@prefix vivo: . @prefix edm: . @prefix ns0: . @prefix dcterms: . @prefix skos: . vivo:departmentOrSchool "Land and Food Systems, Faculty of"@en ; edm:dataProvider "DSpace"@en ; ns0:degreeCampus "UBCV"@en ; dcterms:creator "Leung, Peter Chung Kwor"@en ; dcterms:issued "2010-01-28T21:36:04Z"@en, "1975"@en ; vivo:relatedDegree "Master of Science - MSc"@en ; ns0:degreeGrantor "University of British Columbia"@en ; dcterms:description """White Leghorn pullets were fed goitrogenic rapeseed meal or soybean meal during either or both the growing period and the laying period. Rapeseed meal was fed at a level of 17.5 percent of the starter-grower diet and 19.0 percent of the layer diet. Control rations contained 11.0 and 13.1 percent of soybean meal, respectively. The responses to the dietary treatments were studied when the birds were over 90 weeks of age. Parameters measured included: weights, histology and iodine contents of the thyroid glands, thyroidal I-131 uptake at different time intervals after administration of the radio-iodine, distribution of injected doses of radioiodine in iodinated substances in the thyroid glands, iodine contents of egg yolks, and the thyroidal characteristics of the progeny of these birds at hatching. Birds which were being fed rapeseed meal at the time the thyroid glands were examined showed effects which varied in magnitude according to the length of time over which rapeseed meal had been fed. Thyroid weights, follicle diameters, amounts of epithium and iodine contents of the thyroid glands of rapeseed meal-fed birds increased whereas coupling efficiency declined with time. The effects of overstimulation of the thyroid gland induced by the feeding of goitrogenic rapeseed meal during the growing period persisted for as long as 75 weeks after withdrawal of the source of goitrogen from the diet. Thyroid weights, thyroid iodine contents and radioiodine uptake were all increased in birds which had received goitrogen during the growing period. Follicle diameters were greater and heights of epithelial cells were lower in the thyroid glands of these birds than in those of the control. Eggs from rapeseed meal-fed birds were low in iodine. The thyroid glands of newly-hatched progeny from dams which had been fed goitrogenic rapeseed meal were observed to be hypertrophic. Since analysis of the eggs for goitrin was negative and analysis for iso-thiocyanates indicated similar concentrations in the eggs from birds fed rapeseed meal and from control birds fed soybean meal, the hypothesis was that the low concentration of iodine in the eggs of the goitrous birds was due to diversion of a large proportion of circulating iodine into the thyroid glands with the result that the amounts reaching the developing ova were reduced."""@en ; edm:aggregatedCHO "https://circle.library.ubc.ca/rest/handle/2429/19277?expand=metadata"@en ; skos:note "EFFECTS OF CHRONIC AND EARLY GOITROGENIC .STIMULATION ON THYROIDAL METABOLISM I N THE CHICKEN AND IN - - \" THE PROGENY. . • BY PETER CHUNG KWOR LEUNG A THESIS SUBMITTED IN PARTIAL FULFILMENT OF THE REQUIREMENTS FOR THE DEGREE OF MASTER OF SCIENCE i n t h e Department o f i POULTRY SCIENCE We a c c e p t t h i s t h e s i s as c o n f o r m i n g t o t h e r e q u i r e d s t a n d a r d THE UNIVERSITY OF BRITISH COLUMBIA Augus t , 1975 In present ing th is thes is in p a r t i a l fu l f i lment of the requirements for an advanced degree at the Un ivers i ty of B r i t i s h Columbia, I agree that the L ibrary sha l l make it f ree ly ava i l ab le for reference and study. I fur ther agree that permission for extensive copying of th is thes is for s c h o l a r l y purposes may be granted by the Head of my Department or by h is representa t ives . It is understood that copying or p u b l i c a t i o n of th is thes is for f inanc ia l gain sha l l not be allowed without my wri t ten permiss ion. Department of P o u l t r y S c i e n c e The Un ivers i ty of B r i t i s h Columbia 2075 Wesbrook Place Vancouver, Canada V6T 1W5 Date August 15, 1975 ABSTRACT White Leghorn p u l l e t s were f e d g o i t r o g e n i c r a p e s e e d meal o r soybean meal d u r i n g e i t h e r o r both t h e growing p e r i o d and the l a y i n g p e r i o d . Rapeseed meal was f e d a t a l e v e l o f 17.5 p e r c e n t o f t h e s t a r t e r - g r o w e r d i e t and 19.0 p e r c e n t o f the l a y e r d i e t . C o n t r o l r a t i o n s c o n t a i n e d 11.0 and 13.1 p e r c e n t o f soybean m e a l , r e s p e c t i v e l y . The r e s p o n s e s t o the d i e t a r y t r e a t m e n t s were s t u d i e d when t h e b i r d s were o v e r 90 weeks o f age. P a r a m e t e r s measured i n c l u d e d : w e i g h t s , h i s t o l o g y and i o d i n e c o n t e n t s o f t h e t h y r o i d g l a n d s , t h y r o i d a l 1-131 u p t a k e a t d i f f e r e n t t ime i n t e r v a l s a f t e r a d m i n i s t r a t i o n o f the r a d i o -i o d i n e , d i s t r i b u t i o n o f i n j e c t e d doses o f r a d i o i o d i n e i n i d d i n a t e d s u b s t a n c e s i n t h e t h y r o i d g l a n d s , i o d i n e c o n t e n t s o f egg y o l k s , and t h e t h y r o i d a l c h a r a c t e r i s t i c s o f the progeny o f t h e s e b i r d s a t h a t c h i n g . B i r d s which were b e i n g f e d r a p e s e e d meal a t the t i m e t h e t h y r o i d g l a n d s were examined showed e f f e c t s which v a r i e d i n magnitude a c c o r d i n g t o the l e n g t h o f t i m e o v e r which r a p e s e e d meal had been f e d . T h y r o i d w e i g h t s , f o l l i c l e d i a m e t e r s , amounts o f e p i t h i u m and i o d i n e c o n t e n t s o f t h e t h y r o i d g l a n d s o f r a p e s e e d m e a l - f e d b i r d s i n c r e a s e d whereas c o u p l i n g e f f i c i e n c y d e c l i n e d w i t h t i m e . The e f f e c t s o f o v e r s t i m u l a t i o n o f t h e t h y r o i d g l a n d i n d u c e d by t h e f e e d i n g o f g o i t r o g e n i c r a p e s e e d meal d u r i n g t h e growing p e r i o d p e r s i s t e d f o r as l o n g as 75 weeks a f t e r w i t h d r a w a l o f the s o u r c e o f g o i t r o g e n f r o m t h e d i e t . T h y r o i d w e i g h t s , t h y r o i d i o d i n e c o n t e n t s and r a d i o i o d i n e uptake were a l l i n c r e a s e d i n b i r d s which had r e c e i v e d g o i t r o g e n d u r i n g the growing p e r i o d . F o l l i c l e d i a m e t e r s were g r e a t e r - i i i -and h e i g h t s o f e p i t h e l i a l c e l l s were lower in::the t h y r o i d g l a n d s o f t h e s e b i r d s t h a n i n t h o s e o f t h e c o n t r o l . Eggs from r a p e s e e d m e a l - f e d b i r d s were low i n i o d i n e . The t h y r o i d g l a n d s o f n e w l y - h a t c h e d progeny from dams which had been f e d g o i t r o g e n i c r a p e s e e d meal were o b s e r v e d t o be h y p e r t r o p h i c . S i n c e a n a l y s i s o f the eggs f o r g o i t r i n was n e g a t i v e and a n a l y s i s f o r i s o -t h i o c y a n a t e s i n d i c a t e d s i m i l a r c o n c e n t r a t i o n s i n t h e eggs from b i r d s f e d r a p e s e e d meal and from c o n t r o l b i r d s f e d soybean m e a l , t h e hypo-t h e s i s was t h a t the low c o n c e n t r a t i o n o f i o d i n e i n t h e eggs o f the g o i t r o u s b i r d s was due t o d i v e r s i o n o f a l a r g e p r o p o r t i o n o f c i r c u l a t i n g i o d i n e i n t o the t h y r o i d g l a n d s w i t h the r e s u l t t h a t t h e amounts r e a c h i n g t h e d e v e l o p i n g ova were r e d u c e d . - i v -TABLE OF CONTENTS Page ABSTRACT i i TABLE OF CONTENTS i v LIST OF TABLES v i LIST OF FIGURES v i i ACKNOWLEDGEMENTS v i i i INTRODUCTION i x REVIEW OF LITERATURE BIOSYNTHESIS AND RELEASE OF THYROID HORMONES 1 EFFECTS OF GOITROGENS ON THYROID HORMONE BIO-SYNTHESIS 6 RAPESEED MEAL AS A SOURCE OF GOITROGEN 8 EFFECTS OF GOITRIN ON THYROID METABOLISM ON THE CHICKEN 10 EFFECTS OF LONG-TERM ADMINISTRATION OF RAPESEED MEAL ON BIRDS AND PROGENY ,13 EXPERIMENTAL E x p e r i m e n t 1: EFFECTS OF CHRONIC AND EARLY STIMU-LATION BY GOITROGENIC ,RSM ON WEIGHT AND HISTOLOGY OF THYROID GLAND 15 E x p e r i m e n t 2: EFFECTS OF CHRONIC AND EARLY STIMU-LATION BY GOITROGENIC-RSM ON TOTAL IODINE CONTENT OF THE THYROID GLAND 23 E x p e r i m e n t 3: EFFECTS ON UPTAKE OF A SINGLE DOSE OF 1-131 BY THE THYROID GLAND 6 HOURS AND 22 HOURS AFTER ADMINISTRATION OF THE RADIOIODINE 29 E x p e r i m e n t 4: THE DISTRIBUTION OF A SINGLE DOSE OF 1-131 IN IODINATED SUBSTANCES IN THYROID GLANDS OF SBM- OR RSM-FED BIRDS 33 - V -E x p e r i m e n t 5: EitEECTSi,OF GOITROGENIC RSM ON SUS-CEPTIBILITY TO HYDROLYSIS OF THYROGLOBULIN 39 Ex p e r i m e n t 6A: EFFECTS OF GOITROGENIC RSM ON THE IODINE CONTENT OF EGG YOLK 42 Exp e r i m e n t 6B: EFFECTS OF INCREASED DIETARY IODINE INTAKE ON IODINE DEPOSITION IN EGG YOLKS OF SBM-AND RSM-FED BIRDS 46 Ex p e r i m e n t 7: OVARIAN UPTAKE OF A SINGLE DOSE OF 1-131 BY DEVELOPING FOLLICLES 50 Exp e r i m e n t 8: EFFECTS OF CHRONIC AND EARLY GOI-TROGENIC STIMULATION BY RSM IN THE DAM ON EMBRYONIC THYROID DEVELOPMENT 55 DISCUSSION THYROIDAL RESPONSE TO CHRONIC GOITROGENIC STIMU-LATION BY RSM 60 PERSISTENCE OF EFFECTS OF EARLY GOITROGENIC STIMU-LATION 67 EFFECTS OF ALTERATIONS IN MATERNAL THYROID META-BOLISM ON EMBRYONIC THYROID DEVELOPMENT IN THE CHICK 70 SUMMARY AND CONCLUSIONS 72 BIBLIOGRAPHY 75 APPENDIX LIST OF TABLES T a b l e Page 1 P e r c e n t a g e c o m p o s i t i o n o f d i e t s f e d i n e x p e r i m e n t 1 and RSM g o i t r o g e n i c i t y t e s t 19 2 Comparison o f a v e r a g e t h y r o i d w e i g h t s o f White Leghorn c o c k e r e l s f e d SBM- o r RSM-supplemented d i e t s a t 1 week o r 3 weeks o f t r e a t m e n t from h a t c h i n g 20 3 Average w e i g h t s o f t h y r o i d g l a n d s i n b i r d s f e d RSM o r SBM f o r d i f f e r e n t p e r i o d s o f t i m e i n e x p e r i m e n t 1 21 4 H i s t o l o g i c a l measurements on t h y r o i d g l a n d s from b i r d s ( S h a v e r s t r a i n ) i n e x p e r i m e n t 1 a t 91 weeks o f age 22 5 I o d i n e c o n c e n t r a t i o n i n t h y r o i d g l a n d s o f b i r d s f e d RSM o r SBM f o r d i f f e r e n t p e r i o d s o f t i m e 27 6 A n a l y z e d i o d i n e c o n c e n t r a t i o n s i n SBM- and RSM-supple-mented d i e t s f e d t o b i r d s i n e x p e r i m e n t s 1 and 6B 28 7 C o n c e n t r a t i o n s o f r a d i o i o d i n e i n t h e t h y r o i d g l a n d and l i v e r 6 and 22 hours a f t e r a d m i n i s t r a t i o n 32 8 D i s t r i b u t i o n , 22 hours a f t e r a d m i n i s t r a t i o n , o f a s i n g l e dose o f r a d i o i o d i n e i n t o t h y r o i d a l compounds 37 9 D i s t r i b u t i o n , 22 hours a f t e r a d m i n i s t r a t i o n , o f a s i n g l e dose o f r a d i o i o d i n e i n t o t h y r o i d a l compounds measured i n h y d r o l y ^ a t e s a f t e r d i f f e r e n t t i m e s o f p r o n a s e d i g e s t i o n . . 41 10 I o d i n e c o n c e n t r a t i o n s i n egg y o l k s from b i r d s on d i f f e r e n t d i e t a r y t r e a t m e n t s i n e x p e r i m e n t 6A 45 11 I o d i n e c o n c e n t r a t i o n s i n egg y o l k s from b i r d s on d i f f e r e n t d i e t a r y t r e a t m e n t s i n e x p e r i m e n t 6B 49 12 R a d i o i o d i n e c o n c e n t r a t i o n s i n t h e t h y r o i d g l a n d s and d e v e l o p i n g o v a r i a n f o l l i c l e s 22 hours a f t e r a d m i n i s t r a t i o n o f a s i n g l e dose o f r a d i o i o d i n e t o b i r d s which had been f e d RSM o r SBM f o r d i f f e r e n t p e r i o d s o f time 53 13 T h y r o i d w e i g h t s a t h a t c h i n g o f progeny b i r d s f e d RSM o r SBM f o r d i f f e r e n t p e r i o d s i n e x p e r i m e n t 8 57 14 . T h y r o i d a l c h a r a c t e r i s t i c s i n n e w l y - h a t c h e d progeny o f b i r d s f e d RSM o r SBM f o r d i f f e r e n t p e r i o d s i n e x p e r i m e n t 8. 58 15 Comparison o f t h y r o i d a l c h a r a c t e r i s t i c s o f dams and p r o -geny on d i f f e r e n t d i e t a r y t r e a t m e n t s 59 - v i i -TABLE OF FIGURES F i g u r e Page 1 Paper chromatograms o f t h y r o i d a l m e t a b o l i s m o f b i r d s on d i f f e r e n t d i e t a r y t r e a t m e n t s 38 2 R e g r e s s i o n o f 1-131 u p t a k e on s u r f a c e a r e a o f o v a r i a n f o l l i c l e s o f b i r d s f e d RSM o r SBM f o r d i f f e r e n t p e r i o d s . . . 54 - v i i i -ACKNOWLEDGEMENTS The a u t h o r g r a t e f u l l y acknowledges t h e s u p e r v i s i o n and encouragement o f P r o f e s s o r B.E. March i n a c c o r d w i t h h e r p a r t i c u l a r c o n t r i b u t i o n t o my academic development (leading t o t h e c o m p l e t i o n o f t h i s work, and a l s o h e r many h e l p f u l a d v i c e s i n a r e a s o u t s i d e t he academic f i e l d . A f u l l measure o f a p p r e c i a t i o n i s a l s o e x t e n d e d t o the o t h e r members o f my T h e s i s Committee: Dr. W.D. K i t t s Chairman, Departments o f Animal S c i e n c e and P o u l t r y S c i e n c e Dr. R i C . Fitzsimmons Department o f P o u l t r y S c i e n c e Dr. C.R. K r i s h n a m u r t i Department o f Animal S c i e n c e Dr. A.M. P e r k s Department o f Z o o l o g y In p a r t i c u l a r , Dr. A.M. Perks must r e c e i v e s p e c i a l m e n t i o n i n a c c o r d w i t h h i s h e l p and c o n c e r n i n my programme. S p e c i a l a p p r e c i a t i o n i s g i v e n t o Dr. J . B i e l y ( R e s e a r c h P r o f e s s o r , Department o f P o u l t r y S c i e n c e ) f o r h i s many i n t e r e s t i n g and s t i m u l a t i n g i d e a s t h a t p r o v i d e d f o o d f o r t h o u g h t , and t o Mr. R ^ S o o n g ( P o u l t r y N u t r i t i o n L a b o r a t o r y ) f o r h i s f r e q u e n t a s s i s t a n c e when needed. - I X -INTRODUCTION The t h y r o i d g l a n d i s one o f t h e f i r s t e n d o c r i n e organs s t u d i e d by man, and the e f f e c t s o f i t s hormones on man and a n i m a l s have l o n g been known. Many a s p e c t s o f i t s m e t a b o l i s m , however, s t i l l remain u n c l e a r . A l t h o u g h many i n v e s t i g a t i o n s have been made on the f o r m a t i o n o f t h e t h y r o i d hormones, both i n v i t r o and i n v i v o , the e x a c t mechanism whereby t h y r o i d hormonogenesis t a k e s p l a c e under p h y s i o l o g i c a l c o n d i t i o n s i s s t i l l u n c e r t a i n . The p i c t u r e becomes more c o m p l i c a t e d when the t h y r o i d g l a n d i s i n h i b i t e d by s u b s t a n c e s such as g o i t r o g e n s . T h y r o i d f u n c t i o n and hormone b i o s y n t h e s i s i s e i t h e r a f f e c t d d o r b l o c k e d , dependent on the mode o f a c t i o n o f t h e g o i t r o g e n c o n c e r n e d . W h i l e numerous s t u d i e s have been made on t h e e f f e c t s o f t h e a n t i t h y r o i d compounds on t h y r o i d f u n c -t i o n s o f e x p e r i m e n t a l a n i m a l s , o n l y a few o f t h e s e i n v e s t i g a t i o n s have examined t h e e f f e c t s o f c h r o n i c g o i t r o g e n i c s t i m u l a t i o n on t h e t h y r o i d g l a n d . Much l e s s i s known o f any p o s s i b l e p e r s i s t e n c e o f e f f e c t s o f e a r l y s t i m u l a t i o n ? by g o i t r o g e n s when t h e y a r e l a t e r withdrawn. In t h e p r e s e n t s t u d y , r a p e s e e d meal was f e d t o c h i c k e n s e i t h e r j u s t f o r t h e growing p e r i o d o r o v e r a l o n g p e r i o d o f t i m e , and t h e e f f e c t s o f t h e d i e t a r y t r e a t m e n t s on the t h y r o i d g l a n d s were examined. Rapeseed i s one o f t h e most v a l u a b l e o i l s e e d c r o p s i n Canada, both f o r i t s p o t e n t i a l i n t h e p r o d u c t i o n o f e d i b l e and i n d u s t r i a l o i l s and i n the m a n u f a c t u r e o f h i g h p r o t e i n meals s u i t a b l e f o r s u p p l e m e n t i n g animal d i e t s . I t has been i n d i c a t e d i n t h e l i t e r a t u r e t h a t the mode o f a c t i o n o f t h e r a p e s e e d meal g o i t r o g e n s i s d i f f e r e n t from t h a t o f o t h e r a n t i --x-t h y r o i d compounds. T h e r e f o r e , a l t h o u g h i t i s now p o s s i b l e t o m a n u f a c t u r e r a p e s e e d meals w i t h ^ a n e g l i g i b l e g o i t r o g e n c o n t e n t , t h e e f f e c t s o f c h r o n i c s t i m u l a t i o n by g o i t r o g e n i c r a p e s e e d meal on t h y r o i d h o r monogenesis, compared t o o t h e r a n t i t h y r o i d compounds such as p r o p y l t h i o u r a c i l and t h i o u r e a , a r e s t i l l w orthy o f i n v e s t i g a t i o n from t h e academic s t a n d p o i n t . In a d d i t i o n , t h e p o s s i b i l i t y t h a t e f f e c t s o f e a r l y s t i m u l a t i o n o f t h e t h y r o i d g l a n d may p e r s i s t v a f t e r w i t h d r a w a l o f t h e g o i t r o g e n w a r r a n t s f u r t h e r s t u d i e s i n i t s own r i g h t . - 1 -REVIEW OF LITERATURE BIOSYNTHESIS AND RELEASE OF THYROID HORMONES The i m p o r t a n c e o f i o d i n e i n the p r e v e n t i o n o f g o i t e r has l o n g been r e c o g n i z e d ( H a r t and S t e e n b e c k , 1918; K a l k u s , 1920; Welch, 1928) and i o d i n e m e t a b o l i s m i n t h e body has been t h o r o u g h l y r e v i e w e d ( W i l g u s e t a l , 1953; C h a i k o f f and T a u r g g , 1948; Hemken, 1970; K o u t r a s , 1972). In t h e a l i m e n t a r y t r a c t , d i e t a r y i o d i d e i s r e a d i l y a b s o r b e d i n t o t h e c i r c u l a t i o n and i s m a i n l y c o n c e n t r a t e d w i t h i n t h e t h y r o i d . The r o l e o f the t h y r o i d , t h e n , i s t o i n c o r p o r a t e i n o r g a n i c i o d i d e i n t o t h y r o i d hormones and t o m a i n t a i n a s u f f i c i e n t r e l e a s e o f t h e s e hormones t o t h e p e r i p h e r y t i s s u e s . I n t r a t h y r o i d a l b i o s y n t h e s i s o f t h y r o i d hormones has been ex-t e n s i v e l y r e v i e w e d ( P l a s k e t t et_ al _ , 1963; Rapoport and DeGroot, 1971; D a v i e s , 1972; Ermans e t a l _ , 1972; DeGroot e t a l _ , 1972; Mason and W i l k i n s o n , 1973). T h e r e f o r e i n the p r e s e n t d i s c u s s i o n , we w i l l r e v i e w o n l y t h e more r e c e n t developments i n the p h y s i o l o g y o f t h y r o i d hormone b i o s y n t h e s i s , w i t h s p e c i a l a t t e n t i o n t o such a r e a s t h a t may perhaps be a f f e c t e d by g o i t r o g e n i c a g e n t s . I t i s g e n e r a l l y a g r e e d t h a t f o r m a t i o n o f t h y r o i d hormones i n -v o l v e s t h e f o l l o w i n g i n t e r m e d i a r y s t e p s (DeGroot e t aj_, 1972): 1) t r a n s -p o r t and c o n c e n t r a t i o n o f i o d i d e i n t o t he t h y r o i d , 2) s y n t h e s i s o f t h y r o -g l o b u l i n , 3) g e n e r a t i o n o f an o x i d i z i n g a g e n t , p r o b a b l y hydrogen p e r o x i d e , 4) o x i d a t i o n o f i o d i d e t o a r e a c t i v e h i g h e r v a l e n c e s t a t e , 5) b i n d i n g o f i o d i n e t o t y r o s i n e p r e s e n t i n t h y r o g l o b u l i n , 6) f o r m a t i o n o f i o d o t h y r o n i n e s i n t h y r o g l o b u l i n by c o u p l i n g o f i o d o t y r o s y l r e s i d e s , 7) s t o r a g e and p r o -t e o l y s i s o f t h y r o g l o b u l i n , and 8) r e l e a s e o f t h y r o i d hormones t o t h e c i r c u l a t i o n . - 2 -I o d i d e t r a p p i n g by t h e t h y r o i d i s dependent on adquate p o t a s s i u m c o n c e n t r a t i o n ( W o l f f and H a l m i , 1963) and i s b l o c k e d by i n h i b i t o r s o f o r a g ents t h a t u n c o u p l e o x i d a t i v e p h o s p h o r y l a t i o n such as 2 , 4 - d i n i t r o p h e n o l , c y a n i d e , c a r d i a c g l y c o s i d e s and a n o x i a (Wyngaarden e t a l _ , 1953; F r e i n k e l and I n g b a r , 1955; W o l f f , 1964). T h e r e f o r e i t i s p r o b a b l y an a c t i v e p r o c e s s r e q u i r i n g e n e r g y r e l a t e d w i t h t h e Na-,K +-ATPase-dependent t r a n s p o r t s ystem ( T u r k i n g t o n , 1962). Much a t t e n t i o n has been f o c u s s e d on t h e s i t e o f t h i s i o d i d e pump mechanism, which i s p r o b a b l y a membrane r e c e p t o r s i t e l o c a t e d i n : t h e p e r i p h e r a l b o r d e r o f t h e f o l l i c u l a r c e l l s o f t h e t h y r o i d ( H a l m i , 1961; P i t t - R i v e r s and T r o t t e r , 1953; Tong e t a l _ , 1962). I t has been s u g g e s t e d t h a t a l e c i t h i n p h o s p h o l i p i d i s a c o n s t i t u e n t o f t h e r e c e p -t o r s i t e ( V i l k k i and Jaakonmaki, 1966) and a r o l e o f F e + + has a l s o been i m p l i c a t e d (Dhopeshwarkar, 1968). T h y r o g l o b u l i n , l i k e o t h e r p r o t e i n s , i s s y n t h e s i z e d i n the p o l y r i b o s o m e s o f the e n d o p l a s m i c r e t i c u l u m o f t h e t h y r o i d c e l l s ( N a d l e r et_ a l _ , 1964; Thomson and G o l d b e r g , 1968); i t t h e n t r a v e l s down t o t h e a p i -c a l r e g i o n o f t h e c e l l s t o be a c c u m u l a t e d i n t h e c o l l o i d (Whur e t a l _ , 1969). I t i s a g l y c o p r o t e i n w i t h a m o l e c u l a r w e i g h t o f a b o u t 650,000 ( G o l d b e r g and L e e d , 1965; Robbins and R a i l , 1960; R a i l e t a l _ , 1960) and a s e d i m e n t a t i o n c o e f f i c i e n t o f 19S when i o d i n a t e d . The s i t e o f i t s i o d i n a t i o n has been t h e t o p i c o f much e x t e n s i v e s t u d i e s ( E d e l h o c h , 1962; Nunez, 1966; L i s s i t z k y , e t a l _ , 1969), and i t i s p r o b a b l y a t t h e b o r d e r o f the a p i c a l membrane o f t h e t h y r o i d c e l l and t h e c o l l o i d . The d i s t r i b u t i o n o f v a r i o u s iodoamino a c i d s i n t h e t h y r o i d t i s s u e i s l a r g e l y d e t e r m i n e d by t h e i o d i n a t i o n l e v e l o f the t h y r o g l o b u l i n (DeCrombrugghe e t a l , 1967; Ermans e t a l , 1968; V a l e n t a e t a l _ , 1968). - 3 -Many q u e s t i o n s p e r t a i n i n g t o t h e mechanism o f t h y r o i d hormone f o r m a t i o n a r e s t i l l u n s o l v e d ; t h e y have r e c e n t l y have d i s c u s s e d t h o r o u g h l y by DeGroot e t a l _ (1972). I t i s w i d e l y a c c e p t e d t h a t i n v i v o i o d i n a t i o n s r e q u i r e g e n e r a t i o n , o f H,,02 w h i c h , t o g e t h e r w i t h 0 2 a r e t h e o n l y b i o l o g i c a l o x i d i z i n g a gents w i t h a n . j D x i d a t i o n ^ r e d u c t i o n p o t e n t i a l above t h a t o f the 2 I \" ^ I 2 c o u p l e , i . e . +0.535 v o l t s ( B e r n a r d and DeGroot, 1969; Ahn and Rosenberg, 1970). A l t h o u g h c y t o p l a s m i c DPNH o x i d a s e ( R o s s i e t a l _ , 1969) and monoamine o x i d a s e ( F i s c h e r et_ a l _ , 1966, 1968) a c t i v i t y have been s u g g e s t e d t o be t h e o r i g i n o f H 2 0 2 , t h e c o f a c t o r NADPH i s l i k e l y t o be i n v o l v e d i n t h y r o i d a l H 2 0 2 p r o d u c t i o n , p o s s i b l y v i a r e a c t i o n w i t h NADPH-cytochrome C r e d u c t a s e (DeGroot and D a v i s , 1961; Nagasaka e t a l _ , 1971). The p a r t i c u l a t e - b o u n d enzyme, p e r o x i d a s e , has been i s o l a t e d from t h y r o i d t i s s u e s o f a number o f s p e c i e s ( A l e x a n d e r , 1959; Hosoya and M o r r i s o n , 1967; Mahoney and Igo, 1966; L j u n g g r e n and Adeson, 1968; Naga-saka and DeGroot, 1971). I t i s a heme p r o t e i n c o n t a i n i n g a p r o s t h e t i c group s i m i l a r t o P r o t o p o r p h y r i n IX (Taurog e t a l _ , 1970). In a d d i t i o n t o u t i l i z i n g H 2 0 2 f o r p e r o x i d a t i o n o f i o d i d e t o i o d i n e — p r o b a b l y t o i t s f r e e r a d i c a l f o r m , I 0 ( K l e b a n o f f e t a l _ , 1962; Y i p and H a d l e y , 1966) — p e r o -x i d a s e has a l s o been h e l d r e s p o n s i b l e f o r c a t a l y z i n g t h e o r g a n i f i c a t i o n o f i o d i n e t o t y r o s i n e p r e s e n t i n t h y r o g l o b u l i n . DeGroot e t a l (1972) e n v i s a g e a \"ping-pong\" mechanism o f o r g a n i f i c a t i o n , which presumably i n v o l v e s f o r -m a t i o n o f f r e e r a d i c a l s o f i o d i d e and t y r o s i n e by p e r o x i d a s e complexed w i t h H 2 0 2 . Union o f the i o d i n e and t y r o s i n e f r e e r a d i c a l s t o form i o d o t y r o s i n e r e t u r n s the enzyme t o i t s o r i g i n a l u n o x i d i z e d s t a t e . The s y n t h e s i s o f i o d o t i y r o s i n e s i s a r a p i d p r o c e s s (DeGroot and D a v i s , 1961). D i s t r i b u t i o n o f r a d i o i o d i n e between m o n o i o d o t y r o s i n e (MIT) and d i i o d o t y r o s i n e (DIT) r e a c h e s a c o n s t a n t l e v e l a f t e r v a r i a b l e l e n g t h s - 4 -o f t i m e , and t h e c o n s t a n t MIT/DIT r a t i o i n p h y s i o l o g i c a l c o n d i t i o n s i s c o n s o n a n t w i t h a p r e c u r s o r - p r o d u c t r e l a t i o n s h i p between t h e two i o d o t y r o -s i n e s ( P l a s k e t t e t a l _ , 1963a). When th e i o d i n e c o n t e n t o f t h e t h y r o i d t i s s u e i s r e d u c e d , however, t h e r e a p p e a r s a marked i n c r e a s e i n t h e MIT/DIT r a t i o ( Q u e r i d o e t a l _ , 1957; Ermans e t a l _ , 1961), w h i c h , as m entioned e a r l i e r , i s dependent on the d e g r e e o f i o d i n a t i o n o f t h y r o g l o b u l i n (Ermans e t a l _ , 1968). The 1 coy p i l i n g 1 o f i o d o t y r o s i n e s t o i n t r a t h y r o g l o b u l i n i o d o -t h y r o n i n e s , as f i r s t p r o p o s e d by H a r i n g t o n and B a r g e r (1927), i s a much s l o w e r p r o c e s s ( P i t t - R i v e r s , 1962; P l a s k e t t e t al_, 1963b). A l t h o u g h i t has been a c h i e v e d i n v i t r o ( P i t t - R i v e r s , 1948; Y i p and K l e b a n o f f , 1952), i t s i n v i v o mechanism i s s t i l l u n c e r t a i n . The f a c t t h a t f r e e t h y r o n i n e i s n o t found i n n a t u r e l e a v e s two p o s s i b l e r o u t e s whereby i o d o t h y r o n i n e s can be formed i n t h y r o g l o b u l i n (DeGroot e t a l _ , 1972): 1) by i n t r a m o l e c u l a r r e a r r a n g e m e n t , i . e . c o n f o r m a t i o n a l changes w i t h i n t h e t h y r o g l o b u l i n which a l l o w an i o d o t y r o s y l f r e e r a d i c a l ( p r o d u c e d by an o x i d i z i n g agent such as p e r o x i d a s e - I \" ) t o be t r a n s f e r r e d t o a n o t h e r i o d o t y r o s y l group t o form i o d o -t h y r o n i n e ; o r 2) by o x i d a t i v e - d e a m i n a t i o n o f DIT t o form h y d r o x y p h e n o l -p y r u v a t e ( D I H P P A ) , p e r o x i d a t i o n o f the e n o l form o f which l e a d s t o the f o r m a t i o n o f a DIHPPA-peroxide, which can then be c o u p l e d w i t h a n o t h e r DIT m o l e c u l e i n t h y r o g l o b u l i n t o form t h y r o x i n e ( T o i e t a l _ , 1963, 1965). D e s p i t e e v i d e n c e o f the p r e s e n c e o f n e c e s s a r y enzymes i n the t h y r o i d t i s u s e (Igo e t a l _ , 1968; B l a s i e t a l _ , 1969a,b), t h e e x t e n t o f t h e s e pathways In v i v o t h y r o i d hormone f o r m a t i o n i s s t i l l u n c e r t a i n . T h u s , t h y r o i d hormone i s s t o r e d i n t h e f o l l i c u l a r lumen, as a c o n s t i t u e n t o f t h e t h y r o g l o b u l i n p o l y p e p t i d e . Recent e l e c t r o n m i c r o g r a p h i c a l d a t a show, i n a d d i t i o n t o c o l l o i d d r o p l e t s , two f u n c t i o n a l l y d i f f e r e n t - 5 — t y p e s o f v e s i c l e s i n the a p i c a l r e g i o n s o f t h y r o i d f o l l i c l e c e l l s ( S e l -j e l i d e t a l _ , 1970): e n d o c y t o t i c and e x o c y t o t i c . E x o c y t o t i c v e s i c l e s may be i n v o l v e d i n the t r a n s p o r t o f newly s y n t h e s i z e d t h y r o g l o b u l i n i n t o the f o l l i c l e lumen f o r s t o r a g e , w h i l e the e n d o c y t o t i c v e s i c l e s may be r e s p o n s i b l e f o r t h e r e - e n t r a n c e o f t h y r o g l o b u l i n i n t o t h e c e l l f o r r e l e a s e t o t h e c i r c u l a t i o n . F u r t h e r m o r e , t h e r e i s e v i d e n t f o r t h e \" l a s t come, f i r s t s e r v e d \" p r i n c i p l e o f i o d i n e t u r n o v e r i n the t h y r o i d pool ( S c h n e i d e r , 1964), which i s s u p p o r t e d by t h e s t r u c t u r a l and f u n c t i o n a l h e t e r o g e n e i t y among t h e f o l l i c l e s ( N a d l e r , 1954; S t u d e r e t a l _ , 1972). Once r e - e n t e r e d i n t o t h e t h y r o i d c e l l , r e l e a s e o f t h e t h y r o -g l o b u l i n i s a c h i e v e d by p r o t e o l y s i s by a c i d i c p r o t e a s e s d a r i d ' p e p t i d a s e s c o n t a i n e d i n t h e l y s o s o m e s ( W o l l m a n e t a l , 1964; S e l j e l i d , 1968). The l i b e r a t e d i o d o t y r o s i n e s a r e d e i o n a t e d by i o d o t y r o s i n e d e i o d i n a s e w i t h i n the t h y r o i d c e l l , w h i l e the i o d o t h y r o n i n e s a r e s e c r e t e d i n t o t h e c i r c u -l a t i o n t o a f f e c t t a r g e t t i s s u e s . A l m o s t e v e r y i n t e r m e d i a r y s t e p i n t h y r o i d hormone b i o s y n t h e s i s and r e l e a s e i s r e g u l a t e d one way o r the o t h e r by t h y r o t r o p i n (TSH) from t h e a d e n o h y p o p h y s i s . An e x c e l l e n t r e v i e w o f the a c t i o n o f t h y r o t r o p i n on t h y r o i d m e t a b o l i s m i s g i v e n by Dumont (1971). In a d d i t i o n t o the e x t r a -t h y r o i d a l c l a s s i c a l n e g a t i v e f e e d b a c k c o n t r o l by t h y r o i d hormones and t h e hypothalamic;\\ppn,tr fp:lcby T S H - r e l e a s i n g hormone (TRF) v i a t h e hypothalamo-hypophyseal p o r t a l system ( S c h a l l y e t a l _ , 1969; Burgus and G u i l l e m i n , 1970), t h e r e seems t o be an i n t r a t h y r o i d a l autonomous mechanism whereby t h y r o i d a l i o d i n e m e t a b o l i s m i s r e g u l a t e d ( W o l f f and C h a i k o f f , 1948; Socolow e t a l , 1968; B a r a k a t and I n g b a r , 1965; Golman e t a^, 1966; S t u d e r and G r e e r , 1968). I t a l s o a p p e a r s t o a l t e r t h y r o i d hormone s e c r e t i o n , i n d e p e n d e n t l y o f TSH s u p p l y (Green and I n g b a r , 1962; H a l m i , 1961; Onaya and H a l m i , 1967). S o l o -mon and Dowling (1960) h y p o t h e s i z e d t h a t a u t o r e g u l a t i o n o f t h y r o i d a l i o d i d e - 6 -t r a n s p o r t a l l o w s an i o d i d e - d e f i c i e n t a nimal t o m a i n t a i n i t s i n t r a t h y r o i d a l hormone s t o r e s . The acc u m u l a t e d s u p p l y may then be r e l e a s e d by t h e e x t r a TSH s e c r e t e d i n r e s p o n s e t o lo w e r e d i o d o t h y r o n i n e l e v e l s i n t h e c i r c u l a t i o n . A n a l y s e s o f d i s t r i b u t i o n o f iodoamino a c i d s i n t h y r o i d t i s s u e s has shown l i t t l e d i f f e r e n c e between t h e a v i a n and t h e mammalian g l a n d s ( S h e l l a b a r g e r and P i t t - R i v e r s , 1958; M e l l e n and Wentworth, 1959). Hence a d i v e r s i t y o f mechanisms has n o t been r e c o g n i z e d f o r t h y r o i d hormone b i o -s y n t h e s i s i n the d i f f e r e n t s p e c i e s . However, i t s h o u l d be no t e d t h a t w h i l e i n mammals L - t r i i o d o t h y r o n i n e (Tg) has a h i g h e r p o t e n c y t h a n an e q u i m o l a r q u a n t i t y o f L - t h y r o x i n e ( T ^ ) , t h e i o d o t h y r o n i n e s have d i f f e r e n t r e l a t i v e p o t e n c i e s i n b i r d s ( S h e l l a b a r g e r , 1955; H u t c h i n s and Newcomer, 1966; New-comer, 1957; Robbins and R a i l , 1957). F u r t h e r m o r e , t h e f i n d i n g by T a t a and S h e l l a b a r g e r (1959) t h a t t h y r o x i n e - b i n d i n g g l o b u l i n i s a b s e n t i n c h i c k e n b l o o d p r o b a b l y e x p l a i n s the f a i l u r e o f p r o t e i n - b o u n d i o d i n e (PBI) as a c r i t e r i o n o f measurement o f t h y r o i d hormone m e t a b o l i s m i n the b i r d . EFFECTS OF GOITROGENS ON THYROID HORMONE BIOSYNTHESIS A n t i t h y r o i d a l s u b s t a n c e s and n a t u r a l g o i t r o g e n s have been ex-t e n s i v e l y r e v i e w e d ( V a n E t t e n , 1969). The f o r m e r i n c l u d e , among o i t h e r s , i n o r g a n i c i o n s s u c h as i o d i d e , ( l a r g e doses o f which i n h i b i t t h y r o i d h o r -mone s y n t h e s i s i n the r a t -- the s o - c a l l e d W o l f f - C h a i k o f f e f f e c t ) , t h e t h i o n a m i d e d r u g s such as t h i o u r e a and t h i o u r a c i l , and the s u l f o n a m i d e s such as p a r a - a m i n o b e n z o i c a c i d and s u l f a n i l a m i d e (Aswood, 1943; Astwood e t a l _ , 1945). Among t h e n a t u r a l l y o c c u r r i n g g o i t r o g e n s , ( - ) - 5 - v i n y l - 2 -t h i o - o x a z o l i d o n e i s o f p a r t i c u l a r i m p o r t a n c e because o f i t s p o s s i b l e r o l e i n t h e p a t h o g e n e s i s o f endemic g o i t e r and w i l l be d i s c u s s e d i n more d e t a i l . - 7 -D i s t u r b a n c e s o r i n h i b i t i o n o f t h y r o i d hormone b i o s y n t h e s i s due t o d e f e c t s o f i o d i n e m e t a b o l i s m c a n be c l a s s i f i e d as f o l l o w s ( B l i z z a r d , 1960): 1) ^ d e f e c t i n t r a p p i n g o f i n o r g a n i c i o d i d e from t h e c i r c u l a t i o n ; 2) i n a b i l i t y t o o x i d i z e i o d i d e t o an a c t i v e form; 3) i n a b i l i t y t o h a l o -genate t y r o s i n e i n p r e s e n c e o f e l e m e n t a l i o d i n e ; 4) i n a b i l i t y t o c o u p l e i o d i n a t e d t y r o s i n e s t o form i o d o t h y r o n c i n e s ; 5) i n a b i l i t y t o h y d r o l y z e t h y r o g l o b u l i n ; 6) abnormal f o r m a t i o n o r r e l e a s e o f t h y r o g l o b u l i n o r \" t h y r o g l o b u l i n - l i k e \" s u b s t a n c e s ; 7) i n a b i l i t y t o d e i o d i n a t e f r e e i o d b -t y r o s i n e s f o r r e u t i l i z a t i o n . One o r more o f t h e s e i n t e r m e d i a t e s t e p s o f t h y r o i d hormone b i o s y n t h e s i s can be i n h i b i t e d by a g o i t r o g e n . F o r example, the t h i o n a m i d e s such as t h i o u r a c i l a p p e a r t o i n t e r f e r e w i t h the o r g a n i c b i n d i n g o f i n t r a t h y r o i d a l i o d i d e ( l i n o , 1961), t h u s competing w i t h the i o d i d e w h i l e the p e r o x i d a s e i s a c t i n g (DeGroot and D a v i s , 1962). The s u l f o n a m i d e s i n t e r f e r e w i t h o r g a n i f i c a t i o n o f i o d i n e w i t h o u t i n h i b i t i n g t h e t r a p p i n g o f i o d i d e ( M i l n e and G r e e r , 1962). T h a t t h e i r mechanisms o f a c t i o n a r e d i f f e r e n t i s e v i d e n t by t h e o b s e r v a t i o n t h a t a d d i t i o n o f i o d i n e r e d u c e s g o i t e r s i z e i n a n i m a l s t r e a t e d w i t h t h i o n a m i d e s b u t i n c r e a s e t h y -r o i d h y p e r t r o p h y i n a n i m a l s t r e a t e d w i t h s u l f o n a m i d e s (MacKenzie and M a c K e n z i e , 1943). In a s y s t e m a t i c s t u d y i n which N - i o d o s u c c i n i m d d e was used as a model f o r the a c t i v e i o d i n e o f the t h y r o i d g l a n d ( J i r o u s e k and Soodak, 1973), a l l c l a s s e s o f g o i t r o g e n s t e s t e d were f o u n d t o r e a c t . The e f f e c t s o f t h e v a r i o u s g o i t r o g e n s on i n v i t r o b i o s y n t h e s i s o f t h y r o i d hormones have been d e l i n e a t e d ( l i n o , 1961). T h e i r a b i l i t y t o c o m p l e t e l y i n h i b i t DIT f o r m a t i o n i n d e c r e a s i n g o r d e r a r e : l - m e t h y l - 2 - m e r c a p t o i m i d a z o l e (MIA); t h i o u r a c i l ( T U ) ; p r o p y l t h i o u r a c i l (PTU) and ( ) - ) - 5 - v i n y l - 2 - t h i o - o x a z o l i d o n e ( g o i t r i n ) ; t h i o u r e a and m e t h y l t h i o u r a c i l (MTU). T h e i r a b i l i t y t o i n h i b i t - 8 -MIT f o r m a t i o n i n d e c r e a s i n g o r d e r a r e : MIA; PTU, MTU, TU and t h i o u r e a ; g o i t r i n . A l l o f them seem t o show s i m i l a r mechanisms o f a c t i o n e x c e p t g o i t r i n . T h i s i s i n agreement w i t h e v i d e n c e p r e s e n t e d i n a f o l l o w i n g s e c t i o n w i t h r e g a r d t o g o i t r o g e n i c ! ' t y o f r a p e s e e d meal. Many s t u d i e s have been f o c u s e d on t h e enzyme systems i n the t h y r o i d g l a n d , n o t a b l y on t h y r o i d a l p e r i d a s e ( S e r i f and Kirkwood, 1958) and i t s p o s s i b l e i n t e r a c t i o n w i t h a n t i t h y r o i d compounds. Though p e r -c h l o r a t e a p p e a r s t o have no e f f e c t on t h e enzyme, i t has been shown t h a t a z i d e , c y a n i d e , t h i o u r a c i l , t h i o c y a n a t e , p-aminobenzoate, and 3-amono-1 , 2 , 4 - t r i a z o l e a r e p o t e n t i n h i b i t o r s o f i o d i d e p e r o x i d a s e (Alemander, 1959), and t h a t some n a t u r a l l y o c c u r r i n g g o i t r o g e n s i n c l u d i n g g o i t r i n may a l s o p o s s e s s i n h i b i t o r y a c t i v i t y on t h e v a r i o u s t h y r o i d enzymes such as p e r o x i d a s e , cytochrome C o x i d a s e and DPNH-linked i o d i n a t i n g a c t i v i t y ( L a n g e r and M i c h a j l o v s k i j , 1972). I n s u f f i c i e n t p r o t e o l y s i s o f t h e t h y r o g l o b u l i n and s u p p r e s s i o n o f l y s o s o m a l enzyme a c t i v i t i e s have a l s o been r e p o r t e d as causes o f g o i t e r i n man ( B e c k e r s and D e V i s s c h e r , 1962) and m i c e ( I t k i a w a and Kawada, 1974). RAPESEED MEAL AS A SOURCE OE GOITROGEN The seed meals from rape ( B r a s s i c a napus L.) and t u r n i p r a p e (B. c a m p e s t r i s L . ) , a f t e r t h e o i l s have been e x p e l l e d o r s o l v e n t - e x t r a c t e d , a r e h i g h p r o t e i n - c o n t a i n i n g m a t e r i a l s and used i n animal f e e d s t u f f s (Bowland e t a l _ , 1965; C l a n d i n i n and R o b b l e e , 1966; C l a n d i n i n , 1967; B e l l and B e l z i l e , 1965; R utkowski, 1971). - 9 -P r a c t i c a l l y a l l p l a n t s o f the f a m i l y C r u c i f e r a e , t o which t h e genus B r a s s i c a b e l o n g s , which have been i n v e s t i g a t e d c o n t a i n g l u c o s i n o l a t e s ( V a n E t t e n 1969). In B r a s s i c a p l a n t s , t w e l v e d i f f e r e n t g l u c o s i n o l a t e s a r e known t o o c c u r and a r e d i s t r i b u t e d i n a l m o s t a l l p a r t s o f t h e p l a n t s ( J o s e f s s o n , 1967). M y r o s i n a s e i s p r e s e n t i n the t i s s u e s o f t h e s e g l u c o -s i n o l a t e - p r o d u c i n g p l a n t s as w e l l as i n g a s t r o i n t e s t i n a l t r a c t o f a n i m a l s ( G r e e r , 1962 a , b ) . I t i s p r o b a b l y an -SH dependent enzyme s i n c e i t i s i n a c t i v a t e d by i n h i b i t o r s o f t h a t c h e m i c a l r a d i c a l (Sandberg and H o l l y , 1932). G l u c o s i n o l a t e s a r e b i o l o g i c a l l y i n a c t i v e . D e s t r u c t i o n o f t h e p l a n t c e l l u l a r s t r u c t u r e a l l o w s t h e m y r o s i n a s e enzyme ( o r enzyme system) t o h y d r o l y z e t h e g l u c o s i n o l a t e s , l i b e r a t i n g g l u c o s e and b i s u l p h a t e , w h i l e a c t i v a t i n g t h e compounds i n t o one o f t h r e e g r o u p s : g o i t r i n and r e l a t e d n i t r i l e s ; o r g a n i c i s o t h i o c y a n a t e s ; i n o r g a n i c t h i o c y a n a t e ( E t t l i n g e r and Lundeen, 1957). The g o i t r o g e n i c p r o p e r t i e s o f r a p e s e e d meal a r e l a r g e l y a t t r i -b uted t o ( - ) - 5 - v i n y l - o x a z o l i d i n e t h i o n e ( g o i t r i n ) , a h y d r o l y s i s p r o d u c t o f i t s p r e c u r s o r s , p r o g o i t r i n and e p i - p r o g o i t r i n . I t i s p r o b a b l y n o t p r e f o r m e d i n the g l u c o s i n o l a t e m o l e c u l e but a r i s e s from c y c l i z a t i o n f o l l o w i n g myro-s i n a s e a c t i o n ( C l a n d i n i n e_t a l _ , 1959). The mechanism o f g o i t r i n i n f l u e n c e on t h e t h y r o i d g l a n d i s f a r from b e i n g u n d e r s t o o d . Rutkowski (1971) s t a t e d t h a t g o i t r i n b l o c k s t h e i r r e v e r s i b l e mechanism c o n n e c t e d w i t h o r g a n i c b i n d i n g o f i o d i n e i n t h e t h y r o i d , t h u s p a r t i a l l y s u p p r e s s i n g t h y r o i d hormone b i o s y n t h e s i s . T h i s , however, i s n o t borne o u t by r e p o r t s from o t h e r i n v e s t i g a t o r s (Matsumoto e t a l , 1969; A k i b a and Matsumoto, 1971). C l a n d i n i n e t i l (1961) d e s c r i b e d -10-a h i s t o p a t h o l o g i c a l p i c t u r e o f the r a p e s e e d m e a l - t r e a t e d c h i c k e n t h y r o i d g l a n d , which shows c o n s i d e r a b l e h y p e r t r o p h y and h y p e r p l a s i a o f p a r e n c h y -mal.\" e l e m e n t s and a l m o s t c o m p l e t e l o s s o f f o l l i c u l a r c o l l o i d . T h i s i s f u r t h e r s u p p o r t e d by t h e p h o t o m i o r o g r a p h i c f i n d i n g s o f Rutkowski ( 1 9 7 1 ) , i n which desquamation o f e p i t h e l i u m compressed f o l l i c l e s i n s m a l l r e s t r i c t e d a r e a s and d e c r e a s e i n c o l l o i d c o u l d be r e a d i l y o b s e r v e d i n t h y r o i d s o f c h i c k e n s a f t e r f o u r weeks o f r a p e s e e d meal f e e d i n g . The i s o t h i o c y a n a t e s and t h i o c y a n a t e s have a c o n s i d e r a b l y lower g o i t r o g e n i c e f f e c t than the o x a z o l i d i n e t h i o n e s (Fertman and C u r t i s , 1951; Gmelin and V i r t a n e n , 1960). They b l o c k i o d i n e a c c u m u l a t i o n i n t o t h e t h y -r o i d g l a n d , but t h e i r e f f e c t c o u l d be removed by a d d i t i o n o f i o d i n e t o t h e r a t i o n ( R u t k o w s k i , 1971). A s y n e r g e t i c i n f l u e n c e o f t h e o x a z o l i d i n e -t h i o n e s , i s o t h i o c y a n a t e s and t h i o c y a n a t e s may be p o s s i b l e ( t h e s o - c a l l e d \" B r a s s i c a f a c t o r \" ) , however, s i n c e i t has been shown ( L a n g e r , 1966) t h a t t h e e f f e c t o f the m i x t u r e o f g o i t r o g e n s was c o n s i d e r a b l y h i g h e r than t h e sum o f r e s u l t s o b t a i n e d i n a d m i n i s t r a t i n g them i n d i v i d u a l l y . EFFECTS OF GOITRIN ON THYROID METABOLISM OF THE CHICKEN The problems i n v o l v e d i n t h e f e e d i n g o f rapesteed meal t o l i v e -s t o c k have been e x t e n s i v e l y reviewed ( B e l l , 1955, 1957; Manns and Bowland, 1963; Manns e j t a l _ , 1963). T h y r o i d h y p e r t r o p h y has i n v a r i a b l y been r e p o r t e d i n d o m e s t i c a n i m a l s f e d r a p e s e e d meal ( T u r n e r , 1946;Bla k e l y and A n d e r s o n , 1948 a,b'; W i t z e t al, 1950; K l a i n e t a l _ , 1956) and has been a s c r i b e d t o t h e e f f e c t o f t h e known g o i t r o g e n i c compounds o f r a p e s e e d : ( > - ) - 5 - v i n y l - 2 - t h i o -o x a z o l i d o n e (Astwood e t a l _ , 1949; C a r r o l l , 1949) and a l l y l o r c r o t o n y l i s o t h i o c y a n a t e s (Dow and A l l e n , 1954). -11-( _ ) _ 5 - v i n y l - 2 - t h i o - o x a z o l i d o n e ( g o i t t r i m ) i s t h e most p o t e n t n a t u r a l g o i t r o g e n i c compound i s o l a t e d so f a r ( L a n g e r e t ^ a l _ , 1971). I t was f i r s t i d e n t i f i e d i n r a p e s e e d by Astwood ejt a l _ (1949 a,b) and C a r r o l l (1949) as L - 5 - v i n y l - 2 - t h i o o x a z o l i d o n e and l a t e r renamed ( - ) - 5 - 2 - o x a z o l i d i n e -t h i o n e . I t s s y n t h e t i c a n a l o g u e , , ( + ) - 5 - v i n y l - 2 - o x a z o l i d i n e t h i o n e , was f i r s t made a v a i l a b l e by E t t l i n g e r (1950). The work o f C l a n d i n i n (1962) and C l a n d i n i n e t _ a l _ (1966) i n the e f f e c t s o f s y n t h e t i c g o i t r i n i n t h y r o i d m e t a b o l i s m i n the growing c h i c k s u g g e s t s t h e f o l l o w i n g sequence o f e v e n t s : i n i t i a l l y , i o d i n e uptake i s g r e a t l y r e d u c e d and c o l l o i d s t o r e s i n t h e g l a n d s a r e r a p i d l y d e p l e t e d . C h i c k growth r a t e i s d e p r e s s e d . Rapid e n l a r g e m e n t o f the g l a n d s f o l l o w s w i t h accompanying h y p e r t r o p h y and h y p e r p l a s i a . I o d i n e u p t a k e i n c r e a s e s . C o l l o i d s t o r e s r e t u r n and t h e g l a n d t a k e s on a more normal a p p e a r a n c e . Hence i t would a p p e a r t h a t c h i c k s f e d t h e g o i t r o g e n e v e n t u a l l y r e a c h p h y s i o l o g i c a l e q u i l i b r i u m a t i n c r e a s e d t h y r o i d - t o - b o d y w e i g h t r a t i o s . When comparing t h e g o i t r o g e n i c e f f e c t s o f g o i t r i n , 1-methyl -2 - m e r c a p t o i m i d a z o l e ( m ethimazole) and 6 - m e t h y l - t h i o u r a c i l ( m e t h i o c i l ) i n g r o wing c h i c k s , Matsumoto e t al_ (1968) r e p o r t e d t h a t t h e c h i c k s w hich r e c e i v e d 0.05% g o i t r i n showed t h y r o i d e n l a r g e m e n t , the h i g h e s t t h y r o i d a l u ptake o f 1-131 and t h e slow r e l e a s e o f 1-131, which was i n marked c o n t r a s t t o the m e t h i m a z o l e - o r m e t h i o c i l - f e d c h i c k s . They c o n c l u d e d t h a t t h e i n c o r p o r a t i o n o f b l o o d 1-131 by the t h y r o i d g l a n d i s n o t i n h i b i t e d and t h a t , from t h e s l o w r e l e a s e o f 1-131 and the d e p r e s s e d t h y r o i d hormone i n b l o o d , some s t e p o f t h y r o i d hormone b i o s y n t h e s i s and f u r t h e r m o r e s e c r e t i o n o f t h y r o i d hormone i n t o b l o o d o r d e i o d i n a t i o n o f some iodoamino a c i d s i n t h y r o i d g l a n d may be i n h i b i t e d by t h e t r e a t m e n t w i t h g o i t r i n . -12-T h a t the g o i t r o g e n i c e f f e c t s o f g o i t r i n a r e d i f f e r e n t from t h o s e o f p r o p y l t h i o u r a c i l (PTU) i n the growing c h i c k was f u r t h e r s t u d i e d by Matsumoto e t al_ (1969) and A k l b a and Matsumoto (1971). T h e s e i n v e s t i -g a t o r s showed t h a t w h i l e t h e PTU c h i c k s do n o t m a i n t a i n i n c o r p o r a t e d 1-131 i n t h e t h y r o i d g l a n d b u t r e l e a s e i t i n t o t h e b l o o d e a s i l y , the g o i t r i n - f e d c h i c k ; i s promoted t o m a i n t a i n a l a r g e amount o f i n c o r p o r a t e d 1-131 i n t h e g l a n d and a p p e a r s t o i n h i b i t i t s r e l e a s e . F u r t h e r m o r e , s i n c e the p e r c e n t a g e o f i o d o t h y r o n i n . e s (Tg and T^) i n t h i s l a t t e r group a l s o d e c r e a s e d t o o n e - t h i r d o f the c o n t r o l c h i c k s i n t h e i r d i s t r i b u t i o n o f r a d i o a c t i v e i o d i n a t e d s u b s t a n c e s found i n t h y r o i d homogenates, and t h a t t h e h i g h e s t p e r c e n t a g e was o b s e r v e d i n MIT (65%) and t h e MIT/DIT r a t i o was h i g h e r than t h a t o f the c o n t r o l group, i t i s q u i t e p o s s i b l e t h a t g o i t r i n a l m o s t n e v e r i n h i b i t s t h e m o n o i o d i n a t i o n o f t y r o s i n e , but i t i n h i b i t s the c o u p l i n g o f MIT t o s y n t h e s i z e DIT. The f i n d i n g o f i n c r e a s e d MIT p e r -c e n t a g e i s i n agreement w i t h r e s u l t s o f l i n o e t a l _ (1961) and G r e e r e t a l (1962) i n r a t s g i v e n s m a l l d oses o f PTU. R e p o r t s on t h e e f f e c t o f PTU o r t h i o u r a c i l on t h e i n t r a t h y r o i d a l m e t a b o l i s m o f 1-131 a r e numerous. F o r example, t h i o u r a c i l i n h i b i t s t h e o r g a n i c b i n d i n g o f i o d i n e i n r a t t h y r o i d i n v i v o (Astwood and B i s s e l , 1944) and i n v i t r o , ( F r a n k l i n e t a l _ , 1944), PTU b l o c k s the i o d i n a t i o n o f t y r o s i n e ( P i t t - R i v e r s , 1948; Mayberry and Astwood, 1960) and i n h i b i t s t h e s y n t h e s i s and c o u p l i n g o f DIT ( R i c h a r d and I n g b a r , 1959). Thus i t a p p e a r s t h a t PTU i n h i b i t s t h e s e s t e p s o f t h y r o i d hormone b i o s y n t h e s i s : o x i d a t i o n o f i o d i d e , i o d i n a t i o n o f t y r o s i n e , and i o d o t h y r o n i n e s y n t h e s i s from i o d o t y r o s i n e s . The most d i s t i n c t d i f f e r e n c e between e f f e c t s o f g o i t r i n and PTU i s t h a t the amount o f i o d o t h y r o n i n e s s y n t h e s i z e d i n the g l a n d i s markedly d e p r e s s e d -13-i n PTU-fed c h i c k s but i s i n c r e a s e d i n g o i t r i n - f e d c h i c k s w h i l e the plasma l e v e l o f t h y r o i d hormone i s d e p r e s s e d i n both groups ( A k i b a and Matsumoto, 1971). I t i s s u g g e s t e d t h a t t h e h y d r o l y s i s o f t h y r o g l o b u l i n and s e c r e t i o n o f t h y r o i d hormone from t h e g l a n d may be d e c r e a s e d o r a t l e a s t n o t enhanced by g o i t r i n t r e a t m e n t i n growing c h i c k s . M o r e o v e r , i n t h e i r s t u d y o f t h y r o i d f u n c t i o n o f c h i c k s a f t e r w i t h d r a w a l o f g o i t r i n from the d i e t f o l l o w i n g twenty-one days o f f e e d i n g , A k i b a and Matsumato (1973) r e p o r t e d a r a p i d r e s t o r a t i o n o f normal t h y r o i d m e t a b o l i s m and hormone b i o s y n t h e s i s , d e s p i t e h y p e r t r o p h y o f t h y r o i d g l a n d which p e r s i s t s f o r i r e l a t i v e l y l o n g e r t i m e . A f t e r w i t h d r a w a l o f g o i t r i n , plasma PBI-131 was r a d i c a l l y e l e v a t e d and a l i t t l e i n c r e a s e c o u l d be d e m o n s t r a t e d . T h i s s o - c a l l e d rebound phenomenon i s s i m i l a r t o g o i t r o g e n w i t h d r a w a l i n o t h e r l a b o r a t o r y a n i m a l s such as m i c e , r a t s and g u i n e a p i g s (D'Angelo e t a l _ , 1951, 1954; L i p n e r e t a l _ , 1959; S t u d e r and G r e e r , 1967; L a n g e r, 1968) and can be e x p l a i n e d by an o v e r s t i m u l a t i o n o f TSH on t h e u n b l o c k e d t h y r o i d g l a n d a f t e r r e l e a s e o f g o i t r o g e n i c e f f e c t s by g o i t r o g e n w i t h d r a w a l (D'Angelo, 1961, 1969). The g r a d u a l i n v o l u t i o n i n t h y r o i d w e i g h t a f t e r g o i t r i n w i t h d r a w a l i s i n agreement w i t h t h a t r e p o r t e d f o r PTU w i t h d r a w a l i n c h i c k s ( A k i b a e t al, 1971), r a t s (Shimoda, 1960) and g u i n e a p i g s (D'Angelo e t al, 1954). EFFECTS OF LONG-TERM ADMINISTRATION OF RAPESEED MEAL ON BIRDS AND PROGENY A l t h o u g h a c o n s i d e r a b l e amount o f work has been c a r r i e d o u t on the f e e d i n g o f r a p e s e e d meal t o young a n i m a l s , ' r e l a t i v e l y few e x p e r i m e n t s have been performed w h e r e i n the g o i t r o g e n s a r e f e d c h r o n i c a l l y ( P e l t o l a , -14-1965; Summers e t a l _ , 1969; Vogt e t al_, 1969; C l a n d i n i n and Robblee, 1970; March e t a l _ , 1972). In an e x p e r i m e n t where r a p e s e e d meal was used as a p r o t e i n supplement i n the d i e t s o f l a y i n g hens f o r 252 days ( J a c k s o n , 1969), the h i s t o l o g i c a l p i c t u r e o f t h e t h y r o i d g l a n d s was n o t i d e n t i c a l w i t h t h a t o b t a i n e d i n growing c h i c k s ( C l a n d i n i n and B a y l y , 1960). No e v i d e n c e o f c e l l u l a r i n f i l t r a t i o n was o b t a i n e d , thus s u p p o r t i n g t h e view t h a t t h e i n i t i a l e f f e c t o f the r a p e s e e d g o i t r o g e n s a r e compensated f o r by t h y r o i d e n l a r g e m e n t , and the c h r o n i c a l l y - t r e a t e d b i r d s have normal p h y s i o l o g i c a l f u n c t i o n w i t h r e g a r d t o t h e e g g - l a y i n g p r o c e s s . Some i n v e s t i g a t o r s r e p o r t e d t h e p r e s e n c e o f s o - c a l l e d Psammoma b o d i e s i n t h e t h y r o i d s o f c h r o n i c a l l y s t i m u l a t e d a n i m a l s ( A x e l r a d and L e b l o n d , 1955; Wollman, 1961; Grimm e _ t a l _ , 1970), which a r e p r o b a b l y dense a g g r e g a t i o n s o f t h y r o p r o t e i n r e l a t e d t o t h e o b s e r v a t i o n o f r a d i o a c t i v i t y w i t h a v e r y l o n g b i o l o g i c a l h a l f - l i f e i n t h y r o i d s d e s c r i b e d by Van M i d d l e s -worth (1965). F o l l i s ( 1 9 6 5 ) , however, c o u l d n o t f i n d t h e s e Psammoma b o d i e s i n t h y r o i d s o f hamster, mice o r monkeys g i v e n g o i t r o g e n s f o r p r o l o n g e d p e r i o d s . I t a p p e a r s t h a t g o i t r i n i s n o t t r a n s f e r r e d f o r d e p o s i t i o n i n t h e egg ( R u t k o w s k i , 1971; March et^ a l _ , 1972). The t h y r o i d g l a n d s o f new l y - h a t c h e d progeny of.dams f e d rapes'eed m e a l , however, showed h y p e r t r o p h y , which was g r e a t e r when the d i e t c o n t a i n e d t h e h i g h e r l e v e l o f r a p e s e e d meal (March e t ^ a l . , 1972). A l s o t h e s e c h i c k s a t h a t c h i n g were s l i g h t l y s m a l l e r t h a n t h o s e from b i r d s f e d soybean meal c o n t r o l d i e t . T h i s i s i n agreement w i t h t h e a d v e r s e e f f e c t s o f r a p e s e e d meal i n progeny o f o t h e r s p e c i e s such as g i l t s ( B e l l and B e l z i l e , 1965). -15-E x p e r i m e n t 1: EFFECTS OF CHRONIC AND EARLY STIMULATION BY GOITROGENIC RSM ON WEIGHT AND HISTOLOGY OF THYROID GLAND I t has l o n g been known t h a t the f e e d i n g o f RSM t o the fowl c a u s e s t h y r o i d h y p e r t r o p h y and h y p e r p l a s i a . However, s i n c e most o f t h e h i s t o l o g i c a l d a t a a r e on growing c h i c k s , l i t t l e i s known about t h e a l t e r a t i o n o f c h r o n i c RSM s t i m u l a t i o n on t h y r o i d h i s t o l o g y . F u r t h e r m o r e , t h e e x t e n t t o which a d a p t a t i o n o f t h e t h y r o i d g l a n d i s p o s s i b l e v i a t h e mechanism o f compen-s a t i o n has not been e s t a b l i s h e d . When compensatory f u n c t i o n s o f the h y potha-l a m u s - p i t u i t a r y - t h y r o i d a x i s i n the fowl c e a s e and a t r o p h y o f the g l a n d b e g i n s have n o t been a c c u r a t e l y a s s e s s e d . I t i s n o t c l e a r i f i n v o l u t i o n does o c c u r i n t h e e a r l y - s t i m u l a t e d t h y r o d d g l a n d s when g o i t r o g e n i c s t i m u l a t i o n i s a l l e v i a t e d . Hence t h e f o l l o w i n g l o n g - t e r m e x p e r i m e n t was c o n d u c t e d t o e l u c i d a t e t h e s e e f f e c t s on t h y r o i d w e i g h t and h i s t o l o g y . M a t e r i a l s and Methods: In t h i s and a l l t h e f o l l o w i n g e x p e r i m e n t s ( w i t h t h e e x c e p t i o n s o f e x p e r i m e n t s 5 and 6B), the b i r d s used were on an e x p e r i m e n t d e s i g n e d t o compare p r o d u c t i o n and m o r t a l i t y o f b i r d s f e d soybean meal (SBM) and r a p e s e e d meal (RSM) as the s o u r c e o f s u p p l e m e n t a r y p r o t e i n i n g r owing and/or l a y i n g d i e t s (March e t a l , 1975). D a y - o l d White Leghorn c h i c k s o f two s t r a i n s ( Shaver c h i c k s and DeKalb c h i c k s ) were r e a r e d i n b a t t e r i e s and f e d s t a r t i n g and growing d i e t s c o n t a i n i n g e i t h e r SBM o r RSM as i n d i c a t e d i n T a b l e 1. The s t a r t i n g d i e t s were f e d from 0-8 weeks and t h e growing d i e t s from 8-23 weeks. The b i r d s were v a c c i n a t e d a g a i n s t Marek's d i s e a s e , N e w c a s t l e d i s e a s e and i n f e c t i o u s b r o n c h i t i s . Causes o f m o r t a l i t y were r e c o r d e d . -16-A t 23 weeks o f age t h e b i r d s were t r a n s f e r r e d t o community cages and f e d the l a y i n g d i e t s ( T a b l e 1) c o n t a i n i n g e i t h e r SBM o r RSM. The l a y i n g d i e t s were f e d a c c o r d i n g t o the f o l l o w i n g c r o s s - o v e r d e s i g n . One h a l f t h e b i r d s were c o n t i n u e d on t h e p r o t e i n supplement which t h e y had r e c e i v e d d u r i n g t h e growing p e r i o d and the o t h e r h a l f were t r a n s f e r r e d t o the a l t e r n a t e p r o t e i n supplement. The t r e a t m e n t s were d e s i g n a t e d as S-S, R-S, R-R and S-R, t h e f i r s t l e t t e r o f each p a i r i n d i c a t i n g soybean meal o r r a p e s e e d meal i n t h e growing d i e t and t h e s e c o n d l e t t e r i n d i c a t i n g soybean meal o r r a p e s e e d meal i n t h e l a y i n g d i e t . S i x hundred b i r d s were d i s t r i b u t e d i n t o 24 l o t s o f 25 b i r d s e a c h , i n community cages (2 s t r a i n s x 4 t r e a t m e n t s x 3 r e p l i c a t e s ) . A t 91 weeks, f o r t y b i r d s o f t h e Shaver s t r a i n (10 from each t r e a t m e n t ) were weighed and k i l l e d . The t h y r o i d g l a n d s were d i s s e c t e d f r e e from c o n n e c t i v e t i s s u e and f a t , weighed, and i m m e d i a t e l y f i x e d i n b u f f e r e d 10% f o r m a l i n f o r s u b s e q u e n t h i s t o l o g i c a l p r e p a r a t i o n ( H i s t o l o g y L a b o r a t o r y , Z o o l o g y Department, U.B.C.). The g l a n d s were embedded i n p a r a f f i n , s e c t i o n e d a t a t h i c k n e s s o f 5 j u , and e v e r y 20th s e c t i o n o f t h e l e f t g l a n d was s t a i n e d w i t h h a e m a t o x y l i n and e o s i n . The c e n t r a l s e c t i o n s o f each g l a n d were chosen f o r measurements o f e p i t h e l i a l h e i g h t and volume p r o p o r t i o n o f t h y r o i d a l t i s s u e s u s i n g a m i c r o m e t e r f i t t e d t o the l i g h t m i c r o s c o p e . A t 99 weeks o f age, f o r t y b i r d s o f t h e DeKalb s t r a i n were i n -d i v i d u a l l y weighed and k i l l e d , and t h e i r t h y r o i d g l a n d s were removed f o r measurement o f w e i g h t s o n l y . A l l e x p e r i m e n t a l d a t a o f t h i s and the f o l l o w i n g e x p e r i m e n t s were s u b j e c t e d t o a n a l y s i s o f v a r i a n c e and Duncan's m u l t i p l e range t e s t s ( S n e d e c o r and C o c h r a n , 1956). -17-G o i t r o g e n i c i t y o f the RSM f e d i n the e x p e r i m e n t a t the time t h e b i r d s were k i l l e d was d e t e r m i n e d by a s e p a r a t e e x p e r i m e n t , i n which d a y - o l d White Leghorn c o c k e r e l s were f e d e i t h e r a SBM- o r RSM-supplemented d i e t ( T a b l e 1) s i m i l a r i n c o m p o s i t i o n t o t h o s e f e d i n e x p e r i m e n t 1. S i x c h i c k s from each t r e a t m e n t a t 1 week and t w e l v e c h i c k s from each t r e a t m e n t a t 3 weeks were k i l l e d t o d e t e r m i n e t h e w e i g h t o f t h e t h y r o i d g l a n d s . R e s u l t s : G o i t r o g e n i c i t y o f t h e p r e s e n t l o t o f RSM (Span) was e s t a b l i s h e d by the h y p e r t r o p h y o f t h e t h y r o i d g l a n d s shown b y : t h e RSM-fed c h i c k s a t 1 week o r 3 weeks o f t r e a t m e n t ( T a b l e 2 ) . The a v e r a g e t h y r o i d w e i g h t s (mg/100 g body w e i g h t ) o f the S h a v e r and DeKalb b i r d s a t 91 and 99 weeks o f age a r e p r e s e n t e d i n T a b l e 3 S -The R-R b i r d s showed the most t h y r o i d a l e n l a r g e m e n t o v e r t h a t o f t h e c o n t r o l (10 t o 13 t i m e s i n c r e a s e ) , w h i l e t h e R-S b i r d s a l s o had d o u b l e t h e t h y r o i d s i z e o f t h e S-S b i r d s . H i s t o l o g i c a l d a t a o f t h e t h y r o i d g l a n d s o f t h e Shaver b i r d s a t 91 weeks a r e p r e s e n t e d i n T a b l e 4. T o t a l p e r c e n t a g e t h y r o i d e p i t h e l i u m i n the R-R b i r d s was l e s s than t h a t i n the c o n t r o l (15.1% vs 2 4 . 4 % ) , w h i l e t h e a v e r a g e e p i t h e l i a l h e i g h t o f the R-R t h y r o i d s (10.65 ja) was h i g h e r than t h a t (7.13>i) f o r t h e c o n t r o l . S i m i l a r l y , t h e R-S t h y r o i d s showed a d e c r e a s e i n p e r c e n t a g e e p i t h e l i u m (14.8%) but t h e a v e r a g e e p i t h e l i a l h e i g h t (5.61 ja) was a l s o l o wer than t h a t o f t h e c o n t r o l . -18-C o n s i s t e n t w i t h t he appearance o f e n l a r g e d g l a n d s , t he a v e r a g e f o l l i c l e d i a m e t e r s o f t h e RSM-treated t h y r o i d g l a n d s were a l s o i n c r e a s e d o v e r t h a t o f the c o n t r o l , t h e l a r g e s t f o l l i c l e s b e i n g i n t h e R-R t h y r o i d s . A g a i n , the a v e r a g e f o l l i c l e d i a m e t e r o f t h e R-S t h y r o i d s was a l s o g r e a t e r than t h a t o f t h e c o n t r o l (0.84 mm vs 0.54 mm). I t s h o u l d be noted t h a t , w i t h a l l t h e h i s t o l o g i c a l p a r a m e t e r s measured, t h e S-R t h y r o i d s were c o n s i s t e n t l y i n t e r m e d i a t e between t h e R-R and t h e c o n t r o l g l a n d s . Under l i g h t m i c r o s c o p y , t h e f o l l i c l e ilumerisof t h e t h y r o i d s i n the f o u r t r e a t m e n t s were f i l l e d w i t h s t a i n e d c o l l o i d . The R-R and S-R g l a n d s showed the appearance o f c o l l o i d g o i t e r . Desquamated c e l l s were i n f r e q u e n t but d i d o c c u r i n the t h y r o i d g l a n d s o f t h e c h r o n i c a l l y RSM-fed b i r d s . No e v i d e n c e o f a t r o p h y was seen i n t h e s e g l a n d s . O n l y i n one o r two o c c a s i o n s had t h y r o i d c y s t s been found i n the c h r o n i c a l l y R SM-stimulated b i r d s . -19-T a b l e 1: P e r c e n t a g e c o m p o s i t i o n o f d i e t s f e d i n e x p e r i m e n t 1 and RSM g o i t r o g e n i c i t y t e s t E x p e r i m e n t 1 RSM-Goitro-g e n i c i t y S t a r t e r - G r o w e r L a y e r T e s t Code L e t t e r S R S R S R I n g r e d i e n t Soybean meal 11.0 - 13.1 - 26.3 7.8 Rapeseed meal - 17.5 - 19.0 - 30.0 Ground wheat 66.5 60.0 60.6 53.5 53.6 42.6 Ground c o r n 15.0 15.0 6.0 6.0 10.0 10.0 Ground o a t s - - 10.0 10.0 - -Dehydrated c e r e a l g r a s s 2.0 2.0 2.0 2.0 2.0 2.0 Bone meal 2.0 2.0 2.5 2.5 2.0 2.0 Limestone 1.0 1.0 4.75 4.75 1.3 1.9 I o d i z e d s a l t 0.5 0.5 0.25 0.25 0.5 0.5 F e e d i n g t a l l o w 2.0 2.0 0.5 2.0 3.0 3.0 DL-Methionine - - 0.1 - 0.1 -L - L y s i n e - - - - - 0.1 T r i c a l c i u m phosphate - - - - 1.2 0.1 M i c r o n u t r i e n t s * * ** ** * * */kg: manganese s u l p h a t e 132 mg, c h o l i n e c h l o r i d e 1320 mg, n i a c i n 27 mg, r i b o f l a v i n 4.0 mg, c a l c i u m p a n t o t h e n a t e 9.2 mg, f o l a c i n 0.55 mg, p y r i d o x i n e HC1 2.9 mg, b i o t i n 0.09 mg, v i t a m i n B . 2 13.0 meg, v i t a m i n A 4400 I.U., v i t a m i n D 3 660 I.U., Zn b a c i t r a c i n 9.7 mg, o l e a n d o -mycin 11.0 mg. * * / k g : manganese s u l p h a t e 198 mg, r i b o f l a v i n 3.3 mg, v i t a m i n B , 2 3.3 meg, v i t a m i n A 6600 I . I L , v i t a m i n D 0 660 I.II. -20-T a b l e 2: Comparison o f a v e r a g e t h y r o i d w e i g h t s o f White Leghorn c o c k e r e l s f e d SBM- o r RSM-supplemented d i e t s a t 1 week o r 3 weeks o f t r e a t m e n t f r o m h a t c h i n g T r e a t m e n t s Average t h y r o i d w e i g h t s (mg/100 g.3tiipdy?wgtght) 1 week 3 weeks SBM - d i e t 5.8 ( 6 ) * 5.1 (12) RSM - d i e t 9.4 (6) 9.9 (12) * No. o f d e t e r m i n a t i o n s -21-T a b l e 3: Average w e i g h t s o f t h y r o i d g l a n d s i n b i r d s f e d RSM o r SBM f o r d i f f e r e n t p e r i o d s o f time i n e x p e r i m e n t 1 Ave. t h y r o i d wt. D i e t a r y a t 58 weeks o f age* t r e a t . mg/100 g body wt. EXPERIMENT 1 S t r a i n A Ave. t h y r o i d wt. a t 91 weeks o f age mg/100 g body wt. S t r a i n B Ave. t h y r o i d wt. a t 99 weeks o f age mg/100 g body wt. S-S R-S R-R S-R 1 1 . 0 a A ( 1 2 ) * * 27.1° (12) 67.8° (12) 4 4 . 9 C (12) 1 0 . 7 a A ( 1 0 ) 2 3 . 5 a B ( 1 0 ) 112.5° (10) 84.9° (10) 1 1 . 2 a A ( 1 0 ) 2 8 . 4 a B ( 1 0 ) 149.4 C (10) 103.4° (10) T h i o u r a c i l i n s t a r t i n g d i e t u n t i l a K D / i o \\ 4 wks o f a g e : 1 7 . 7 a D B U ^ L e v e l o f s i g n i f i c a n c e 5% 1% 1% * u n p u b l i s h e d d a t a from an e a r l i e r e x p e r i m e n t i n t h i s l a b o r a t o r y ** number o f d e t e r m i n a t i o n s p er t r e a t m e n t a D c d V a l u e s w i t h t h e same s u p e r s c r i p t a r e n o t s i g n i f i c a n t l y d i f f e r e n t . AB S i g n i f i c a n t l y d i f f e r e n t a t t h e 1% l e v e l o f s i g n i f i c a n c e by s e p a r a t e c o m p a r i s o n . -22-T a b l e 4: H i s t o l o g i c a l measurements on t h y r o i d glands, from b i r d s ( S h a v e r s t r a i n ) i n e x p e r i m e n t 1 a t 91 weeks o f age. Ave.* Ave. Ave. % Wt. Ave. no. D i e t a r y t h y r o i d e p i t h e l i a l f o l l i c l e e p i t h e l i u m e p i t h e l i u m f o l l i c l e s t r e a t m e n t wt. g, h t . / i diam. mm mg a c r o s s gland S-S 0 . 1 9 a A 7.13 a 0.108 a 24.4 C 4 1 . l a 67 R-S 0 . 5 2 a B 5.72 a 0.167° 14.8 a 7 6 . 5 a 51 R-R 2.30 c 10.65° 0.227 c 1 5 . 1 a b 334.6° 58 S-R 1.39° 10.11° 0 . 1 9 6 ° C 2 0 . 1 a b c 277.4° 67 L e v e l o f s i g n i f i c a n c e 1% 1% 5% 5% 1% * f i v e g l a n d s examined per t r e a t m e n t a D c d v a l u e s w i t h t h e same s u p e r s c r i p t a r e n o t s i g n i f i c a n t l y d i f f e r e n t . AB s i g n i f i c a n t l y d i f f e r e n t a t the 1% l e v e l o f s i g n i f i c a n c e by s e p a r a t e comparison, -23-E x p e r i m e n t 2: EFFECTS OF CHRONIC AND EARLY STIMULATION BY GOITROGENIC RSM ON TOTAL IODINE CONTENT OF THE THYROID GLAND S i n c e the h i s t o l o g i c a l p i c t u r e o f t h e R-R and S-R t h y r o i d s showed t h e h e a l t h y a p p e a r a n c e o f c o l l o i d g o i t e r i n s t e a d o f some degree o f a t r o p h y due t o c h r o n i c RSM s t i m u l a t i o n , t h e f o l l o w i n g e x p e r i m e n t was c o n d u c t e d t o d e t e r m i n e t h e t o t a l i o d i d e c o n t e n t o f t h e t h y r o i d t i s s u e as a f f e c t e d by the t r e a t m e n t s . M a t e r i a l s and Methods: F i v e Shaver b i r d s o f each o f the f o u r d i e t a r y t r e a t m e n t s (S-S, R-S, R-R and S-R as i n e x p e r i m e n t 1) were k i l l e d and t h e i r t h y r o i d g l a n d s removed and weighed. Each p a i r o f g l a n d s was homogenized a P o t t e r - E l v e h j e m g l a s s - h o m o g e n i z e r t o g i v e a 25 ml s u s p e n s i o n w i t h d e i o n i z e d w a t e r . The a n a l y t i c a l method f o r t h e measurement o f t o t a l i o d i n e c o n t e n t was an a d a p t a t i o n o f the B a r k e r p r o c e d u r e f o r serum ( B a r k e r , 1948). The c o n c e n t r a t i o n o f i o d i n e was d e t e r m i n e d by m e a s u r i n g i t s c a t a l y t i c e f f e c t on t h e r a t e o f r e a c t i o n between e e r i e i o n and a r s e n i o u s a c i d , which i n v o l v e d t h e c o n v e r s i o n o f t h e y e l l o w e e r i e i o n t o the c o l o r l e s s c e r o u s i o n . The c o l o r o f t h e e e r i e i o n was d e t e r m i n e d p h o t o m e t r i c a l l y twenty m i n u t e s a f t e r t h e r e a c t i o n s t a r t e d , and t h e i o d i n e c o n e n t r a t i o n was r e l a t e d i n v e r s e l y t o the l o g a r i t h m o f t h e o p t i c a l d e n s i t y . 1 ml o f t h e t h y r o i d t i s s u e homogenate was weighed i n t o Kimax 16 x 125 mm c u l t u r e t u b e s . 1 ml o f 4N sodium c a r b o n a t e was added t o each tube and t h e c o n t e n t was mixed t h o r o u g h l y by a s h a k e r . The t u b e s were d r i e d i n n -24-an oven a t 100°C t o complete d r y n e s s , a f t e r which t h e y were ashed i n a m u f f l e f u r n a c e a t 625 +_ 25°C f o r 4 h o u r s . The tubes were then removed and a l l o w e d t o c o o l . Four ml o f mixed a c i d s o l u t i o n (7N h^SC^ + 2N HC1; 1:1) were added t o each t u b e . The c o n t e n t s were mixed and a l l o w e d t o s t a n d f o r a p p r o -x i m a t e l y 10 m i n u t e s t o d i s s o l v e t h e a s h . T h r e e ml o f d e i o n i z e d water were added, t h e c o n t e n t s o f the tube a g a i n mixed. 10>ul o f t h i s m i x t u r e was added t o 8 ml o f d e i o n i z e d water i n a n o t h e r s e t o f 12 ml g l a s s - s t o p p e r e d t u b e s . A b l a n d and t h r e e i o d i n e s t a n d a r d s (.01, .05 and .10 jug) were a n a l y z e d i n d u p l i c a t e w i t h each g r o u p o f samples. A l l t h e t u b e s ( s t a n d a r d , b l a n k and sample) c o n t a i n e d 8 ml o f s o l u t i o n . To each g l a s s - s t o p p e r e d t u b e ( b l a n k s , s t a n d a r d s and samples) was added 0.5 ml o f 0.1N sodium a r s e n i t e s o l u t i o n . The tubes were p l a c e d i n a s h a k i n g water b a t h a t 39 +_ 1°C f o r 10 m i n u t e s t o a l l o w f o r t h e r m a l e q u i l i b r i u m . C e r i c ammonium s u l f a t e s o l u t i o n (.02N) was a l s o warmed i n the same water b a t h . U s i n g a C e n t a u r m i c r o l i t e r p i p e t t e , 1 ml o f c e r i c ammonium s u l f a t e s o l u t i o n was added t o each g l a s s - s t o p p e r e d tube a t p r e c i s e l y 1 minute i n t e r v a l s . A s t o p watch was used t o e n s u r e a c c u r a t e t i m i n g . The c o n t e n t o f t h e tube was i m m e d i a t e l y shaken t o e n s u r e t h o r o u g h m i x i n g . Twenty mi n u t e s a f t e r t h e c e r i c ammonium s u l f a t e s o l u t i o n was added t o t h e f i r s t t u b e , measurements o f o p t i c a l d e n s i t y a t 420 nm were begun, u s i n g an Unicam SP1800 s p e c t r o p h o t o m e t e r . These r e a d i n g s were made a t i n t e r -v a l s o f 1 minute i n o r d e r t h a t the l a p s e d t i m e f o r a l l o f t h e samples were c o n s t a n t . Measurements o f the o p t i c a l d e n s i t y o f t h e b l a n k and t h e s t a n d a r d i o d i n e s o l u t i o n s were p l o t t e d on s e m i l o g paper. The c o n c e n t r a t i o n s o f i o d i n e i n t he unknown samples were r e a d from t h e graph and m u l t i p l i e d by t h e d i l u t i o n -25-f a c t o r o f 700 ( t o compensate f o r the 10 u l a l i q u o t t a ken from t h e 7 ml a c i d s o l u t i o n o f t h e ashed s a m p l e ) . T o t a l i o d i n e c o n t e n t i n the p a i r o f t h y r o i d g l a n d s = i o d i n e c o n c e n t r a t i o n i n t h e sample x 25. The i o d i n e c o n c e n t r a t i o n s i n the d i e t s were a l s o a n a l y z e d by t h e same p r o c e d u r e , as m o d i f i e d by Sunderman (1963). 0.1 t o 0.2 g o f t h e sample was weighed i n t o c u l t u r e t u b e s . A f t e r a d d i t i o n o f 1 ml o f 4N sodium c a r -bonate and subsequent d r y i n g a t 100 C, the samples were ashed a t 625 + 25 C f o r 12-16 h o u r s . Upon c o o l i n g and m i x i n g w i t h 4 ml o f mixed a c i d s o l u t i o n ,; and 3 ml o f d e i o n i z e d w a t e r , the c o n t e n t s o f the tubes were passed t h r o u g h a Pyrex h i g h form f r i t t e d g l a s s f i l t e r o f F ( f i n e ) p o r o s i t y t o remove any i n s o l u b l e m a t e r i a l . A 3 ml a l i q u o t o f the f i l t r a t e from each ashed sample was p i p e t t e d i n t o a n o t h e r s e t o f g l a s s - s t o p p e r e d tubes t o which 5 ml o f d e i o n i z e d were added t o make up a t o t a l o f 8 ml i n each t u b e . O p t i c a l den-s i t i e s o f t h e unknown samples were r e a d o f f from a s t a n d a r d graph and mul-t i p l i e d by a d i l u t i o n f a c t o r o f 7/3 t o compensate f o r t h e 3 ml a l i q u o t t a k e n from t h e 7 ml a c i d s o l u t i o n o f t h e ashed sample. R e s u l t s : The a v e r a g e a b s o l u t e g l a n d w e i g h t s , i o d i n e c o n c e n t r a t i o n and t o t a l i o d i n e c o n t e n t o f t h e t h y r o i d s o f t h e b i r d s a f t e r c h r o n i c d i e t a r y t r e a t m e n t s a r e g i v e n i n T a b l e 5. The a b s o l u t e t h y r o i d w e i g h t s were i n the same o r d e r as the r e l a t i v e w e i g h t s ; the l a r g e s t a g a i n b e i n g t h e R-R g l a n d s (2.08 g) which were 10 t i m e s the s i z e o f t h e c o n t r o l . -26-The i o d i n e c o n c e n t r a t i o n (mg p e r g t h y r o i d t i s s u e ) was l o w e s t i n R-R b i r d s (0.89) compared t o 2.65 mg/g o f t h e c o n t r o l . However, because o f t h e much e n l a r g e d s i z e , the R-R t h y r o i d s had an a v e r a g e t o t a l i o d i n e c o n t e n t o f 1.72 mg i n the p a i r o f g l a n d s , w h i l e t h a t i n t h e c o n t r o l was o n l y 0.55 mg. L i k e w i s e , t h e R-S t h y r o i d s had an a c c u m u l a t i o n o f 1.25 mg i o d i n e i n a p a i r o f g l a n d s , which was more than d o u b l e t h a t o f the c o n t r o l . A g a i n , t h e i o d i n e c o n c e n t r a t i o n o r t o t a l i o d i n e c o n t e n t o f t h e S-R t h y r o i d s were i n t e r m e d i a t e between t h o s e o f t h e R-R and S-S g l a n d s . The i o d i n e c o n c e n t r a t i o n s o f t h e i . r e s p e c t i v e / i d i e t s a r e shown i n T a b l e 6. Both SBM- and RSM-supplemented r a t i o n s had i o d i n e v a l u e s above t h a t o f 0.3 mg/kg, which i s t h e NRC s t a t e d r e q u i r e m e n t f o r i o d i n e by p o u l t r y . -27-T a b l e 5: I o d i n e c o n c e n t r a t i o n i n t h y r o i d g l a n d s o f b i r d s f e d RSM o r SBM f o r d i f f e r e n t p e r i o d s o f timei; Ave. i o d i n e c o n c e n t r a t i o n D i e t a r y Ave. wt. o f i n g l a n d t r e a t m e n t t h y r o i d g l a n d s * g mg/g S-S 0.20 a 2.65 a 0.55 a R-S 0.47 a 2.73 a 1.25 b R-R 2.08 c 0.89 b 1.72 c S-R 1.51 b 1.08 b 1.65 c L e v e l o f s i g n i f i c a n c e 5% 1% 5% * 5 d e t e r m i n a t i o n s per t r e a t m e n t . abed v a l u e s w i t h the same s u p e r s c r i p t a r e n o t s i g n i f i c a n t l y d i f f e r e n t . T o t a l i o d i n e i n g l a n d mg \\ -28-T a b l e 6: A n a l y z e d i o d i n e c o n c e n t r a t i o n s i n SBM- and RSM-supplemented d i e t s f e d t o b i r d s i n e x p e r i m e n t s 1 and 6B. A n a l y z e d i o d i n e c o n c e n t r a t i o n s R a t i o n s mg/kg In e x p e r i m e n t s 1 and 6B: SBM-supplemented 0.85 RSM-supplemented 0.91 In e x p e r i m e n t 6B o n l y : SBM-supplemented + K I 0 3 1.10 RSB-supplemented + KIO., 1.35 -29-E x p e r i m e n t 3: EFFECTS ON UPTAKE OF A SINGLE DOSE OF 1-131 BY THE THYROID GLAND 6 HOURS AND 22 HOURS AFTER ADMINISTRATION OF THE RADIOIODINE R a d i o i d d i n e has l o n g been used i n a s s e s s i n g t h y r o i d i o d i n e me-t a b o l i s m ( C h a i k o f f and T a u r o g , 1948; T u r n e r e t a l _ , 1959; P i t t - R i v e r s and R a i l , 1961). S i n c e t h e g o i t r o u s t h y r o i d g l a n d s o f t h e c h r o n i c a l l y RSM-s t i m u l a t e d b i r d s a p p e a r e d t o be a c t i v e l y a c c u m u l a t i n g i o d i n e (which c o u l d be t h r e e t i m e s as much as i n the c o n t r o l g l a n d s ) , i t would be o f i n t e r e s t t o compare the p a t t e r n s o f t h y r o i d a l r a d i o i o d i n e u p t a k e among the b i r d s o f \\ the d i f f e r e n t t r e a t m e n t s . The f o l l o w i n g e x p e r i m e n t s was c o n d u c t e d t o d e t e r m i n e the p e r c e n t a g e uptake o f a dose o f 1-131 by the t h y r o i d g l a n d 6 and 22 hours a f t e r a d m i n i s t r a t i o n . M a t e r i a l s and Methods: (A) 1-131 u p t a k e a t 6 hours a f t e r a d m i n i s t r a t i o n : F i v e b i r d s ( S h a v e r s t r a i n ) o f each o f t h e f o u r t r e a t m e n t s as i n e x p e r i m e n t 1 were moved t o i n d i v i d u a l cages i n a s e p a r a t e room r e s e r v e d f o r r a d i o i s o t o p e s t u d i e s . The b i r d s were g i v e n t h e i r r e s p e c t i v e d i e t s and w a t e r ad l i b i t u m . 1-131 was p u r c h a s e d from Atomic Energy o f Canada, Ottawa, 131 i n t h e f o r m o f Na I i n sodium s u l f a t e s o l u t i o n . 0 . 5 / i C i o f 1-131 i n 0.5 ml p h y s i o l o g i c a l s a l i n e was i n j e c t e d i n t o the l e f t b r a c h i a l v e i n o f each b i r d . The,time o f each i n j e c t i o n was n o t e d . A t e x a c t l y 6 hours a f t e r each 1-131 a d m i n i s t r a t i o n , t h e b i r d s were k i l l e d . The t h y r o i d g l a n d s were removed-; and weighed, and c o u n t e d f o r r a d i o a c t i v i t y i n a N u c l e a r - C h i c a g o 8166 w e l l -t y p e s c i n t i l l a t i o n c o u n t e r . The c o u n t r a t e s (cpm) were c o r r e c t e d f o r back-ground and c o i n c i d e n c e l o s s a c c o r d i n g t o t h e e q u a t i o n i n Wang and W i l l i s ( 1 9 6 5 ) : -30-1 - mT where n i s the c o r r e c t e d cpm, m i s t h e o b s e r v e d cpm, and T i s t h e r e s o l v i n g t i m e o f t h e c o u n t i n g assembly. The c o r r e c t e d cpm was c a l c u l a t e d back t o i n j e c t i o n ( z e r o ) t i m e by c o r r e c t i o n w i t h t h e decay c o n s t a n t o f 1-131, and e x p r e s s e d as a p e r c e n t a g e o f t h e c o r r e c t e d c o u n t r a t e o f a s t a n d a r d o r i g i n a l dose ( 0 . 5 / i C i ) . (B) 1-131 uptake a t 22 hours a f t e r a d m i n i s t r a t i o n : T h i s p a r t o f t h e e x p e r i m e n t was c a r r i e d o u t i n c o n j u n c t i o n w i t h e x p e r i m e n t 4. A n o t h e r twenty White Leghorn l a y e r s ( S h a v e r ) , f i v e o f each t r e a t m e n t , from t h e same s t o c k were s e p a r a t e l y caged. 50 u C i o f 1-131 i n normal s a l i n e was i n j e c t e d i n t o t h e l e f t wing v e i n o f each b i r d and t h e ti m e o f i n j e c t i o n was r e c o r d e d . The b i r d s were g i v e n t h e i r r e s p e c t i v e d i e t s and water ad l i b i t u m . Twenty-two hours a f t e r 1-131 a d m i n i s t r a t i o n , t h e b i r d s were k i l l e d , and t h e i r t h y r o i d s were removed and homogenized i n a P o t t e r -Elvehjem. g j a s s - h o m o g e n i z e r i n T r i s - H C l b u f f e r t o g i v e a s o l u t i o n o f t i s s u e c o n c e n t r a t i o n 40 mg/ml. 100 u l o f t h i s s u s p e n s i o n was ta k e n f o r c o u n t i n g o f r a d i o a c t i v i t y i n a w e l l - t y p e s c i n t i l l a t i o n c o u n t e r . A g a i n , t h e c o u n t r a t e s were c o r r e c t e d f o r background, c o i n c i d e n c e l o s s and d e c a y , and f i n a l l y e x p r e s s e d as p e r c e n t a g e s o f t h e c o r r e c t e d cpm o f t h e o r i g i n a l dose ( 5 0 y u C i ) o f r a d i o i o d i n e . In a d d i t i o n t o t h e t h y r o i d g l a n d s , p o r t i o n s o f t h e l i v e r o f each b i r d s were e x c i s e d , weighed and c o u n t e d f o r r a d i o a c t i v i t y . -31-R e s u i t s : The a v e r a g e a b s o l u t e t h y r o i d w e i g h t s , t o t a l p e r c e n t a g e t h y r o i d u p t a k e , p e r c e n t a g e u p t a k e p e r 100 mg t h y r o i d t i s s u e , and e s t i m a t e d p e r c e n t a g e uptake p e r u n i t e p i t h e l i a l w e i g h t o f 1-131 f o r both 6 h and 22 h i n t e r v a l s a r e p r e s e n t e d i n T a b l e 7. The marked i n c r e a s e o f a b s o l u t e t h y r o i d 1-131 uptake by t h e c h r o n i c a l l y RSM-fed b i r d s were c o m p a t i b l e w i t h t h e i r enhanced a c c u m u l a t i o n o f t h y r o i d a l i o d i d e as d e t e r m i n e d by c h e m i c a l a n a l y s i s . When the' * I-131 u p t a k e / g t h y r o i d ' d a t a were c a l c u l a t e d w i t h t h e 'average p e r c e n t a g e e p i t h e l i u m ' o f t h e r e s p e c t i v e g l a n d s as d e t e r m i n e d i n e x p e r i m e n t 1, e s t i m a t e d v a l u e s o f '1-131 u p t a k e / w e i g h t e p i t h e l i u m ' were o b t a i n e d . F u r t h e r m o r e , i t can be n o t e d t h a t w h i l e t o t a l t h y r o i d 1-131 uptake had dropped from 6 hours t o 22 hours i n the S-S and R-S b i r d s , i t remained h i g h i n t h e R-R and S-R g r o u p s . T h e r e d i d n o t a p p e a r t o be any d i f f e r e n c e i n t h e l i v e r 1-131 c o n c e n t r a t i o n 22 hours a f t e r a d m i n i s t r a t i o n o f t h e r a d i o i o d i n e ( T a b l e 7) among the b i r d s from t h e d i f f e r e n t d i e t a r y t r e a t m e n t s . -32-T a b l e 7: C o n c e n t r a t i o n s o f r a d i o i o d i n e i n the t h y r o i d g l a n d and 1 l i v e r 6 and 22 hours a f t e r a d m i n i s t r a t i o n . T o t a l t h y r o i d T h y r o i d E s t i m a t e d t h y . r a d i o a c t i v i t y r a d i o a c t i v i t y r a d i o a c t i v i t y L i v e r D i e t a r y % i n j e c t e d dose % i n j e c t e d dose % i n j e c t e d dose r a d i o a c t i v i t y t r e a t m e n t /100 mg t h y r o i d /mg e p i t h e l i u m a t 22 h 6 h 22 h 6 h 22r.h 6 h 22 h cpm/g S-S 9.1 a 7.8 a 4.7 b 4.2 b c .194 .172 556 R-S 13.6 3 10.9 a 2.3 a 3.6 b c .156 .247 397 R-R 35.4 b 40.6 b 1.5 a 2.2 a .104 .143 694 S-R 30.1 b 42.4 b 2.7 a 3.0 a b .134 .147 382 L e v e l o f s i g n i f i c a n c e 1% 1% 1% 5% v a l u e s w i t h t he same s u p e r s c r i p t a r e n o t s i g n i f i c a n t l y d i f f e r e n t . -33-Experiment 4: THE DISTRIBUTION OF A SINGLE DOSE OF 1-131 IN IODINATED SUBSTANCES IN THYROID GLANDS OF SBM- OR RSM-FED BIRDS S i n c e t h e c h r o n i c a l l y R SM-stimulated t h y r o i d g l a n d s showed such marked enhancement o f 1-131 uptake when compared t o t h e c o n t r o l , i t might be p e r t i n e n t t o know how t h i s a c c u m u l a t e d r a d i o i o d i n e i s d i s t r i b u t e d among t h e v a r i o u s iodo-compounds i n the t h y r o i d t i s s u e . The d e t e r m i n a t i o n o f l a b e l e d iodoamino a c i d d i s t r i b u t i o n i n t h y r o g l o b u l i n s i s c u r r e n t l y a common measure o f t h y r o i d hormone b i o s y n t h e s i s under p h y s i o l o g i c a l o r p a t h o l o g i c a l c o n d i t i o n s ( M e l l e n and Wentworth, 1959; G a t t e r e a u e t a l _ , 1971; Kobayashi and G r e e r , 1971, 1973; B u r g i e_t al_, 1974; A k i b a and Matsumoto, 1973; S o r i m a c h i and U i , 1974). The p r e s e n t e x p e r i m e n t was c a r r i e d o u t t o d e t e r m i n e t h e e f f e c t s o f c h r o n i c and e a r l y g o i t r o g e n i c s t i m u l a t i o n by RSM on t h e d i s t r i b u t i o n o f 1-131 r a d i o a c t i v i t y i n t h y r o i d h y d r o l y s a t e s a f t e r p ronase d i g e s t i o n . M a t e r i a l s and Methods: F i v e b i r d s (same as i n e x p e r i m e n t 3B) o f each o f f o u r t r e a t m e n t s (S-S, R-S, R-R and S-R) f e d e x p e r i m e n t a l d i e t s as i n e x p e r i m e n t 1 were s e -p a r a t e d from t h e r e s t o f the p o p u l a t i o n . 50,uCi o f 1-131 (Atomic Energy, Ottawa) i n 0.5 ml normal s a l i n e was i n j e c t e d i n t r a v e n o u s l y t o each b i r d s . Twenty-two hours a f t e r i n j e c t i o n , the t h y r o i d g l a n d s were removed. The g l a n d s were weighed and homogenized i n a P o t t e r - E l v e h j e m . g l a s s - h o m o g e n i z e r i n i c e - c o l d T r i s ( h y d r o x y m e t h y l ) a m i n o m e t h a n e - H C l b u f f e r , pH 8.6 (Lamas and DeEscobar, 1972), c o n t a i n i n g 10 M t h i o u r a c i l ( S t u d e r and G r e e r , 1968), t o g i v e a s o l u t i o n o f t i s s u e c o n c e n t r a t i o n 40 mg/ml. -34-The method f o r t h e measurement o f I - 1 3 1 - l a b e l e d m e t a b o l i t e s i n t h y r o i d t i s s u e f o l l o w e d was a m o d i f i c a t i o n o f t h a t o f Agerbaek (1972). Two ml o f w e l l - m i x e d homogenate, t i s s u e c o n c e n t r a t i o n 40 mg/ml, was p l a c e d i n a Thunberg tube and 4 mg Pronase ( s u b t i l o p e p t i d a s e - A , K and K F i n e C h e m i c a l s ) p l u s 2 drops o f t o l u e n e as b a c t e r i o s t a t i c agent were added. The tubes were e v a c u a t e d and f l u s h e d w i t h p u r i f i e d N 2 s e v e r a l t i m e s , n i t r o g e n f i n a l l y b e i n g a l l o w e d t o remain i n t h e tube a t a t m o s p h e r i c p r e s s u r e . D u r i n g t h e e v a c u a t i o n p r o c e d u r e i t was n e c e s s a r y t o t a p t h e t u b e b r i s k l y t o c o n t r o l f r o t h i n g . The d i g e s t i o n t o o k p l a c e i n t h e s t o p p e r e d tube under n i t r o g e n p l a c e d i n a s h a k i n g water bath a t 37 t fC f o r 21-24 h o u r s . F o r t y / J 1 o f d i g e s t s u s p e n s i o n was removed i m m e d i a t e l y a f t e r v i g o r o u s s h a k i n g o f the d i g e s t i o n tube and was a p p l i e d on Whatmann 3 MM chromatography paper s t r i p s (4.5;.em w i d e ) . A p p l i c a t i o n t o o k p l a c e i n a s t r e a m o f c o l d , a t m o s p h e r i c a i r a f t e r which the chromatography paper was suspended i n an a i r p r o o f and t e m p e r a t u r e i s o l a t e d j a r where i t was e q u i l i b r a t e d w i t h s o l v e n t f o r about 12 hours b e f o r e chromatography was s t a r t e d . F r e s h l y p r e p a r e d chromatography s o l v e n t n - b u t a n o l ( r e d i s t i l l e d ) - e t h a n o l - a m m o n i a IN (^5:1:2) was used. The a s c e n d i n g chromatogram was d e v e l o p e d f o r about 16 hours a t room t e m p e r a t u r e i n the d a r k (Lamas and D e E s c a r b a r , 1972). The chromatogram was then s p r a y e d l i g h t l y w i t h f e r r i c h l o r i d e - p o t a s s i u m f e r r i c y a n i d e -a r s e n i c a c i d (Gmelin and V i r t a n e n , 1959) so t h a t t h e c h r o m a t o g r a p h i c p r o c e s s c o u l d be v i s u a l l y m o n i t o r e d . The s e c t i o n s o f t h e chromatograms c o r r e s p o n d i n g t o MIT, DIT, T^ and T 3 > t o g e t h e r w i t h the o r i g i n and i n t e r m e d i a t e a r e a s , were e x c i s e d and c o u n t e d i n a w e l l - t y p e s c i n t i l l a t i o n c o u n t e r . O r i g i n 1-131, as used i n t h i s s t u d y , r e f e r r e d not o n l y t o r a d i o a c t i v i t y r e m a i n i n g a t the s i t e : , -35-o f a p p l i c a t i o n , but t o a l l 1-131 r a d i o a c t i v i t y up t o t h e DIT band. The 1-131 c o r r e s p o n d i n g t o each i n d i v i d u a l iodoamino a c i d was e x p r e s s e d as a p e r c e n t a g e o f t h e t o t a l 1-131 r a d i o a c t i v i t y on t h e chromatogram. A r e f e r e n c e chromatogram was a l s o o b t a i n e d by s p o t t i n g 1 u l o f a s t a n d a r d s o l u t i o n ( a p p r o x i m a t e l y 0.6%) o f s t a b l e K I , MIT, DIT, T^ and T^ (Sigma C h e m i c a l s ) t o g e t h e r w i t h a s m a l l amount o f d i g e s t . The p a t t e r n o f e l u t i o n o f t h e s e compounds i n the chromatogram was i d e n t i c a l t o t h a t g i v e n by Agerbaek (1972). S e v e r a l d i f f e r e n t s o l v e n t systems were t e s t e d such as n - b u t a n o l -a c e t i c a c i d - w a t e r (4:1:5) used by Kobayashi and G r e e r ( 1 9 7 1 ) , c o l l i d i n e - 3 N ammonium h y d r o x i d e (3;1) used by Inoue and T a u r o g ( 1 9 6 7 ) , n - b u t a n o l - a b s o l u t e e t h a n o l - 0 . 2 5 N ammonium h y d r o x i d e (5:1:2) used by Kobayashi and G r e e r ( 1 9 7 1 ) , and n - b u t a n o l - e t h a n o l - e t h a n o l - a i n r n o n i a 0.4N (5:1:2) used by Agerbaek ( 1 9 7 2 ) , b e f o r e a f i n a l d e c i s i o n was made on n - b u t a n o l - e t h a n - 1 N ammonia (5:1:2) which had been used by M o u r i z e t al_ (1966) and Lamas and DeEscobar (1972). Moreover, i t was found n e c e s s a r y t o r e d i s t i l l n - b u t a n o l and t o use a f r e s h l y - p r e p a r e d s o l v e n t system f o r each chromatography r u n . R e s u l t s : E f f e c t s o f t h e d i f f e r e n t d i e t a r y t r e a t m e n t s upon t h e d i s t r i b u t i o n o f r a d i o a c t i v e i o d i n a t e d s u b s t a n c e s found i n t h y r o i d g l a n d s 22 hours a f t e r i n j e c t i o n o f 1-131 a r e p r e s e n t e d i n T a b l e 8. A q u a n t i t a t i v e l y r e v e r s e d r e l a t i o n s h i p between t h e p e r c e n t a g e s o f r a d i o a c t i v i t y i n the i o d o t y r o s i n e s o f t h e c h r o n i c a l l y R S M - s timulated t h y r o i d t i s s u e s was e v i d e n t : In the S-S and R-S t h y r o i d s , t h e m a j o r i t y o f t h e 1-131 was f o u n d i n the DIT band, whereas i n t he R-R and S-R chromatograms, most o f t h e r a d i o a c t i v i t y was l o c a t e d a t t h e -36-MIT band. T h e r e was a c c o r d i n g l y an i n c r e a s e i n t h e MIT/DIT r a t i o s o f the RSM- s t i m u l a t e d t h y r o i d s . F u r t h e r m o r e , i t appeared t h a t RSM-stimulated b i r d s had more r a d i o a c t i v i t y r e s i d i n g i n t h e i o d i d e band and a t t h e o r i g i n , which p r o b a b l y r e p r e s e n t e d d e g r a d a t i v e l o s s o r h e t e r o g e n e i t y o f l a b e l e d i o d o p r o t e i n w i t h r e g a r d t o h y d r o l y s i s (Inoue and T a u r o g , 1967; Kobayashi and G r e e r , 1975). Upon s p r a y i n g w i t h f e r r i c h l o r i d e - p o t a s s i u m f e r r i c y a n i d e - a r s e n i c a c i d , t h e bands i n t h e chromatograms a p p e a r e d c l e a r l y v i s i b l e e x c e p t f o r Tg. D e s p i t e q u e s t i o n s as t o t h e p r e s e n c e o f t r i i o d o t h y r o n i n e i n t h y r o g l o b u l i n o f th e aves (Vlig'm, 1958; S o r i m a c h i and U i , 1974), 1-131 r a d i o a c t i v i t y was d e f i n i t e l y l o c a t e d i n the r e g i o n c o r r e s p o n d i n g t o t h e Tg band o f t h e r e f e r e n c e chromatogram. The amounts o f Tg and T^ p r e s e n t were, however, r e l a t i v e l y s m a l l and have t h e r e f o r e been combined i n t h e t a b l e ( T a b l e 8 ) . Th e r e appeared t o be an a d d i t i o n a l band i n a l l t h e chromatograms o f t h e S-S and R-S g r o u p s , which was a b s e n t from t h e R-R and S-R p a t t e r n s ^ ( F i g u r e 1 ) . T h i s band was c l o s e b u t d i s t i n c t l y s e p a r a t e d from t h e I \" band, between t he I \" and t h e DIT l e v e l s . In t h e absence o f i n d e n t i f i c a t i o n , i t was d e s i g n a t e d by (?) i n T a b l e 8. I t mi g h t be a h i t h e r t o u n r e c o g n i z e d iodoamino a c i d ( d i i o d o t h y r o n i n e , T2?) i n the chromatogram o r a more e x p e r i m e n t a l a r t e -f a c t o f t h e chromatography system. C o u p l i n g e f f i c i e n c y , as c a l c u l a t e d by Tg + T^/ ( M I T / D I T ) , was lower f o r t h e RSM-stimulated b i r d s . Once a g a i n , as c o n s i s t e n t w i t h t h e o t h e r p a r a m e t e r s o f t h y r o i d m e t a b o l i s m measured, t h e o r d e r o f i n h i b i t i o n i n t h y r o i d hormone b i o s y n t h e s i s , as r e p r e s e n t e d by t h e p a t t e i t i o f d i s t r i b u t i o n o f l a b e l e d m e t a b o l i t e s i n t h e t h y r o i d t i s s u e s , appeared i n t h e f o l l o w i n g o r d e r : R-R>S-R>R-S>S-S. -37-T a b l e 8: D i s t r i b u t i o n , 22 hours a f t e r a d m i n i s t r a t i o n , o f a s i n g l e dose o f r a d i o i o d i n e . i n t o t h y r o i d a l compounds. D i e t a r y % o f t o t a l t h y r o i d a l a c t i v i t y T3+T4 t r e a t m e n t O r i g i n * I \" MIT DIT ? T 3 & T 4 MIT/DIT MIT/DIT S-S 5.0 1.2 40.7 47.9 1.9 3.3 0.85 3.89 R-S 5.3 1.4 42.7 45.3 2.0 3.3 0.95 3.43 R-R 8.3 2.8 49.1 36.8 - 3.0 1.34 2.28 S-R 6.4 2.1 49.0 39.5 - 2.9 1.27 2.39 R e p r e s e n t s t o t a l r a d i o a c t i v i t y from l i n e o f a p p l i c a t i o n t o band o f DIT. -38-F i g u r e 1: Paper chromatograms o f t h y r o i d a l m e t a b o l i t e s o f b i r d s on d i f f e r e n t d i e t a r y t r e a t m e n t s . C h r o m a t o g r a p h i c s o l v e n t : n-butanol-ethanol-ammonia IN (5:1:2). A b b r e v i a t i o n s used: 0 ( o r i g i n ) , DIT ( 3 , 5 - d i i o d o t y r o s i n e ) , MIT ( 3 - m o n o i o d o t y r o s i n e ) , S - S R - S R - R S - R -39-E x p e r i m e n t 5: EFFECTS OF GOITROGENIC RSM ON SUSCEPTIBILITY TO HYDROLYSIS OF THYROGLOBULIN H e t e r o g e n e i t y o f t h e t h y r o i d f o l l i c l e s has o f t e n been d i s c u s s e d i n t h e l i t e r a t u r e w i t h r e g a r d t o t h e r a t e o f i o d i n e t u r n o v e r ( S n e i d e r , 1964), and h e t e r o g e n e i t y o f t h e i o d o p r o t e i n s i n the t h y r o i d g l a n d have l o n g been r e c o g n i z e d ( T a r u t a n i and U i , 1969a). More r e c e n t l y , i t has been s u g g e s t e d t h a t t h y r o g l o b u l i n s s t o r e d i n t h e c o l l o i d may perhaps a l s o be heterogenous w i t h r e g a r d t o the r e l e a s e o f t h y r o i d hormones (Simon e t a l _ , 1966; T a r u t a n i and U i , 1969b; Nunez e t al_, 1966; Koyayashi and G r e e r , 1971; K o h l e r e t a l _ , 1971; Rosenberg et_ a l _ , 1964). S i n c e o u r f i n d i n g ( e x p e r i m e n t 4) o f a l a r g e r p e r c e n t a g e o f 1-131 r a d i o a c t i v i t y r e s i d i n g i n the I \" and o r i g i n r e g i o n s o f t h e chromato-grams o f t h e R S M - s t i m u l a t e d t h y r o i d t i s s u e s , i t was c o n s i d e r e d r e l e v e n t t o t e s t whether t h e r e was i n d e e d a d i f f e r e n c e i n the s u s c e p t i b i l i t y t o p r o t e o l y s i s among t h e i o d o p r o t e i n s o f t h e v a r i o u s t r e a t m e n t g r o u p s . The f o l l o w i n g e x p e r i -ment was d e s i g n e d t o d e m o n s t r a t e any h e t e r o g e n e i t y i n the t h y r o i d t i s s u e s o f RSM- and SBM-fed b i r d s w i t h r e f e r e n c e t o the r a t e o f h y d r o l y s i s by p r o n a s e . M a t e r i a l s and Methods: White Leghorn c o c k e r e l s , 8 weeks o f age, f e d t h e SBM- o r RSM-s upplemented d i e t s same as t h o s e f o r t h e RSM g o i t r o g e n i c i t y t e s t i n e x p e r i m e n t 1 were used. S i x b i r d s each o f t h e two t r e a t m e n t s (SBM and RSM) were put i n s e p a r a t e cages and g i v e n w a t e r and t h e i r r e s p e c t i v e d i e t s ad 1 i b i t u m . 5 0 / j C i o f 1-131 i n 0.5 ml normal s a l i n e was i n j e c t e d i n t o t h e l e f t b r a c h i a l v e i n o f e a c h . Twenty-two hours a f t e r r a d i o i o d i n e a d m i n i s t r a t i o n , t h r e e b i r d s -40-from each t r e a t m e n t were k i l l e d and t h e i r t h y r o i d s removed. Each p a i r o f g l a n d s was homogenized i n t h e same manner as i n e x p e r i m e n t 4, and s u b j e c t e d t o pronase d i g e s t i o n i n a water bath a t 37°C. Four hours a f t e r i t s commence-ment h y d r o l y s i s o f s i x samples was s t o p p e d ( t h r e e from each t r e a t m e n t ) and 4 0 / j l o f the d i g e s t s u s p e n s i o n was t r a n s f e r r e d f o r chromatography as d e s c r i b e d i n e x p e r i m e n t 4. The o t h e r s i x samples were s i m i l a r l y t r e a t e d a f t e r 24 hours o f pronase d i g e s t i o n . R e s u l t s : P e r c e n t d i s t r i b u t i o n o f r a d i o a c t i v i t y i n t h y r o i d t i s s u e s o f SBM- o r RSM-treated c o c k e r e l s a f t e r f o u r o r t w e n t y - f o u r hours o f p r o n a s e d i g e s t i o n a r e g i v e n on T a b l e 9. T h a t d i g e s t i o n was n o t c o m p l e t e by 4 hours was e v i d e n t by t h e h i g h p e r c e n t a g e o f r a d i o a c t i v i t y r e m a i n i n g a t t h e o r i g i n o f the chromatograms. However, t h e r e was no d i f f e r e n c e i n the p e r c e n t a g e s o f r a d i o a c t i v i t y r e m a i n i n g a t t h e o r i g i n . A f t e r 24 hours o f h y d r o l y s i s , t h e MIT/DIT r a t i o o f t h e RSM-fed b i r d s was m a r k e d l y i n c r e a s e d o v e r t h a t o f t h e c o n t r o l . T h e r e a l s o d i d n o t appear t o be any d i f f e r e n c e i n t h e p e r c e n t a g e s o f 1-131 r a d i o a c t i v i t y c o r r e s p o n d i n g t o t h e i o d o t h y r o n i n e s o r the ' o r i g i n ' bands i n the chromatograms o f t h e t h y r o i d t i s s u e o f t h e RSM- o r SBM-fed b i r d s . The d i s t r i b u t i o n o f r a d i o a c t i v i t y i n i o d i n a t e d s u b s t a n c e s i n t h e t h y r o i d t i s s u e o f young c o c k e r e l s i n the p r e s e n t e x p e r i m e n t was comparable t o t h a t r e p o r t e d f o r growing c h i c k s by o t h e r i n v e s t i g a t o r s ( e . g . A k i b a and Matsumoto, 1973). -41-T a b l e 9: D i s t r i b u t i o n , 22 hours a f t e r a d m i n i s t r a t i o n , o f a s i n g l e dose o f r a d i o i o d i n e i n t o t h y r o i d a l compounds measured i n h y d r o l y z a t e s a f t e r d i f f e r e n t times o f p r o n a s e d i g e s t i o n . D i e t a r y % o f t o t a l t h y r o i d a l r a d i o a c t i v i t y t r e a t m e n t O r i g i n I \" MIT DIT T 3 & T 4 MIT/DIT A f t e r 4 hours pronase h y d r o l y s i s : SBM 16.2 6.1 29.7 41.3 6.7 0.72 RSM 15.0 6.5 43.8 24.3 10.4 1.81 A f t e r 24 hours p r o n a s e h y d r o l y s i s : SBM 7.0 6.0 33.2 46.8 9.0 0.72 RSM 8.1 7.2 44.3 28.2 12.2 1.58 -42-E x p e r i m e n t 6A: EFFECTS OF GOITROGENIC RSM ON THE IODINE CONTENT OF EGG YOLK A d i r e c t r e l a t i o n s h i p between t h e i o d i n e c o n t e n t o f t h e dam's d i e t and t h e i o d i n e c o n c e n t r a t i o n i n t h e egg has l o n g been known (Hercus and R o b e r t s , 1927; Simpson and S t r a n d , 1930; W i l d e r et_ a l _ , 1933; Asmundson ejt a l _ , 1936) and the e f f e c t s o f i o d i n e d e f i c i e n c y on embryo development and h a t c h a b i l i t y have been c l e a r l y e s t a b l i s h e d ( H o l l a n d e r and R i d d l e , 1943, 1946; Wil g u s e t a l _ , 1953; B e n n e t t and Adams, 1952; V i d a l , 1952; Adams and Buss, 1952; K i n g s b u r y e t a l _ , 1955; Romanoff and L a u f e r , 1956; Perdomo et_ al _ , 1966). T h y r o i d h i s t o l o g y o f t h e n e w l y - h a t c h e d progeny showed h y p e r t r o p h y o f t h e e p i -t h e l i u m and a l a c k o f c o l l o i d , which c o u l d be r e v e r s e d by a d d i t i o n o f p o t a s s i u m i o d i d e t o t h e dam's d i e t . T h y r o i d a l h y p e r t r o p h y was r e p o r t e d from t h i s l a b o r a t o r y i n t h e progeny from dams c h r o n i c a l l y f e d a RSM-supplemented d i e t (March et_ a l _ , 1972). The g o i t r o u s c o n d i t i o n was a l l e v i a t e d when the young c h i c k s were f e d a d i e t n o r m a l l y supplemented w i t h i o d i n e . The resemblance between t he progeny from the c h r o n i c a l l y RSM-treated b i r d s t o t h o s e from dams f e d an i o d i n e - d e f i c i e n t d i e t l e d t o the assumption o f an i o d i n e d e f i c i e n c y i n the eggs o f the f o r m e r group. T h i s was f u r t h e r s u p p o r t e d by t h e f a i l u r e t o d e m o n s t r a t e any d e p o s i t i o n o f g o i t r i n i n the eggs o f the RSM-fed b i r d s o r any d i f f e r e n c e i n i s o t h i o -c y a n a t e c o n t e n t o f t h e eggs o f t h e s e b i r d s when compared t o c o n t r o l (March e t a l , 1972). A n a l y s i s o f i o d i n e c o n t e n t o f t h e d i e t s , however, d i d not r e v e a l any d e f i c i e n c y o f t h i s e l e m e n t i n t h e RSM-supplemented r a t i o n ( e x p e r i m e n t 1 ) . S i n c e the egg p r o d u c t i o n and h a t c h a b i l i t y o f t h e s e b i r d s f e d c h r o n i c a l l y w i t h -43-the RSM-supplemented d i e t were a f f e c t e d (March et_ al_, 1975) and t h e progeny from them c o n s i s t e n t l y showed t h y r o i d h y p e r t r o p h y a t h a t c h i n g ( e x p e r i m e n t 8 ) , t h e f o l l o w i n g e x p e r i m e n t was c o n d u c t e d t o d e t e r m i n e i o d i n e c o n c e n t r a t i o n i n the eggs from t h e d i f f e r e n t t r e a t m e n t s and h o p e f u l l y t o e l u c i d a t e t h e materno-'emb.r'yonic-rela.tionsh.ip. :i'(e*-re*• • p e r i m e n t 3 ) . T h i s conforms w i t h t h e f i n d i n g o f Matsumoto e t a l _ (1968) pra the e f f e c t s o f g o i t r i n i n growing c h i c k s . A c c o r d i n g t o t h e s e i n v e s t i g a t o r s , i n a d d i t i o n t o m a i n t a i n i n g a l a r g e amount o f i n c o r p o r a t e d 1-131 i n t h e t h y r o i d g l a n d , the g o i t r i n - f e d c h i c k s a l s o show a slow t h y r o i d a l r e l e a s e o f t h e i n -j e c t e d r a d i o i o d i n e . T h i s may perhaps e x p l a i n our f i n d i n g t h a t i n b i r d s ' o f c h r o n i c RSM t r e a t m e n t s uptake o f a s i n g l e dose o f r a d i o i o d i n e a t 22 hours was g r e a t e r t h a n a t 6 h o u r s . T h i s was i n c o n t r a s t t o t h e b i r d s on t r e a t m e n t s S-S and R -S-in which r a d i o a c t i v i t y had, by 22 h o u r s , d e c l i n e d from t h e amount p r e s e n t 6 hours a f t e r i n j e c t i o n . The t ime o f maximum r a d i o a c t i v i t y , f o l l o w i n g , a d m i n i s t r a t i o n o f r a d i o i o d i n e , which i s d e t e r m i n e d by t h e r a t e o f r e l e a s e o f -62-i o d i n e r e l a t i v e t o the r a t e o f u p t a k e , o c c u r r e d l a t e r i n t h e RSM-fed b i r d s t h a n i n the SBM-fed b i r d s . R e l a t e d t o t h e a c t i v e p r o c e s s o f i o d i n e a c c u m u l a t i o n o f t h e t h y r o i d g l a n d o f RSM-fed b i r d s i s i t s h i s t o l o g i c a l p i c t u r e . The f u n c t i o n a l h y p e r a c t i v i t y i s d e m o n s t r a t e d by t h e m a s s i v e c o n t e n t o f e p i t h e l i u m i n t h e i r h y p e r t r o p h i c f o l l i c l e s . The a v e r a g e e p i t h e l i a l h e i g h t o f t h e s e g l a n d s i s a l s o s i g n i f i c a n t l y h i g h e r t h a n t h a t o f t h e c o n t r o l . The c o l l o i d i n t h e f o l l i c u l a r lumen o f t h e s e g l a n d s appear t o be b r i g h t l y s t a i n e d , which i s i n agreement w i t h t h e i r h i g h i o d i n e c o n t e n t . A l t h o u g h t h e o c c u r r e n c e o f e p i t h e -l i a l c e l l s s l o u g h i n g o f f i n t o the lumen a r e h i g h e r i n the c h r o n i c a l l y RSM-fed b i r d s t h a n i n the c o n t r o l , no s i g n o f f o l l i c u l a r a t r o p h y can be f o u n d . Thus i t can be s a i d t h a t t h e s e b i r d s a r e h y p e r a c t i v e i n the t h y r o i d a l c l e a r a n c e o f i o d i n e from t h e b l o o d c i r c u l a t i o n . G o i t r o g e n i c i n h i b i t i o n o f c o n v e r s i o n o f i o d i d e a c c u m u l a t e d by t h e t h y r o i d i n t o i t s a c t i v e f o r m and f u r t h e r t o MIT and DIT has been e x t e n s i v e l y i n v e s t i g a t e d ( S t a n b u r y e t a l _ , 1955; Haddad and S i d b u r y , 1959; G l a y t o n e t a l , 1958; F l o y d e t a l _ , i 9 6 0 ; P a r k e r and B e i e r w a l t e s , 1961; B a s c h i e r i et_ a l _ , 1963). A c c o r d i n g t o t h e s e i n v e s t i g a t o r s , such an i o d i d e o r g a n i z a t i o n d e f e c t i s c h a r a c t e r i z e d by a predominance o f i n o r g a n i c i o d i n e and s c a r c i t y o f i o d o -amino a c i d s when the t h y r o i d t i s s u e h y d r o l y z e d , p o s s i b l y due t o f a i l u r e o f i o d i n a t i o n o f t y r o s i n e by t h y r o i d a l p e r o x i d a s e . In our p r e s e n t i n v e s t i g a t i o n i n which t h e p r e - l a b e l e d t h y r o i d g l a n d s o f t h e c h r o n i c a l l y RSM-fed b i r d s were h y d r o l y z e d and the iodoamino a c i d s f r a c t i o n a t e d ( e x p e r i m e n t 4 ) , t h e f r a c t i o n s o f MIT and DIT were l a r g e and t h e r e was l i t t l e i n o r g a n i c i o d i n e 22 hours a f t e r a d m i n i s t r a t i o n o f r a d i o i o d i n e . Thus t h e i o d i d e o r g a n i z a t i o n p r o c e s s i s p r o -b a b l y n o t d i s t u r b e d i n the RSM-fed b i r d s . -63-T h a t t h y r o i d hormone b i o s y n t h e s i s i s i n d e e d i n h i b i t e d i n the RSM-fed b i r d s i s e v i d e n t by t h e r e s u l t s o b t a i n e d on f r a c t i o n a t i o n o f t h e iodoamino a c i d s i n t h y r o i d h y d r o l y s a t e s . The d i s t r i b u t i o n o f 1-131 r a d i o -a c t i v i t y among v a r i o u s t h y r o i d m e t a b o l i t e s shows t h a t t h e p r o p o r t i o n ' o f r a d i o i o d i n e p r e s e n t as MIT, 22 hours a f t e r a d m i n i s t r a t i o n , i s g r e a t e r i n t h e RSM-fed b i r d s . The su b s e q u e n t i n c r e a s e i n MIT/DIT r a t i o i s h e l d commonly as an i n d i c a t i o n o f d e f e c t ( s ) i n t h y r o i d hormonogenesis. C o u p l i n g e f f i c i e n c y i . e . Tg + T 4 / ( M I T / D I T ) , as a measure o f t h e combined r a t e s a t which d i o d o -t y r o s i n e and t h y r o i d hormones a r e pro d u c e d i s , i n o r d e r , SS>RS>SR>RR and i s i n v e r s e l y p r o p o r t i o n a l t o t h e t o t a l amount o f i o d i n e i n t h e t h y r o i d g l a n d . However, s i n c e t h e p e r c e n t a g e s o f t h y r o i d i o d o t h y r o n i n e s i n the RSM-fed b i r d s a r e n o t lower t o any s i g n i f i c a n t e x t e n t when compared t o t h e c o n t r o l , i t i s no t l i k e l y t h a t t h e c o u p l i n g mechanism o f MIT and DIT t o i o d o t h y r o n i n e s i s i n h i b i t e d p e r se by t h e f e e d i n g o f g o i t r o g e n i c RSM. T h i s seems t o be i n agreement w i t h t h e f i n d i n g o f A k i b a and Matsumoto (1971) t h a t a l t h o u g h t h e s y n t h e t i c a c t i v i t y o f t h e i o d o t h y r o n i n e s i n g o i t r i n - f e d c h i c k s i s ab o u t two-t h i r d o f the c o n t r o l a t 24 hours a f t e r dosage o f r a d i o i o d i n e , t h e amount o f i o d o t h y r o n i n e s s y n t h e s i z e d i n t h e s e c h i c k s i s about two times t h a t o f t h e c o n t r o l . In the p r e s e n t s t u d y , t h e s l i g h t l y l o w e r p e r c e n t a g e s o f t h y r o i d a l i o d o t h y r o n i n e s i n the RSM-fed birds,,;:when m u l t i p l i e d by t h e 10- t o 1 3 - f o l d i n c r e a s e i n the w e i g h t s o f t h e s e g l a n d s o v e r t h e c o n t r o l , w i l l g i v e e s t i m a t e d i o d o t h y r o n i n e v a l u e s l a r g e r t h a n t h a t o f t h e c o n t r o l . An i n h i b i t o r y s i t e would a p p e a r t o be a t t h e f o r m a t i o n l e v e l o f i o d o t y r o s i n e s , s i n c e i t i s shown t h a t t h e MIT/DIT r a t i o i s i n c r e a s e d i n the t h y r o i d s o f t h e RSM-fed b i r d s , w h i l e t h e c o u p l i n g d e f i c i e n c y i n t h e R-R and S-R groups i s a l s o d e p r e s s e d . Whatever t h e s i t e o f i n h i b i t i o n , however, i t ap p e a r s t h a t t h e s e c h r o n i c a l l y s t i m u l a t e d b i r d s have f u l l y compensated f o r t h e d e f e c t by t h y r o i d h y p e r t r o p h y and h y p e r p l a s i a , most l i k e l y i n v o l v i n g t h e -64-s e c r e t i o n s o f t h e h y p o t h a l a m u s - p i t u i t a r y - t h y r o i d a x i s . An u n r e p o r t e d ex-p e r i m e n t u s i n g t h e c h i c k method o f TSH b i o a s s a y ( B a t e s , 1962) and h i s t o l o g i c a l e x a m i n a t i o n o f the p i t u i t a r y g l a n d s o f t h e s e b i r d s f a i l e d t o be c o n c l u s i v e i n t h i s r e s p e c t . F u r t h e r i n v e s t i g a t i o n s i n t h i s a r e a a r e n e c e s s a r y b e f o r e the neurohormonal r e l a t i o n s h i p s c o n t r o l l i n g t h y r o i d m e t a b o l i s m o f t h e RSM-f e d b i r d s w i l l be e l u c i d a t e d . In the normal t h y r o i d , s y n t h e s i s and s e c r e t i o n a r e t a k i n g p l a c e under \" s t e a d y s t a t e \" c o n d i t i o n s n e a r t h e b o r d e r o f t h e e p i t h e l i a l c e l l i n t h e f o l l i c l e s . T h i s e q u i l i b r i u m i s d i s t u r b e d a l m o s t i m m e d i a t e l y a f t e r t h e i n t r o -d u c t i o n o f a g o i t r o g e n . Under t h e i n f l u e n c e o f t h e e a r l y r i s e i n TSH, new f r a c t i o n s o f t h y r o g l o b u l i n t a k e p a r t i n t h e hormonal exchange and t h e 1-131 mixes w i t h a d i f f e r e n t pool than i n t h e \" r e s t i n g \" , i o d i n e - r i c h t h y r o i d s . T h y r o i d s e c r e t i o n under t h e s e c o n d i t i o n s o f i o d i n e d e f i c i e n c y c o u l d t h u s f o l l o w r a t h e r c o m p l i c a t e d rules-. T h e r e f o r e , a more p r o b a b l e s i t e o f i n h i b i t i o n i s a t the r e l e a s e and s e c r e t i o n o f i o d o t h y r o n i n e s i n t o t h e c i r c u l a t i o n , and t h i s i s s u p p o r t e d by t h e work o f A k i b a and Matsumoto (1971) on the e f f e c t o f g o i t r i n on the t h y -r o i d hormone b i o s y n t h e s i s i n growing c h i c k s . P a r t i c u l a r l y , i t i s presumed t h a t t h e s e c r e t i o n o f t h y r o i d hormone from t h e g l a n d by h y d r o l y s i s o f t h y r o g l o b u l i n i s d e p r e s s e d o r t h a t t h e r e i s no i n c r e m e n t o f h y d r o l y s i s o f t h y r o g l o b u l i n by t h e a p p a r e n t i n c r e a s e d s e c r e t i o n o f TSH from t h e p i t u i t a r y caused by t h e f e e d i n g o f g o i t r i n . Under t h e s e c i r c u m s t a n c e s the t h y r o i d g l a n d s were c a p a b l e o f c o n t i n u o u s h y p e r p l a s i a f o r the d u r a t i o n o f t h e e x p e r i m e n t . The s i t u a t i o n may be s i m i l a r t o t h a t when a s m a l l amount o f t h y r o x i n e i s a d m i n i s t e r e d t o b i r d s i n which t h y r o i d hormone p r o d u c t i o n i s b l o c k e d by a d m i n i s t r a t i o n o f t h i o u r a c i l . S h u l t z e and T u r n e r (1945) found t h a t t h e g o i t r o g e n i c i t y o f t h i o u r a c i l i s en-hanced when a s m a l l amount o f t h y r o x i n e i s a d m i n i s t e r e d s i m u l t a n e o u s l y w i t h t h e t h i o u r a c i l . In t h e p r e s e n t s t u d y , t h e h y d r o l y s i s r a t e by e x t r a n e o u s -65-p r o n a s e does n o t appear t o be any s l o w e r i n the t h y r o i d l o b e s o f the RSM-fed b i r d s t h a n the c o n t r o l ( e x p e r i m e n t 5 ) . How t h i s can be a p p l i e d t o endogenous p r o t e o l y s i s o f t h y r o g l o b u l i n by t h y r o i d enzymes, however, i s u n d e t e r m i n e d . Hence the i n h i b i t o r y e f f e c t o f g o i t r o g e n i c RSM on the r e l e a s e o f hormones from t h y r o g l o b u l i n and s ubsequent s e c r e t i o n from t h e t h y r o i d g l a n d remains a p o s s i -b i l i t y . Due t o a d e f e c t i n i o d o t y r o s i n e d e i c d i n a s e , MIT o r DIT may n o t be d e i o d i n a t e d and a l a r g e amount o f i o d o t y r o s i n e may be l o s t i n t h e u r i n e , r e -s u l t i n g i n a d i s t u r b a n c e i n r e u t i l i z a t i o n o f i n o r g a n i c i o d i n e i n i o d i d e o r -g a n i z a t i o n ( S t a n b u r y , 1957; S t a n b u r y and M o r r i s , 1958). Such a p o s s i b l e e f f e c t by g o i t r o g e n i c RSM on t h e d e i o d i n a t i o n p r o c e s s has n o t been i n v e s t i g a t e d i n the p r e s e n t s t u d y . A l t h o u g h y c o u n t i n g o f e x c i s e d p i e c e s o f l i v e r 22 hours a f t e r a d m i n i s t r a t i o n o f r a d i o i o d i n e has n o t r e v e a l e d any n o t a b l e d i f f e r e n c e i n t h e 1-131 c o n c e n t r a t i o n o f t h e l i v e r s o f t h e b i r d s on t h e d i f f e r e n t d i e t a r y t r e a t -ments, t h e r e l a t i v e r a t e s o f removal o f i o d i n a t e d compounds by t h e s e organs c a n n o t be known from t h e s e d a t a . However, a d e f e c t i n t h e d e i o d i n a t i o n p r o c e s s due t o the f e e d i n g o f g o i t r o g e n i c RSM i s u n l i k e l y , a t l e a s t f o r t r i i o d o t h y r o n i n e . U n p u b l i s h e d d a t a from t h i s l a b o r a t o r y has shown t h a t when a s i n g l e dose o f 1-131-1abeled Tg was a d m i n i s t e r e d i n t o RSM-fed b i r d s , t h e r a t e o f 1-131 t u r n -o v e r , measured by r a d i o a c t i v i t y o f t h e e x c r e t a 24 hours o r 48 hours a f t e r i n j e c t i o n , i s n o t d i f f e r e n t from t h a t o f t h e c o n t r o l . Hence t h e d e i o d i n a t i o n c a p a c i t y i n t h e RSM-fed b i r d s does n o t a p p e a r t o be i n h i b i t e d . In c h r o n i c e x p e r i m e n t s w i t h hamster, i t was f o u n d t h a t i n s e v e r e i o d i n e d e f i c i e n c y a p a r t o f the s t o r e d r a d i o a c t i v i t y i n t h e t h y r o i d c o u l d n o t be d i s c h a r g e d even a f t e r TSH s t i m u l a t i o n and PTU t r e a t m e n t . T h i s i n d i c a t e d a l t e r e d , a b n o r m a l l y s t r o n g , p r o t e i n b i n d i n g o f t h e i n t r a t h y r o i d a l iodoamino a c i d s under c e r t a i n c o n d i t i o n s (Van M i d d l e s w o r t h , 1965; F o l l i s , 1965). In the p r e s e n t c a s e , t h e p o s s i b i l i t y t h a t t h e d e f e c t s i n t h y r o i d f u n c t i o n due t o -66-f o r m a t i o n o f abnormal i o d o p r o t e i n i n t h e t h y r o i d g l a n d i s u n l i k e l y . T h i s abnormal i o d o p r o t e i n o t h e r t h a n t h y r o g l o b u l i n i n t h e t h y r o i d i s w a t e r - s o l u b l e 'and b u t a n o l - i n s o l u b l e , so t h a t on paper chromatography w i t h b u t a n o l , i t w i l l n o t m i g r a t e from t h e o r i g i n ( S t a n b u r y and M c G i r r , 1957; DeGroot e t a l _ , 1958; DeGroot and S t a n b u r y , 1959; Van Wyk ejt a l _ , 1962). No such p r o t e i n was d e t e c t e d i n the t h y r o i d gliands o f the p r e s e n t s t u d y . The p e r c e n t a g e s o f r a d i o a c t i v i t y r e m a i n i n g a t the o r i g i n on t h e chromatograms o f t h e t h y r o i d h y d r o l y s a t e s a r e s i m i l a r and w i t h i n normal ranges f o r a l l b i r d s on the f o u r d i e t a r y t r e a t m e n t s . I t i s t h e r e f o r e h y p o t h e s i z e d t h a t c h r o n i c f e e d i n g o f g o i t r o g e n i c RSM t o b i r d s may i n t e r f e r e w i t h t h y r o i d f u n c t i o n , p r o b a b l y a t an i n t e r m e d i a r y s t e p i n v o l v i n g t h e f o r m a t i o n o f i o d o t y r o s i n e s and/or t h e r e l e a s e o r s e c r e t i o n o f t h e t h y r o i d hormones. One o r more enzyme systems such as t h y r o i d p e r o x i d a s e i n t h e t h y r o i d g l a n d may be i n v o l v e d . D i s o r d e r s o f t h e s e systems have been c o n s i d e r e d t o cause d i s t u r b a n c e s o f t h y r o i d hormone p r o d u c t i o n and provoke compensatory e n l a r g e m e n t o f t h e t h y r o i d w i t h h i g h u p t a k e o f i o d i n e (Fukase e t a l _ , 1967). Thus, i n t h e c h r o n i c a l l y RSM-fed b i r d s , a d i s t u r b a n c e o f t h y -r o i d hormone b i o s y n t h e s i s and/or r e l e a s e due t o an i n t r i n s i c enzyme a l t e r a t i o n i s s t r o n g l y s u g g e s t e d . The h i s t o l o g i c a l p i c t u r e and r a d i o i o d i n e m e t a b o l i s m i n the t h y r o i d s o f t h e s e b i r d s s u g g e s t h y p e r a c t i v i t y , and t h i s conforms w i t h t h e h y p o t h e s i s t h a t i n c r e a s e s i n enzyme a c t i v i t i e s t e n d t o o c c u r when t h e enzymes a r e s t i m u l a t e d o v e r a p e r i o d o f time (such as t h a t may o c c u r i n c h r o n i c RSM t r e a t m e n t ) , and t h a t once t h e enzyme p a t t e r n c h anges, t h e p a t t e r n s o f s u b s t r a t e d i s p o s i t i o n t e n d t o become f i x e d (Chance ejt a l _ , 1965). T h i s i d e a on t h e \" b i o c h e m i c a l i m p r i n t i n g \" o f m e t a b o l i c e x p e r i e n c e i n c e l l s s i g n i f i e s a d a p t i v e i n c r e a s e s i n enzyme a c t i v i t i e s t h a t o c c u r a l o n g s p e c i f i c m e t a b o l i c pathways when t h e r e i s a l a r g e and s u s t a i n e d i n c r e a s e i n s u b s t r a t e t r a f f i c t h r o u g h them, and may w e l l e x p l a i n t h e h y p e r f u n c t i o n o f t h e t h y r o i d o f t h e -67-R-R and S-R b i r d s which c o n t i n u o u s l y a c c u m u l a t e i o d i n e o f the c i r c u l a t i o n f o r o r g a n i f i c a t i o n . The e x p e r i e n c e o f a c comodating l a r g e amounts o f s u b s t r a t e i s o f t e n r e f l e c t e d i n a c h a r a c t e r i s t i c i m p r i n t on the enzyme p a t t e r n o f t h e c e l l . M o r eover, changes i n enzyme p a t t e r n p r o b a b l y c o n s t i t u t e a l o n g - r a n g e m e t a b o l i c c o n t r o l d e v i c e which i s super-imposed on o t h e r r e g u l a t o r y mechanisms such as the h y p o t h a l a m u s - p i t u i t a r y - t h y r o i d a x i s o r t h e f e e d b a c k mechanisms, which o p e r a t e w i t h i n a much s h o r t e r time s c a l e . PERSISTENCE OF EFFECTS OF EARLY GOITROGENIC STIMULATION The v a r i o u s p arameters o f t h y r o i d a c t i v i t y show t h a t t h e e f f e c t s o f e a r l y g o i t r o g e n i c s t i m u l a t i o n o f t h e t h y r o i d g l a n d remained a p p a r e n t i n l a t e r l i f e . Thus t h y r o i d g l a n d s remained e n l a r g e d i n b i r d s which had been f e d RSM d u r i n g t h e growing p e r i o d and i n b i r d s g i v e n t h i o u r a c i l f o r o n l y t h e f i r s t f o u r weeks o f l i f e ( T a b l e 3 ) . T h y r o i d g l a n d s examined from b i r d s 67 weeks a f t e r w i t h d r a w a l o f RSM from t h e d i e t had t h e h i s t o l o g i c a l a p p e a r a n c e o f c o l l o i d g o i t e r . In t h e s e b i r d s the t h y r o i d f o l l i c l e s were l a r g e r and the h e i g h t o f t h e e p i t h e l i a l c e l l s l o w e r . T h i s p e r s i s t e n c e o f e f f e c t o f e a r l y o v e r - s t i m u l a t i o n o f t h e t h y r o i d i s i n agreement w i t h t h e f i n d i n g s o f Bakke et_ a T (1970) i n t h e r a t . In t h e r a t b r i e f p e r i n a t a l h y p o t h y r o i d i s m i n d u c e d by a d m i n i s t r a t i o n o f p r o p y l t h i o u r a c i l t o t h e p r e g n a n t dam and/or t o t h e neonate r e s u l t e d i n a p e r s i s t e n t l y e n l a r g e d t h y r o i d g l a n d w i t h l a r g e r f o l l i c l e s and l o w e r e p i t h e l i a l h e i g h t . These i n v e s t i g a t o r s r e p o r t e d a l s o t h a t p e r i n a t a l h y p o t h y r o i d i s m c a u s e d a d e l a y i n p u b e r t y . E a r l i e r e x p e r i m e n t s i n t h i s l a b o -r a t o r y has r e v e a l e d no e v i d e n c e t h a t t h e i n g e s t i o n o f g o i t r i n by c h i c k e n s up t o 24 weeks o f age a f f e c t e d t h e time o f s e x u a l m a t u r i t y o r r e p r o d u c t i v e c a p a c i t y when RSM was withdrawn a t t h i s t i m e (March e t _ a l _ , 1975). T h y r o i d a l e n l a r g e m e n t was r e l a t e d t o t h e l e n g t h o f t i m e o v e r which t h e g o i t r o g e n i c RSM -68-was f e d . The f o l l i c l e s i z e and p r o p o r t i o n o f e p i t h e l i a l t i s s u e were l o w e r i n t h e t h y r o i d g l a n d s o f t h e R-R b i r d s t h a n i n t h e S-R b i r d s a t 91 weeks o f age. The t o t a l amount o f i o d i n e i n the g l a n d o f t h e R-S b i r d s was g r e a t e r than t h a t i n the c o n t r o l S-S b i r d s b ut t h e c o n c e n t r a t i o n p er u n i t w e i g h t o f g l a n d was s i m i l a r ( e x p e r i m e n t 2 ) . C o n s i s t e n t w i t h t h i s o b s e r v a t i o n was t h e g r e a t e r t o t a l u p t a k e o f a s i n g l e dose o f r a d i o i o d i n e by t h e t h y r o i d g l a n d s o f t h e R-S b i r d s b u t a l o w e r uptake when e x p r e s s e d r e l a t i v e t o t h e w e i g h t o f g l a n d ( e x p e r i m e n t 3 ) . T h e r e f o r e i t can be s a i d t h a t , a l t h o u g h t he s i z e o f t h e t h y r o i d g l a n d i s p e r m a n e n t l y a f f e c t e d by e a r l y g o i t r o g e n i n t a k e , t h e t h y r o i d a c t i v i t y i s n o r m a l . In t h e e n l a r g e d g l a n d t h e e p i t h e l i a l t i s s u e r e sponds n o r m a l l y t o t h y r o t r o p i n and t h e c a p a b i l i t y f o r t h y r o i d hormone s y n t h e s i s t o meet t h e r e q u i r e m e n t s o f t h e b i r d s i s n o t a d v e r s e l y a f f e c t e d . The p r e s e n c e o f i n -c r e a s e d numbers o f e p i t h e l i a l c e l l s i n t h e t h y r o i d g l a n d s o f t h e b i r d s f e d g o i t r o g e n i c RSM d u r i n g t h e growing p e r i o d makes i t p o s s i b l e , once t h e i n h i -b i t i o n o f t h y r o i d hormone p r o d u c t i o n and/or s e c r e t i o n i s withdrawn, f o r t h e h y p e r p l a s t i c g l a n d t o meet t h e r e q u i r e m e n t s o f t h e body f o r t h y r o i d hormone a t a lower l e v e l o f a c t i v i t y ( i . e . a t a lower l e v e l o f t h y r o t r o p i n s t i m u l a t i o n ) . I t has been shown t h a t t h y r o i d a l e p i t h e l i a l c e l l s , f o l l o w i n g e l i m i n a t i o n o f t h y r o t r o p i n s t i m u l a t i o n , c o n t i n u e t o t r a p i o d i n e and t o s e c r e t e t h y r o i d hormone i n t o t h e f o l l i c u l a r lumen f o r some t i m e a t l e a s t , whereas t h e r e l e a s e o f t h y -r o i d hormone i n t o t he c i r c u l a t i o n s t o p s a l m o s t i m m e d i a t e l y (Bjorkman e t a l , 1974). The f a c t t h a t t o t a l i o d i n e was h i g h e r and t h a t t h e r e were more t h y -r o n i n e s p r e s e n t i n t h e g l a n d s o f RSM-fed b i r d s than i n SBM-fed b i r d s i n d i c a t e s t h a t e n d o c y t o s i s and r e l e a s e o f t h y r o i d hormone i n t o t h e c i r c u l a t i o n were r e t a r d e d i n the f o r m e r . In R-S b i r d s w i t h d r a w a l o f g o i t r i n a t 24 weeks o f age -69-was f o l l o w e d by c o l l o i d g o i t e r because t h e i o d i n e c o n c e n t r a t i o n by t h e g l a n d c o n t i n u e d a t a f a s t e r r a t e than d i d t h e r e l e a s e o f i o d i n e from t h e g l a n d . T h i s i s d i f f e r e n t from t h e f i n d i n g o f A k i b a and Matsumoto (1973) on t h e complete r e c o v e r y o f t h y r o i d f u n c t i o n w i t h i n a s h o r t time a f t e r w i t h d r a w a l o f g o i t r i n i n growing c h i c k s . A p e r s i s t e n t d i f f e r e n c e i n i o d i n e m e t a b o l i s m i n d u c e d by t h e f e e d i n g o f RSM t o b i r d s d u r i n g t h e growing p e r i o d was s t i l l e v i d e n t more t h a n a y e a r l a t e r i n t h e l e s s e r amount o f a s i n g l e dose o f r a d i o i o d i n e d e p o s i t e d i n the d e v e l o p i n g ova o f t h e s e b i r d s compared t o c o n t r o l b i r d s which had neven been f e d a g o i t r o g e n i c d i e t . T h i s a s p e c t o f t h e s t u d y ( t o be d i s c u s s e d l a t e r ) p r o v i d e s a d d i t i o n a l e v i d e n c e t h a t the t h y r o i d g l a n d m a i n t a i n s a h e i g h t e n e d c a p a c i t y f o r uptake o f i o d i n e as a r e s u l t o f e a r l y o v e r - s t i m u l a t i o n . The mechanism o f the p e r s i s t e n c e o f e a r l y s t i m u l a t i o n j-by g o i t r o -gens o f t h e t h y r o i d g l a n d i s unknown. The W o l f f - C h a i k o f f e f f e c t , which com-monly e x p l a i n s t h e b l o c k i n g e f f e c t o f i n t r a t h y r o i d a l i o d i d e o n i b i o s y n t h e s i s o f t h e t h y r o i d hormones when i o d i n e - d e f i c i e n t r a t s a r e r e - f e d a h i g h i o d i n e d i e t , i s c h a r a c t e r i z e d by a v e r y h i g h i o d i d e p e r c e n t a g e o f i n t r a t h y r o i d a l r a d i o a c t i v i t y , h i g h l a b e l e d MIT/DIT r a t i o , and absence o f l a b e l e d i o d o t h y r o n i n e s . N o r m a l l y t h i s b l o c k i n g e f f e c t o f l a r g e c o n c e n t r a t i o n s o f i o d i d e on hormone s y n t h e s i s i s a t r a n s i e n t phenomenon. T h e r e i s p r o b a b l y an autonomous a d a p t a -t i o n o f the t r a n s p o r t mechanism t o t h e h i g h l e v e l o f i o d i d e i n which t h e r e i s a r a p i d l o s s o f t h y r o i d a l c a p a c i t y t o c o n c e n t r a t e i o d i d e . In t h e R-S b i r d s i n t h e p r e s e n t s t u d y , i s i t p o s s i b l e t h a t t h i s a d a p t a t i o n mefchanism i s i m p a i r e d by g o i t r o g e n i c RSM so t h a t c o l l o i d g o i t e r r e s u l t s ? A l t h o u g h t h e W p l f f -C h a i k o f f e f f e c t i s not e v i d e n t i n t h e t h y r o i d h y d r o l y z a t e s o f t h e s e b i r d s , d e f e c t s i n t h e a u t o r e g u l a t i o n system i n t h e s e g l a n d s may s t i l l r emain a pos-s i b l e mechanism whereby e f f e c t s o f e a r l y g o i t r o g e n i c s t i m u l a t i o n m a y . p e r s i s t . -70-I t i s t h e r e f o r e c o n c l u d e d t h a t t h e e f f e c t s o f e a r l y g o i t r o g e n i c s t i m u l a t i o n o f t h e t h y r o i d g l a n d p e r s i s t a f t e r w i t h d r a w a l o f t h e g o i t r o g e n . The t h y r o i d g l a n d which has been o v e r - s t i m u l a t e d c o n t a i n s l a r g e r t h a n normal numbers o f e p i t h e l i a l c e l l s and a c c o r d i n g l y l e s s a c t i v i t y o f i n d i v i d u a l c e l l s i s r e q u i r e d t o m a i n t a i n normal l e v e l s o f hormone s e c r e t i o n . EFFECTS OF ALTERATIONS IN MATERNAL THYROID METABOLISM ON EMBRYONIC THYROID DEVELOPMENT IN THE CHICK S i n c e eggs l a i d by b i r d s f e d RSM c o n t a i n e d l e s s i o d i n e than when SBM was f e d ( e x p e r i m e n t 6A), t h e h y p o t h e s i s was t e s t e d t h a t , i n t h e g o i t r o u s b i r d s , the i n c r e a s e d t h y r o i d a l uptake o f d i e t a r y i o d i n e from t h e c i r c u l a t i o n l i m i t e d t h e amount a v a i l a b l e f o r d e p o s i t i o n i n the d e v e l o p i n g o v a . To e l i m i n a t e t h e p o s s i b i l i t y t h a t some component o f RSM i n t e r f e r e d w i t h a b s o r p -t i o n o f d i e t a r y i o d i n e from the i n t e s t i n e t h e d i s t r i b u t i o n o f i n j e c t e d r a d i o i o d i n e between t h e o v a r i a n f o l l i c l e s and the t h y r o i d g l a n d was compared i n t h e b i r d s f e d t h e SBM and RSM d i e t s ( e x p e r i m e n t 7 ) . The d a t a i n T a b l e 12 c l e a r l y show t h a t the d e v e l o p i n g ova i n the RSM-fed b i r d s c o n t a i n e d l e s s o f t h e i n j e c t e d r a d i o i o d i n e t h a n ova i n the SBM-fed b i r d s . I t i s a p p a r e n t from F i g u r e 2 t h a t t h e r a t e o f i o d i n e u p t a k e by t h e d e v e l o p i n g f o l l i c l e i s p r o -p o r t i o n a l t o t h e t o t a l r a t e o f t r a n s f e r o f a l l y o l k components from t h e c i r c u l a t i o n i n t o the f o l l i c l e . The amount o f i o d i n e p r e s e n t i n t h e f o l l i c l e i s t h e r e f o r e a f u n c t i o n o f both t h e r a t e o f t h e f o l l i c u l a r growth and t h e c o n c e n t r a t i o n o f i o d i n e i n t h e plasma. In c o n t r a s t t o t h e d e v e l o p i n g o v a , t h e t h y r o i d g l a n d s o f t h e RSM-fed b i r d s c o n t a i n e d more r a d i o i o d i n e 22 hours a f t e r i n j e c t i o n than d i d t h o s e o f t h e SBM-fed b i r d s . D e t a i l s o f the d i s t r i -b u t i o n o f the r a d i o a c t i v i t y among d i f f e r e n t i o d i n a t e d t h y r o i d a l compounds have a l r e a d y been d i s c u s s e d . - 7 1 -P r e l i m i n a r y t e s t s i n t h i s l a b o r a t o r y ( u n p u b l i s h e d d a t a ) have i n d i c a t e d t h a t most o f the i o d i n e p r e s e n t i n eggs i s i n i t s i n o r g a n i c f o r m . I f t h i s i s t r u e , i t w i l l be i n agreement w i t h t h e p o s t u l a t i o n t h a t when more i o d i n e i s t r a p p e d by t h e t h y r o i d g l a n d from t h e e x t r a t h y r o i d a l i n o r g a n i c p o o l , l e s s o f t h i s element w i l l be a v a i l a b l e f o r d e p o s i t i o n i n the d e v e l o p i n g ova. To t e s t t h i s h y p o t h e s i s , i t i s t h o u g h t t h a t i f the d i e t a r y i o d i n e i n t a k e o f t he RSM-fed b i r d s i s i n c r e a s e d t o o v e r t h a t can be a c c u m u l a t e d by t h e t h y r o i d g l a n d , t h e n e t d e p o s i t i o n o f t h i s element i n t h e eggs w i l l be f u r t h e r enhanced d e s p i t e t he i n h i b i t o r y e f f e c t o f t h e g o i t r o g e n i n t h y r o i d m e t a b o l i s m . R e s u l t s f r o m e x p e r i m e n t 6B seem t o b e a r t h i s o u t . T h y r o i d w e i g h t s a t h a t c h i n g were compared i n t h e progeny o f c h i c k e n s t h a t had been f e d g o i t r o g e n i c RSM and SBM r e s p e c t i v e l y i n the growing and/or l a y i n g d i e t s ( e x p e r i m e n t 8). The t h y r o i d g l a n d s were h e a v i e s t i n the progeny o f b i r d s which had been f e d RSM i n both t h e growing and t h e l a y i n g d i e t s . T h e r e was, however, a c a r r y o v e r o f t h e e f f e c t o f f e e d i n g RSM d u r i n g t h e growing p e r i o d when t h e b i r d s were s h i f t e d t o a SBM d i e t a t m a t u r i t y . A l t h o u g h t h e a v e r a g e w e i g h t s o f t h e g l a n d s o f t h e progeny o f dams r e c e i v i n g SBM i n t h e l a y i n g and f e d RSM d u r i n g t h e growing p e r i o d were n o t s i g n i f i c a n t l y h e a v i e r than i n t h o s e from dams f e d SBM t h r o u g h o u t l i f e , t h e p r o p o r t i o n o f e p i t h e l i u m i n t h e g l a n d s was g r e a t e r from t he RSM-fed b i r d s . In o t h e r words, t h e s t i m u l u s t o the t h y r o i d g l a n d was e v i d e n c e d by an i n c r e a s e i n t h e r a t i o j o f e p i t h e l i a l t i s s u e t o c o l l o i d w i t h i n t h e t h y r o i d g l a n d s but was n o t s u f f i c i e n t t o r e s u l t i n i n c r e a s e d t h y r o i d w e i g h t . On the o t h e r hand, progeny from dams f e d RSM d u r i n g t he l a y i n g p e r i o d o n l y , i had s m a l l e r t h y r o i d g l a n d s than d i d t h o s e f e d RSM t h r o u g h o u t l i f e . The p e r c e n t a g e o f e p i t h e l i a l t i s s u e was s i m i l a r i n t h e g l a n d s o f the progeny from t h e s e two t r e a t m e n t s . -72-SUMMARY AND CONCLUSIONS From t h e d a t a o f t h e v a r i o u s : e x p e r i m e n t s d i s c u s s e d above, t h e f o l l o w i n g c o n c l u s i o n s c o n c e r n i n g t h e c h r o n i c e f f e c t s o f f e e d i n g g o i t r o g e n i c RSM t o b i r d s can be drawn. A n a l y s e s . o f J h e 7 , t h y r o i d g l a n d s showed t h a t t h e i o d i n e c o n t e n t o f t h e RSM-fed b i r d s ( e x p e r i m e n t 2) was s i g n i f i c a n t l y h i g h e r than t h a t i n t h e c o n t r o l . The c h r o n i c a l l y s t i m u l a t e d (R-R and S-R) b i r d s had a t h r e e - f o l d i n c r e a s e i n t h e i r t h y r o i d a l i o d i n e c o n t e n t w h i l e even t he R-S group (which had o n l y been f e d g o i t r o g e n i c RSM d u r i n g t h e growing p e r i o d ) had o v e r t w i c e the t h y r o i d a l i o d i n e c o n t e n t o f the c o n t r o l b i r d s . T h i s was m a i n l y due t o t h e g o i t r o u s s t a t e o f the RSM-stimulated g l a n d s . The c h r o n i c a l l y - s t i m u l a t e d t h y r o i d g l a n d s showed a s e v e n - t o t e n - f o l d h y p e r t r o p h y , w h i l e t h e R-S group had an a v e r a g e t h y r o i d w e i g h t d o u b l e t h a t o f t h e c o n t r o l ( e x p e r i m e n t 1 ) . H i s t o l o g i c a l d a t a ( e x p e r i m e n t 1) c o n f i r m e d t h e a c t i v e l y f u n c -t i o n i n g s t a t e o f t h e t h y r o i d g l a n d s o f t h e b i r d s on c h r o n i c RSM and/or SBM t r e a t m e n t s . The t h y r o i d s o f the R-R and S-R b i r d s showed c o l l o i d g o i t e r r a t h e r t h a n a t r o p h y t h a t would sometimes be e x p e c t e d from c h r o n i c g o i t r o g e n i c t r e a t m e n t s , c o u p l e d w i t h i n c r e a s e d a v e r a g e e p i t h e l i a l h e i g h t s and f o l l i c l e d i a m e t e r s i n t h e t h y r o i d g l a n d s o f b i r d s i n t h e s e g r o u p s . T h i s gave s u p p o r t t o t h e h y p o t h e t i c a l p i c t u r e o f a s t i m u l a t e d t h y r o i d g l a n d o f the c h r o n i c a l l y R S M - s t imulated b i r d s a c t i v e l y a c c u m u l a t i n g i o d i n e from t he b l o o d . On-the.etfcer hand, t h e t h y r o i d g l a n d s o f the b i r d s f e d c h r o n i c a l l y w i t h g o i t r o g e n i c RSM were found t o be a c c u m u l a t i n g r a d i o i o d i n e , when 1-131 upt a k e was measured a t d i f f e r e n t t ime i n t e r v a l s a f t e r a d m i n i s t r a t i o n o f a -73-s i n g l e dose i n t r a v e n e o u s l y ( e x p e r i m e n t 3). T h i s f o u r - t o f i v e - f o l d i n c r e a s e i n t h y r o i d a l r a d i o i o d i n e u p t a k e c o u l d a c c o u n t f o r t h e d e c r e a s e i n i o d i n e d e p o s i t i o n i n t h e eggs o f the RSM-fed b i r d s . The s i t e o f i n h i b i t i o n by g o i t r o g e n i c RSM on i n t r a t h y r o i d a l i o d i n e m e t a b o l i s m o f t h e c h r o n i c a l l y t r e a t e d b i r d s was i n v e s t i g a t e d ( e x p e r i -ment 4). I t was f o u n d t h a t a l t h o u g h t h e r e a p p e a r e d t o be i n h i b i t i o n a t an i n t e r m e d i a r y s t e p ( s ) i n t h y r o i d hormone b i o s y n t h e s i s as d e m o n s t r a t e d by an a l t e r a t i o n o f the i o d o t y r o s i n e r a t i o , t h e p r o d u c t i o n o f i o d o t h y r o n i n e s was e s t i m a t e d t o be adequate as j u d g e d from t h e i r p e r c e n t a g e s i n t h e g o i t r o u s t h y r o i d g l a n d s o f the RSM-fed b i r d s . A more p r o b a b l e i n h i b i t o r y s i t e m i g ht be a t the r e l e a s e mechanism o f t h e s e hormones from the t h y r o g l o b u l i n . An e x p e r i m e n t ( e x p e r i m e n t 5) t o d e t e r m i n e t h e r e s p e c t i v e r a t e s o f t h y r o g l o -b u l i n h y d r o l y s i s by t h e e x t r a n e o u s enzyme pronase f a i l e d t o r e v e a l any d i f f e r e n c e between the SBM- and RSM-fed b i r d s . However, the p o s s i b i l i t y t h a t t h e r e m i g h t be a d i f f e r e n c e i n t h e t h y r o g l o b u l i n p r o t e o l y s i s r a t e s by endogenous t h y r o i d enzymes between the two groups o f b i r d s i s s t i l l open t o f u r t h e r i n v e s t i g a t i o n . From t h e above d a t a , i t was h y p o t h e s i z e d t h a t t h e g o i t r o u s t h y r o i d g l a n d s o f the c h r o n i c a l l y RSM-fed b i r d s were a c t i v e l y a c c u m u l a t i n g i o d i n e from the c i r c u l a t i o n because o f a p o s s i b l e i n h i b i t o r y s t e p i n t h y r o i d hormone b i o s y n t h e s i s and/or r e l e a s e , so t h a t l e s s o f t h e element was a v a i l a b l e f o r d e p o s i t i o n t o t h e eggs. The y o l k o f eggs from c h r o n i c a l l y RSM-fed b i r d s was i n d e e d f o u n d t o be s i g n i f i c a n t l y l o w er i n t o t a l i o d i n e c o n t e n t than t h o s e from t h e c o n t r o l ( e x p e r i m e n t 6A). S i n c e t h e i o d i n e c o n c e n t r a t i o n i n t h e a l b u m i n was r e g a r d e d as t o o low t o be o f s i g n i f i c a n c e i n the p r e s e n t i s s u e , i t was c o n c l u d e d t h a t -74-the c h r o n i c a l l y RSM-fed b i r d s d e p o s i t e d l e s s i o d i n e i n t h e i r eggs. T h i s was f u r t h e r s u p p o r t e d by a r a d i o i o d i n e s t u d y ( e x p e r i m e n t 7) which demons-t r a t e d a d e c r e a s e i n the d e p o s i t i o n o f 1-131 i n t h e d e v e l o p i n g o v a r i a n f o l l i c l e s o f t h e s e b i r d s compared t o the c o n t r o l . S i n c e t h e r e was no i n h i b i t i o n by g o i t r o g e n i c RSM on i o d i n e a b s o r p t i o n i n t h e g a s t r o i n t e s t i n a l t r a c t because t h e r a d i o i o d i n e was a d m i n i s t e r e d i n t r a v e n o u s l y , t h i s d i f f e r e n c e i n i o d i n e d e p o s i t i o n i n the eggs o f t h e RSM- and SBM-fed b i r d s c o u l d n o t be due t o a d i f f e r e n c e i n i o d i n e l e v e l p i i r r r i d i e t a r y i n t a k e . The RSM-supplemented d i e t a c t u a l l y c o n t a i n e d a s l i g h t l y h i g h e r c o n c e n t r a t i o n o f i o d i n e t h a n t h e SIBM-supplemented r a t i o n f e d t o t h e b i r d s ( e x p e r i m e n t 2 ) . T h a t t h i s d e c r e a s e o f i o d i n e d e p o s i t i o n i n the eggs o f the RSM-fed b i r d s c o u l d be due t o a more r a p i d e x c r e t i o n o f t h i s e l e m e n t was a l s o u n l i k e l y , s i n c e u n p u b l i s h e d d a t a from t h i s l a b o r a t o r y showed t h a t t h e r e was p r a c t i c a l l y no d i f f e r e n c e i n the d e i o d i n a t i o n o r e x c r e t i o n r a t e J . when e x t r a n e o u s r a d i o a c t i v e t r i i o d o t h y r o n i n e was a d m i n i s t e r e d t o t h e SBM-o r RSM-fed b i r d s . T h y r o i d a l h y p e r t r o p h y and a l t e r e d t h y r o i d a l c h a r a c t e r i s t i c s were found i n t h e progeny o f the RSM-fed dams. I t i s c o n c l u d e d t h e r e f o r e t h a t t h e t h y r o i d a l e n l a r g e m e n t o b s e r v e d i n the progeny o f c h i c k s from dams f e d g o i t r o g e n i c RSM i s due t o a low c o n c e n t r a t i o n o f i o d i n e i n t h e eggs. 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E x p e r i m e n t 1: S t a t i s t i c a l a n a l y s i s comparing t h e a v e r a g e w e i g h t s o f t h y r o i d g l a n d s i n White Leghorn p u l l e t s (Shaver s t r a i n ) f e d RSM and/or SBM, 91 weeks o f age. i ) A n a l y s i s o f v a r i a n c e : S o u r c e o f v a r i a t i o n d f MS F P Among groups 3 23719.7 48.0 < 0 . 0 l W i t h i n groups 36 493.9 T o t a l 39 i i ) D u n c a n ! s m u l t i p l e range t e s t : D i e t a r y t r e a t m e n t S-S R-S S-R R-R Ave. t h y r o i d wt. mg/100 body wt. 10.7 23.5 84.9 112.5 Duncan's comparison P < 0 . 0 1 ! a a b c i i i ) S e p a r a t e comparison o f S-S and R-S g r o u p s : Source o f v a r i a t i o n d f MS F P Among groups 1 819.2 39.4 < 0 . 0 i t W i t h i n groups 18 20.8 T o t a l 19 -96-T a b l e 2 ( A ) . E x p e r i m e n t 1: S t a t i s t i c a l a n a l y s i s comparing t h e a v e r a g e w e i g h t s o f t h y r o i d g l a n d s i n White Leghorn p u l l e t s ( D e K a l b - s t r a i n ) f e d RSM and/or SBM, 99 weeks o f age. i ) A n a l y s i s o f v a r i a n c e : S o u r ce o f v a r i a t i o n d f MS F P Among groups 3 35005.0 20.7 <0.01 W i t h i n groups 36 1693.0 T o t a l 39 i i ) Duncan's m u l t i p l e range t e s t : D i e t a r y t r e a t m e n t S-S R-S S-R R-R Ave. t h y r o i d wt. mg/100 g body wt. 11.2 28.4 103.4 149.4 Duncan's co m p a r i s o n P < 0 . 0 l „ a a b c i i i ) S e p a r a t e comparison o f S-S and R-S g r o u p s : Source o f v a r i a t i o n d f MS F P Among groups 1 1477.5 69.7 * 0 . 0 L W i t h i n groups 18 21.2 T o t a l 19 -97-T a b l e 3 ( A ) . E x p e r i m e n t 1: S t a t i s t i c a l a n a l y s i s comparing t h e av e r a g e t h y r o i d w e i g h t s (g) o f White Leghorn p u l l e t s ( S h a v e r s t r a i n ) a t 91 weeks o f age. i ) A n a l y s i s o f v a r i a n c e : S o u r c e o f v a r i a t i o n d f MS F P Among groups 3 4.5067 44.4 <0.0J.. W i t h i n groups 16 0.1016 T o t a l 19 i i ) Duncan's m u l t i p l e range t e s t : D i e t a r y t r e a t m e n t S-S R-S S-R R-R Ave. t h y r o i d w t . ( g j 0.1877 0.5175 1.3930 2.3027 Duncan's c o m p a r i s o n P < 0.0'J : a a b c i i i ) S e p a r a t e comparison o f S-S and R-S g r o u p s : S o u r c e o f v a r i a t i o n d f MS F P Among groups 1 0.2720 11.29 <0.0-L W i t h i n groups 8 0.0241 T o t a l 9 -98-T a b l e 4 ( A ) . Ex p e r i m e n t 1: S t a t i s t i c a l a n a l y s i s comparing t h e a v e r a g e e p i t h e l i a l h e i g h t s i n t h y r o i d g l a n d s o f White Leghorn p u l l e t s a t 91 weeks o f age. i ) A n a l y s i s o f v a r i a n c e : S o u r c e o f v a r i a t i o n d f MS F P Among groups 3 28.050 11.53 <0.01 W i t h i n gorups 16^ 2.433 T o t a l 19 i i ) Duncan's m u l t i p l e range t e s t : Di e t a r y t r e a t m e n t R-S S-S S-R R-R Ave. e p i t h e l i a l h e i g h t s , ja 5.61) 7.13 10.11 10.65 Duncan's comparison P < 0 . 0 i : a a b b -99-T a b l e 5 ( A ) . Ex p e r i m e n t 1: S t a t i s t i c a l a n a l y s i s comparing t h e aver a g e f o l l i c l e d i a m e t e r i n t h y r o i d g l a n d s o f White Leghorn p u l l e t s a t 91 weeks o f age. i ) A n a l y s i s o f v a r i a n c e : Source o f v a r i a t i o n d f MS F P Among groups 3 12905 11.44 <0.0i»! W i t h i n g roups 16_ 1128 T o t a l 19 i i ) Duncan's m u l t i p l e range t e s t : D i e t a r y t r e a t m e n t S-S R-S S-R R-R Ave. f o l l i c l e d i a m e t e r , mm .108 .167 .196 .227 Duncan's comparison P< 0.05'.\". a b bc c -100-T a b l e 6 ( A ) . E x p e r i m e n t 1: S t a t i s t i c a l a n a l y s i s comparing t h e p e r c e n t a g e e p i t h e l i u m i n t h y r o i d g l a n d s o f White Leghorn p u l l e t s a t 91 weeks o f age. i ) A n a l y s i s o f v a r i a n c e : S o u r c e o f v a r i a t i o n d f MS F P Among groups 3 111.44 2.99 ^ 0 . 0 5 W i t h i n groups 16_ 37.32 T o t a l 19 i i ) Duncan's m u l t i p l e range t e s t : D i e t a r y t r e a t m e n t R-S R-R S-R S-S P e r c e n t a g e e p i t h e l i u m 14.2 15,1 20.1 24.4 Duncan's comparison a ab abc c P< O.OB: -101-T a b l e 7 ( A ) . Ex p e r i m e n t 1: S t a t i s t i c a l a n a l y s i s comparing t h e aver a g e e p i t h e l i a l w e i g h t s i n t h y r o i d g l a n d s o f White Leghorn p u l l e t s a t 91 weeks o f age. i ) A n a l y s i s o f v a r i a n c e : S o u r ce o f v a r i a t i o n d f MS F P Among groups 3 105615 18.92 <0.0-lI W i t h i n groups li5 5580 T o t a l 19 i i ) Duncan's m u l t i p l e range t e s t : Di e t a r y t r e a t m e n t S-S R-S S-R R-R Weight o f e p i t h e l i u m , mg 41.1 76.5 277.4 334.6 Duncan's comparison a a b b P^O.OII -102-T a b l e 8 ( A ) . E x p e r i m e n t 2: S t a t i s t i c a l a n a l y s i s comparing t h e aver a g e w e i g h t s o f t h y r o i d g l a n d s o f White Leghorn p u l l e t s f e d RSM o r SBM f o r d i f f e r e n t p e r i o d s o f t i m e . i ) A n a l y s i s o f v a r i a n c e : S o u r ce o f v a r i a t i o n : d f MS F P Among groups 3 3.8623 23.09 < 0 . 0 l W i t h i n groups 16_ 0.1673 T o t a l 19 i i ) Duncan's m u l t i p l e range t e s t : D i e t a r y t r e a t m e n t S-S R-S S-R R-R Ave. t h y r o i d w e i g h t s , g 0.2039 0.4733 1.5115 2.9781 Duncan's co m p a r i s o n P<0.0'5 a a b c -103-T a b l e 9 ( A ) . E x p e r i m e n t 2: S t a t i s t i c a l a n a l y s i s comparing t h e a v e r a g e i o d i n e c o n c e n t r a t i o n s i n t h y r o i d g l a n d s o f White Leghorn p u l l e t s . i ) A n a l y s i s o f v a r i a n c e : S o u r ce o f v a r i a t i o n d f MS F P Among groups 3 4.879 29.25 <0.01 W i t h i n groups 16^ 0.1668 T o t a l 19 i i ) Duncan's m u l t i p l e range t e s t : D i e t a r y t r e a t m e n t R-R S-R S-S R-S Ave. i o d i n e c o n c e n t r a t i o n , mg/g 0.89 1.08 2.65 2.73 Duncan's comparison P< 0.0.1':, a a b b -104-T a b l e 1 0 ( A ) . Experiment 2: S t a t i s t i c a l a n a l y s i s comparing t h e t o t a l i o d i n e c o n t e n t s i n t h y r o i d g l a n d s o f White Leghorn p u l l e t s . i ) A n a l y s i s o f v a r i a n c e : S o u r c e o f v a r i a t i o n d f MS F P Among groups 3 1.4305 25.8 < 0 . 0 l W i t h i n groups 16^ 0.0554 T o t a l 19 i i ) Duncan's m u l t i p l e range t e s t : D i e t a r y t r e a t m e n t S-S R-S S-R R-R T o t a l i o d i n e i n g l a n d , mg 0.55 1.25 1.65 1.72 Duncan's c o m p a r i s o n P^O.OSc a b c c -105-T a b l e 1 1 ( A ) . E x p e r i m e n t 3: S t a t i s t i c a l a n a l y s i s comparing t h e t o t a l t h y r o i d r a d i o a c t i v i t y 6 hours a f t e r a d m i n i s t r a t i o n o f t h e r a d i o i o d i n e . i ) A n a l y s i s o f v a r i a n c e : S o u r c e o f v a r i a t i o n d f MS F P Among groups 3 851.00 18.4 < 0.01 W i t h i n groups 16 46.26 T o t a l 19 i i ) Duncan's m u l t i p l e range t e s t : D i e t a r y t r e a t m e n t S-S R-S S-R R-R P e r c e n t a g e o f i n j e c t e d dose 9.08 13.55 30.14 35.36 Duncan's comparison P<0.01' a a b b -106-T a b l e 1 2 ( A ) . E x p e r i m e n t 3: S t a t i s t i c a l a n a l y s i s comparing t h e t o t a l t h y r o i d r a d i o a c t i v i t y 22 hours a f t e r a d m i n i s t r a t i o n o f t h e r a d i o i o d i n e . i ) A n a l y s i s o f v a r i a n c e : S o u r ce o f v a r i a t i o n d f MS F P Among groups 3 1731.54 40.5 <.0.01 W i t h i n groups 16. 42.72 T o t a l 19 i i ) Duncan's m u l t i p l e range t e s t : D i e t a r y t r e a t m e n t S-S R-S R-R S-R P e r c e n t a g e p f i n j e c t e d dose 7.83 10.87 40.57 42.40 Duncan's comparison P < 0 . 0 i : a a b b -107-T a b l e 1 3 ( A ) . Experiment 3: S t a t i s t i c a l a n a l y s i s comparing t h e t h y r o i d r a d i o a c t i v i t y (% i n j e c t e d dose/g t h y r o i d ) 6 hours a f t e r a d m i n i s t r a t i o n o f t h e r a d i o i o d i n e . i ) A n a l y s i s o f v a r i a n c e : S o u r c e o f v a r i a t i o n d f MS F P Among groups 3 933.54 10.40 <0.01 W i t h i n groups 16 89.73 T o t a l 19 i i ) Duncan's m u l t i p l e range t e s t : D i e t a r y t r e a t m e n t R-R R-S S-R S-S % i n j e c t e d dose per g t h y r o i d 15.41 23.13 26.98 47.18 Duncan's comparison P<0.01-' a a a b -108-T a b l e 1 4 ( A ) . E x p e r i m e n t 3: S t a t i s t i c a l a n a l y s i s comparing the t h y r o i d r a d i o a c t i v i t y {% i n j e c t e d dose/g t h y r o i d ) 22 hours a f t e r a d m i n i s t r a t i o n o f t h e r a d i o i o d i n e . i ) A n a l y s i s o f v a r i a n c e : S o u r c e o f v a r i a t i o n d f MS F P Among groups 3 376.34 3.88 <0.G5 W i t h i n groups _16 96.91 T o t a l 19 i i ) Duncan's m u l t i p l e range t e s t : D i e t a r y t r e a t m e n t R-R S-R R-S S-S % i n j e c t e d dos>e per g t h y r o i d 21.61 29.55 35.58 41.96 Duncan's comparison P < 0 . 0 5 ; a ab bc bc -109-T a b l e 1 5 ( A ) . E x p e r i m e n t 6A: S t a t i s t i c a l a n a l y s i s comparing t h e i o d i n e c o n c e n t r a t i o n s i n egg y o l k s from b i r d s on d i f f e r e n t d i e t a r y t r e a t m e n t s . i ) A n a l y s i s o f v a r i a n c e : S o u r ce o f v a r i a t i o n d f MS F P Among groups 2 50473 22.02 <0.01... W i t h i n g roups 18_ 2292 T o t a l 20 i i ) Duncan's m u l t i p l e range t e s t : D i e t a r y t r e a t m e n t S-R S-S R-S I o d i n e cone, i n egg y o l k , ppb 144 273 286 Duncan's comparison P < 0 . 0 l - a b b -110-T a b l e 1 6 ( A ) . E x p e r i m e n t 6B: S t a t i s t i c a l a n a l y s i s comparing t h e i o d i n e c o n c e n t r a t i o n s i n egg y o l k s from b i r d s on d i f f e r e n t d i e t a r y t r e a t m e n t s . i ) A n a l y s i s o f v a r i a n c e : S o u r c e o f v a r i a t i o n d f MS F P Among groups 3 43093 4.321 <0.05 W i t h i n g r o u p s 20_ 9973 T o t a l 23 i i ) Duncan's m u l t i p l e range t e s t : D i e t a r y t r e a t m e n t RSM RSM+I SBM SBM+I I o d i n e cone, i n egg y o l k , ppb 41 73 105 241 Duncan's c o m p a r i s o n P<0.01 a a a b i i i ) S e p a r a t i o n c o m p a r i s o n o f SMB and RSM g r o u p s : S o u r c e o f v a r i a t i o n d f MS F P Among groups 1 12352 10.78 < 0 . 0 l W i t h i n groups l f j 1145 T o t a l 11 T a b l e 1 7 ( A ) . E x p e r i m e n t 7: S t a t i s t i c a l a n a l y s i s comparing t h e av e r a g e t o t a l r a d i o i o d i n e i n 5 l a r g e s t o v a r i a n f o l l i c l 22 hours a f t e r a d m i n i s t r a t i o n . i ) A n a l y s i s o f v a r i a n c e : S o u r c e o f v a r i a t i o n d f MS F P Among groups 3 21.5 39.09 <0.01 W i t h i n groups 16_ 0.55 T o t a l 19 i i ) Duncan's m u l t i p l e range t e s t : D i e t a r y t r e a t m e n t S-R R-R R-S S-S % i n j e c t e d dose i n 5 l a r g e s t o v a r i a n f o l l i c l e s 2.624 2.830 5.796 6.717 Duncan's comparison P<0.01 a a b b i-3-ttIHDS—PERMCA3IL IT V A?i|) f-CUl. ICuL A'* 317F R E G R E S S I O N A N A L Y S I S , T H L OFPE'M.'C'. f V A ^ I A H I F IS L O T , PERM CONt»rri!|t . / f iM TO T IPLE REGRfcSSI 0\" I »:r-l'.(;LM)i M V A I' I A h L f; Hr GRFSSI ON CCEhT J C U NT S T A N D A R D E R R O R Of- T H L : RtGR. C O E F F . \" • P i - T T i l . Cut f F I C 11 'N r O F DfcTfcRuI'NATIOM VARIANCE RATIO (K} S.AKTA L (.1 r; A W r. A •0.982Sn3F-01 0.2708SSK+01 INTERCEPT STANDARD ERROR OF ESTIMATE -RESIDUAL \"VARIANCE MULTIPLE CORRELATION\" COEFFICIENT R SQUARED -VARrA^CE-RATTO—CF1 \" THE VARIABLE TO 8E 0 * I T T l D IS 0. 1 1 1586C-01 0, J.286Q4E + 00 •10.27 : -0.119S75E+0 1 = o.i634H00 0.923H8E-02 •275,2h 6 9 S . 3 0 -34 r.SQ' 23,u94 17.740 ~I 0 , 7 / i INTERCEPT TANHARD\"ERROR OF EST IHATE RESIDUAL VARIANCE MULTIPLE CORRELATION COEFFICIENT -R—STJARED VARIANCE RATIO CF) THE VARIABLE TO BE OMITTED IS -0 . 7 f i 8 5 6 5 E + 00 \".r E RATIO CF) = - 0 . 9 7 3 1 C 6 E + 0 0 = 0.1S6303E+UO ~s—\"0\"- 2 W S 0 7 E - ' 0 1 _ = 0 . 6 6 7 9 5 = 0 . 7 ^ 3 3 5 -= •-= S . A R E A WTT H 2 —AND 2D DEC R ETS~ 5 4 . 3 6 6 61 .060 H (t> r-» SO REGRESSION A N A L Y S I S , THE DEPENDENT VARIABLE IS LOG PERM C O N T R I B U T I O N TO M U L T I P L E R E G R E S S I O N INDEPENDENT — VARIABLE\" r L O G AREA REGRESSION \" C O E F F I C I E N T ' 0 .253555E + 00' INTCRCEPT STANDARD ERROR OF ESTIMATE -RESIDUAL\"VARIANCE\" MULTIPLE CORRELATION C O E F F I C I E N T R SQUARED -VXR17TNCE RATIO ( D : = - 0 . 8 0 1 9 9 4 E - 0 1 •-= 0 .294138E+00 \" = 0 . 8 6 5 1 7 I E - 0 1 ~\"T = 0 . 2 8 7 8 3 = 0 ,09284 = r.\"\"897 T I T H -5SIOW-7rN\"ATTSTS7 T^ C\" DEF»CEDENT vART*BLE\"\"rS—PERM-INDEPENDENT VARIABLE REGRESSION C O E F F I C I E N T S'.AREA A R F A * * 2 ~AREA**3\" 0 .561084E+00 - 0 . 2 7 8 9 2 1 E - 0 1 ~~0;3S\"2982EiT;r\" STANDARD ERROR OF THE — R E E R , - - C O E F F — \" 0 .184100E+00 PARTIAL COEFFIC IENT \"OF \"OE'TERMINAT I OF! . . . 8 . 2 6 VARIANCE RATIC\"CF} 1 . 897 T 3TN13 2T DTn5Rnrs~DF~FREED'OH— CONTRiauTICN TO MULTIPLE REGRESSION \"ST AWAR D ERHTJR-CF THE REGR, C O E F F , PTRTTAL\" COEFF IC IENT OF DETERMINATION V A R I A \\ C E RATIO CF) 0 .213811E+00 0, 152627E-01 \"0,3T3TJ2r-0T\" - 4 4 . 0 6 197 .33 - -99753~ 6 ,886 3 .340 1,271 W X -tf-a H S rt -sv-09 n (D rn CO H -O 3 Co 03 CO INTCRCEPT \"STAND A R D~C R R O R - O F ^ E S T I H A T E - RESIDUAL VARIANCE MULTIPLE CORRELATION C O E F F I C I E N T ~R~S OU'A R ETT : VARIANCE RATIO (F ) THE VARIABLE TO BE OMITTED IS = - 0 , S 9 2 9 5 S E + 0 0 -=— 0 .73 61 07E+ 0 0-= 0 ,617964E+00 0 , 7 3 3 1 2 \"= 0Y53747\" = A R E A * * 3 7', 359 WITH AND 19 DEGREES OF FREEDOM 1 f S-n-Bjkrys REGRESSION ANALYSIS, THE DEPENDENT VARIAHI.K i s L „ G PERM CONTRJhuT IPM TO ruLTCPLt\" REGRESSION I NOEPErJOEN T V A R I A rt L F S'.AREA L O G AREA REGRESSION COEF (• ICIb NT -0.H5704'Jfc-01 0.216407E+01 'STANDARD E R R O R S OF THE . REGR. COEFF, P'A»TT AC COEFFICIENT OF DETER\"I NATION V A R I A N C E R A T I O (F) INTCRCEPT STANDARD ERROR OF ESTIMATE -RrstntTAr-VA-RrATgcrr 0.141453E-01 0.363090E+00 37.95 25.20 MULTIPLE CORRELATION COEFFICIENT R SQUARED \"VAWTflNTC RATIO (h ) THE VARIABLE TO SE OMITTED IS = - 0 , 123046F_tyi = 0.250676EtU0 - 07628394F-TJT~ = 0.79407 = 0.63150 36,710 35.523 LOG AREA -ffTTH 2 AW 22—TTFGRCEt) Or FRETOTna-R E G R E S S I O N A N A L Y S I S , T H E D E P E N D E N T V A R I A H L F . I S L O G PERM CONTRIBUTION TO M I J U U P L F REGRESSION INDEPENDENT \"VTRTA-OCE— S . A R E A I N T E R C E P T STANDARD ERROR OF ESTIMATE ~ESlDUAL\" VATrrffwCE-REGRFSSION '\"CTjEFrrcTeMT— -0.823994E-02 STANDARD ERROR OF THE RETTR7 COEFT\"J— . PARTIAL COEFF- ICIENT OF •Ut.ItKMTKJtTTCm- VARIANCE RA i iu u y 0.832869E-02 MULTIPLE CORRELATION COEFFICIENT R SQUARED V T R T A N C E R A i i u c n = -0,117142C+00 = 0.396438F+00 = 0.157I63E+00 = 0.19103 0.03649 = 0T87T 3 , 6 5 0,871 T T H ~ 1 ^A'NO 2 3 D E G R E E S UF FREED07T REGRE^SrOfr-Am-YSIS-TTHC D E P E N D E N T V A R I A B L E ' S - P E R M CONTRIBUTION TO MULTIPLE REGRESSION INDEPENDENT VARIABLE REGRESSION C O E F F I C I E N T \"STAND A RT)—ERRCFT OF THE REGR, COEFF, S'.AREA A R K A A * 2 _ARE\"A i\"*3 -PSR7TSL~ COEFFICIENT OF DETERMINATION VARIANCE RATIO ( F ) I N T E R C E P T - S T A N D A R D - E R R O R \" O F ' T S T I M A T E R E S I D U A L VARIANCE M U L T I P L E CORRELATION C O E F F I C I E N T ~R~S CUT* R ED -V A R I A N C E RA T I O CF) THE V A R I A B L E TO BE OMITTED I S 0.216527E+00 •0.114076K-01 \"tr7T5'*nr29r-Trr 0.857160C-01 0.598289E-02 '57119723E-TJ3--120.55 324.45 ~-T517T9-= -0,625384E-U1 = 0.3n9C37E+00 = 0,l3fa780EtoO = 0.71212 = ~ r f 5 0 7 1 r 6 . 3 B 1 3 . 6 3 6 l.oOO s AREA * *3 7,202 *ITH AND 21 DEGREES OF FREEDOM PAGE H Co Tr-io O X T3 H 3 -»-3 rt 09 a -co— p. 3 To\"\" 3 0> CO CO REGRESSION A N A L Y S I S , THE DEPENDENT VAkJ ArtL.E I S LIT, PERM CONTRIBUTION -T ti v-; i L T J P L E RttiKr.SS-lON I N D E P E N D E N T V AH I A B L E S . A R E A L O G A R E A REGRESSION C O E F F I C I E N T • O . l C S l u E + O C 0 .279482E+01 I N T E R C E P T STANDARD ERROR OF ESTIMATE - R E S IDU AL~~V ARI\"ANCE~ M U L T I P L E CORRELATION C O E F F I C I E N T R SQUARED \"-VARIANCE RATTtT^fF) - 0 . 144323E+01 0 . 159572E + 00 0 . 2 5 4 6 3 3 E - 0 1 0 . 9 0 9 6 8 0 , 8 2 7 5 2 \\ STANDARD\" ERROR OF THE RFGR. C O E F F . 0 . 106617E-01 0 t 281865L+0O \" ? A R T T A L CUf.r F ICIENT OF DL T t R'' I (•' AT 101>' C.f-.95 15.80 THE V A R I A B L E TO BE OMITTED I S T27777-L0G AREA WITH AND ' - D E G R F F S \" O F \" F R E E D O M \" V A K I A N C F P A T ] D f F ) 105 .123 9 8 . 5 1 7 REGRESSION A N A L Y S I S , THE DEPENDENT V A R I A B L E I S LOG PERM INDEPENDENT - — V A R I A X L E -S'. A R E A R E G R E S S I O N C O E F F I C I E N T \" -• 0 . 9 2 6 4 3 2 E - 0 2 INTERCEPT' \" \\ . STANDARD ERRORS OF ESTIMATE - R E S T DO At V ARTA~NCE MULTIPLE CORRELATION C O E F F I C I E N T R SQUARED VARIANCE RATIO CF} = 0 . 2 0 0 2 2 5 E - 0 1 = 0 , 3 6 4 9 6 9 E + 0 0 \"=\"—OT 1 3 a 0 , 2 3 8 2 1 = 0 , 0 5 6 7 4 C O N T R I T I O N TO M U L T I P L E R E G R E S S I O N \"T73B\"3 WITH S T A N D A R D E R R O R Or THE \" R E G R ; \" c o E r F ' i 0 , 7 8 7 6 0 5 E - 0 2 PARTIAL COF.FF IC1F.NT \"OF \"DETER'1 IN ATI ON \" 5 .67 VARIANCE RATIO\"-CF\"} 1.384 T A\"NTT :3 mrcTfLEs UF FREEUDH~ -CO-\"DElTrNDENT VARTABtTE TS PERW-I N U E P E N D E N T V A R I A B L E R E G R E S S I O N C O E F F I C I E N T I N T E R C E P T -STANDARD-ERROR—OF E S T I M A T E \" R E S I D U A L VARIANCE M U L T I P L E CORRELATION C O E F F I C I E N T R SQUARED VARIANCE RATIO ( F } THE V A R I A B L E TO BE OMITTED I S S'.AREA A R E A * * 2 - A K E A W 3 -0.443172E+00 • 0 . 2 6 3 6 0 8 E - 0 1 -0-.4IB8\"03t-0\"3--0 .542931E+U0 O\",3S3969F + 00 0 . 147432F + 00 0 .30780 -0TG525JI C O N T R I B U T I O N TO M U L T I P L E R E G R E S S I O N \"\"STAND A PTJ—\"E RR D R\"~ OF T H E R E G R , C O E F F , 0. 1 14627E + 00 0.814305E-02 -0\",\"-ir>72^BE\"-0T-T X R T T\"A E -COEFFICIENT OF DETERMINATION • 2 4 6 . 1 8 64 t .31 •T27TCT-V'ARlAr.CE RATIO CF) 13 .146 WITH ANO = A R t A**3 DEGREES OF FREEDOM 14.947 10,480 \"5 .212 -116-T a b l e 2 2 ( A ) . Experiment 8: S t a t i s t i c a l a n a l y s i s comparing t h e a v e r a g e t h y r o i d w e i g h t s o f progeny when dams were 91 weeks o f age. i ) A n a l y s i s o f v a r i a n c e : S o u r ce o f v a r i a t i o n d f MS F P Among groups 3 .0000175 0.99 n.s. W i t h i n groups 76_ .0000177 T o t a l 79 -117-T a b l e 2 3 ( A ) . E x p e r i m e n t 8: S t a t i s t i c a l a n a l y s i s comparing t h e a v e r a g e p e r c e n t a g e e p i t h e l i u m i n t h y r o i d g l a r i d s o f progeny a t h a t c h i n g . i ) A n a l y s i s o f v a r i a n c e : S o u r ce o f v a r i a t i o n d f MS F P Among groups 3 179.50 3.77 <0.05 W i t h i n groups 16_ 47.58 T o t a l 19 i i ) Duncan's m u l t i p l e range t e s t : Di e t a r y t r e a t m e n t o f dam S-S R-S S-R R-R % t h y r o i d e p i t h e l i u m i n progeny 43.0 49.2 55.1 55.8 Duncan's c o m p a r i s o n P<0.05 a ab b b -118-T a b l e 2 4 ( A ) . E x p e r i m e n t 8: S t a t i s t i c a l a n a l y s i s comparing t he av e r a g e h e i g h t s o f t h y r o i d e p i t h e l i u m o f progeny o f b r i d s f e d RSM o r SBM f o r d i f f e r e n t p e r i o d s . i ) A n a l y s i s o f v a r i a n c e : S o u r c e o f v a r i a t i o n d f MS F P Among groups 3 1.369 2.894 <0.10 W i t h i n groups 16 0.473 T o t a l 19 i i ) Duncan's m u l t i p l e range t e s t : D i e t a r y t r e a t m e n t o f dam;.; S-S R-R R-S S-R Ave. h e i g h t s o f e p i t h e l i u m , JJ 3.864 4.708 4.761 5.092 Duncan's comparison P 4.0.05 a ab ab b R-S -120-Photnricrographs: (I60x) Thyroid gland R-R "@en ; edm:hasType "Thesis/Dissertation"@en ; edm:isShownAt "10.14288/1.0093250"@en ; dcterms:language "eng"@en ; ns0:degreeDiscipline "Poultry Science"@en ; edm:provider "Vancouver : University of British Columbia Library"@en ; dcterms:publisher "University of British Columbia"@en ; dcterms:rights "For non-commercial purposes only, such as research, private study and education. Additional conditions apply, see Terms of Use https://open.library.ubc.ca/terms_of_use."@en ; ns0:scholarLevel "Graduate"@en ; dcterms:title "Effects of chronic and early goitrogenic stimulation on thyroidal metabolism in the chicken and in the progeny"@en ; dcterms:type "Text"@en ; ns0:identifierURI "http://hdl.handle.net/2429/19277"@en .