THE RELATIONSHIP OF THYROID ACTIVITY TO LACTATION, GROWTH, AND SEX IN SHEEP BY QNKAR NATH SINGH A THESIS Submitted to the School of Graduate Studies of Michigan State College of Agriculture and Applied Science in partial fulfillment of the requirements for the degree of DOCTOR OF PHILOSOPHY Department of Animal Husbandry Year 1954 ProQ uest Number: 10008428 All rights reserved IN FO R M ATIO N TO ALL USERS The quality o f this reproduction is dependent upon the quality o f the copy subm itted. In the unlikely event that the author did not send a com plete m anuscript and there are m issing pages, these will be noted. Also, if m aterial had to be removed, a note will indicate the deletion. uest ProQ uest 10008428 Published by ProQ uest LLC (2016). Copyright of the Dissertation is held by the Author. All rights reserved. T his w ork is protected against unauthorized copying under Title 17, United States Code M icroform Edition © ProQ uest LLC. ProQ uest LLC. 789 East Eisenhower Parkway P.O. Box 1346 Ann Arbor, Ml 4 8 1 0 6 - 1346 ACKNOWLEDGMENTS The author is deeply indebted to Dr. H. A. Henneman, Assistant Professor of Animal Husbandry, Reineke, and Dr. E. P. Professor of Physiology and Pharmacology, for their continued interest and suggestions during the course of this investigation. The writer wishes to express his sincere appreciation to Dr. R. H. Nelson, Head of the Department of Animal Hus­ bandry, for his valuable help and criticism in the preparation of this manuscript. He is grateful to Michigan State College for awarding the Graduate Council and Berckmans* deWeert Foundation Fellowships which enabled him to complete this investigation. ONKAR NATH SINGH Candidate for the degree of DOCTOR OF PHILOSOPHY Final Examination: June 17, 1954, 1:30 P.M., Room 201, Agricultural Hall. Dissertation: The Relationship of Thyroid Activity to Lacta­ tion, Growth and Sex in Sheep. Outline of Studies: Major subject: Animal Breeding Minor Subjects: Physiology, Statistics Biographical Items: Born, January 11, 1931, Farahada, Shahabad, Bihar, G.B.V.C., Bihar Veterinary College, Patna, India. India, 1950. Sahitya Ratna, Rastriya Vishwavidyalay, Nagpur, 1950. India, P.G. (An. Hus.), Indian Veterinary Research Institute, Izatnagar, 1951. M.S. (An. Hus.), Michigan State College, East Lansing, U.S.A., 1953. Experience: Veterinary Asst. Surgeon, Govt, of Bihar, India, 19 51. Graduate Council Fellowship, Michigan State College, 1952-53. Berckmans' de Weert Foundation Research Fellowship, Michigan State College, 1953-54. Member of American Society of Animal Production, of Sigma X i . and Society TABLE OF CONTENTS PAGE ................................................. 1 OBJECTIVES ................................................... 4 INTRODUCTION REVIEW OF LITERATURE ...................................... EXPERIMENTAL PROCEDURE .................................... 5 15 RESULTS AND DISCUSSION: A. Refinement of a Technique for Measuring Thyroid Secretion Rate ................................. B. The Relationship of Thyroid Output Rate to Lac­ tation in Ewes as Measured by Growth Rate of the Lambs ........................................ C. Thyroid Secretion Rate and G r o w t h .............. 22 57 46 D. Thyroid Secretion Rate and S e x ................... 51 ................................... 55 SUMMARY AND CONCLUSIONS A P P E N D I X ............................................. LITERATURE CITED ............................................ 57 74 LIST OF TABLES PAGE Table Table Table Table 1. Effect of 1-Thyroxine Administration on the External Thyroid Counts of Shropshire Wether 3 8 0 ...................................... 29 E. Effect of 1-Thyroxine Administration on the External Thyroid Counts of Shropshire Wether 3 0 E ..... .................. *............ 30 3. Effect of 1-Thyroxine Administration on the External Thyroid Counts of Shropshire Wether 3 7 1 ....... 31 4. Effect of 1-Thyroxine Administration on the External Thyroid Counts of Shropshire W e t h e r s ..................................... 3E Table 5. Correlation and Regression Coefficients between 1 - Thyroxine Administration and Per­ cent of Previous Count of Thyroid Radioac­ tivity Taken D a i l y ......................... 33 Table 6. Correlation and Regression Coefficients between 1 - Thyroxine Administration and Per­ cent of Previous Count of Thyroid Radioac­ tivity Taken Alternate Day. ...... 34 Table 7. Correlation and Regression Coefficients between 1-Thyroxine Administration and Per­ cent of Previous Count of Thyroid Radioac­ tivity Taken Every Third D a y .............. 35 Table 8. The Thyroid Output Rate (t-^) of 1^31 Lac tating Shropshire Ewes and Gain in Weight of Their Lambs Up to Three Weeks of A g e ..... 42 Table 9. Daily 1-Thyroxine Secretion Rate of Lambs and Their Gain in Weight During One Month Period. 47 10. Daily 1-Thyroxine Secreted per 100 Pounds of Body Weight by Ewe, Ram, and Wether Lambs.... 52 Table Table 11. Analysis of Variance...................... 53 LIST OF AFFENDIX TABLES PAGE Table Table Table 1. Data on the Thyroid Radioactivity of Rats in Response to Different Levels of 1-Thyroxine Administration - Experiment 1 ................. 57 2. Data on the Thyroid Radioactivity of Rats in Response to Different Levels of 1 - Thyroxine Administration - Experiment I I ............... 59 3. Data on the Thyroid Radioactivity of Rats in Response to Different Levels of 1-Thyroxine Administration - Experiment III.............. 61 Table 4. Data on the Thyroid Radioactivity in Response to Different Levels of 1-Thyroxine Administra­ tion to Ewe L a m b s ............................... 62 Table 5. Data on the Thyroid Radioactivity in Response to Different Levels of 1-Thyroxine Adminis­ tration to Wether Lambs........................ 69 6. Data on the Thyroid Radioactivity in Response to Different Levels of 1 - Thyroxine Adminis­ tration to Ram Lambs ............................ 71 Table LIST OF FIGURES PAGE Figure 1. Monitor with a Scintillation and GeigerMueller T u b e ................................... 17 Figure 3. External Foreleg C o u n t ................ . ..... 18 Figure 3, External Thyroid C o unt ...................... 19 Figure 4. Effect of 1-Thyroxine Injections on Percent of Previous External Thyroid Counts......... 24 5. Effect of 1-Thyroxine Injections on Percent of Previous External Thyroid Coun t s ......... 34 6. Effect of 1-Thyroxine Injections on Percent of Previous External Thyroid Counts......... 26 Figure Figure Figure 7. External Thyroid Counts on Shropshire Ewes.. 38 Figure 8. External Thyroid Counts on Shropshire Ewes.. 39 Figure 9. External Thyroid Counts on Shropshire Ewes.. 39 Figure 10. External Thyroid Counts on Shropshire E w e s . . 40 Figure 11. External Thyroid Counts on Shropshire Ewes.. 41 Figure 12* External Thyroid Counts on Shropshire Ewes.. 41 13* Regression Line of Gain in Weight of Suck­ ling Twin Lambs up to Three Weeks of Age on the Output tts of ll31 in the Thyroid of Lactating D a m s ................................ 43 14. Regression Line of One Month's Gain in Weight on Daily 1-Thyroxine Secretion of Ewe Larnibs...................................... 49 Figure Figure INTRODUCTION Reported research indicates that the thyroid gland is associated with almost every organ and tissue in the body. Therefore, a change in its functional level, hyper-thyroidism, such as hypo- or reflects demonstrable changes in many func­ tional processes of an Individual. One of the most notable effects of its varying activity is the accompanying change in the basal metabolic rate. However, other processes in the body, viz. growth, lactation, sexual activity etc., are found to be related with it, directly or indirectly, to a varying extent. Growth is one of the most important characteristics of a living organism. It is either the basis of or closely associated with many productive processes in livestock. Meat, milk and wool production are dependent upon or closely related to growth. The relationship of thyroid activity and growth rate has received very little attention and therefore it was made a part of this study. In all farm animals where the newly born animal is not artificially reared, the young depends upon the milk of its dam for adequate nourishment during the early growth period. Though the hereditary growth potentialities of the individual are predetermined at conception, the expression of it is sub­ ject to the influence exerted by environmental factors. The most important or these factors during the early stages after birth is the dam's milk. Therefore, the quantity of milk produced by the ewe is of fundamental economic importance for the success of fat-lamb production. In this study an attempt has been made to correlate thyroid activity with the milk production of the ewe as measured by lamb gains up to 3 weeks of a g e . The classical methods of thyroid research, designed to produce a deficiency state by removal of the thyroid or by thiouracil administration, and to produce a hyper-functioning state by administration of glandular extract or synthetic products have not shed any light on the normal function of the thyroid in relation to the existing variation in the various economic characters of farm animals. Moreover, the results of the studies with added thyroactive substances and its effect on growth and other characters have been found to be contradictory. Some of the conflicting reports may be partly due to the large amount of variation in the hyperthy­ roidism induced. It seems, therefore, of utmost importance to obtain the actual thyroid hormone secretion rate of animals under normal conditions. It is obvious that the most desirable and practical technique for determining thyroid secretion rate will be the one that can be used on live animals. There is some indication that thyroid activity is related to the problem of summer sterility in rams and also may have an effect on the seasonal breeding habit of the ewe. Before addi­ tional studies can be conducted on the addition of thyroactive substances, a basic knowledge of thyroxine secretion rate in the different sexes would be helpful. If we learn the differ­ ences occuring under normal conditions in the hormone secre­ tion rate, we may be able to breed more consistently the type of animal capable of high producing ability. This study has been designed to obtain such basic information on the thyroid hormone secretion rate in relation to certain characters in sheep. 4 OBJECTIVES 1. To refine an extrapolation technique for determining the thyroid secretion rates in life sheep which would be practical for other livestock. 2. To determine the relationship between the thyroid activity of lactating ewes and growth of their suckling lambs during the early stages of life. 3. To determine the relationship of thyroid secretion rate to growth and sex in sheep. 5 REVIEW' OF LITERATURE Studies of the thyroid gland have shown that it exercises a profound influence on the growth and development of all the higher vertebrates. Curling (1850) observed an absence or atrophy of the thyroid in two typical cases of cretinism in the human. Pfaundler (1934) also observed marked dwarfism in the human resulting from a non-functional thyroid gland. Schiff (1856) studied the effect of thyroidectomy in dogs and observed a stasis of growth in all cases. Similar results of thyroidectomy in dogs were reported by Dott (1933), and Binswanger (1936). The effects of thyroidectomy in laboratory animals are analogous to those observed in the human. Kojimi (1917) thyroidectomised growing rats and observed retarded growth, reduced calcium and nitrogen retention, intake. and decreased feed Salmon (1938) observed the characteristics of cretinism in rats due to the effect of thyro-parathyroidectomy in new-born individuals. Hughes (1944) studied the effect of thiouracil administration to rats from the time of birth and reported that it resulted in a marked retardation of growth, ment and other changes similar to cretinism. arrested develop­ Similar effects of thyroidectomy have been observed for many other laboratory animals, such as, the mouse (Davenport and Swingle, 1927), b guinea pig (Silberberg and Silberberg, 1940; Williams et. al. 1941), rabbit (Basinger 1916, Kunde 192,6), and monKey Fleischmann, Schumaker and Straus 1943), Numerous investigations have also demonstrated that thyroid­ ectomy has greatly depressed growth in farm animals. Simpson (1924) reported the effect of thyroidectomy on triplet female goats. Two of them were thyroidectomised at the age of 22 days while the third was kept as a control. About three months following the operation, he observed marked cretinism of the two kids compared to the normal growth of the control. Moreover, he (1913 and 1924) observed similar results after thyroid abla­ tion in sheep, and found that the age at thyroidectomy had a marked influence on its effect on the individual. He reported that the lambs thyroidectomised at one to two months of age exhibited marked stunted growth, whereas practically no ill effect on growth was found in those thyroidectomised at 6 to 8 months of age. Subsequently, Mangold (1932), Marston and Peirce (1932), Todd and Wharton (1934), and Todd, Wharton and Todd (1938) experimenting with sheep; and Reingtee and Turner (1941), Reineke, Bergman and Turner (1941) using goats; con­ firmed the reported cretin condition as observed by Simpson (1913 and 1924). Only a few experimental reports are available regarding the effects of thyroidectomy on the growth of cattle. Brody and Frankenbach (1942) reported that thyroidectomy of a Jersey heifer at 54 days of age reduced the mature body weight by over 50 percent. The characteristic symptoms of hypothyroid­ ism began to appear about a month after the thyroidectomy. 7 The observations of* Spielman et al (1954) were similar to those or Brody and Frankenbach (1942.). They stated that thyroidectomy of the immature bovine was followed the normal growth processes. by a serious impairment of There was complete in height and severe retardation of stasis of gain gain in body weight. Marked differential response in the pattern of growth d e ­ pression due to thyroidectomy has been observed by various workers. Todd and Wharton (1934) studied the disturbed growth patterns of the skull in thyroidectomised lambs and reported a delayed eruption of the teeth, defective growth of the frontal bones, nasal bones and of the upper jaw. Demonstrable defects In the growth of the brain case proper and mandibles were not observed. Deficient growth of the epiphysis, defective devel­ opment of age characters in the epiphysis and shaft, and dimin­ ished velocity of the growth of shaft were also observed in the early thyroidectomised sheep by Todd, Wharton and Todd (1938). Marston and Peirce (1932) reported their findings on the growth of wool in Merino sheep following thyroidectomy. They observed a considerable decrease in the wool production and the degree of greasiness of the fleece. No appreciable difference was found in the fibre diameter. The reports reviewed indicate the analogous effects of thyroidectomy in all mammals and in general the results are more severe if the function is impaired in early life. Pathological conditions resulting from thyroid removal in human patients led workers to attempt replacement therapy. The first reported attempt was made by Schiff (1884) in the 8 dog. He stated that part of the effects of thyroidectomy could be relieved by transplanting the thyroid from another dog. Murray (1891) reported that myxedema in humans was relieved by the injection of sheep thyroid extract. of young, Resumption of growth thyroidectomised animals has been observed by the replacement therapy with thyroxin or thyroprotein by Brody and Frankenbach (1943), Reineke and Turner (1941), and Spielman et al (1945). An increase in the thyroid functional level in the initially normal animal, as a result of thyroid administration, may result in either acceleration (Cameron and Carmichael, or retardation of growth (Hammett, 1924). 1930) Variability of the effect was mainly due to the size of the dose administered and its relation to the physiologically optimal thyroid functional level. If the amount administered did not increase total metabolism to the extent where catabolism predominated or was not above the optimum level, growth in weight was hastened. If the amount administered was large enough, optimum level, that is above the growth was retarded. Some evidence indicates that mild hyperthyroidism may be conducive to rapid growth. Irwin, Reineke and Turner (1943) fed several levels of thyroprotein to White Plymouth Rock and Rhode Island Red chicks for 3 months and reported that 36 grams per cwt. of feed was the best dose for feathering and growth. Wheeler, Hoffman and Graham (1948) reported a significant increase in body weight of male Rhode Island Red chicks at 12 weeks of age that were fed 10 grams of thyroprotein per hun- 9 dred pounds of feed but observed no significant difference in weight of the females. There was a marked stimulation of early feather growth in the treated birds. Quisenberry and Krueger (1948) studied the effect of f e e d ­ ing protamone (iodinated casein) up to 6 weeks of age on 800 chicks of New Hampshire and White Plymouth Hock breeds, and reported that the rate of gain and feed utilisation were improved. Distortion of body proportions produced by increased thyroid activity has been observed by various workers. Cameron (1922) reported that thyroid feeding in rats and rabbits resulted in hypertrophy of the internal organs which tended to recover following the discontinuance. Silberberg and Silberberg (1938) found that the feeding of thyroid sub­ stance stimulated the growth of the epiphyseal cartilage and accelerated its further development into hypertrophic carti­ lage in immature guinea pigs. In fowl also, definite struc­ tural alterations have been reported from feeding thyroid materials (Nevalonnyi 1927, Horning and Torrey 1927). The interrelationship between the thyroid and the gonads has been indicated by much research, but its nature is not very clearly understood. The early studies of Marine and Kendall (1917), Levin (1921), and others showed that the incidence of thyroid d i s ­ turbance was greater in girls and women than in boys and men. Evans and Long (1921a, 1921b) observed no effect of thyroidectomy in rats on the onset of puberty or the length 10 of* the estrous cycle. Moreover, no marked effect on the estrous cycle was found as a result of feeding fresh thyroid gland. Lee (1925) reported that thyroidectomy in rats did not affect materially the age of puberty but lengthened the estrous cycle. Hammett (1929) also observed no relationship between thyroid activity and the development of the reproduc­ tive system. He suggested that the reported disturbed sex conditions were secondary to the disturbance in general growth. The work of Brody et al (1942) suggested that the reported absence of a relationship may be due to the differ­ ences in age at thyroidectomy. Brody and Frankenbach (1942) studied the effect of thyroidectomy on a Jersey heifer and reported that at the age of 40 months, pletely undeveloped sexually. the heifer was com­ Moreover, the administration of thyrolactin (iodinised milk protein) after 40 months of age stimulated the sexual development. Da Costa and Carlson (1933) reported that the adminis­ tration of large doses of desiccated thyroid to white rats resulted in the retardation of the sexual maturation of both sexes, while small doses of thyroid tended to accelerate it. Schockart (1931) observed a greater hypertrophy of the male accessory sex glands after thyroidectomy than before when anterior pituitary emulsions were fed. Marine (1937) observed the enlargement of the thyroid gland during puberty and pregnancy. However, he (1935) sug­ gested that such an effect on the thyroid was perhaps caused by thyrotropic hormone stimulation. The effect of estrogenic hormone on thyroid activity in animals has been studied by many workers. Laqueur, Hart and de Jongh (1926) reported that the injection of estrogenic hormone increased the basal metabolism of both normal and ovariectomised rats from 15 to 20 percent two or three days after administration. Tagliaferro (1933) observed hyperac­ tivity of the thyroid after 5 to 10 days of estrin adminis­ tration. He further reported that continued administration of estrin for 20 days or more resulted in the opposite effect. Kunde et al (1930) found marked hyperplasia of the thyroid in the dog as a result of estrogen administration. Aron and Benoit (1932) reported that hyperthyroidism produced by thyro­ tropic hormone injection was prevented by the administration of large doses of estrin. Their observation was later con­ firmed by Spence (1936), and Elmer, Giedosz and Scheps (1938). Farbman (1944) reported that the administration of large doses of estrogenic hormone seemed to have some inhibitory effect on the thyroid of humans. Epstein (1950) studied the effect of exogenous estrogen administration on the thyroid of immature chicks. He stated that the administration of estrogen caused no effect on the size of the thyroid gland but depressed the iodine turnover rate in normal animals. Thyroid has been reported to influence the fertility of farm animals and fowls. During summer months, rams of some breeds have been observed to produce low quality semen as indicated by decreased sperm concentration and volume of semen, with an increased number: Berliner 1937 of abnormal spermatozoa (McKenzie and , Bogart and Mayer 1946, and Gunn et al 1942). Berliner and Warbritton (1937) reported that thyroidectomised rams produced semen of low quality and administration of thyroxine to such animals resulted in increased spermatogenesis. Moreover, rams with low fertility also showed marked improve­ ment in semen quality when treated with thyroxine. therefore, They, suggested that the summer sterility might be indi­ rect and due to a thyroid deficiency. Bogart and Mayer (1946) observed similar results with thyroprotein feeding. But the results obtained by Warwick et al (1948) did not support the observations of Berliner and Warbritton (1937) and Bogart and Mayer (1946). Schultze and Davis (1946), and Reineke (1946) treated dairy bulls with varying amounts of thyroprotein for different lengths of time and reported that improvement was observed in the quality of semen and to some extent in the conception rate* Winchester (1939), and Taylor and Burmester (1940), w ork­ ing with chickens, observed a marked decline in egg production following thyroidectomy which was found to be increased after replacement therapy. Shaffner and Andrews (1948) found reduced semen quality in fowls as a result of hypothyroidism. They concluded that thiouracil impaired the ability of the gonads to produce viable sperms which were capable of surviving normal lengths of time in the oviduct of hen. Both qualitative and quantitative methods have been used to determine the secretory activity of the thyroid gland. The qualitative methods are based upon the changes which occur in the thyroid gland in conjunction with the changes in its sec­ retory activity. By these methods, the thyroid function is determined by: (i) observation of the changes in mitotic activity, (ii) measurement of the diameter of the thyroid follicles, (iii) measurement of the follicular epithelium, and (iv) determination of increased or decreased amounts of protein bound iodine in the thyroid and/or in the blood. Until recently, the quantitative determination of thyroid activity was based on replacement therapy. By this method, the amount of thyroactive material required to maintain thyroi­ dectomised animals in a normal state was determined. Dempsey and Astwood proposed a new method, In 1943, the goitrogenic method, for the determination of the thyroid secretion rate. For this, a goitrogenic drug, e.g., thiouraeil, was adminis­ tered simultaneously with graded doses of thyroxine for a period of t i m e , and the amount of thyroxine required to m a i n ­ tain the normal weight of the thyroid was considered to be the normal secretion rate. Though this method is in general use today In the study of thyroid function of laboratory animals, its application for the determination of the normal thyroid secretion rate of large animals Is not practical. Therefore, studies have been directed towards developing a technique suitable for large animal research. The use of radioiodine (1-^-**) for thyroid studies is based on the principle that the thyroid has a great affinity for iodine, and its hormone (thyroxine) contains about 65 per- cent of iodine. Hertz and coworkers were the first to employ radioiodine In 1938 as an indicator of the thyroid function. Their works (1938-1941) on rabbits demonstrated the selective concentration of iodine by the thyroid, which was greatly increased by stimulation of the thyroid through thyrotropin administration. Henneman (1953) developed an extrapolation technique for measuring the thyroid secretion rate. Sheep were used as the experimental animal and they were injected with one microcurie of 1131 per pound of body weight. A base count was taken over the thyroid gland 7 days after injection of I ^ l . After this period, different levels of 1 - thyroxine were injected subcutaneously daily for 3 consecutive days, might adjust its secretion to that dose. so that the thyroid Another count was taken after this period, and expressed In terms of the percent of previous count. The estimated thyroid secretion rate was the level of 1 - thyroxine which gave 100 percent of previous count, and was predicted by the regression equation. When the technique was used for the determination of individual 1 - thy­ roxine secretion, consistent results were obtained, but due to the smaller number of observations on each individual sheep, statistically significant values were not consistently obtained. To obtain a significant value of 1-thyroxine secretion for an individual animal, it was realized that more observations were necessary in response to varying doses of 1 - thyroxine administration on the radioactivity of the thyroid. in view, an attempt was made to refine the technique. With this EXPERIMENTAL PROCEDURE There are mainly two ways to study the accumulation of radioiodine in the thyroid: (i) direct, and (il) indirect. The direct method is the measure of gamma radiation by apply­ ing the Geiger-Mueller or scintillation counter externally at the thyroid. Indirect measurement is the estimation of thyroid uptake by the determination of total urinary excretion of iodine in a specified period. For this, it is assumed that the thyroid uptake is equal to the difference between the dose administered and the amount excreted (Keating et al 1949, Shipley et al 1950, and Fields and Le Roy 1952). In general, direct measurement is preferred over the indirect or excretory one. The method used in this study for the determination of thyroid activity in sheep was the direct one. injected with I1S1 subcutaneously pound of body weight. The animals were at the rate of 1 uc. per Theamount of radioactivity of llsl at the time of administration was computed by the formula: N = where N = No = No e the activity at time, t the activity when t = 0 e = base of natural logrithm ^ decay constant of I ^ l = t z time A Geiger-Mueller or scintillation counter, shown in figure 1, was used for measuring the radioactivity in sheep. Several counts were taken over the thyroid gland, which was presumed to he in the area of the neck where the highest count was obtained, at regular time intervals by the external a ppli­ cation of the Geiger-Mueller or scintillation tube (figure 2). The body background was taken on the outside of the foreleg of the sheep as shown in figure 3. A shield was used an the Geiger-Mueller or scintillation tube when taking counts so that only gamma rays were measured. Moreover, the scintilla­ tion tube was wrapped with cloth to facillitate its handling. Throughout the experiment the same amount of pressure was used on holding the tube against the neck and foreleg as far as possible. The wool was clipped from the thyroid region of the neck and foreleg of all the sheep before the administra­ tion of I-k^l. A standard source of Co60 was counted at the time counts were taken on the sheep to check the monitor. readings obtained on the sheep were corrected The to the standard to eliminate any error due to the variable efficiency of the counter. The counts obtained on the thyroid of the sheep were further corrected for the body background and physical decay and plotted on a semi-log scale. There are various ways to compare the individuals regard­ ing their thyroid activity from the curves obtained. They m a y be compared with respect to the rapidity of uptake or the rapidity of output. In either case, they can be compared by the computed values for k. b, or biological t^. For this Figure 1. A Monitor with a Scintillation and Geiger-Mueller Tube. 17 Figure 2. External Thyroid Count 18 Figure 3. External Foreleg Count. 19 20 study, the regression coefficient (b) of the thyroid counts on different time intervals for each sheep was used and o b ­ tained by the formula: byx * _S(X_- X)(I - Y) S (X - X) where S r sum X s time Y s counts per minute A straight line was fitted to the output portion of the plotted curve of thyroid activity, by solving the regression equation: Y s a 4 bX; where a - Y - bX. The thyroid output half-time of I131 for each sheep was obtained from the turnover portion of the curve. Though the individuals are compared for their thyroid activity by the obtained values of ti of radioiodine, the estimate of actual amount of thyroid hormone secretion cannot be made. For a quantitative estimate of the daily secretion rate of the thyroid, the method used was the one developed by Henneman (1953) and modified in this study. Quantitative determination of the thyroid secretion has been made in terms of either d, 1-thyroxine or 1 - thyroxine. These two estimates are interchangeable, since Reineke and Turner (1945) have reported that 1-thyroxine is twice as active physiologically as d,1-thyroxine. Harington and Salter (1930) have shown that 1 - thyroxine is the form of thyroxine produced by the thyroid gland. In this study, 1 - thyroxine was used to determine the thyroid secretion rate. It was supplied by Glaxo Laboratories, Greenford, Middlesex, England, further purified by Dr. E. P. Reineke. and was The dry thyroxine was weighed on an analytical balance and dissolved in distilled water. NaOH, This solution was first made slightly alkaline with and then enough acid (HC1) was added to make it slightly cloudy. It was then stored in the refrigerator and used within a two week period. £2 RESULTS AND DISCUSSION A. Refinement of a Technique for Measuring Thyroid Secretion Rate To refine the technique, developed by Henneman (1953), for measuring the thyroid secretion rate of live sheep, four experiments were conducted. In three experiments rats were used because they were inexpensive and easy to handle and the results obtained may be applicable in other species. The fourth experiment was conducted on the wether lambs of the Shropshire b r e e d . Experiment I and II The first and second experiments were conducted on four and six female rats respectively, maintained on an ordinary laboratory diet at a room temperature of 74° F. They were injected intraperitoneally with 40 micro curies of 1 1^1 an 1-Thyroxine Tnjactions on Percent of Previous External Thyroid Oounts. 0 1 25 Experiment III To check the secretion rate estimated by the method used in the first two experiments, on 24 female rats. a third experiment was conducted All the rats were injected with 40 u c . of pl31 and the base count was taken 120 hours after injection. A scintillation counter was used in this experiment. The rats were divided into 5 groups of 5 each, except for the control w h i c h .included 4 rats. All five groups had approxi­ mately the same average weight. Each of the four groups under treatment was injected subcutaneously with a different level of 1-thyroxine once daily for 7 days. The doses used were 1.0, 1.5, 2.0, and 2.5 ug. per 100 gm. body weight. The second or final count was taken 7 days after the base count. The data were corrected for physical decay and body back­ ground as in the other two experiments. The data are presented in figure 6. The coefficients of correlation and regression obtained between the levels of 1 - thyroxine administration and percent of previous count were 0.942 and 23.8, respectively. These values are statistically significant at the 1 percent level. The average daily secre­ tion rate of 1 - thyroxine, as estimated by the regression equation Y = 44.3 - 23.8X, was 2.340 ug. per 100 gms. of body weight. With any new method and in the absence of similar deter­ minations for comparison, there is always the possibility of some error being introduced by misunderstood factors involved in the method. The results obtained by this method were «• 36 100 90 Y = 44.3+23.8 X 70 OP PREVIOUS COURT 80 40 0 0.5 1.0 2.0 1.5 2.5 1-THYROXINE ADMINISTERED (ug.)/ 100 GRAMS Fig. 6. Effect of 1-Thyroxine Injections on Percent of Previous External Thyroid Counts. 27 fairly repeatable as found in the determination of 1 - thyroxine secretion rates of different groups of rats, maintained under similar environmental conditions, at different periods. How­ ever , the estimated secretion rate was markedly lower than the one reported by Perry (1951) which may be attributed to the probable difference in the quality of 1 - thyroxine adminis­ tered. The estimated 1 - thyroxine secretion rate in this study was close to the one determined by the goitrogen tech­ nique. Monroe and Turner (1946) reported that the thyroid secretion rate of growing female rats ranged from 2.82 to 4.63 micrograms of d,1-thyroxine per hundred grams body weight. EXPERIMENT IV In this experiment, breed were used. 5 wether lambs of the Shropshire They were injected subcutaneously with I ^ l at the rate of 1 uc. per pound of body weight, and the radio­ activity of the thyroid was measured by a Geiger-Mueller tube between the 7th and the 22nd day after the injection. The wethers were divided into two groups with three animals in the first group (A) and two in the second group (B). On the output portion of the curve of thyroid radioactivity, the wethers in group A were Injected daily for two consecutive days with a quantity of 1-thyroxine, whereas those in group B were injected daily for three days. The level of 1-thyroxine administration was changed after this period. In group A, the thyroid radioactivity was determined daily, while in group B, it was determined every third day prior to the admin­ istration of a different level of 1-thyroxine. The data obtained are presented in tables 1, 2, 3, and 4. As in the other experiment, it was assumed that the level of 1-thyroxine administration, which held the count at one h u n ­ dred percent of previous count, was the amount to replace the normal thyroid secretion. Hence, the regression equation was computed for each individual lamb and was used to estimate the rate of 1 - thyroxine secretion. The daily 1 - thyroxine secretion rates of 3 wether lambs were estimated to be 0.107, 0.095, and 0.106 mg. per 100 pounds of body weight based on the daily measurement of thyroid radioactivity. lambs, they were 0.108, 0.096, For the same and 0.106 mg. when the radioac­ tivity was measured on alternate days. The secretion rate of the other two lambs were 0.110 and 0.118 mg. when the counts were taken every third day. Tables 5, 6, and 7 show the correlation and regression coefficients between the level of 1 - thyroxine administration and the percent of previous count, when the counts were taken daily, alternate days and every third day. None of the coef­ ficients, except 0.646 and 1.002, obtained for the counts taken daily on individual wether, is statistically signifi­ cant. The obtained high but insignificant correlation values on the individual animals suggest the necessity of more obser­ vations because by using the intra^wether statistical treatment the coefficients are highly significant for the counts taken every day and also for counts taken on alternate days, significant when taken every day. and 29 TABLE 1 EFFECT OF 1-THYROXINE ADMINISTRATION ON THE EXTERNAL THYROID COUNTS OF SHROPSHIRE WETHER 380 Days Following 1131 Admini str at i on Mg. or 1-Thyroxine Administered per 100 Lbs. Body Weight Thyroids Counts per Minute Percent of Previous Count 7 (Zero time) 8 1022 9 1196 10 1255 11 1133 90.3 968 IS 0.025 1052 92.9 13 0.025 988 92.6 14 0.050 908 91.9 15 0.050 876 96.5 16 0.075 731 83.4 17 0.075 707 96.5 18 0.100 671 94.9 19 0.100 668 99.6 702 105.1 20 Percent of Previous Day Before Count ^Corrected for background and physical decay 83.8 86.3 80.5 91.8 104.6 30 TABLE E EFFECT OF 1 - THYROXINE ADMINISTRATION ON THE EXTERNAL THYROID COUNTS OF SHROPSHIRE WETHER 302 Days Following x131 Administration Mg. of 1-Thyroxine Administered per 100 Lbs. Body Weight Thyroid* Counts per Minute Percent of Previous Count 7 CZero time) a 2631 9 3588 10 3816 11 3572 93.6 3343 12 0.025 3394 95.0 13 0.025 3300 97.2 14 0.050 3107 94.2 15 0.050 2932 94.4 16 0.075 2602 88.7 17 0.075 2542 97.7 ia 0.100 2538 99.8 19 0.100 2453 96.7 2584 105.3 20 Percent of Previous Day Before Count •^Corrected for background and physical decay 88.9 91.5 83.7 97.5 101.8 31 TABLE 3 EFFECT OF 1-THYROXINE ADMINISTRATION ON THE EXTERNAL THYROID COUNTS OF SHROPSHIRE WETHER 371 Days Following 1131 Administration Mg. of 1-Thyroxine Administered per 100 Lbs. Body Weight Thyroid* Counts per Minute Percent of Previous Count 7 (Zero time) 8 2240 9 2362 10 2503 11 2310 92.3 2293 IS 0.025 2247 97.2 13 0.025 2105 93.7 14 0.050 1993 94.7 15 0.050 1902 95.4 16 0.075 1771 93.1 17 0.075 1706 97.4 18 0.100 1755 102.9 19 0.100 1672 95.3 1774 106.1 20 Percent of Previous Day Before Count ^•Corrected for background and physical decay 89.8 88.7 88.9 100.2 101.1 32 TABLE 4 EFFECT OF 1-THYROXINE ADMINISTRATION ON THE EXTERNAL THYROID COUNTS OF SHROPSHIRE WETHERS Days Following jizx Adminis­ tration M g . of 1-Thyroxine Administered per 100 Lbs. Body Weight 7 (Zero time ) 10 13 0.025 14 0.025 15 0.025 16 0.050 17 0.050 is 0.050 19 0.100 20 0.100 21 0.100 22 Wether 325 Thyroid* Counts per Minute Percent of Previous Count Wether 0 Thyroid* Percent of Counts per Previous Count Minute 3291 2127 4225 2171 3790 89.7 1564 72.0 3129 82.6 1200 76.7 2662 85.1 1103 91.9 2697 101.3 1047 94.9 ^Corrected for background and physical decay 33 TABLE 5 CORRELATION AND REGRESSION COEFFICIENTS BETWEEN 1-THYROXINE ADMINISTRATION AND PERCENT OF PREVIOUS COUNT OF THYROID RADIOACTIVITY TAKEN DAILY • Wether Number Sum of Squares & Products D.F. , Sx2 Sxy Sy2 b r i 380 9 0 .01250 0.01253 0.03010 0.646* 1.002* 302 9 0.01250 0.00823 0.01709 0.563 0.658 371 9 0.01250 0.00893 0.01764 0.601 0.714 wither 27 0.03750 0.02969 0.06483 0. 602**0 .792** -^Significant at 5 percent level ^ S i g n i f i c a n t at 1 percent level 34 TABLE 6 CORRELATION AND REGRESSION COEFFICIENTS BETWEEN 1 - THYROXINE ADMINISTRATION AND PERCENT OF PREVIOUS COUNT OF THYROID RADIOACTIVITY TAKEN ALTERNATE DAY Wether Number D .F. Sum of Squares & Products Sx2 1 Sxy \ I r b Sy3 380 4 0.00625 0.01178 0.03570 0.788 1.884 30 a 4 0 *00625 0.00795 0.02027 0.706 1.272 371 4 0.00625 0.00853 0.01603 0.852 1.364 Intrala wether 0.01875 0.02826 0.07200 0.769** 1.507* •^Significant at 1 percent level 35 TABLE 7 CORRELATION AND REGRESSION COEFFICIENTS BETWEEN 1-THYROXINE ADMINISTRATION AND PERCENT OF PREVIOUS COUNT OF THYROID RADIOACTIVITY TAKEN EVERY THIRD DAY Wether Number Sum of Squares & Products D* F . , Sac2 Sacy I U Sy2 525 3 0.00547 0.00757 0.02061 0.713 1.384 No tag 3 0.00547 0.01324 0.03785 0.921 8.422 Intrawether 6 0.01094. 0.02081 ^Significant at 5 percent level 0.05846 0.823 1.902 As there was not much difference between the uncorrected count when radioactivity was measured every day, the effect of errors in the readings could be a high percentage of the difference between the counts. There was a much greater dif­ ference between the counts taken on alternate days and there­ fore the percentage effect of normal errors in obtaining the thyroid counts was materially reduced. Hence, on the basis of such general observation as well as the results obtained from the statistical treatment, it was decided to administer the same level of 1-thyroxine for four consecutive days and measure the thyroid radioactivity on alternate days which w ould provide two counts for each level of 1 - thyroxine administration. tions, From the data to be presented in other sec­ it is apparent that this technique gave a good estimate of the thyroid hormone secretion. However, the observation of the last experiment of the studies indicates that more than two counts may be obtained for each level of 1 - thyroxine administration by the use of a scintillation tube. The scin­ tillation tube was much more sensitive than the G.M. tube to gamma radiation. Therefore thyroid counts significantly greater than the background count could be obtained for a muc h greater length of time after I131 injection when the scintillation tube was substituted for the G.M. tube. 37 B. The Relationship of Thyroid Output Rate to Lactation in Ewes as Measured by Growth Rate of the Lambs Eleven Shropshire ewes were used to determine the rel a ­ tionship between the thyroid output rate (tjJ of i131 and milk 2 production as measured by the growth rate of the nursing lambs u p to three weeks of age. The data on the external thyroid counts of the ewes are graphically shown in figures 7, 8, 9, 10, 11 and 12. The out­ put half-time was computed for each individual ewe and they are presented in table 8 along with the lamb gains. The thyroid output rate varied between 144.7 hours to 358.3 hours. These values of output half-time are much lower than those obtained by Terry (1951) for non-lac tat ing adult ewes which was 32 days. This difference may be due to increased thyroid activity during lactation. Rough (1951) reported that the process of lactation protected the thyroid of the mouse from radiation damage indicating a higher output of radioiodine from the thyroid. Henneman (1953) reported a significantly greater thyroid activity in lactating ewes than in non-lactating ewes. However, Monroe and Turner (1946) observed no significant dif­ ference in the thyroid activity of rats due to lactation. The coefficient of correlation between the output ti of seven lactating ewes and the gain of their suckling twin lambs was -0.553. level, This is statistically significant at the 5 percent indicating a relationship between the thyroid activity of the dam and the growth and development of lambs during the first three weeks following birth. sa 5000 4000 3000 2000 EWE 924 *“< 1000 s 800 « w cu to EWE. 911 500 S3 400 o o 300 200 100 0 50 100 150 200 Fig. 7. External Thyroid Counts on Shropshire Ewes 250 300 39 7000 4000 COUNTS PER MINUTE 9000 1000 0 50 100 150 HOURS 200 250 300 250 300 COUNTS PER MINUTE Fig, 8. External Thyroid Counts on Shropshire Ewes. 9000 7000 5000 4000 3000 2000 10001 0 50 100 150 HOURS 200 Fig. 9. External Thyroid Counts on Shropshire Ewe 40 10000 6000 6000 5000 EWE 546 3000 2000 EWE 552 1000 H 800 (u 700 600 CO E-, 500 2 » 400 o o 300 200 100 0 50 100 150 HOURS 200 Fig. 10. External Thyroid Counts on Shropshire Ewes. 250 300 41 20000 10000 ;6000 ,4000 os EWE H30 CO 2000 o o 1000 100 150 HOURS 200 250 300 250 300 Fig. 11. External Thyroid Counts on Shropshire Ewes. 10000 ^ 8000 ^ 6000 *"* 5000 4000 EWE 215 co £3 2000 o o 1000 100 150 200 Fig. 12. External Thyroid Counts on Shropshire Ewes. 42 TABLE 8 THE THYROID OUTPUT RATE (tj) of I131 IN LACTATING SHROPSHIRE EWES AND GAIN IN WEIGHT OF THEIR LAMBS UP TO THREE WEEKS OF AGE Ewe N o . and Year of Birth Lamb Thyroid Number Output and Sex ti (hrs.) Birth Weight 21-Day Weight (lbs.) ( lbs.) Correction Gain for in Sex Weight (l b s .) Corrected Gain in Weight ( l b s .) 12 9 911-50 152.0 734 E 735 E 8.0 9.0 20 18 924-50 184.7 739 a 740 E 8.0 7.0 16.5 14.5 8.5 7.5 657-49 212.0 676 E 677 E. 7.5 8.0 16.25 16.0 8.75 8.0 644-49 358.3 678 E 679 R 8.5 8.5 16.0 17.75 7.5 9.25 645-49 235.2 743 E 744 E 7.0 6.5 14.25 15.0 7.25 8.5 552-48 161.8 741 R 742 E 7.5 7.0 21.0 15.0 13.5 8.0 546-48 144.7 682 E 683 E 8.0 8.0 18.5 18.0 10.5 10.0 10.5 10.0 215-47 271.3 738 E 8.5 22.25 13.75 13.75 212-47 338.2 746 E 8.0 17.5 9.5 9.5 232-47 281.3 674 R 9*0 23.5 14.5 H30-47 178.1 685 E 9.5 23 •25 13.75 12 9 -2 6.5 7.5 8.75 8.0 -2 7.5 7.25 7.25 8.5 -2 -1 11.5 8.0 13.5 13.75 43 12 10 Y = H . Ul\ - 0.013 X fc. o CO Q 100 200 300 400 500 OUTPUT HALF-TIME ( Hours ) Fig. 13. Regression Line of Gain in Weight of Suckling Tjriy Lambs up to Three Weeks of Age on the Output 1 1 of I in the Thyroid of Lactating Dams. 44 The regression coefficient, of the gain of suckling twin lambs on the turnover t^r of ewes was found to be -0.013. in the thyroid of lactating The regression line was fitted to the data and is shown in figure 13. Schultze and Turner (1945) studied the effect of lacta­ tion on thyroid secretion in goats. They reported that 6 lactating goats, producing an average of 2.6 pounds of milk per day had a daily d, 1-thyroxine secretion rate of 1425 ug., whereas the same goats in advanced lactation, producing an average of 1.8 pounds of milk per day, secreted 1000 ug. Various workers have reported the effect of feeding thyroid materials on lactation and of lactation on the rate of early post-natal growth. The literature on the response to thyroid feeding on lactation has been reviewed by Blaxter et al (1949). Graham (1934a, 1934b), Beineke (1943), and Archibald (1945) observed an increase in milk yield of dairy cows by thyroid feeding or thyroxine injection. Bonsma (1944) reported that differences in the level of milk production of ewes has a marked influence on the early post-natal growth rate of lambs, mainly up to six weeks of age. These reports give support to the observation of this study that the lactat­ ing ewe with greater thyroid activity secretes a larger quan­ tity of milk resulting in increased growth of suckling lambs. The data on the thyroid activity of four lactating ewes and growth of their single lambs in the first three-week period are presented in table 8. Inspection of these data shows that there is no apparent relationship between the two 45 variables. It may be due to the small amount of data or that the ewe with a low thyroid activity was producing enough milk for the rearing of a single lamb. Out of eleven ewes used in this study, four were 6 years old, two were 5, three were 4, and two were 3. Table 8 shows no significant influence of the age of ewes on the thyroid activity during lactation. It may be due to the small amount of data or lactation probably has a greater effect on the function of the thyroid than age. C. Thyroid Secretion Rate and Growth For the study on the growth of lambs in relation to the thyroid activity, twelve Shropshix*e ewe lambs were used. They were fed in the barn for three weeks after weaning (approxi­ mately 120 days of age) to accustom them to barn feeding. At the end of this period, an initial weight was taken and the lambs remained on feed for one month (August 34 to September 33, 1953). month. The final weight was taken at the end of the The daily 1-thyroxine secretion rate of Individual lambs was estimated during this period. For this, different levels of 1-thyroxine were injected, each for 4 days and counts were obtained on the alternate days. A scintillation counter was used to measure the thyroid radioactivity. A regression line was fitted to the ascending portion of the curve, and the dose of 1-thyroxine holding 100 percent radio­ iodine in the thyroid was predicted and considered to be the daily thyroid secretion rate of the individual. The estimated thyroid secretion rate of individual lambs and their growth during a one-month period are presented in table 9. The degree of relationship between the two variables-thyrold secretion rate and growth of ewe lambs--was measured by the correlation and regression coefficients. A correlation of 0.792 was obtained between the daily 1-thyroxine secretion rate and the growth of ewe lambs. This Is statistically sig­ nificant at the 1 percent level, and indicates that the lambs with a higher thyroid activity gained faster than those with a lower thyroid activity. 47 TABLE 9 DAILY 1-THYROXINE SECRETION RATE OF LAMBS AND THEIR GAIN IN WEIGHT DURING ONE MONTH PERIOD Ewe Lamb No. Daily 1-Thyroxine Secretion per 100 lbs. Body Weight -0?g.) _ .. Initial Weight Final Weight (lbs.) ( l b s .) Total Gain (lbs.) 735 0.058 53.0 57.0 4.0 720 0.088 67.0 78.0 11.0 678 0.059 67.0 71.0 4.0 738 0.083 88.5 94.5 6.0 719 0.084 75.0 82.0 7.0 718 0.040 78.5 81.0 2.5 740 0.105 52.5 66.0 13.5 682 0.086 67.5 76.5 9.0 684 0.134 76.5 87.5 11.0 743 0.058 64.0 70.5 6.5 744 0.080 63.5 69.0 5.5 746 0.112 77.5 86.0 8.5 48 The regression of growth of lambs on the daily secretion rate of 1-thyroxine was 97,35. The regression line was fitted to the data, by solving the equation: Y : a 4- bX, as shown in figure 14, Hence, the pounds of gain of ewe lambs (Y) for the period would be estimated by multiphying the mg. of daily 1-thyroxine secretion rate of an individual (X) with the regression coefficient (b) and adding the product to 'a' which is -0.59. The results obtained are in line with some reported research. Glazener and Shaffner (1948) observed rapid growing strains of Barred Plymouth Rock and New Hampshire Red chickens to have higher thyroid activity than slow growing strains. Conversely, the slow5 growing Plymouth Rocks showed a greater growth response to iodinated casein than the rapid growing strain. However, no differences were found between fast and slow feathering strains of Rhode Island Red chicks by Boone, Davidson and Reineke (1950). More recently, Kunkel et al (1953) studied the relationship of the level of serum proteinbound iodine to the rates of gain in beef calves, and reported that there was an optimum level of protein-bound iodine associ­ ated with fast gaining rate of the animals. In the light of their observations the result of this study indicates that the optimum level of thyroid secretion rate was not passed or reached in this study. Moreover, the protein-bound iodine level and the thyroid secretion as determined in the present study are not necessarily directly comparable. The P.B.I. represents the balance between the rate of hormone secretion 49 Y = -0.59-/*97*25 X 12 POUNDS OP GAIN 10 *.025 0.050 0.075 0.100 0.125 l-THYROXINE SECRETION (mg.) Fig. 14* Regression Lino of Ono Month Gain in Weight on Daily 1-Thyroxine Secretion of Ewe Lambs. 0.150 by the thyroid and the rate of its metabolism and excretion. The direct estimate of thyroid secretion gives a measure of the total daily output by the thyroid. 51 D * Thyroid Secretion Hate and Sex In studying differences in the thyroid secretion rate of sheep due to the effect of sex, the group of 20 Shropshire lambs, consisting of 12 ewes, 5 rams, and 4 wethers, was used. While selecting the lambs for this study, age was considered an important factor and held as nearly equal as possible. This study was conducted along with the one on the thyroid secretion rate and growth of ewe lambs under similar feeding and management. The estimated thyroid secretion rates of lambs are pre­ sented in table 10. On the average, the daily 1-thyroxine secretion rate per 100 pounds of body weight in different sexes of lambs was found to be: ewe 0.082 mg., ram 0.066 mg., and wether 0.048 mg. To study the difference in thyroid secretion rate between sexes, in table 11. The the analysis of variance was used as shown *F’ value indicates a significant difference between at least two of the means at the five percent level. By lt ‘ test, the difference between the mean secretion rate of ewe and wether lambs was found to be significant at the five percent level, which Indicates that the ewe lambs on the average secrete more 1-thyroxine than the wether lambs. The differences between the average secretion rate of ewe and ram lambs, and ram and wether lambs were not significant. Studies on the effect of sex on thyroid function has been made for several animal species. In the pigeon, Riddle (19B9) reported that the thyroid of the female was larger than that of the male. Marza and Blinov (1936) made histological 52 TABLE 10 DAILY 1-THYROXINE SECRETED PER 100 POUNDS OF BODY WEIGHT BY EWE, RAM, AND WETHER LAMBS i i • o S3 EWE RAM 1-Thyroxine i Secretion No. (mg. ) 1-Thyroxine i Secretion (mg.) WETHER No. 1-Thyroxine Secretion (mg.) 735 0.058 731 0.065 737 0.068 720 0.088 739 0.051 733 0.042 678 0.059 732 0.052 680 0.031 738 0.083 686 0.076 741 0.052 719 0.084 688 0.085 718 0.040 740 0.105 682 0.086 684 0.134 743 0.058 744 0.080 746 0.112 Average: 0.082 <6: 0.066 0.048 examinations of the thyroid of the two sexes in the pigeon, and stated that the female pigeon had a higher secretory activity than the male. Schultze and Turner (1945) studied the thyroid secretion rate of males and females of the White Plymouth Rock breed of chickens, and reported that the thyroid of the female was more active than that of the male. female, In the the rate ranged from 1.5 ug. to 2.01 ug. of d, 1-thy­ roxine per 100 grams body weight while in the male, from 1.44 ug. to 1.98 ug. it was Monroe and Turner (1946) studied the daily thyroid secretion rate for growing rats and reported that it varied from 2.83 to 4.63 micrograms of d, 1-thyroxine in females and 3.29 to 3.64 micrograms of d ,1-thyroxine in males per hundred grams body weight. TABLE 11 ANALYSIS OF VARIANCE Source of Variation D.F. Total Between sex Error S.S. M.S. 20 0.01297 2 0.00371 0.00186 18 0.00926 0.00051 F. 3.647* •fcSIgnificant at the 5 percent level. Results obtained on the effects of castration on thyroid function are somewhat controversial. Chouke, Friedman and Loeb (1930), working with guinea pigs, observed no significant difference in the mitotic activity of the thyroid as a result of castration. Zalesky (1935) also did not find any gross or microscopic effect of castration on the thyroid in the 13- 54 lined ground squirrel. Schultze and Turner (1945) reported that the thyroid secretion rate or the castrated male fowl was about 16 percent less than that of the normal. Sherwood, Savage and Hall (1933) stated that the metabolism rate in rats was decreased after castration. As the activity of the thyroid gland is governed by the thyrotropic hormone secretion of the anterior pituitary, studies have been made on the effect of castration in this regard. In cattle, thyrotropic hormone content of the pitui­ tary was found to be lowered due to castration (Bates el al 1935, Reece and Turner 1937). Turner and Cupps (1940) also observed a similar result in rats. 55 SUMMARY AND CONCLUSIONS 1. A study was made to determine the relationship of* thyroid activity to lactation, growth and sex in sheep. Experiments were conducted to refine an extrapolation technique for measuring the thyroid secretion rate of live sheep. 2. The thyroid adjusted its secretion to the adminis­ tration of a certain level of 1-thyroxine within two days. 3. The sensitivity of the scintillation tube was found to be greater than the Geiger-Mueller in this study. 4. Using this technique, the estimated daily 1-thyroxine secretion rate of female rats per 100 gms. body weight in two experiments was 2.566 0.145 ug. and 2.768 ± 0.271 ug. These values are in agreement with the secretion rate obtained by the goitrogen technique. 5. The turnover rate (ti) of i^-^l 2 ^he thyroid of eleven lactating Shropshire ewes was found to vary from 144.7 hours to 358.3 hours. 6. The coefficients of correlation and regression obtained between biological ti of seven lactating ewes and gain of suckling twin lambs for the first three-week period were -0*552 and -0.0X3 respectively, which are significant at the 5 percent level. These values indicate that the lac­ tating ewe with a greater thyroid activity probably secretes a larger quantity of milk, resulting in an increased growth of suckling lambs. 56 7. There was no apparent relationship between biologi­ cal ti of four lactating ewes and gain of their suckling single lambs during the first three weeks of lactation. This may be due either to the limited amount of data or to the fact that ewes with low thyroid activity were producing enough milk for the rearing of a single lamb. 8. The effect of thyroid secretion rate on the growth of sheep was studied on twelve ewe lambs, about five months old. The correlation coefficient between daily 1-thyroxine secretion rate and growth of ewe lambs was 0.792, and the regression coefficient was 97.25, both significant at the 1 percent level. This indicates that lambs with higher thyroid activity gained faster than those with lower secre­ tion r a t e . 9. The relationship of sex in sheep to thyroid activity was studied on 12 ewe lambs, of the Shropshire breed. 5 ram lambs, and 4 wether lambs On the average, the daily secretion rate per 100 pounds of body weight in different sexes of lambs was: ewe 0.082 mgm. , ram 0.066 mgm. , and wether 0.048 mgm. Only the difference in secretion rate between ewe and wether lambs is significant at the 5 percent level. 57 APPENDIX I TABLE 1 DATA ON THE THYROID RADIOACTIVITY OF RATS IN RESPONSE TO DIFFERENT LEVELS OF 1-THYROXINE ADMINISTRATION EXPERIMENT I Day Rat 1 1 - Thyroxine Thyr o id* Per cent 1-Thyroxine per 100 Gffl. Count of per 100 Grin. Body Weight per Previous Body Weight (ug.) (ug. ) Minute Count 0 4799 Rat 2 Thyroid* Percent Count or per Previous Minute Count 2432 2 0.42 4034 84.1 0.44 2029 83.4 4 0.85 3621 89.8 0.89 1774 87.4 6 1.27 3340 92.2 1.33 1697 95.7 8 1.69 3116 93.3 1.73 1628 95.9 10 2.12 2927 93.9 2.22 1618 99.4 12 2.54 3023 103.3 2.67 1613 99.7 1518 2874 13 15 2.97 2880 100.2 3.11 1545 101.8 17 3.39 2987 103.7 3.56 1495 96.8 19 3.81 2995 100.3 4.00 1513 101.2 21 4.24 2810 93.8 4.44 1275 84.3 23 8.47 2558 91.0 8.39 123 6 95.3 56 TABLE 1 - continued 1 Day Bat 3 Rat 4 1-Thyroxine Thyroid* Percent 1-Thyroxine Thyroid* Percent per 100 Gm. Count of Count of per 100 Gm. Previous Body Weight Body Weight per per Previous (ug,) Count (ug.) Minute Minute Count 4781 4400 0 2 0,43 3499 79.5 0.37 3552 74.3 4 0.87 3154 90.1 0.74 2941 82.8 6 1.30 2927 92.8 1.11 2493 84.8 8 1.74 2810 96.0 1.48 2036 81.7 10 2.17 2749 97.8 1.85 1737 85.3 12 2.61 2812 102.3 2.22 1613 92.9 1565 2566 13 15 3.04 2474 96.4 2.59 1545 98.7 17 3.48 2263 91.5 2.96 1583 102.4 19 3.91 2434 107.5 3.33 1654 104.5 21 4.35 1894 77.8 3.70 1367 82.7 23 8.70 1789 94.5 7.41 1121 82.0 ^Corrected for background and physical decay 59 TABLE 2 DATA ON THE THYROID RADIOACTIVITY OF RATS IN RESPONSE TO DIFFERENT LEVELS OF 1-THYROXINE ADMINISTRATION EXPERIMENT II i Day 1-Thyroxine per XOO On. Body Weight (ug.) Rat 1 Thyroid* Percent Count per Min. Previous Count Rat 2 Thyroid* Percent of Count per Min. Previous Count 0 0 5610 I 0 4613 82.2 3223 80.6 3 1 3832 83.1 2540 78.8 5 2 3514 91.7 2449 96.4 7 3 3223 91.7 2467 100.7 9 4 3216 99.8 2300 93.2 11 6 3081 95.8 2444 106.3 13 8 3043 98.8 2473 101.2 4000 TABLE 2 - continued Day 1 - Thyroxine per 100 Gm. Body Weight Cug.) Hat 3 Thyroid** Percent Count of per Min. Previous Count 0 0 5130 1 0 4624 90.1 2171 76.9 3 1 4332 93.7 1708 78.7 5 2 4427 102.2 1655 97.1 7 3 4474 101.1 1601 96.7 9 4 4318 96.5 1489 93.0 11 6 4347 100.7 1537 103.2 13 8 4457 102.5 1540 100.2 Rat 4 Thyroid-* Percent Count of Previous per Min. Count 2822 Rat 6 Rat 5 6022 0 0 4540 1 0 3545 78.1 5172 85.9 3 1 2847 80.3 4585 88.7 5 2 2757 96.8 4431 96.6 7 3 2683 97.3 4515 101.9 9 4 2404 89.6 4251 94.2 11 6 2415 100.5 4464 105.0 13 8 2398 95.2 4444 99.6 -^Corrected for background and physical decay TABLE 3 DATA ON THE THYROID RADIOACTIVITY OF RATS IN RESPONSE TO DIFFERENT LEVELS OF 1-THYROXINE ADMINISTRATION EXPERIMENT III Rat No. 1 2 3 4 5 6 7 a 9 10 n 12 13 14 15 16 17 18 19 20 21 22 23 24 1-Thyroxine Admini stered Daily/100 Gm. (ug.) 0 0 0 0 1.0 1.0 1.0 1.0 1.0 1.5 1.5 1.5 1.5 1.5 2.0 2.0 2.0 2.0 2.0 2.5 2.5 2.5 2.5 2.5 0 Day* 7 th Day* Thyroid Thyroid Count/Min. Count/Min. 10987 8987 10000 6395 9582 6095 7464 9372 15799 10145 9645 9777 8927 8095 8074 13827 7671 22709 8084 9429 8503 7022 10073 5981 4363 3486 4548 2549 5748 4220 6090 4341 11842 8808 8647 8288 7922 7743 6895 13447 7252 21859 7832 8762 8581 6897 10069 5765 •^Corrected for background and physical decay Percent of Previous Count 39.7 38.8 45. 5 39.9 60.0 71.2 56.5 65.0 74.9 86.8 89.7 84.8 88.7 95.7 85.4 97.3 94.5 96.3 96.9 92.9 100.9 98.2 100.0 96.4 62 TABLE 4 DATA ON THE THYROID RADIOACTIVITY IN RESPONSE TO DIFFERENT LEVELS OF 1-THYROXINE ADMINISTRATION TO EWE LAMBS Day 1 -Thyroxine per 100 lbs. Body Weight (mg.) Ewe 735 Thyroid Count per Minute Percent of Previous Count 0 0 S2090 2 0 20448 92.6 4 0 .05 18962 92.7 6 0.05 18547 97.8 8 0.10 17760 95.8 10 0.10 19516 109.9 IS 0.15 2XS37 108.8 14 0.15 18655 87.8 18800 100.8 16 TABLE 4 - continued 1 -Thyroxine per 100 lbs. Body Weight Day (mg# ) Ewe 720 Thyroid* Count per minute Percent of Previous Count 0 0 24217 2 0 21679 89.5 4 0.05 19318 89.1 6 0.05 17741 91.8 Q 0.10 16840 94.9 10 0.10 18088 107.4 12 0.15 17574 97.2 14 0.15 17563 99.9 16 0.20 18680 106.4 18 0.20 16898 90.5 16187 95.8 20 64 TABLE 4- continued Day 1-Thyroxine Ewe 678 Ewe 738 per 100 lbs. Body Weight Thyroid* Percent of Thyroid* Percent od Count Previous Previous Count (mg.) per Min. Count Count per Min. 0 0 22182 2 0 21135 95.3 23551 93.3 4 0.05 20393 96.5 22825 96.9 6 0.05 20110 98.6 22764 99.7 8 o H• o 19440 96.7 21240 93.3 10 0.10 20516 105.5 21111 .99.4 12 0.15 21150 103.1 21894 103.7 14 0.15 18655 88.2 22781 104.1 16 0.20 18280 98.0 20320 89.2 IS 0.20 17965 98.3 20856 102.6 18476 102.8 21809 104.6 20 25242 65 TABLE 4 - continued 1-Thyroxine per 100 lbs. Body Weight Day (mg,) Ewe 719 Ewe 718 Thyroid* Percent of Thyroid* Percent of Count Previous Count Previous 'per Min, Count Count per Min. 0 Q 23249 2 0 22367 96.2 16885 93.3 4 0,05 19648 87.8 14454 85.6 6 0,05 19454 99.0 15204 105.2 8 0.10 18200 93.6 14900 98.0 0.10 19326 106.2 15232 102.2 12 0.15 18747 97.G 14768 97.0 14 0.15 17664 94.2 15312 103.7 16 0.25 18680 105.8 13480 88.0 18 0.25 18533 99.2 12964 96.2 17120 92.4 13221 102.0 20 18102 66 TABLE 4 - continued 1-Thyroxine per 100 Lbs. Body Weight Day (mg.) Ewe 740 Ewe 682 Thyroid*!'- Percent of Thyroid* Percent of Count Previous Count Previous 'per Min. Count per Min. Count 0 0 20668 2 0 18177 87.9 25121 95.6 4 0.05 15283 84.1 23183 92.3 6 0.05 14398 94.2 22142 95.5 8 0.10 13360 92.8 21300 96.2 10 0.10 13447 100.7 21039 98.8 12 0.15 13107 97.5 21866 103.9 14 0.15 12617 96.3 22404 102.5 16 0.25 11680 92.6 22080 98.6 18 0.25 11471 98.2 20453 92.6 11498 100.2 22205 108.6 20 26273 67 TABLE 4- continued 1-Thyroxine per 100 Lbs. Body Weight Day (mg.) Ewe 684 Ewe 743 Thyroid* Percent of Thyroid* Percent of Count Previous Count Previous per Min. Count per Min. Count 0 0 19514 2 0 16993 87.1 16993 94.0 4 0.05 15885 93.5 15197 89.4 6 0.05 14112 88.8 14734 97.0 8 0.10 13320 94.4 14700 99.8 XO 0.10 13376 100.4 14875 101.2 12 0.15 12794 95.6 15112 101.6 14 0.15 11595 90.6 16027 106.1 16 0.30 11120 95.9 13280 82.9 18 0.30 11421 102.7 14125 106.4 10283 90.0 13504 95.6 20 18082 68 TABLE 4 - continued Day 1-Thyroxine per 100 Lbs* Body Weight (mg.) 1 Ewe 744 Thyroid* Percent of Count Previous per Min. Count Ewe 746 Thyroid* Percent of Count Previous per Min. Count 0 0 27736 2 0 24808 89.4 24156 90.0 4 0.05 22997 92.7 21580 89.3 6 0.05 22848 99.4 19488 90.3 a 0.10 20720 90.7 18300 93.9 10 0.10 20563 99.2 18207 99.5 12 0.15 21982 106.9 18144 99.7 14 0 *15 20120 91.5 17734 97.7 16 0.30 20480 101.8 18120 102.2 18 0.30 20998 102.5 15761 87.0 22261 106.0 16611 105.4 20 26839 ^-Corrected for background and physical decay 69 TABLE 5 DATA ON THE THYROID RADIOACTIVITY IN RESPONSE TO DIFFERENT LEVELS OF 1-THYROXINE ADMINISTRATION TO WETHER LAMBS 1-Thyroxine per 100 Lbs. Body Weight Day (mg. ) Wether 737 Wether 733 Thyroid* Percent of Thyroid* Percent of Count Previous Previous Count per Min. per Min. Count Count 28783 0 0 13118 2 0 12077 92.1 27754 96.4 4 0.05 10984 90.9 25352 91.3 6 0.05 9744 88.7 27754 109.5 8 0.10 10200 104.7 26120 94.1 10 0.10 11160 109.4 22284 85.3 12 0.15 11167 100.1 23830 106.9 10114 90.6 20563 86.3 14 70 TABLE 5 - continued Day 1-Thyroxine 1 Wether 680 Wether 741 per 100 Lbs. Body Weight Thyroid* Percent of Thyroid* Percent of Previous Count Count Previous Count Count per Min. (mg.) 1per Min. 0 0 29819 2 0 28925 97.0 15127 93.6 4 0*05 26085 90.2 13141 86.9 6 0,05 28241 108.3 12768 97.2 3 0.10 29160 103.3 12800 100.3 10 0.10 28020 96.1 13528 105.7 12 0.15 24929 89.0 13705 101.3 25872 103.8 12768 93.2 14 16158 ■^Corrected for background and physical decay 71 TABLE 6 DATA ON THE THYROID RADIOACTIVITY IN RESPONSE TO DIFFERENT LEVELS OF 1-THYROXINE ADMINISTRATION TO RAM LAMBS Day 1-Thyroxine ' Ram 731 Ram 739 per 100 Lbs* Body Weight Thyroid* Percent or Thyroid* Percent of Count Previous Count Previous (mg. ) per Min. Count per Min. Count 0 0 13585 3 0 13174 96.7 30593 94.8 4 0.05 10998 90.3 17343 84.3 6 0.05 10668 97.0 17430 100.5 3 0.10 10480 98.3 17300 98.7 10 0.10 10677 101.9 17936 104.3 13 0.15 9503 89.0 16074 89.6 9106 95.8 16531 103.8 14 21719 TABLE 6 - continued Day 1 - Thyroxine Bam 732 Ram 686 per 100 Lbs. Body Weight Thyroid* Percent of* Thyroid* Percent of Count Previous Count Previous (mg. ) per Min. Count per Min. Count 0 0 12108 2 0 10363 85.6 19445 93.8 4 0.05 9842 95.0 17766 91.4 6 0.05 10248 104.1 17421 98.1 8 0.10 9640 94.1 16800 96.4 10 0.10 8744 90.7 16003 95.3 12 0.15 8122 92.9 16074 100.4 8198 100.9 15926 99.1 14 20725 TABLE 6 - continued Day 1-Thyroxine per 100 Lbs. Body Weight (mg.) Ram 688 Thyroid* Count per minute Percent of* Previous Count 0 0 14716 2 0 12766 86.7 4 0.05 11393 89.2 6 0.05 10248 89.9 8 0.10 10280 100.3 10 0.10 10434 101.5 12 0.15 9588 91.9 8837 92.2 14 ^Corrected for background and physical decay LITERATURE CITED Archibald, 1945 J. G. Some Effects of Thyroprotein on the Composi­ tion of Milk. Jour. Dairy Sci. 28: 941. Aron, M. and J. Benoit. 1932 Action Antagoniste de le Thyreo-stimuline Prehypophysaire et de la Folliculine Ovarienne sur le Fonctionnement Thyroidien. Compt. Rend Soc. de Biol. 109: 923. Basinger, 1916 H. R. Bates, R. 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