. F., 1943. The influence of different amounts
of illumination upon the production of semen in the
fowl. J. Exp. 2001. 94:73-95.

Latimer, H. B. and M. F. Landwer, 1925, The relative
volumes and the arrangement of the cortical and the
medullary cells of the suparenal gland of the chicken.
Anat. Rec., 29, 389.

Law, G. R. J. and I. L. Kosin, 1958. Seasonal reproductive
ability of male domestic turkeys as observed under two
ambient temperature. Poultry Sci. 37:1034-1047.

M

102

Long, J. A., 1983. The adrenal. In: Histology, Cell and
Tissue Biology. ed:Weiss, L. Elsevier Biomedical, New
York, New York.

 

Mann, T., 1964. The Biochemistry gf Semen and pf the Male
Reproductive Tract. Methuen and Co., Ltd., London.

 

 

 

McDougald, L. R., and T. E. McQuistion, 1980. Mortality
from heat stress influenced by anticoccidal drugs.
Poultry Sci. 59:2421-2423.

Moore, C. R., 1924. Properties of the gonads as controllers
of somatic and psychical characteristics. VI.
Testicular reactions in experimental cryptorchidism.
Am. J. Anat. 34:269-316.

Moore, C. R., 1951. Experimental studies on the male
reproductive system. J. Urol. 65:497-512.

Nir, I,. D. Yam and M. Perek, 1975. Effects of stress on
the corticosterone content of the blood plasma and
adrenal gland of intact and bursectomized Gallus
Domesticus. Poult. Sci. 54:2101-2110.

 

Oakberg, E. F., 1951. Genetic differences in quantitative
histoloy of the adrenal, organ weights, and interorgan
correlations in White Leghorn chickens. Growth 15:57-
78.

Oluyemi, J. S., and F. A. Roberts 1979. Poultry Production
13 Warm Wet Climates. The Macmillan Press Ltd. London

 

 

 

Renden, J. A., and G. R. McDaniel, 1984. Reproductive
performance of broiler breeders exposed to cycling high
temperature from 17 to 20 weeks of age. Poultry
Sci. 63:1481-1488.

Rhynes, W. E., and L. L. Ewing, 1973. Testicular endocrine
function in herford bulls exposed to high ambient
temperature. Endocrinology 92:509-515.

Rock, R., and D. Robinson, 1965. Effect of induced
intrascrotal hyperthermia on testicular function in
man. Am. J. Obstet. Gynecol. 93:793-809.

Sa

Se

rn

103

Salisbury, G. W. and J. R. Lodge, 1962. Metabolism of
spermatozoa. Advances in Enzymology 24:35-104.

Salt, W. R. 1954. The structure of the cloacal
protuberance. Auk. 71:64-73.

Sauer, F. C., and H. B. Latimer, 1931. Sex differences in
the proportion of cortex and medulla in the chicken
suprarenal. Anat. Rec. 50:289.

Schmidt, I. G., and L. H. Schmidt, 1938. Variations in the
structure of adrenals and thyroids produced by
thyroxine and high environmental temperatures.
Endocrinology 23:559-565.

Scott, M. L., M. C. Nesheim, and R. J. Young, 1976. In:
Nutrition 9: the Chicken. M. L. Scott and Associates.
Ithaca, New York.

 

 

Siegel, H. S., and P. B. Siegel, 1961. The relationship of
social competition with endocrine weights and activity
in male chickens. Animal Behav. 9:151-158.

Siller, W. G., P. W. Teague, and G. M. Mackenzie, 1975. The
adrenal cortico—medullary ratio in the fowl. Br.
Poult. Sci. 16:335-342.

Skinner, J. D., and G. N. Louw, 1966. Heat stress and
spermatogenesis in Bos indicus and Bos taurus cattle.
J. Appl. Physiol. 21:1784-1790.

 

 

Steinberger, E. and W. J. Dixon, 1959. Some observations on
the effect of heat on the testicular germinal
epithelium. Fertility and Sterility 10:578-595.

Sturkie, P. D., 1965. Avian Physiology. Comstock Publishing
Associates, Ithica, N.Y.

 

Taher, A. I., E. W. Gleaves, and F. B. Mather, 1985.
Feeding pattern responses to changes in dietary energy
or environmental temperature in the domestic fowl.
Poultry Sci. 64:986-990.

104

Takeda, A. 1982. Studies on reversible inactivation of cock

spermatozoa by temperature: 1. Effect of several
factors on revesible inactivation. Biol. Abstr., 74
:8126.

Tepperman, J., F. L. Engel, and C. N. H. Long, 1943. A
review of adrenal cortical hypertrophy. Endocrinolgy
32:373-402.

Unsicker, K., 1973. Fine structure and innervation of the
avian adrenal gland. I. Fine structure of adrenal
chromaffin cells and ganglion cells. Z. Zellforsch.
Mikrosk. Anat. 145:389.

Vo, K. V., M. A. Boone, B. L. Hughes, and J. F. Knechtges,
1980. Effect of ambient temperature on sexual maturity
in chickens. Poultry Sci. 59:2532-2535.

Volcani, R., 1953. Seasonal variations in spermatogenesis
of some farm animals under the climatic conditions of
Israel. Bull. Res. Council of Israel. 3:123-126.

Weibel, E. R., S. K. Ganzagne and F. S. Water, 1966.
Practical stereological methods for morphometric
cytology. J. Cell Biol. 30:23-28.

Whelen, R. F. and I. S. De La Lande, 1963. Action of
adrenalin on limb blood vessels. Brit. Med. Bull.
19:125-131.

Whittow, G. C. 1965. Avian Physiology, Cornell Univ. Press,
Ithica, New York. pp. 186-271.

 

Wilson, E. K., F. W. Pierson, P. Y. Haster, R. L. Adams, and
W. J. Stadelman., 1980. The effects of high
environmental temperature on feed passage time and
performance traits of White Pekin ducks. Poultry Sci.,
59:2322-2330.

 

Yousef, M. K. and H. D. Johnson, 1985. Endocrine system and
thermal environment. In: Stress Physiology 13
Livestock, Ed: Yousef, M. K., Vol 1. CRC Press, Inc.
Boca Raton, Florida.