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An analysis of the thyroid role in juvenile steelhead (Salmo Gairdneri Richardson) and factors responsible.. Eales, John Geoffrey 1963

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AN ANALYSIS OF THE THYROID ROLE IN JUVENILE STEELHEAD (SALMO GAJJKDNERI RICHARDSON) AND FACTORS RESPONSIBLE FOR ITS SEASONAL FLUCTUATION IN ACTIVITY  by  JOHN GEOFFREY EALES B.A., Oxford U n i v e r s i t y , 1959 M.Sc,  University  of B r i t i s h Columbia, 1961  A THESIS SUBMITTED IN PARTIAL FULFILMENT OF THE REQUIREMENTS FOR THE DEGREE OF DOCTOR OF PHILOSOPHY i n the Department of Zoology We accept t h i s t h e s i s as conforming t o t h e r e q u i r e d standard  Members o f t h e Department THE UNIVERSITY OF BRITISH COLUMBIA A p r i l , 1963  In the  this thesis  r e q u i r e m e n t s f o r an  British  mission  for. r e f e r e n c e  for extensive  p u r p o s e s may  be  of  w i t h o u t my  written  Department  of  by  in partial  d e g r e e at  the  the  this thesis  Head o f my  i s understood  permission.  Columbia,.  fulfilment  of  University  of  s h a l l make i t f r e e l y  I further  agr.ee t h a t for  or  c o p y i n g , or  shall  per-  scholarly  Department  that  for f i n a n c i a l gain  The U n i v e r s i t y o f B r i t i s h V a n c o u v e r 8, Canada.  the  Library  study.  copying of  It  this thesis  that  and  granted  representatives..  cation  advanced  Columbia, I agree  available  his  presenting  not  be  by publi-  allowed  The U n i v e r s i t y o f B r i t i s h Columbia ' FACULTY OF GRADUATE STUDIES PROGRAMME OF THE FINAL ORAL  PUBLICATIONS  EXAMINATION  FOR THE DEGREE OF A comparative study o f t h y r o i d f u n c t i o n i n migrant j u v e n i l e salmon. Can. J . Zool 41: ( i n p r e s s ) . 1963. The  DOCTOR OF PHILOSOPHY  of  t h y r o i d g l a n d and low temperature r e s i s t a n c e o f g o l d f i s h . Can. J . Z o o l . , 41: ( i n p r e s s ) , w i t h W.S. Hoar. 1963  JOHN GEOFFREY EALES  B.A., O x f o r d U n i v e r s i t y , 1959 M . S c , The U n i v e r s i t y o f B r i t i s h Columbia, 1961  TUESDAY, May 7, 1963 at 9:30 A.M. IN ROOM 3332, BIOLOGICAL SCIENCES .BUILDING  COMMITTEE,IN  CHARGE  Chairman: F.H. Soward J.R. Adams D.H. C h i t t y I. McT. Cowan W.S. Hoar , External  W.N. Holmes W.D. K i t t s V.J. Okulitch N.J. Wilimovsky.  Examiner: E.J.W. B a r r i n g t o n  Department  o f Zoology,  Nottingham  University  AN ANALYSIS OF THE THYROID ROLE IN JUVENILE STEELHEAD (SALMO GAIRDNERI RICHARDSON) AND FACTORS RESPONSIBLE . FOR ITS SEASONAL FLUCTUATION IN ACTIVITY ABSTRACT  GRADUATE STUDIES  I n v e s t i g a t i o n of f a c t o r s c o n t r o l l i n g s e a s o n a l changes i n t h y r o i d a c t i v i t y of j u v e n i l e s t e e l h e a d t r o u t (Salmo • g a i r d n e r i Richardson) i n f r e s h water r e v e a l e d p o s i t i v e c o r r e l a t i o n s between temperature and r a d i o i o d i n e a s s e s s ments of t h y r o i d a c t i v i t y . Temperature and t h y r o i d e p i t h e l i a l h e i g h t , however, showed n e g a t i v e c o r r e l a t i o n s . These c o r r e l a t i o n s were v e r i f i e d e x p e r i m e n t a l l y . In y e a r l i n g s , the t h y r o i d was r e f r a c t o r y to i n c r e a s i n g p h o t o p e r i o d (January to J u l y ) but two-year-old p o t e n t i a l migrants showed a p o s i t i v e response at the same season. Thus, the h i g h smolt t h y r o i d a c t i v i t y i s induced by the combined i n f l u e n c e o f r i s i n g temperature and i n c r e a s i n g spring photoperiod. Body mass ( l o g a r i t h m ) was i n v e r s e l y r e l a t e d to v a r i o u s 1^31 parameters ( l o g a r i t h m ) , so that s m a l l f i s h had h i g h er t h y r o i d a c t i v i t i e s than large f i s h . P r e c o c i o u s sexu a l m a t u r a t i o n of two-year-old male p a r r , i n c r e a s e d swimming e x e r c i s e and i n c r e a s e d s a l i n i t y were a s s o c i a t e d w i t h higher thyroid a c t i v i t y . Increase i n ambient 1*27 d_ p r e s s e d t h y r o i d a c t i v i t y and i n d i c a t e d that the a c t i v i t y of the g l a n d ( a s s e s s e d by current methods) i s p a r t l y a compensation f o r low I•'•27 a v a i l a b i l i t y . The h i g h t h y r o i d a c t i v i t y o f the smolt may be due p a r t l y to endemic g o i t r e e  Since low temperature and 8-hour day l e n g t h i n h i b i t e d the t h y r o i d a c t i v i t y of p o t e n t i a l migrants but d i d not prevent s i l v e r i n g , the r o l e of t h y r o x i n i n guanine depos i t i o n under n a t u r a l c o n d i t i o n s i s doubted; Possible radiohormone c a t a b o l i c s i t e s were l o c a t e d i n m e t a b o l i c a l l y a c t i v e t i s s u e s i n c l u d i n g gut, kidney, l i v e r and brain. These f i n d i n g s suggest a g e n e r a l r a t h e r than a t i s s u e - s p e c i f i c r o l e of t h y r o x i n i n metabolism. I t i s concluded that t h y r o x i n may have no s t i m u l a t o r y r o l e i n s m o l t i f i c a t i o n but r e f l e c t s i n s t e a d the t o t a l m e t a b o l i c demands on the t i s s u e s .  F i e l d of Study:  Zoology  Q u a n t i t a t i v e Methods i n Zoology Comparative P h y s i o l o g y Comparative E t h o l o g y Comparative I n v e r t e b r a t e Embryology Biology of Fishes Endocrinology Marine F i e l d Course F i s h e r i e s Seminar  Other  P.A. Larkin W.S. Hoar W.S. Hoar M.D.F. Udvardy C.V. Finnegan C.C., L i n d s e y W.N. Holmes P.A. Dehnel Staff  Studies:  Biochemistry  Faculty  of M e d i c i n e  ABSTRACT Investigation thyroid activity Richardson)  of f a c t o r s  of  juvenile  i n fresh water  c o n t r o l l i n g seasonal steelhead  revealed  trout  changes  (Salmo  positive  gairdneri  correlations  between  temperature  and r a d i o i o d i n e a s s e s s m e n t s o f t h y r o i d  ivity.  Temperature  and t h y r o i d e p i t h e l i a l h e i g h t ,  showed n e g a t i v e  correlations.  These  in  act-  however,  c o r r e l a t i o n s were  verified  experimentally. In yearlings photoperiod migrants  t h e t h y r o i d was r e f r a c t o r y  to  (January t o J u l y ) but t w o - y e a r - o l d  showed a p o s i t i v e  response  at the  increasing potential  same s e a s o n .  the high smolt t h y r o i d a c t i v i t y  i s i n d u c e d by t h e  influence  and i n c r e a s i n g  of r i s i n g temperature  Thus,  combined  s p r i n g photon  period. B o d y mass J13>1  parameters  (logarithm)  (logarithm),  thyroid activities of two-year-old increased  was i n v e r s e l y so t h a t  than large  male p a r r ,  fish.  to  s m a l l f i s h had Precocious  increased  s a l i n i t y were a s s o c i a t e d  related  various higher  sexual  maturation  swimming e x e r c i s e  and  with higher thyroid  activity.  127  Increase  i n ambient  indicated that methods)  I  ' depressed  the a c t i v i t y of  the g l a n d  of the  and  ( a s s e s s e d by  i s p a r t l y a compensation f o r low l  The h i g h t h y r o i d a c t i v i t y endemic  thyroid activity  1 2  7  current  availability.  s m o l t may be due p a r t l y  to  goitre.  Since  low temperature  and 8 - h o u r d a y l e n g t h (  i n h i b i t e d the  thyroid activity silvering,  of p o t e n t i a l m i g r a n t s but d i d not  the r o l e of t h y r o x i n i n guanine  n a t u r a l conditions i s doubted. s i t e s were l o c a t e d gut,  kidney,  l i v e r and b r a i n .  is  concluded that  tissues.  deposition  under  tissues  These f i n d i n g s role  catabolic  including  suggest a  general  of t h y r o x i n i n metabolism.  t h y r o x i n may h a v e no s t i m u l a t o r y r o l e  s m o l t i f i c a t i o n but r e f l e c t s on t h e  prevent  P o s s i b l e radiohormone  in metabolically active  rather than a t i s s u e - s p e c i f i c It  i i i -  instead the t o t a l metabolic  in  demands  ACKNOIiVLEDGEMENTS  I wish t o express my Hoar, P. R. S. C , c r i t i c i s m and  s i n c e r e g r a t i t u d e t o P r o f e s s o r W.  S.  Department of Zoology, f o r h i s s t i m u l a t i o n ,  e n t h u s i a s t i c guidance throughout t h i s p r o j e c t .  I am a l s o indebted t o P r o f e s s o r s J . R. Adams, W.  N. Holmes,  N. J . Wilimovsky (Department of Zoology) and P r o f e s s o r W.  D.  Kitts  I  (Animal S c i e n c e )  for their constructive criticism.  f u r t h e r wish t o acknowledge the s e v e r a l graduate students from, whom I gained  so much d u r i n g i n f o r m a l d i s c u s s i o n s .  Dr. K. A. E v e l y n of the S t r o n g Laboratory, General H o s p i t a l , supplied.the r a d i o i o d i d e . Smith and J . G. Terpenning arranged  Vancouver  Messrs. S.  B.  ( B r i t i s h Columbia P i s h and Game)  f o r the p r o v i s i o n of steelhead from the C u l t u s Lake  Hatchery, where the experimental months by Mr.  f i s h were tended f o r s e v e r a l  Pat M a r t i n and h i s s t a f f .  A g r e a t number of  h i s t o l o g i c a l p r e p a r a t i o n s were m e t i c u l o u s l y made by Miss S. Tabata.  The  s t a t i s t i c a l a n a l y s e s were f a c i l i t a t e d by use  IBM Computing Centre, U n i v e r s i t y of B r i t i s h Columbia and c o - o p e r a t i o n of the s t a f f there was a l l these people  g r e a t l y appreciated.  I wish t o express my  t h e i r various services;  without  of the To  deepest a p p r e c i a t i o n f o r  them t h i s  study would have  been i m p o s s i b l e . P i n a n c i a l a s s i s t a n c e i n the form of a F i s h e r i e s Research Board S t u d e n t s h i p  enabled me  t o c a r r y out t h i s  study.  TABLE OF CONTENTS PAGE INTRODUCTION  1  MATERIALS AND METHODS  4  1. L i v i n g M a t e r i a l s  4  2.  Radioiodide I n j e c t i o n  4  3.  Blood Sampling and S e p a r a t i o n o f P r o t e i n bound and Inorganic R a d i o i o d i n e . . .  6  4.  Body and T h y r o i d  8  5.  Thyroid Histology  6.  S t a t i s t i c a l Treatment  .  .  .  .  .  .  .  .  9 11  RESULTS ' I.  12 COMPARISON  OF YEARLING STEELHEAD WITH DIFFERENT  RATES OF RADIOIODINE METABOLISM  .  .  .  .  12  i  1.  E x t r a t h y r o i d a l and T h y r o i d a l Clearance o f Plasma ±131  13  2.  Experimental Measurement of T h y r o i d Accumulation  21  3.  ll?l  E s t i m a t i o n o f Radiohormone Output Using 27  the C o n v e r s i o n R a t i o II.  SEASONAL CHANGES IN RADIOIODINE METABOLISM AND THE HISTOLOGICAL APPEARANCE OF THE THYROID IN JUVENILE STEELHEAD HELD IN FRESH WATER  .  .  30  1.  I ? ! Excretion  30  2.  T h y r o i d Uptake  34  3.  Conversion R a t i o  34  4.  H i s t o l o g i c a l Changes  33  1  V  -  TABLE OF CONTENTS  (Continued) PAGE  III.  EXPERIMENTAL INVESTIGATION OF THE INFLUENCE OF TEMPERATURE ON THE HISTOLOGICAL APPEARANCE OF THE THYROID AND RADIOIODINE METABOLISM IN 36  YEARLING STEELHEAD PARR IV.  EXPERDIENTAL INVESTIGATION OF TEMPERATURE AND PHOTOPERIOD INFLUENCE ON THYROID CELL HEIGHT AND RADIOIODINE METABOLISM OF STEELHEAD  V. VI.  YEARLINGS (January t o J u l y )  44  EFFECT OF BODY MASS ON RADIOIODINE METABOLISM  35  EXPERIMENTAL INVESTIGATION OF THE INFLUENCE OF SEASONAL CHANGE IN INCREASING PHOTOPERIOD  AND  INCREASING TEMPERATURE ON THE RADIOIODINE METABOLISM AND THYROID CELL HEIGHT OF TWO-YEAR-  VII.  OLD STEELHEAD  6l  1.  l l ? l Excretion  62  2.  T h y r o i d Uptake o f I ? !  64  3.  Conversion R a t i o  67  4.  C e l l Height  70  1  INFLUENCE OF PRECOCITY OF MALE PARR ON THYROID 70  ACTIVITY VIII.  INFLUENCE OF EXERCISE ON I ? METABOLISM OF 1  1  72  STEELHEAD IX.  THE INFLUENCE OF CHEMICAL CHANGES (SALINITY AND IODINE LEVELS OF THE MEDIA) ON I ^ ! METABOLISM 1  OF STEELHEAD AND CHUM SALMON (Oncorhynchus keta) 74  - v iTABLE OF CONTENTS  (Continued) PAGE  X.  XI.  THE RELATIONSHIP BETWEEN RADIQIODINE METABOLISM AND SILVERING  85  PERIPHERAL SITES OF RADIOHORMONE CATABOLISM  89  DISCUSSION I.  95  COMPARISON OF PARAMETERS FOR MEASURING THYROID ACTIVITY AND THE EFFECT OF TEMPERATURE ON THESE PARAMETERS  II.  FACTORS CONTRIBUTING TO. SEASONAL CHANGES IN THYROID ACTIVITY IN JUVENILE STEELHEAD  III.  95  .  .  102  THE ROLE OF THE THYROID IN THE STEELHEAD.  .  112  SUMMARY AND CONCLUSIONS  116  BIBLIOGRAPHY  119  LIST OP FIGURES FIGURE 1.  2.  3.  4.  PAGE  Seasonal changes i n water temperature i n the l a b o r a t o r y and i n two streams near Vancouver, B r i t i s h Columbia  -5  Plasma c l e a r a n c e f o l l o w i n g a s i n g l e J.131 i n j e c t i o n i n f i s h h e l d at 3° 0 (O) and 10° C (•). Each p o i n t r e p r e s e n t s a mean of 3 t o 7 f i s h . (Covariance a n a l y s i s , Table I)  14  T o t a l body c l e a r a n c e (O) and body e x c l u d i n g t h y r o i d c l e a r a n c e (•) f o l l o w i n g a s i n g l e 1^-31 i n j e c t i o n a t 5 ° C ( i n a c t i v e t h y r o i d ) and 1 0 ° C a c t i v e t h y r o i d ) . Equations c a l c u l a t e d separately f o r each phase. Each p o i n t r e p r e s e n t s a mean of 5 t o 7 f i s h (Covariance a n a l y s e s , T a b l e s I I and III)  18  (A) Percentage accumulation o f 1^51 by the t h y r o i d f o l l o w i n g a s i n g l e i l ? i n j e c t i o n at 1 0 ° C ( a c t i v e ) and 5 ° C ( i n a c t i v e ) . Standard d e v i a t i o n s shown. Each p o i n t r e p r e s e n t s a mean o f 5 t o 7 fish. (B) TUF v a l u e s f o r the same f i s h as ( A ) . (C) T/S v a l u e s f o r t h e same'fish as (A). The d i p h a s i c tendency i s shown i n the a c t i v e s t a t e (Covariance a n a l y s i s , T a b l e IV)  23  Change i n CR f o l l o w i n g a s i n g l e 1.131 i n j e c t i o n i n f i s h a t 10° C ( a c t i v e ) and 5 ° C ( i n a c t i v e ) . Each p o i n t r e p r e s e n t s a mean o f 5 t o 7 f i s h . .  28  1  5.  6.  7.  Seasonal change i n plasma T.131, % dose l ! 3 1 i n body, T/S, TUF, CR, mean e p i t h e l i a l cell''height and percentage o f f o l l i c l e s c o n t a i n i n g b l u e c o l l o i d i n s t e e l h e a d p a r r and smolts. Seasonal change i n temperature i s shown. Standard d e v i a t i o n s a r e shown f o r c e r t a i n parameters . . .  J>2  C o r r e l a t i o n s between water temperature and f o u r l l 3 1 parameters. A l l v a l u e s drawn from seasonal data ( F i g . 6 ) . A (f. dose i n body); B(CR); C(T/S); D(TUF) . .  33  -  viii  LIST OF FIGURES (Continued) FIGURE 8.  9,  10.  11.  12.  13.  14.  15.  PAGE  E x p e r i m e n t a l demonstration o f the i n f l u e n c e o f temperature on percentage accumulation o f l l 3 1 i n the t h y r o i d 4 and 8 days a f t e r a s i n g l e l ! 3 1 i n j e c t i o n , on mean c e l l h e i g h t (O) and on percentage of f o l l i c l e s c o n t a i n i n g b l u e c o l l o i d (•)  38  E x p e r i m e n t a l demonstration of the i n f l u e n c e o f temperature on 1"131 l e v e l s i n plasma and t o t a l body 4 days a f t e r a s i n g l e l " 1 3 l i n j e c t i o n . Each p o i n t r e p r e s e n t s 4 t o 7 i n d i v i d u a l s (Covariance a n a l y s i s , T a b l e VI)  40  E x p e r i m e n t a l demonstration o f the i n f l u e n c e o f temperature on T/S v a l u e s 4 and 8 days a f t e r a single l ! 3 l injection. Each p o i n t r e p r e s e n t s a mean o f 4 t o 7 i n d i v i d u a l s  42  E x p e r i m e n t a l demonstration o f the i n f l u e n c e o f temperature on CR v a l u e s 4 and 8 days a f t e r a s i n g l e 1131 i n j e c t i o n . Each p o i n t r e p r e s e n t s a mean of 4 t o 7 i n d i v i d u a l s (Covariance a n a l y s i s , T a b l e VI)  43  E x p e r i m e n t a l demonstration of the combined i n f l u e n c e o f temperature and photoperiod on plasma 1131 l e v e l s and percentage dose i n t o t a l body 4 days a f t e r a s i n g l e J.131 i n j e c t i o n i n 14 t o 18month s t e e l h e a d . Each p o i n t r e p r e s e n t s a mean of 8 t o 12 f i s h ( S t a t i s t i c s , T a b l e V I I I ) . . . .  46  E x p e r i m e n t a l demonstration of the combined i n f l u e n c e o f p h o t o p e r i o d and temperature on mean c e l l h e i g h t and T/S r a t i o s 4 days a f t e r a s i n g l e 1131 i n j e c t i o n i n 14 t o 18-month s t e e l h e a d . C e l l h e i g h t - each p o i n t r e p r e s e n t s a mean o f 3 t o 4 f i s h ; T/S - each p o i n t r e p r e s e n t s a mean of 8 t o 12 f i s h ( S t a t i s t i c s , Table VIII) 48 E x p e r i m e n t a l demonstration o f the combined i n f l u e n c e of p h o t o p e r i o d and temperature on plasma clearance following a s i n g l e l"l31 i n j e c t i o n i n 18-month s t e e l h e a d . Each p o i n t r e p r e s e n t s a mean of 5 t o 7 f i s h (Covariance a n a l y s i s , T a b l e I X ) .  51  E x p e r i m e n t a l demonstration of the combined i n f l u e n c e of p h o t o p e r i o d and temperature on T/S v a l u e s i n 18-month s t e e l h e a d f o l l o w i n g a s i n g l e 1131 i n j e c t i o n . Each p o i n t r e p r e s e n t s a mean of 5 t o 7 f i s h (Covariance a n a l y s i s , T a b l e I X ) . .  52  - ix LIST OF FIGURES  (Continued)  FIGURE 16.  17.  18.  19.  PAGE  E x p e r i m e n t a l demonstration of the combined i n f l u e n c e of photoperiod and temperature on CR v a l u e s f o l l o w i n g a s i n g l e 1131 i n j e c t i o n i n 18month s t e e l h e a d . Each p o i n t r e p r e s e n t s a mean of 5 to 7 i n d i v i d u a l s  53  I n f l u e n c e of body mass on T/S v a l u e s and percentate of l l 3 1 i n the t o t a l body 84 hours a f t e r a s i n g l e 1131 i n j e c t i o n i n 19-month s t e e l h e a d . .  56  I n f l u e n c e of body mass (gram) on plasma 1.131 l e v e l s and T/S v a l u e s (4 days a f t e r i n j e c t i o n ) and CR v a l u e s (8 days a f t e r i n j e c t i o n ) . Means and standard d e v i a t i o n s are shown f o r l a r g e , medium and s m a l l s i z e groups. No s i g n i f i c a n t d i f f e r e n c e was noted between medium and l a r g e s i z e groups f o r any 1131 parameter, but both d i f f e r e d from the s m a l l group f o r a l l parameters (p<.01)  59  R e l a t i o n s h i p between the l o g a r i t h m of body mass and the l o g a r i t h m of v a r i o u s 1131 parameters.  T/S i 20.  21.  22.  (A), r = - 0 . 5 7 ;  body (O),  CR (•), r = - 0 . 5 3 ;  r - -0.67  E x p e r i m e n t a l demonstration of the combined i n f l u e n c e of temperature and p h o t o p e r i o d on plasma 1131 l e v e l s and percentage dose i n t o t a l body 4 days a f t e r J.131 i n j e c t i o n i n 26 t o 50-month s t e e l head ( p o t e n t i a l m i g r a n t s ) . Each p o i n t r e p r e s e n t s a mean of 6 i n d i v i d u a l s ( S t a t i s t i c s , T a b l e X I ) .  60  63  - E x p e r i m e n t a l demonstration of the combined i n f l u e n c e of temperature and p h o t o p e r i o d on T/S v a l u e s 4 and 8 days a f t e r J.131 i n j e c t i o n i n 26 to 30-month s t e e l h e a d ( p o t e n t i a l m i g r a n t s ) . Each p o i n t r e p r e s e n t s a mean of 6 i n d i v i d u a l s ( S t a t i s t i c s , TABLE XI)  65  E x p e r i m e n t a l demonstration of the combined i n f l u e n c e of temperature and p h o t o p e r i o d on TUF v a l u e s 4 and 8 days a f t e r l ! 3 1 i n j e c t i o n i n 26 to 30-month s t e e l h e a d ( p o t e n t i a l m i g r a n t s ) . Each p o i n t r e p r e s e n t s a mean of 6 i n d i v i d u a l s ( S t a t i s t i c s , T a b l e XI)  66  X LIST OF FIGURES  (Continued) PAGE  FIGURE 23.  24.  25.  26.  E x p e r i m e n t a l demonstration of the combined i n f l u e n c e of temperature and p h o t o p e r i o d on mean c e l l h e i g h t and CR (8 days a f t e r 1.131 injection) i n 26 t o 30-month s t e e l h e a d ( p o t e n t i a l m i g r a n t s ) . C e l l h e i g h t - means ( A O ) and i n d i v i d u a l v a l u e s ( A O ) shown; CR - each p o i n t r e p r e s e n t s a mean of 6 i n d i v i d u a l s . .  68  TUF (open bars) and CR (shaded bars) f o r precoc i o u s two-year-old male (U0T and immature 0^ s t e e l h e a d i n e a r l y March under 4 combined c o n d i t i o n s of temperature and p h o t o p e r i o d . . . .  71  I n f l u e n c e of i n c r e a s e d swimming a c t i v i t y on v a r i o u s aspects of 1.131 metabolism i n 19-month s t e e l h e a d . Each p o i n t r e p r e s e n t s a mean of 6 i n d i v i d u a l s (Covariance a n a l y s i s , T a b l e 2 1 1 ) . .  73  E f f e c t of t r a n s f e r of 20-month s t e e l h e a d to sea water (25o/oo s a l i n e ) and i o d i d e - r e i n f o r c e d f r e s h water on v a r i o u s aspects of 1.131 metabolism measured 4 and 8 days a f t e r 1131 i n j e c t i o n . Mean v a l u e s r e p r e s e n t e d (SW,X ; FW + J.1 7,A ; 2  27.  28.  29.  30.  E f f e c t of t r a n s f e r of 30-month smolts and p a r r t o sea water (25 0 / 0 0 s a l i n e ) on v a r i o u s aspects of 1131 metabolism. Mean v a l u e s (bar) and i n d i v i d u a l v a l u e s shown E f f e c t of t r a n s f e r of u n d e r y e a r l i n g p o t e n t i a l migrant chum salmon t o sea water (25 0 / 0 0 s a l i n e ) and i o d i d e - r e i n f o r c e d f r e s h water. Mean v a l u e s (bar) and i n d i v i d u a l v a l u e s shown. X s < 0 . 0 5 ; Demonstration of the l a c k of a r e l a t i o n s h i p between s i l v e r i n g and any aspect of l ! 3 1 metabol i s m (8 days a f t e r i n j e c t i o n ) . Mean v a l u e s (bar) and i n d i v i d u a l values shown Change i n the t i s s u e : blood r a d i o a c t i v i t y i n a v a r i e t y of t i s s u e s from 19-month s t e e l h e a d . Each p o i n t r e p r e s e n t s a mean of 14 t o 16 individuals  81  83  88  91  - xi L I S T OF FIGURES  (Continued)  FIGURE 31.  32.  PAGE P e r c e n t a g e change ( r e l a t i v e t o t h e v a l u e 3 days a f t e r i n j e c t i o n ) of t i s s u e : b l o o d r a t i o s 6 and 9 d a y s a f t e r i n j e c t i o n i n 19-month s t e e l h e a d . .  93  Diagrammatic r e p r e s e n t a t i o n of the hypothesis t h a t t h y r o x i n u t i l i z a t i o n and p r o d u c t i o n r e s pond d i f f e r e n t l y t o t e m p e r a t u r e , x^ a n d X^Q r e p r e s e n t the d i f f e r e n c e s between u t i l i z a t i o n a n d p r o d u c t i o n a t 5 and 10° C . S i n c e x^ i s g r e a t e r t h a n XT_Q i"t w o u l d i m p l y g r e a t e r r e l e a s e o f TSH a t l o w t e m p e r a t u r e s a n d a l s o a g r e a t e r c e l l height  99  LIST OF TABLES TABLE I.  II.  -  PAGE  A n a l y s i s o f covariance f o r plasma l ! 3 1 (plasma 1131 B i o l o g i c a l C o n c e n t r a t i o n Coefficient/100) and time (hr) f o r a c t i v e ( 1 0 ° C) and i n a c t i v e (50 c) t h y r o i d s t a t e s  15  A n a l y s i s o f covariance f o r I ^ c l e a r a n c e from the t o t a l body'and time ( h r ) ; and body e x c l u d ing t h y r o i d and time ( h r ) . S l o p e g - S l o p e ^ =' 1  1  T t  Rate constant III.  IV.  V. VI.  VII.  VTII.  IX.  X.  f o r t h y r o i d T!31 uptake  (Slope-,).  19  A n a l y s i s of covariance f o r T131 c l e a r a n c e from t o t a l body and time (hr) and body e x c l u d i n g t h y r o i d and time (hr) f o r a c t i v e (10° C) and i n a c t i v e ( 5 ° C) t h y r o i d s t a t e s . The a n a l y s i s i n d i c a t e s the d i f f e r e n c e i n e x c r e t i o n r a t e s between the two temperatures i n phase I I .  20  A n a l y s i s of covariance f o r T/S and time (hr) f o r a c t i v e (10© C) and i n a c t i v e (5° C) t h y r o i d states  24  A summary o f r a d i o i o d i n e parameters and t h e i r significance  31  A n a l y s i s o f covariance f o r the r e l a t i o n s h i p between v a r i o u s 1131 parameters and temperature (° 0)  41  Summary o f the f o u r combined temperature and photoperiod conditions. Symbols shown a r e used in figures  45  Summary of s t a t i s t i c a l l y s i g n i f i c a n t d i f f e r e n c e s f o r s e v e r a l J.131 parameters between d i f f e r e n t c o n d i t i o n s o f temperature and p h o t o p e r i o d i n y e a r l i n g s i n June. White-Wilcoxon non-parametric r a n k i n g t e s t used  49  A n a l y s i s o f covariance performed on plasma 1131 and time ( h r ) , and T/S and time (hr) under f o u r d i f f e r e n t p h o t o p e r i o d and temperature regimes.  54  Summary of i n j e c t i o n d e t a i l s f o r s i z e experiment  58  - xiii L I S T OF TABLES  -  (Continued)  TABLE XI.  XII.  XIII.  PAGE Summary o f s t a t i s t i c a l l y s i g n i f i c a n t d i f f e r ences f o r s e v e r a l l ! 3 1 parameters between d i f f e r e n t c o n d i t i o n s o f t e m p e r a t u r e and p h o t o period i n two-year-olds (potential migrants) i n l a t e May ( W h i t e - W i l c o x o n t e s t ) . . . .  69  A n a l y s i s of c o v a r i a n c e f o r the r e l a t i o n s h i p b e t w e e n v a r i o u s 1^31 p a r a m e t e r s a n d t i m e ( h r ) f o r p a r r h e l d i n f a s t and s l o w c u r r e n t s . .  75  The e f f e c t o f d i f f e r e n t t e m p e r a t u r e a n d p h o t o p e r i o d r e g i m e s on t h e p e r c e n t a g e i n c i d e n c e o f s m o l t i f i c a t i o n i n t w o - y e a r - o l d s t e e l h e a d between 27 A p r i l a n d 29 M a y , 1962  87  INTRODUCTION Seasonal a variety  changes i n t h y r o i d a c t i v i t y have been r e c o r d e d by  of techniques  i n several  include Misgurnus fo3silis (Hagen,  (Zaitzev,  Plecoglossus (Berg,  (Lieber,  1936); P h o x i n u s p h o x i n u s  Esox l u c i u s  altivelis  species.  1936); A n g u i l l a  These anguilla  ( B a r r i n g t o n and M a t t y ,  1955); Salmo t r u t t a  (Swift,  (Honma, 1 9 5 9 ) ; F u n d u l u s  Gorbman a n d K o b a y a s h i ,  (Hickman, 1962).  teleost  1955,  1959);  heteroclitus  1959)» a n d P l a t i c h t h y s  Comprehensive  1934);  stellatus  reviews r e l a t i n g to these  have been w r i t t e n by Hoar (1959), M a t t y (i960) and S w i f t Most s t u d i e d o f a l l have been t h e s e a s o n a l thyroid activity  i n anadromous  salmonids.  f r e s h water b u t , except f o r spawning, sea.  the r i v e r s  t o t h e sea and a t t h i s  changes i n morphology, obvious  change,  behaviour  smoltification,  These f i s h h a t c h i n  young  (1939).  this  has been p l a c e d  and metabolism.  T h e most  i s the loss  of black parr  changes i n t h e u n d e r l y i n g p h y s i o l o g y  initial  on t h e r o l e  hypertrophy  i960).  marks  l i v e r y of the smolt.  discovery,  first  (Salmo s a l a r )  of the t h y r o i d during  Robertson  and h y p e r p l a s i a  (1948) o b s e r v e d  One at  noted  by Hoar  more a n d more  p r i m a r i l y by F o n t a i n e and h i s c o - w o r k e r s , ( F o n t a i n e , and F o n t a i n e ,  descend  characteristic  i n t h e A t l a n t i c salmon  Since  i n the  salmon  s m o l t i f i c a t i o n i s a marked t h y r o i d h y p e r a c t i v i t y histologically  life  time undergo  and a c q u i s i t i o n o f t h e s i l v e r y guanine o f t h e most emphasized  (i960).  changes i n  spend a d u l t  A t a c e r t a i n p e r i o d i n development,  changes  emphasis  smoltification 1954; L e l o u p  a similar  i n the thyroid of smolting  rainbow  - 2 troujt (Salmo g a i r d n e r l ) guanine d e p o s i t i o n  and  a l s o claimed t h a t t h y r o x i n  (Robertson, 1949).  always dependent upon t h y r o x i n . Oncorhvnchus). the t h y r o i d s  of s i l v e r i n g coho and  are r e l a t i v e l y i n a c t i v e (Hoar and r o l e of the  t h y r o i d i n the  salmon i s , t h e r e f o r e , speculations  S i l v e r i n g , however, i s not  In P a c i f i c salmon (genus  moderately a c t i v e at m i g r a t i o n but  The  stimulated  are  those of pink, s p r i n g and B e l l , 1950;  Eales,  s m o l t i f i c a t i o n and  f a r from e s t a b l i s h e d .  have been made, the  sockeye  1961,  chum  1963).  m i g r a t i o n of  •  Though c e r t a i n  c o n t r o l of these seasonal  t h y r o i d f l u c t u a t i o n s i s almost e n t i r e l y u n i n v e s t i g a t e d .  The  matter i s f u r t h e r complicated by the d i s c r e p a n c y between h i s t o l o g i c a l and  radiochemical determinations.  With these u n r e s o l v e d p o i n t s l o g i c a l and  r a d i o c h e m i c a l i n v e s t i g a t i o n of t h y r o i d f u n c t i o n  attempted i n the This  i n mind, a comprehensive h i s t o -  steelhead trout  s p e c i e s i s anadromous and  was  (Salmo g a i r d n e r i R i c h a r d s o n ) .  shows an e a s i l y r e c o g n i z a b l e  parr-  smolt t r a n s f o r m a t i o n . The of  i n v e s t i g a t i o n was  divided  i n t o f o u r i n t e r r e l a t e d areas  study: (a)  A preliminary  comparative a n a l y s i s  metabolism i n s t e e l h e a d w i t h low (b)  and  of  high t h y r o i d  Seasonal o b s e r v a t i o n s on h i s t o l o g y  and  radioiodine activity. radioiodine  metabolism from 17 months p r i o r t o m i g r a t i o n up t o seaward m i g r a t i o n i t s e l f t o determine s e a s o n a l changes. (c)  E x p e r i m e n t a l i n v e s t i g a t i o n of environmental f a c t o r s  and  i n t e r n a l changes i n f l u e n c i n g these s e a s o n a l f l u c t u a t i o n s . (d)  An  i n v e s t i g a t i o n of p o s s i b l e  c a t a b o l i s m i n s t e e l h e a d and  s i t e s of radiohormone  the r e l a t i o n s h i p between  radioiodine  m e t a b o l i s m and s i l v e r i n g . From t h e s e i n v e s t i g a t i o n s factors and a l s o  contributing to  i t was h o p e d t o d e t e r m i n e  seasonal  t o g a i n knowledge  changes i n t h y r o i d  activity  concerning the r o l e of the  p a r t i c u l a r l y at the time of m i g r a t i o n .  the  thyroid,  MATERIALS AND METHODS 1.  Living  Materials  Steelhead  were o b t a i n e d as y e a r l i n g s  or two-year-olds  t h e B r i t i s h C o l u m b i a F i s h a n d Game H a t c h e r y a t C u l t u s B r i t i s h C o l u m b i a , where and k e p t  i n large  t h e y were f e d  concrete  B r i t i s h Columbia.  received  fresh  an abundant  The t e m p e r a t u r e  c o n t a i n the  switch regulated  to the n a t u r a l outside  and a l s o  It  departed  i n onset  i n general  i n a manner s i m i l a r  (Fig. l ) .  Illumination  via a photoelectric Thus,  length  cell  a changing  exposed  seasonal  o u t s i d e p h o t o p e r i o d was i m p o s e d u p o n  from the n a t u r a l c o n d i t i o n i n not  or offset  They  tapwater  a n d was c o n t r o l l e d i n  illumination.  conforming to the  the f i s h .  lamps)  on  t o V a n c o u v e r known t o  s p e c i e s under i n v e s t i g a t i o n  by an automatic  daylength  close  of  but w i t h l i t t l e d i u r n a l  was f o u n d t o a l t e r  was a r t i f i c i a l ( f l u o r e s c e n t  gradual  temperature  found i n c e r t a i n streams  they  Lake C i t y , U t a h ) .  supply of dechlorinated  v a r y i n g markedly i n seasonal  food  University  H e r e t h e y w e r e h e l d i n d o o r s and f e d ( J . R. C l a r k , S a l t  to that  At intervals  of Zoology,  commercial f i s h food  change.  Lake,  on d r i e d c o m m e r c i a l  outdoor tanks.  were t r a n s p o r t e d t o t h e Department  from  (i.e.  t h e r e was no t w i l i g h t  being  period)  intensity.  F i s h were removed f r o m t h e s e c o n d i t i o n s f o r a v a r i e t y experiments  i n which they were s u b j e c t e d  and c h e m i c a l e n v i r o n m e n t s .  These  to different  Radioiodide Carrier-free  physical  c o n d i t i o n s a r e many a n d v a r i e d  a n d may be c o n s i d e r e d m o s t c o n v e n i e n t l y w i t h t h e 2.  of  results.  Injection sodium i o d i d e  (Nal^l)  w  as  diluted with d i s t i l l e d  - 5 -  F i g . 1. S e a s o n a l c h a n g e s i n w a t e r t e m p e r a t u r e i n t h e l a b o r a t o r y a n d i n two s t r e a m s n e a r V a n c o u v e r , B r i t i s h Columbia.  - 6 water and i n j e c t e d volume between syringe  i n t r a p e r i t o n e a l l y v i a t h e d o r s a l muscle i n a  0 . 0 2 a n d 0.05 m l u s i n g a 0 . 2 5 - m l  a n d g a u g e 30 n e e d l e .  Duplicates  tuberculin  of the injected  dose  w e r e d i l u t e d t o 100 m l w i t h 3% p o t a s s i u m i o d i d e s o l u t i o n i n 100-ml v o l u m e t r i c f l a s k s injected  dose).  The f i s h were n o t a n a e s t h e t i z e d  j e c t i o n but q u i c k l y l i f t e d out of t h e water and t h e n i n j e c t e d to the water.  stated,  injection.  i n a f i n e mesh n e t  operation took a matter a l l f i s h were k e p t  of  returned seconds.  i n running water  A p p r o x i m a t e l y 0.1^uc I ^ ! p e r g r a m b o d y  weight  1  was a d m i n i s t e r e d t o each 3•  during i n -  through t h e n e t t i n g and immediately  The e n t i r e  Unless otherwise after  ( l ; m l - 1% of  and used as s t a n d a r d s  fish.  B l o o d S a m p l i n g and S e p a r a t i o n of P r o t e i n - b o u n d and I n o r g a n i c Radioiodine B l o o d samples were o b t a i n e d by c u t t i n g t h r o u g h t h e c a u d a l  peduncle  and drawing the b l o o d by c a p i l l a r y a c t i o n i n t o a  heparinized glass tube. t h i s procedure  to facilitate  anaesthetized. by  f i s h were a n a e s t h e t i z e d  handling;  for  s m a l l f i s h were not  W i t h i n seconds o f s a m p l i n g a l l f i s h were  killed  decerebration. The t e c h n i q u e  radioactive  However,  f o r determining the organic  constituents  been d e s c r i b e d  here  Very large  fine  o f s m a l l samples  and i n o r g a n i c  of f i s h plasma has  ( H i c k m a n , 19&0; H o a r a n d E a l e s ,  19&3; E a l e s ,  i n view of s l i g h t m o d i f i c a t i o n , the technique  is  1963). given  in full. On w i t h d r a w a l o f t h e b l o o d s a m p l e ,  t u b e was p l u g g e d  the heparinized  a t one end w i t h " p l a s t i c e n e "  and t h e  capillary corpuscles  s e p a r a t e d from t h e plasma by c e n t r i f u g a t i o n .  The plasma  was  s e p a r a t e d from t h e c o r p u s c l e s by b r e a k i n g the tube a t t h e i r junction.  The tube p l u s plasma c o n t e n t s were weighed and t h e  plasma blown i n t o 4 ml o f 12.5f° t r i c h l o r o a c e t i c a c i d i n a t h i c k - w a l l e d p y r e x ' c e n t r i f u g e tube. T  12-ml  The empty c a p i l l a r y tube  was t h e n reweighed t o determine t h e plasma mass.  The  plasma  p r o t e i n p r e c i p i t a t e was broken up w i t h a g l a s s r o d , c e n t r i f u g e d and t h e s u p e r n a t e decanted o f f and saved.  Two ml o f 2.5f»  t r i c h l o r o a c e t i c a c i d were added and t h e p r e c i p i t a t e a g a i n broken up and c e n t r i f u g e d .  T h i s p r o c e d u r e was r e p e a t e d f o r a second  t i m e g i v i n g a t o t a l o f 8 ml o f s u p e r n a t e c o n s t i t u t i n g t h e nonp r o t e i n components o f t h e plasma and i n c l u d i n g I ^ . 1  r e m a i n i n g p r o t e i n s were d i s s o l v e d i n 4 ml o f 1.5 N h y d r o x i d e (NaOH).  1  The  sodium  T h i s c o n s t i t u t e d the p r o t e i n f r a c t i o n of the  plasma and i n c l u d e d t h e hormonal o r p r o t e i n - b o u n d r a d i o i o d i n e (PBI^l).  The 4-ml sample o f d i s s o l v e d P B I ^ 1  1  was  transferred  t o a g l a s s c o u n t i n g tube and counted f o r 3000 c o u n t s . t h e 1^31  wash d e r i v e d from t h e same plasma sample was  H a l f of pipetted  t o a s i m i l a r tube and counted f o r 3000 c o u n t s o r l o n g e r depending on i t s a c t i v i t y .  By d o u b l i n g t h i s count t h e t o t a l I ? ! c o n t e n t  o f t h e sample was f o u n d .  1  A l s o counted f o r 30,000 c o u n t s were  4 ml (4%) o f t h e s t a n d a r d s o l u t i o n .  Thus e i t h e r f r a c t i o n c o u l d  be e x p r e s s e d as a p e r c e n t a g e of t h e i n j e c t e d dose.  A l l counts  were made i n a N u c l e a r C h i c a g o W e l l C o u n t e r (Model DS5 S c i n t i l l a t i o n Detector).  Versatile  From t h e s e d a t a two parameters were  determined(i) 1963).  C o n v e r s i o n r a t i o . OR  (Hickman, I960, 1962; E a l e s ,  l?6l,  OR  8 -  count/min x  =  p g j l 3 1 count/min + l ! 3 1 _  organic  100 count/min  radioiodine(hormonal)  t o t a l plasma r a d i o a c t i v i t y (ii)  B i o l o g i c a l concentration c o e f f i c i e n t f o r I  t h e plasma.  1  ^ in  T h i s i s d e r i v e d from t h e b i o l o g i c a l c o n c e n t r a t i o n  c o e f f i c i e n t , which expresses  plasma 1^-31  p e r c e n t a g e of t h e dose i n j e c t e d  c o n c e n t r a t i o n as a  (Comar, 1955).  I t allows f o r  t h e f a c t t h a t the same dose put i n t o a f i s h h a l f the mass of a n o t h e r f i s h w i l l have t w i c e as much r a d i o a c t i v i t y p e r mass of plasma, and p e r m i t s comparison of plasma I 3 1 x  between f i s h o f d i f f e r e n t  sizes.  b i o l o g i c a l concentration coefficient % of i n j e c t e d dose as l ! 3 1  concentration  =  i n plasma sample x body w e i g h t ( g )  mass o f sample(g) The  b i o l o g i c a l c o n c e n t r a t i o n c o e f f i c i e n t has always been  by  100.  4.  Body and  divided  Thyroid  I m m e d i a t e l y a f t e r d e a t h the f i s h was  weighed and t h e  basi-  Tbranchial r e g i o n cut out f r o m the f r o n t o f a r c h I ( a n t e r i o r ) t h e end of a r c h I I I . t i s s u e was  T h i s r e g i o n i n c l u d i n g the e n t i r e  to  thyroid  t h e n dropped i n t o a c o u n t i n g t u b e c o n t a i n i n g B o u i n s T  f i x a t i v e and counted f o r 30,000 counts i n a N u c l e a r C h i c a g o W e l l Counter.  Two  4-ml  standards  (0.04% of t h e i n j e c t e d dose) were  counted s i m u l t a n e o u s l y and t h e p e r c e n t a g e of t h e i n j e c t e d dose i n the t h y r o i d a r e a was  determined.  The  t h y r o i d s were counted  i n Bouin's f l u i d so t h a t t h e y c o u l d be s e c t i o n e d l a t e r . d i s c r e p a n c y may  have been i n t r o d u c e d by c o u n t i n g  Some  essentially  - 9 a point source, well,  against  undigested  the t h y r o i d r e s t i n g at the  a 4-ml s t a n d a r d .  t h y r o i d s and t h e  i n the  comparison  same t h y r o i d d i g e s t e d  then d i l u t e d t o 4 ml revealed measured  However,  bottom of the  negligible  same c o u n t i n g s y s t e m  between  w i t h . NaOH a n d  differences  (Wiggs,  counting  when This  1962).  is  p r o b a b l y due t o t h e l o w a b s o r p t i o n o f t h e h a r d ^ r a d i a t i o n o f jl31  T h y r o i d u p t a k e was e x p r e s s e d  #  t i o n of the  injected  as t h e percentage  accumula-  dose.  T h e b o d y l a c k i n g t h y r o i d was s l i c e d a n d t h e p i e c e s t o f o r m a 5-cm s q u a r e whose fish.  By c o u n t i n g t h i s  end-probe  crystal,  an i d e n t i c a l d i s t a n c e These  a distance  an estimate  i n t h e b o d y c o u l d be made.  to a percentage of the  cut to the  injected  of the  dose,  5»  this  radioactivity  s t a n d a r d s were c o u n t e d inverted  dose.  as the  of  absorbent  samples,  w i t h 10 m l 10% o f  B o d y r e t e n t i o n o f 1.131 was m e a s u r e d ,  considerable  injected  in I ^ 1  1  dose.  of  8.08  and samples  the  therefore,  T h i s meant t h a t  with a h a l f - l i f e  at  end-probe.  Each standard then represented  e n t i r e l y e l i m i n a t e d as b o t h s t a n d a r d s same  radioactivity  To c o n v e r t  same s o l i d g e o m e t r y  as a p e r c e n t a g e o f the decay,  of  total  and p o s i t i o n f r o m the  standard s o l u t i o n .  injected  size  o f 10 cm f r o m a n  s t a n d a r d s were formed by p e r m e a t i n g p i l e s  paper, of  "body" at  v a r i e d w i t h the  s c i n t i l l a t i o n c o u n t e r w i t h a 45 mm d i a m e t e r a n d 38.5 mm  t h i c k N a l (Tl) left  thickness  arranged  physical  days,  decayed  was at  the  rate. Thyroid Histology Thyroid tissues  serially  sectioned  fixed i n Bouin's f l u i d  (10 u)  were  i n the r e g i o n of the  decalcified,  second  branchial  - 10 a r c h and then s t a i n e d by t h e Azan technique Theoretically inactive  1954;  this  s t a i n renders  g l a n d and b l u e  Barrington,  and t h e lowest  gland  (Pickford,  c e l l heights  (tallest  + lowest/2  absent.  Since the t h y r o i d of t e l e o s t s i s v a r i a b l e , t o randomize the f o l l i c l e s measured. from at l e a s t  The  t o one o f f o u r c a t e g o r i e s - b l u e , r e d ,  m i x t u r e o f r e d and b l u e o r  branchial region.  examined  were measured u s i n g an o c u l a r s c a l e .  c o l l o i d was a s s i g n e d  tissue  1953,  F i f t y unbroken f o l l i c l e s were  and t h e t a l l e s t  * mean c e l l h e i g h t )  1953).  the f o l l i c l e c o l l o i d r e d i n an  i n an active  1963).  (Gurr,  three  I f three  different  i t was  necessary  T h i s was done b y  selecting  areas of the second  s l i d e s were a v a i l a b l e  from t h i s  area,  17 f o l l i c l e s w o u l d be e x a m i n e d o n s l i d e I , 17 o n s l i d e I I a n d 16 on s l i d e I I I .  Randomization of f o l l i c l e s  one s e c t i o n was a c h i e v e d tissue  by c e n t r i n g t h e o b j e c t i v e  s t r u c t u r e was d i f f i c u l t  t o h i g h power.  to detect,  The f o l l i c l e c l o s e s t  t h e h i g h p o w e r was t h e n t h e f i r s t follicle  and t h e n  switching  to the ocular scale  t o be a s s e s s e d .  under  The n e x t  The f o l l i c l e i m m e d i a t e l y t o t h e l a t t e r ' s  was e x a m i n e d t h i r d a n d s o o n u n t i l exhausted.  follicles treated  individual  t o be e x a m i n e d was t h e one i m m e d i a t e l y t o i t s l o w e r  left-hand margin.  was  on t h e t h y r o i d  u n d e r v e r y l o w power m a g n i f i c a t i o n s u c h t h a t  follicle  left  t o be e x a m i n e d o n a n y  I f t h i s procedure  before  the quota f o r the s l i d e  resulted  t h e quota were r e a c h e d ,  i n exhaustion of  another  s i m i l a r l y i n t h e same s l i d e b u t a t l e a s t  posteriorly.  Since  s e c t i o n was 150 jx  f o l l i c l e d i a m e t e r s were u s u a l l y f a r l e s s  t h a n 150 u , r e p e t i t i v e  measurement  lower  o f f o l l i c l e s was a v o i d e d .  - 11 6.  S t a t i s t i c a l Treatment In s t u d i e s of r a t e s of change of I  body compartments,  1  ^ fractions i n various 1  l i n e a r r e g r e s s i o n s were c a l c u l a t e d and e i t h e r  t h e s l o p e s o r a d j u s t e d means compared ( S t e e l and T o r r i e , i 9 6 0 ) .  by a n a l y s i s o f c o v a r i a n c e  I n comparing s m a l l samples ( f r e q u e n t l y  below 12), which i n many i n s t a n c e s appeared t o depart from a normal d i s t r i b u t i o n , t h e Wilcoxon-White non-parametric r a n k i n g t e s t was employed  (Snedecor, 1956).  - 12 RESULTS I.  COMPARISON OE YEARLING STEELHEAD WITH DIFFERENT RATES OF RADIOIODINE METABOLISM R e l i a b l e radioiodide estimates  possible  of t h y r o i d a c t i v i t y  o n l y when a t h o r o u g h s t u d y o f r a d i o i o d i n e m e t a b o l i s m  h a s b e e n made w i t h t h e a n i m a l i n v a r i o u s activity. activity  Despite extensive in teleosts,  i n a n y one s p e c i e s  lower vertebrates, it  states of  are r a r e .  of r a d i o i o d i n e metabolism  L e l o u p and F o n t a i n e (i960)  i s d i f f i c u l t to reconstruct  m e t a b o l i s m f o r a n y one s p e c i e s .  it  covered  is  so  vast  a complete p i c t u r e of  1^31  The w o r k b y H i c k m a n (1959,  i s an e x c e p t i o n .  of r a d i o i o d i n e metabolism at b e e n made i n t h e s t u d y ,  have  survey of r a d i o i o d i n e metabolism i n  b u t t h e number o f s p e c i e s  on P l a t i c h t h y s s t e l l a t u s  thyroid  use of r a d i o i o d i n e t o study t h y r o i d  intensive studies  presented a comprehensive  that  are  different  S i n c e many  levels  estimates  of a c t i v i t y  i s considered necessary  to  1962)  have  present  a comparison between r a d i o i o d i n e m e t a b o l i s m i n f i s h w i t h  active  and i n a c t i v e g l a n d s .  I  I n t h i s instance the d i f f e r e n c e  m e t a b o l i s m was i n d u c e d w i t h a 5° C t e m p e r a t u r e holding f i s h at The o b j e c t  5 a n d 10° of t h i s  a s i n g l e d o s e o f 1^31  difference  f o r a p e r i o d of s e v e r a l days.  teleost  This could on t h e  same i n d i v i d u a l s f o r t h e e n t i r e  p r e s e n t e d t h e p r o b l e m o f k e e p i n g f i s h a l i v e f r o m one  diffuse  by  s t u d y was t o f o l l o w t h e m e t a b o l i s m o f  Use of the  m i n a t i o n to the next.  ^  1  C.  b e done i d e a l l y b y m e a s u r i n g t h e d e s i r e d p a r a m e t e r s individuals.  in  Besides  period  deter-  the d i f f i c u l t y of counting  thyroid i n vivo i n small f i s h ,  same  t h e r e was  the the  - 13  -  g r e a t e r d i f f i c u l t y of e x t r a c t i n g adequate blood samples r e p e t i t i v e l y without d e p l e t i n g the blood volume. these d i f f i c u l t i e s , 55 f i s h  (8-25  g, av = 1 2  To  obviate  g) from each  temperature regimen (5 and 10° 0) were " s i m u l t a n e o u s l y " w i t h 1131  and 5 t o 7 f i s h from each group sampled at the  r e q u i r e d time i n t e r v a l s Each f i s h was of I ^ 1  1  injected  ( 1 2 , 3 6 , 6 0 , 8 4 , 1 0 8 , 1 3 2 , 1 5 6 , 2 0 4 and 276 h r ) .  weighed and plasma I ^ 1  dose i n body, percentage  c o n v e r s i o n r a t i o recorded.  concentration,  1  of 1^31  By s e r i a l l y  percentage  dose i n t h y r o i d and sampling  from the popu-  l a t i o n s i n t h i s manner and d e r i v i n g the mean v a l u e f o r each sample a t a g i v e n time under a g i v e n c o n d i t i o n , i t was p o s s i b l e t o f o l l o w the changes i n 1^31  c o n c e n t r a t i o n s i n s e v e r a l compart-  ments as an average of the whole p o p u l a t i o n . 1.  I 31  E x t r a t h y r o i d a l and T h y r o i d a l Clearance of Plasma  1  R a d i o i o d i n e i n j e c t e d i n t r a p e r i t o n e a l l y i s r a p i d l y absorbed (Hickman, 1 9 5 9 ) .  i n t o the blood stream  The plasma l e v e l i s  then c o n t i n u a l l y d e p l e t e d due t o l o s s v i a a v a r i e t y of r o u t e s (Eig.  2).  A s m a l l percentage  of the dose i s e v e n t u a l l y  accumulated by the t h y r o i d but the g r e a t e r p a r t i s e x c r e t e d . Although  plasma I^31  the d i f f e r e n c e was  w  a  s  l o s t f a s t e r a t the h i g h e r  not s t a t i s t i c a l l y s i g n i f i c a n t  temperature, (Table I ) . A l l  the mean v a l u e s f o r 5° C f i s h were above those f o r 10° however, the a d j u s t e d means were s t a t i s t i c a l l y (p<0.01).  C fish,  different  T h i s shows c o n v i n c i n g l y t h a t the temperature e l e v a -  t i o n of 5° 0 had caused  an i n c r e a s e i n 1^31  by one or more r o u t e s .  Of f u r t h e r i n t e r e s t was  tendency a t the h i g h e r temperature.  l o s s from the plasma the  curvilinear  This i s considered i n greater  - 14 -  F i g . 2. Plasma c l e a r a n c e f o l l o w i n g a s i n g l e I inj e c t i o n i n f i s h h e l d a t 5° C (O) and 10° C (•). Each p o i n t r e p r e s e n t s a mean of 5 t o 7 f i s h (Covariance a n a l y s i s , Table I ) . p  i  - 15 -  TABLE I  A n a l y s i s of covariance f o r plasma I - ^ l (plasma I 3 1 b i o l o g i c a l c o n c e n t r a t i o n c o e f f i c i e n t / 1 0 0 ) and time (hr) f o r a c t i v e ( 1 0 ° C) and i n a c t i v e ( 5 ° C) t h y r o i d s t a t e s x  CONDITION  SLOPE  INACTIVE  -0.0127  ACTIVE  -0.0158  ?s  df  2.08  and  1  101  P  0.05  F m  df  x  39.3  1 and 104  P  0.01 •  F F  g  - Variance r a t i o f o r slopes. = Variance r a t i o f o r adjusted  means.  - 16 detail  below.  A n e q u a l l y r e l i a b l e a n d p e r h a p s more d i r e c t m e t h o d determining I ^ ! excretion i s 1  injected various the  t o measure  dose r e m a i n i n g i n t h e body time i n t e r v a l s .  injected  Immediately  Thus,  II).  demonstrates  of the e x c r e t i o n d a t a . explanation.  (Day 1-4  curvilinear  a  phase  As yet  I I and I I I .  ( 1 9 6 2 ) , who  Applied to these data  initial  explanation is  i f an i s o t o p e  it  suggested by a  curvilinear  i n one c o m p a r t m e n t  t o t w o o r more c o m p a r t m e n t s  Thus t h e c o m p o s i t e  as  2 t o 3 days and t h e n a  curvilinear  may b e a c o m b i n a t i o n o f a r a p i d a n d a s l o w T131 routes.  observed  i t may be due t o a n  Oomar (1955) who d e m o n s t r a t e d m a t h e m a t i c a l l y t h a t  different  differences  satis-  A more p l a u s i b l e  rates.  and  t h i s h a s no c o m p l e t e l y  diuresis for  at  two  i n d u c e d by " l a b o r a t o r y d i u r e s i s "  would suggest a s i g n i f i c a n t  different  have  nature  of experimental handling.  b e i n g d i l u t e d b y movement  they  (curvilinear)  A c c o r d i n g t o Wiggs  r e l a t i o n s h i p i s expected  I  significant  are given i n Tables the d i p h a s i c  rapid ion loss  stabilization.  the  of  are  under both c o n d i t i o n s ,  a s i m i l a r phenomenon i n s t i c k l e b a c k s ,  a result  is  The s t a t i s t i c a l l y  these r e l a t i o n s h i p s  and g e n e r a l  and the p e r c e n t a g e  l i n e a r p l o t s w i t h an i n f l e x i o n  f o r each parameter  • phase  Table.II  factory  at  evident  r e l a t i o n s h i p s have been c o n s i d e r e d  between  (excluding thyroid)  To s i m p l i f y t h e a n a l y s i s ,  been approximated t o d i p h a s i c  D a y 3-11  of  This parameter  aspect of both parameters.  3|- d a y s .  the percentage  dose i n t h e t o t a l body ( i n c l u d i n g t h y r o i d )  shown i n F i g u r e 3«  of  at  is markedly  relationship loss  via  two  A c c o r d i n g t o Comar, t h e r a p i d r e m o v a l would  - 17  -  p r e d o m i n a t e i n i t i a l l y b u t w o u l d be r e p l a c e d l a r g e l y b y t h e removal rate at low I ? 1  routes  levels.  1  from the steelhead  body,  In considering' I ? 1  the g i l l  slower  removal  1  could conceivably  be  t h e more r a p i d a n d t h e k i d n e y t h e s l o w e r r o u t e s i n c e t h e r e  is  evidence t o suggest t h a t I - ^ l the g i l l s  t,  removed p r e d o m i n a n t l y v i a  e  (Hickman, 1959).  The d i p h a s i c n a t u r e o f t h e s e c u r v e s does n o t i n f l u e n c e rate  of t h y r o i d l!31  j.131  a c c u m u l a t i o n have been o b t a i n e d by s u b t r a c t i n g the  age d o s e  accumulation.  for thyroid percent-  i n body s l o p e f r o m t h e s l o p e d e r i v e d f r o m t h e body and  t h y r o i d combined (Table I I ) . the  Rate constants  same o v e r b o t h p h a s e s  A t 5° C t h e c o n s t a n t  of the curve  a v - 0.00555) a n d t h e same a p p l i e s a t  is  essentially  (6.0054 a n d 0 . 0 0 5 7 ; 10° C (0.0075 a n d 0 . 0 0 7 7 ;  av = 0.0076), a l t h o u g h t h e r e a r e v e r y marked d i f f e r e n c e s mean c o n s t a n t s It  f r o m t h e two t e m p e r a t u r e s  i s concluded that  on t h e t h y r o i d j l 3 1 constant rates  (i)  temperature has a s i g n i f i c a n t  accumulation rate constant  and ( i i )  i s a r e l i a b l e index of t h y r o i d a c t i v i t y under  of l!31  s t a t i s t i c a l differences a d j u s t e d mean v a l u e s  different  (p=0.01).  at higher  influence  between s l o p e s  (Table I I I ) .  temperatures.  the  However, between  T h i s supports the e a r l i e r evidence l i k e t h y r o i d 1.131  on  demonstrate  showed a s i g n i f i c a n t d i f f e r e n c e  l o s s from the body,  active  this  E l i m i n a t i o n a p p e a r e d more r a p i d a t  h i g h e r t e m p e r a t u r e b u t a g a i n i t was d i f f i c u l t t o  1131  influence  excretion.  excretion rates.  two r e g i m e s  between  (0.00555 a n d O.OO76).  O f f u r t h e r s i g n i f i c a n c e was t h e t e m p e r a t u r e 1.131  the  the  that  a c c u m u l a t i o n , i s more  - 18 -  F i g . 3. T o t a l b o d y c l e a r a n c e (O) a n d b o d y e x c l u d i n g t h y r o i d c l e a r a n c e (•) f o l l o w i n g a s i n g l e 1131 i n j e c t i o n a t 30 c ( i n a c t i v e t h y r o i d ) a n d 10° C ( a c t i v e t h y r o i d ) . Equat i o n s c a l c u l a t e d s e p a r a t e l y f o r each phase. Each point r e p r e s e n t s a mean o f 5 t o 7 f i s h ( C o v a r i a n c e a n a l y s e s , T a b l e s I I and I I I ) .  TABLE I I A n a l y s i s o f covariance f o r I ^ l clearance from the t o t a l body and time ( h r ) ; and body e x c l u d i n g t h y r o i d and time ( h r ) . Slope™ ~ - S l o p e = Rate constant f o r t h y r o i d i l j l uptake fSlope-} a  % DOSE BODY AND % DOSE THYROID Condition  INACTIVE  ACTIVE  Phase  Sloperri  P  +B  I  -0.0161  II  -0.0018  I  -0.0204 '  II  -0.0035  .  9;92** -  14.17**  fo DOSE BODY  df  Slope  1 and 56  -0.0215}  1 and 57  -0.0279 '  F  B  18.5**  -0.0075)  -0.0112  .  df 1. and 62  1 9 . 8 0 * * and 58  Slope  T  0.0054 0.0057  0.0075 0.0077  Denotes s t a t i s t i c a l d i f f e r e n c e between compared s l o p e s (p = 0 . 0 1 ) .  TABLE I I I A n a l y s i s o f covariance f o r I ? clearance from t o t a l body and time (hr) and body excluding t h y r o i d and time ( h r ) , f o r a c t i v e ( 1 0 ° C) and i n a c t i v e (5 0) t h y r o i d s t a t e s . The a n a l y s i s i n d i c a t e s the d i f f e r e n c e i n e x c r e t i o n r a t e s between the two temperatures i n phase I I 1  1  f, DOSE BODY + fo DOSE THYROID  f. DOSE BODY PHASE  CONDITION  INACTIVE  I  ACTIVE  INACTIVE  II  E F  ACTIVE  s m  SLOPE  df  F  m  -0.0161?  1 and -0.0204* 40 -0.0018' J 1 . and 72 -0.0035,  SLOPE  1.175  = Variance r a t i o f o r s l o p e s . s Variance r a t i o f o r adjusted  58.50**  F  s  1 • and 42 -0.0279J  0.80  1 and - 0 . 0 1 1 2 j 68  3.89  -0.0215?  0.697  -O.OO75]  means.  ** denotes h i g h l y s i g n i f i c a n t d i f f e r e n c e ( p ^ O . O l ) . Almost shows s i g n i f i c a n c e between slopes ( p = 0 . 0 5 ) . Other "F" values show no d i f f e r e n c e between s l o p e s . ?  df  F  ?  m  29.87**  - 21 2.  E x p e r i m e n t a l Measurement of T h y r o i d I ^ ! Accumulation 1  I d e a l l y , a l l these I ^ ! movements would he represented i n 1  terms o f r a t e c o n s t a n t s .  I n s m a l l fish,where s e r i a l  f o r determination of regressions  sampling  f o r r a t e constants would  involve  many f i s h , an attempt was made t o determine the extent o f excretion  and t h y r o i d uptake on the b a s i s  measurements. the  of s i n g l e  terminal  F o r comparison these samples were always taken a t  same time a f t e r  injection.  Due t o the i n f l u e n c e  o f temperature on  extrathyroidal  removal o f I - ^ l j mere measurement o f the percentage of the dose accumulated i n the t h y r o i d would be u n r e l i a b l e Relative  t o the standard d e v i a t i o n s  (Fig. 4A).  the d i f f e r e n c e s  between t h e  t h y r o i d uptake means a r e v e r y s l i g h t and a r e e v e n t u a l l y At a s l i g h t l y f a s t e r r a t e o f e x t r a t h y r o i d a l  reversed.  I ^ ! clearance i n 1  f i s h w i t h the a c t i v e gland, the percentage o f the dose accumulated i n the t h y r o i d would have been the same or lower than that  i n the i n a c t i v e g l a n d .  Thus t h y r o i d uptake o f I 3 1 x  measured i n t h i s way i s v e r y dependent on the e x t r a t h y r o i d a l c l e a r a n c e r a t e , and i n any s i n g l e t e r m i n a l uptake t h i s blood l o s s o f 1 ^ 1  m  ust  assessment o f t h y r o i d  be taken i n t o account.  Allowances f o r t h i s have been made i n a v a r i e t y o f ways. Hoar and E a l e s (1963) measured the T h y r o i d Uptake F a c t o r (TUF) where rmT-rr,  -  TUF  =  dose t h y r o i d • % dose t h y r o i d + % dose body f*  x  100  I t measures a t death t h a t percentage o f the non-excreted I ^ ! t h a t has been p a r t i t i o n e d from the body compartment i n t o the 1  -  thyroid.  TUT  v a l u e s f o r the  shown i n F i g u r e 4B. that  -  22  i n a c t i v e and  A far better  active thyroids  s e p a r a t i o n i s now  obtained from t h y r o i d uptake v a l u e s .  t h a t whereas the  l i n e a r r e l a t i o n s h i p was  a f t e r i n j e c t i o n i n the  f i s h w i t h the  shown than  I t i s to be  noted  maintained u n t i l 12  slower r a t e of  Further attention  U n l i k e other 1^21  1  rather  a l s o be noted at t h i s stage how  i s to i n vivo  studies  t h y r o i d a l t i s s u e can Leloup and T/S  i t is  below.  arithmetically  than e x p o n e n t i a l l y r e l a t e d t o time a f t e r i n j e c t i o n .  t h e o r e t i c a l e x p l a n a t i o n f o r t h i s has may  the  i s drawn to t h i s p o i n t  parameters s t u d i e d ,  providing be  not  days  I ^!  metabolism, i t f e l l markedly a f t e r 6 days i n f i s h w i t h active thyroid.  are  been attempted.  applicable  the TUF  adequate s c r e e n i n g of  A It  measure extra-  obtained.  F o n t a i n e ( i 9 6 0 ) have used the  thyroid/serum or  r a t i o / t h y r o i d r a d i o a c t i v i t y / g ^ f o r measuring t h y r o i d serum r a d i o a c t i v i t y / g  uptake,  In a s s e s s i n g t h y r o i d I - ^ l c o n c e n t r a t i o n r e l a t i v e t o serum concentration,  t h i s method i s s i m i l a r t o t h y r o i d  (Hickman, 1959;  Baggerman, i 9 6 0 ) .  a c e r t a i n time a f t e r I ^ 1  i n the  T/S  i s a d i r e c t measure, a t  i n j e c t i o n , of the I '  1  1  t h y r o i d r e l a t i v e to t h a t  was  found convenient to use  T/S  ratio;  i n the  5 1  serum.  accumulation  In t h i s study i t  a r a t i o v e r y s i m i l a r t o the  i t i s r e f e r r e d t o as the T/S  I t i s expressed  clearance  usual  r a t i o i n t h i s study.  as t h y r o i d uptake plasma ll3>l c o n c e n t r a t i o n  =  i plasma I 3 1 x  of i n j e c t e d I  1  5  1  in  thyroid  b i o l o g i c a l concentration c o e f f i c i e n t  100  -  23 -  F i g . 4. (A) Percentage accumulation of I by t h e t h y r o i d f o l l o w i n g a s i n g l e i l ^ l i n j e c t i o n a t 10° C ( a c t i v e ) and 5° 0 ( i n a c t i v e ) . Standard d e v i a t i o n s shown. Each p o i n t r e p r e s e n t s a mean o f 5 t o 7 f i s h . (B)  TUF v a l u e s f o r the same f i s h as ( A ) .  (C) T/S v a l u e s f o r the same f i s h as ( A ) . The d i p h a s i c tendency i s shown i n the a c t i v e s t a t e (Covariance a n a l y s i s , Table I V ) .  (  - 24 -  TABLE IV  A n a l y s i s o f c o v a r i a n c e f o r T/S and time (hr) f o r a c t i v e ( 1 0 ° 0) and i n a c t i v e (5° C) t h y r o i d s t a t e s  CONDITION  INACTIVE  ACTIVE  ** -  PHASE  SLOPE  df  II  0.0255 7 1 V and 0.0281 \ 56  I  0.0362  I  II  1 .and 0.0100 50  P  df  1.44  and 54  | 1 26.44** }and \ 52  F  22.5**  J  9.06*^  Denotes s i g n i f i c a n t d i f f e r e n c e between s l o p e s (p<0.01).  - 25 A b s o l u t e accumulation  r e l a t i v e t o the plasma was  higher i n  the more a c t i v e group over the e n t i r e p e r i o d of study and t h e r e f o r e showed a more complete s e p a r a t i o n of the two t h y r o i d than d i d the TUF  ( F i g . 4C).  Most i n f o r m a t i o n can be  levels  gained,  however, by a mathematical a n a l y s i s of the curves s i n c e , w h i l e the i n a c t i v e g l a n d showed a s t r i c t l y e x p o n e n t i a l r i s e w i t h  time,  the a c t i v e gland showed a c u r v i l i n e a r or d i p h a s i c r e l a t i o n s h i p . I n the a c t i v e gland an i n f l e x i o n of the curve was  apparent  between 4 and 5 days a f t e r i n j e c t i o n and consequently  two  r e g r e s s i o n s were compared s e p a r a t e l y f o r both the a c t i v e inactive states.  and  Thus the e x p o n e n t i a l r e l a t i o n s h i p s between  12 and 108 hours (phase I) and 108  and  276 hours (phase I I )  were computed f o r each temperature (Table I V ) . analysis revealed ( i ) a s t a t i s t i c a l l y  The  mathematical  significant reduction i n  slope i n the a c t i v e gland from the f i r s t  t o the second phase,  but no change i n the i n a c t i v e gland between phases, and ( i i ) t h a t phase I of the a c t i v e gland had a s l o p e  statistically  d i f f e r e n t from phase I of the i n a c t i v e g l a n d . The d i f f e r e n c e i n slope d u r i n g phase I of the two  holding  c o n d i t i o n s i s c o n s i d e r e d due t o g r e a t e r b u i l d - u p of I / ^ l  l  n  the  t h y r o i d r e l a t i v e t o t h a t i n the blood of the f i s h w i t h more a c t i v e glands.  The  change i n s l o p e of the a c t i v e gland i n  phase I I c o u l d mean e i t h e r t h a t the a f f i n i t y of the t h y r o i d f o r i o d i n e had been reduced  or t h a t t h e r e had been a l o s s of r a d i o -  a c t i v i t y from the g l a n d .  The  t h y r o i d uptake a c t i v i t y had the experiment.  Reference  former seems u n l i k e l y u n l e s s the  suddenly  changed i n the middle  t o F i g u r e 4A,  however, shows a  of  - 26 s i g n i f i c a n t drop i n the percentage of the dose i n the t h y r o i d between 8 and 12 days i n the a c t i v e but not i n the i n a c t i v e gland.  I t i s tempting t o i d e n t i f y t h i s l o s s o f r a d i o a c t i v i t y  w i t h radiohormone There  output from the a c t i v e g l a n d .  i s , however, another p o s s i b i l i t y .  Although the net  r e s u l t i s an accumulation of i o d i n e by the t h y r o i d , t h e r e are f l u x e s of I *? and I ^ 12  1  the i n f l u x o f I ^ 1  and plasma l  1  2  there i s l i t t l e  I  s a  1  5  i 1  s  high.  o  The e f f l u x , however, i s low  i n the g l a n d .  As the I ? ! ; ! ! ? 1  2  up i n the t h y r o i d and becomes lowered i n the plasma, t i o n i s r e v e r s e d and the net tendency f o r 1^31 the g l a n d i s i n c r e a s e d . slope.  Initially  i s h i g h s i n c e the r a t i o between plasma I ^ l  1  7 i  both i n t o and out of the g l a n d .  1  since  ratio  builds  the s i t u a -  t o come out of  T h i s c o u l d account f o r the change i n  The reduced i n c r e a s e i n the T/S  r a t i o i s , however,  p a r t l y due t o an output of r a d i o a c t i v e hormone as shown by s t u d i e s of the plasma P B I ^ l . From these d a t a and t h e o r e t i c a l c o n s i d e r a t i o n i t seems e v i d e n t t h a t a T/S after injection  or TUF measurement taken a t some f i x e d  time  (4 days or i n some i n s t a n c e s 8 days) should  g i v e a r e l i a b l e i n d i c a t i o n o f the e f f i c i e n c y of the t h y r o i d " i o d i d e pump  ,,  and t h a t these i n d i c e s are f a r more r e l i a b l e than  t h y r o i d uptake a l o n e .  In t h i s study most t h y r o i d measurements  were done at 4 days a f t e r i n j e c t i o n as shown i n F i g u r e 4C. t h i s same f i g u r e the i n d i v i d u a l v a l u e s of T/S are  f o r 84 and 108  shown f o r both the a c t i v e and i n a c t i v e s t a t e s .  time i n t e r v a l s the a c t i v e and i n a c t i v e T/S statistically different  In hours  At both  v a l u e s were  (p«C0.01, Wilcoxon r a n k i n g t e s t ) ,  and  t h e r e i s every reason t o b e l i e v e t h a t s i n g l e t e r m i n a l assessments  - 27 of  T/S  or TUF  f a c t o r s a t 4 days g i v e an a c c u r a t e i n d i c a t i o n of  the o v e r a l l t h y r o i d I ? ! accumulating 1  3.  efficiency.  E s t i m a t i o n of Radiohormone Output Using the Conversion As i n d i c a t e d above, i t would be l o g i c a l t o expect  Ratio  a  s i g n i f i c a n t b u i l d up i n radiohormone ( P B I  131) i n the plasma  a f t e r about 5 days i n the f i s h w i t h more a c t i v e t h y r o i d g l a n d . F i g u r e 5 shows the change i n the c o n v e r s i o n r a t i o w i t h time where, P B I ^ count/min + I ^ count/min „ . .. P B I ^ count/min x 100 Conversion r a t i o T h i s r a t i o does indeed ascend s t e e p l y at 4 days i n the a c t i v e 1  1  1  1  1  1  0  g l a n d and a f t e r 4 days the r i s e conforms t o an relationship.  exponential  That the e x p o n e n t i a l r e l a t i o n s h i p o n l y becomes  e v i d e n t a f t e r 4 days i s probably an a r t i f a c t caused by  ( i ) the  very s m a l l amounts of r a d i o a c t i v e hormone l i b e r a t e d by the gland over the i n i t i a l  p e r i o d and  ( i i ) a t e c h n i c a l l i m i t a t i o n due  the f a c t t h a t t h e r e i s always imperfect washing of P B l l 3 1 f r a c t i o n such t h a t the c o n v e r s i o n r a t i o below 0.5%. PBI 3 1  1  from the i s rarely  Thus at low l e v e l s o f radiohormone output the t r u e  l e v e l i s masked by I 3 1  1  of the P B I 3  contamination  1  f r a c t i o n , the e x p o n e n t i a l nature of the P B I 3 1  obscured  (CR)  1  and a f a l s e p i c t u r e of a l a g phase i s  output from the g l a n d and  PBr^"** removal  suggested. factors-  from the blood  due t o p e r i p h e r a l u t i l i z a t i o n and e x c r e t i o n of hormone. l a t t e r e f f e c t would cause the CR  1  build-up i s  T h i s e x p o n e n t i a l r i s e of the CR i s i n f l u e n c e d by two  T h i s was  to  exponent t o f l a t t e n w i t h  not e v i d e n t d u r i n g the 12 day o b s e r v a t i o n .  Very  The time. little  - 28 -  F i g . 5. Change i n CR f o l l o w i n g a s i n g l e I ^ injection i n f i s h a t 10° C ( a c t i v e ) a n d 3° 0 ( i n a c t i v e ) . Each p o i n t r e p r e s e n t s a mean o f 3 t o 7 f i s h . 1  1  /  /  /  /x. 20  LOG Y=LOGrf)l6+OOOI8X  /  e  Y=CR  X=HRS  /  CR  /  /  / /  IO  /  /  /  A  / /  /  y /  W /  /  /  / / /  /'x / /  /  /  /  /  4  6  DAYS  •  8  -&  • IO  i s known o f t h e r a t e f i s h and u n t i l  o f r e m o v a l o f hormone f r o m t h e b l o o d i n  t h i s has been d e t e r m i n e d i t s  c a n n o t be a s s e s s e d .  It  c a n be p o i n t e d o u t ,  on t h e CR  however,  that  r e l a t i v e l y large  quantities  of P B T A ^ P B I  i n t h e b l o o d w o u l d be s o s m a l l t h a t t h e  of P B I ^ 1  ''  1 2  of P B I ^  effect  1  1  h a v e b u i l t up t h e  until  ratio chance  b e i n g d e p l e t e d due t o p e r i p h e r a l u t i l i z a t i o n w o u l d be  1  proportionately small. would represent  Thus t h e i n i t i a l phase  l a r g e l y the r a t e  o f t h e CR c u r v e  of radiohormone p r o d u c t i o n .  131 As w i t h o t h e r a s p e c t s of I  y  m e t a b o l i s m , change  i n plasma  PBI131 l e v e l s  o o u l d p r o b a b l y be r e p r e s e n t e d most p r e c i s e l y  terms of r a t e  constants.  extent  In this entire  s t u d y , however,  o f PB-T131 p r o d u c t i o n was a s s e s s e d b y k i l l i n g t h e  8 days a f t e r sample.  in  the fish  i n j e c t i o n a n d t h e n m e a s u r i n g t h e CR o f a p l a s m a  At t h i s  time  PBI131 l e v e l s , are becoming  readily  detectable  i n t h e plasma and a l s o r e v e a l t h e v e r y marked  difference  between the a c t i v e  It  is  and i n a c t i v e t h y r o i d s  5).  i m p o r t a n t t o n o t e t h a t t h e CR o n l y i n d i c a t e s t h e  o f radiohormone p r o d u c t i o n and not the r a t e production.  (Fig.  To i n d i c a t e t h e l a t t e r ,  t h e I ^ l must  e q u i l i b r i u m w i t h I^ 7  throughout  repetitive  to maintain a constant  2  injections  c e r t a i n l y not achieved  of stable  the body.  i n these studies  This  rate  hormone  attain  requires  I ^ l l e v e l and  as a r e s u l t  of a  is  single  injection. I n summary, b y m a k i n g r o u t i n e m e a s u r e m e n t s in  of TI^I  s e v e r a l body compartments a t f i x e d t i m e i n t e r v a l s ,  levels  a great  d e a l c a n be l e a r n e d a b o u t r a d i o i o d i n e m e t a b o l i s m a n d t h y r o i d function.  As a r e s u l t  of these p r e l i m i n a r y . i n v e s t i g a t i o n s  it  -  30 -  was d e c i d e d t o d e t e r m i n e r o u t i n e l y t h e p a r a m e t e r s T a b l e V at 4 and 8 days a f t e r •II.  i n j e c t i o n of  shown i n  I ^ . 1  1  SEASONAL CHANGES I N RADIOIODINE METABOLISM AND THE H I S T O LOGICAL APPEARANCE OF THE THYROID I N J U V E N I L E STEELHEAD HELD I N FRESH WATER F o l l o w i n g the p r e l i m i n a r y i n v e s t i g a t i o n of I ^ ! metabolism 1  c e r t a i n of the parameters seasonally. measured.  Mean c e l l h e i g h t a n d c o l l o i d These d a t a ,  temperature, !•  I ? 1  together  c o l o u r were  with seasonal  also  changes i n water  a r e s u m m a r i z e d i n F i g u r e 6.  Excretion  1  Plasma i l ^ l (4 days) (4 d a y s ) ,  and percentage  o f dose  revealed d e f i n i t e seasonal trends.  pre-migrant year u n t i l negative  shown i n T a b l e V w e r e d e t e r m i n e d  i n t o t a l body  F r o m May o f  s m o l t i f i c a t i o n t h e r e was a  the  general  c o r r e l a t i o n b e t w e e n t h e s e two p a r a m e t e r s a n d  temperature,  i n d i c a t i n g more r a p i d r e m o v a l o f 1-131 at higher This negative  c o r r e l a t i o n i s d e m o n s t r a t e d i n F i g u r e 7 where  mean s e a s o n a l v a l u e s against  temperatures.  temperature  for total ( r = -0.77,  r e t e n t i o n have been p l o t t e d 26df, p<£ 0.001).  portant to note that these seasonal  It  c h a n g e s i n 1"131  i s imrate  of  e x c r e t i o n were n o t o n l y d e m o n s t r a t e d by t o t a l p l a s m a  clearance  but a l s o by e x t r a t h y r o i d a l c l e a r a n c e  indicates  that the l o s s  ( F i g . 7).  o f I ? ! f r o m t h e plasma b o t h t o t h e t h y r o i d and 1  t o t h e o u t s i d e medium was i n c r e a s e d a t h i g h e r These  This  observations  investigations.  temperatures.  support the data from the p r e l i m i n a r y  TABLE V A summary o f r a d i o i o d i n e parameters and t h e i r s i g n i f i c a n c e ABBREVIATION USED  PARAMETER I  II  Plasma l!31 b i o l o g i c a l concentration coefficient/100  Measures plasma I ^ concentrat i o n and a l l o w s estimates of t o t a l blood c l e a r a n c e o f 1131 1  Plasma 1"131  % i n j e c t e d l ! 3 1 dose i n body (excluding t h y r o i d )  % body  I I I fo i n j e c t e d 1"131 dose i n ?' whole body ( i n c l u d i n g thyroid)  IT  SIGNIFICANCE  f» body + fo t h y r o i d  i injected i W dose i n t h y r o i d Plasma H 3 1  1  Measures t o t a l c l e a r a n c e o f 1131 from the body Measures the f» o f i n j e c t e d dose i n whole body. 100 fo - fo i n whole body = % l o s s due t o e x t r a t h y r o i d a l clearance (excretion) A s s e s s e s b u i l d - u p o f 1^31 j _ the t h y r o i d r e l a t i v e t o plasma l l 3 1 . — measures t h e a f f i n i t y o f the t h y r o i d f o r I 3 l and allows f o r e x c r e t i o n r a t e n  T/S  x  V  % dose t h y r o i d % dose body + f» dose t h y r o i d  VI  Conversion r a t i o Plasma P B I ? 1  1  T h y r o i d uptake factor TUF  CR  x 100  Plasma l ! 3 1 + plasma PBI 31 X  Measures that 1 o f the unexcreted 1"131 t h a t i s p a r t i t i o n e d i n t o the t h y r o i d A s s e s s e s extent o f c o n v e r s i o n of 1131 t o P B l i 3 1  -  32  -  F i g . 6. S e a s o n a l c h a n g e i n p l a s m a I ? ! , % d o s e I ? , T / S , T U F , CR, mean e p i t h e l i u m c e l l h e i g h t and p e r c e n t a g e o f f o l l i c l e s c o n t a i n i n g b l u e c o l l o i d i n s t e e l h e a d p a r r and s m o l t s . Seasonal change i n t e m p e r a t u r e i s shown. S t a n d a r d d e v i a t i o n s a r e shown for c e r t a i n parameters. 1  1  1  - 33 -  F i g . 7. C o r r e l a t i o n s b e t w e e n w a t e r t e m p e r a t u r e a n d f o u r 1131 p a r a m e t e r s . A l l v a l u e s drawn f r o m s e a s o n a l d a t a ( F i g . 6). A (% d o s e i n b o d y ) ; B (CR); C (T/S); D (TUF).  66 40t-  r-=o-77  T - O . 7 8  T/S  4 0 h  8  5*  20t-  t  •. " •% • • •  32  O  B T  60(-  =ae9  c O - 4 6  TUF CR 2 0 U  —1 IO  12  8  IO  °c  L. 12  - 34 O f f u r t h e r s i g n i f i c a n c e was t h e d e p a r t u r e o f c e r t a i n g r o u p s of f i s h from the general c o r r e l a t i o n .  These f e l l  c a t e g o r i e s - f i s h b e l o w 20 g i n w e i g h t  ( u s u a l l y 18 m o n t h s o r  younger) fish,  and s m o l t s  (30-31 m o n t h s ) .  the low r e t e n t i o n of I ^ 1  1  I n t h e younger and s m a l l e r  indicated differences  be a t t r i b u t a b l e t o s i z e o r g r o w t h . later. total  Over the p e r i o d of clearance  i n t o two  These f i s h a r e  that  could  reconsidered  s m o l t i f i c a t i o n , however,  both  the  a n d t h e e x t r a t h y r o i d a l c l e a r a n c e w e r e more  r a p i d t h a n a t any o t h e r t i m e of y e a r .  At the time of  anticipated  m i g r a t i o n i n May t h e w a t e r t e m p e r a t u r e was 8 t o 9° C a n d a t t h i s t e m p e r a t u r e t h e p l a s m a a n d b o d y I ^ ! l e v e l s w e r e much 1  l o w e r t h a n t h e p l a s m a a n d b o d y I ^ ! r e t e n t i o n s a t 12 t o 13° C 1  i n p r e - m i g r a n t s i n September of the p r e v i o u s y e a r . indicate that  some f a c t o r  i n f l u e n c i n g t h e I 31 x  2.  This would  o t h e r t h a n t e m p e r a t u r e m i g h t be  excretion rate  i n steelhead  at s m o l t i f i c a t i o n .  Thyroid Uptake T / S a n d TUF v a l u e s a l s o showed t e m p e r a t u r e c o r r e l a t i o n s  w i t h t h e same t w o e x c e p t i o n s The t h y r o i d I ^ 1  1  ( s m a l l f i s h and s m o l t s )  but 6).  (Fig.  uptake c o r r e l a t e d p o s i t i v e l y w i t h temperature  o v e r t h e s e a s o n a l r a n g e o f 4 t o 13°  C at h i g h l y  significant  s t a t i s t i c a l l e v e l s ( F i g . 7 ) ( T / S , r = 0.78, 26df, p<0.001;  T U F , r = 0.89, 26df, p < 0 . 0 0 l ) . 3.  Conversion R a t i o CR v a l u e s  ( r = 0.46,  showed a g e n e r a l i n c r e a s e w i t h t e m p e r a t u r e  rise  l8df, p<0.05), b u t t h e e f f e c t was n o t a s m a r k e d a s  w i t h t h y r o i d uptake  ( F i g s . 6 and 7).  Of p a r t i c u l a r n o t e w e r e  - 35 the v e r y h i g h CR v a l u e s o f smolts a t 8 t o 9° C.  These v a l u e s  were c o n s i d e r a b l y h i g h e r than any recorded d u r i n g the e n t i r e study.  No CR estimates were a v a i l a b l e on y e a r l i n g f i s h below  20 grams. 4.  H i s t o l o g i c a l Changes Whereas a l l l ! 3 1 parameters were p o s i t i v e l y c o r r e l a t e d w i t h  temperature and suggested  a p o s s i b l e temperature dependence, t h e  c e l l h e i g h t parameter i n d i c a t e d no such g e n e r a l c o r r e l a t i o n (Fig.  6).  I t was c o n c l u s i v e l y demonstrated t h a t i n y e a r l i n g  s t e e l h e a d t h e lowest c e l l h e i g h t s occurred a t the h i g h e s t and not t h e lowest temperature,  and much o f the: seasonal data  i n d i c a t e d a tendency f o r h i g h e r c e l l h e i g h t a t lower  temperatures.  In other words, t h e r a d i o i o d i n e and c e l l h e i g h t parameters showed o p p o s i t e t r e n d s w i t h r e s p e c t t o temperature.  Of some  importance,  however, was the s i g n i f i c a n t n e g a t i v e c o r r e l a t i o n between mean v a l u e s f o r c e l l h e i g h t and mean v a l u e s f o r percentage o f follicles  c o n t a i n i n g blue c o l l o i d  ( r = -0.66, l l d f , p<0.05).  T h i s i n d i c a t e d t h a t the c e l l h e i g h t data were not i n agreement w i t h t h e data obtained from c o l l o i d c o l o u r , s i n c e i n c r e a s i n g c e l l h e i g h t i s a s s o c i a t e d w i t h an a c t i v e gland, and absence o f b l u e c o l l o i d i n d i c a t e s decreased It  thyroid  activity.  i s e v i d e n t from t h i s b r i e f survey o f 'seasonal changes  i n 1131 metabolism and h i s t o l o g i c a l change i n s t e e l h e a d t h a t t h e r e i s no a b s o l u t e p i o t u r e o f what might be a f f e c t i n g t h e thyroid. I ^! 1  Temperature seems t o be c o r r e l a t e d w i t h changes i n  metabolism.  But, i s there a c a u s a l r e l a t i o n s h i p between  t h y r o i d a c t i v i t y and changing  temperature?  Furthermore, the  - 36 smolts and s m a l l f i s h showed d e p a r t u r e s from t h i s g e n e r a l w h i l e the c e l l height  trend,  data r e v e a l e d a c o r r e l a t i o n p o s s i b l y i n  the r e v e r s e d i r e c t i o n , and showed a t r e n d o p p o s i t e obtained  by the c o l l o i d s t a i n i n g .  seasonal  v a r i a b l e s including photoperiod.  t o that  There a r e a l s o other  important  C y c l i c a l changes i n  r a d i o i o d i n e metabolism independent o f environmental i n f l u e n c e might a l s o be envisaged.  The r o l e s o f v a r i o u s f a c t o r s i n f l u e n c i n g  the t h y r o i d t h e r e f o r e appear u n c e r t a i n and confused.  In the  remainder o f t h i s study, an attempt has been made t o i s o l a t e experimentally  t h e v a r i o u s components t h a t might i n f l u e n c e the  t h y r o i d and t o e x p l a i n as f u l l y as p o s s i b l e t h e seasonal described IT I.  changes  above. EXPER MENTAL INVESTIGATION 0? THE INFLUENCE OF  TEMPERATURE ON THE HISTOLOGICAL APPEARANCE OF THE THYROID AND RADIOIODINE METABOLISM IN YEARLING STEELHEAD PARR In t h e seasonal  study o f t h y r o i d f u n c t i o n c o r r e l a t i o n s  between r a d i o i o d i n e metabolism and temperature were e s t a b l i s h e d . From such c o r r e l a t i o n s i t might be p r e d i c t e d t h a t temperature causes changes i n r a d i o i o d i n e metabolism.  I n t h i s aspect  o f the  study i t was hoped t o t e s t t h i s p o i n t by h o l d i n g f i s h under c o n d i t i o n s which were d i f f e r e n t i n temperature but s i m i l a r i n a l l other  respects.  To a s s e s s e x p e r i m e n t a l l y  the e f f e c t s o f temperature on  t h y r o i d f u n c t i o n , 12 f i s h were put i n t o an open system o f running water a t temperature, regimes o f 6, 9, 12, 13 and 18° C, a l l h e l d t o +0.05° C.  The h o l d i n g temperature p r i o r t o t h e a c c l i m a t i o n  - 37 was  9 + 0.5°  -  G and the a c c l i m a t i o n temperature was  not  imposed  suddenly but b u i l t up over a p e r i o d of s e v e r a l hours.  On  18  J u l y , 14 days a f t e r i n i t i a t i o n t o the regimes, a l l the f i s h were injected with I - ^ l . days and  S i x were k i l l e d from each c o n d i t i o n at f o u r  d at e i g h t days.  t o 11 and The  i n Table  The  r e s u l t s are shown i n F i g u r e s 8  VI.  e f f e c t of temperature on c e l l h e i g h t and r a d i o i o d i n e  metabolism of Salmo g a i r d n e r i has  been i n v e s t i g a t e d by  (1955a, b ) .  found to be h i g h e s t between  9 and  T h y r o i d a c t i v i t y was  12° C u s i n g r a d i o c h e m i c a l t e c h n i q u e s .  c r i t e r i a , g r e a t e s t a c t i v i t y was tures.  and h i s t o l o g i c a l  I n the present t o t h a t claimed was  With h i s t o l o g i c a l  i m p l i e d at the lower tempera-  A t 20° C the t h y r o i d a c t i v i t y was  chemical  Olivereau  low by both r a d i o -  criteria.  study,  c e l l h e i g h t showed a t r e n d  by O l i v e r e a u .  By t h i s c r i t e r i o n , the  identical gland  s i g n i f i c a n t l y more a c t i v e at lower temperatures ( r = -0.84,  9df, p<0.01) ( F i g . 8 ) .  However, t h e r e was  a  statistically  s i g n i f i c a n t p o s i t i v e c o r r e l a t i o n between i n c r e a s i n g blueness c o l l o i d and  of  i n c r e a s i n g temperature, i n d i c a t i n g a more a c t i v e  t h y r o i d at h i g h e r temperatures by t h i s c r i t e r i o n  (r -  0.71,  9df, p<0.02) ( F i g . 8 ) . In r e g a r d t o the uptake o f r a d i o i o d i n e ( F i g . 8 ) , both 4 and  8 day  estimates  uptake at 12° C. autoradiography, obtained  showed a very s i m i l a r p a t t e r n w i t h maximum  O l i v e r e a u (1955a) measured I ^ l uptake by which i s e s s e n t i a l l y the same technique,  a similar result.  I t has a l r e a d y been p o i n t e d  and out,  however, t h a t l A ^ l uptake by the t h y r o i d i s only a v a l i d measure  - 38 -  F i g . - 8. E x p e r i m e n t a l d e m o n s t r a t i o n o f t h e i n f l u e n c e o f t e m p e r a t u r e on p e r c e n t a g e a c c u m u l a t i o n o f I ? i n the t h y r o i d 4 a n d 8 d a y s a f t e r a s i n g l e 1.131 i n j e c t i o n , on mean c e l l h e i g h t (O) a n d on p e r c e n t a g e o f f o l l i c l e s c o n t a i n i n g b l u e c o l l o i d (•). 1  1  - 39 o f t h e t h y r o i d a f f i n i t y f o r i o d i n e i f a l l o w a n c e i s made f o r rate  of extrathyroidal I ^ 1  on m a x i m a l I ^ 1  1  uptake at  1  clearance.  Thus t h e above  the  conclusions  C c o u l d be i n v a l i d a s a n i n d i c a t i o n  12°  of t h y r o i d a c t i v i t y . F i g u r e 9 shows t h e e f f e c t  o f t e m p e r a t u r e on i - ^ l r e m o v a l  f r o m t h e body as a whole and f r o m t h e plasma a t d i f f e r e n t eratures  (4 a n d 8 d a y s ) .  I ^ l retention falls  exponentially  w i t h temperature.  Some a t t e m p t a t a n a n a l y s i s o f t h e s e  has been p r e s e n t e d  (Table V I ) .  4 and 8 day e s t i m a t e s not d i f f e r  curves  I t would appear that both  of I / ^ l r e t e n t i o n i n d i c a t e slopes  s i g n i f i c a n t l y f r o m each o t h e r .  correlation coefficients  temp-  the  that  do  However, the h i g h  emphasize a s i g n i f i c a n t i n c r e a s e  in  ll31 l o s s w i t h t e m p e r a t u r e . F i g u r e 10 shows t h e a p p r o x i m a t e e x p o n e n t i a l i n c r e a s e T/S w i t h t e m p e r a t u r e .  T h e CR v a l u e s f o r t h e same f i s h  a s i m i l a r e x p o n e n t i a l change coefficients difference It  ( F i g . 11).  in  showed  High correlation  w e r e f o u n d f o r t h e r e l a t i o n s h i p a n d no s t a t i s t i c a l  was e v i d e n t b e t w e e n t h e 4 a n d 8 d a y s l o p e s  i s concluded that temperature accelerates  (Table V I ) .  a l l aspects  o f i l ^ l m e t a b o l i s m b e t w e e n 6 a n d 18° C a n d t h a t r a d i o c h e m i c a l ^ the t h y r o i d i s not maximally a c t i v e suggested. reverse  9 t o 12°  C as  Olivereau  Where h i s t o l o g i c a l c r i t e r i a a r e c o n c e r n e d ,  seems t o be t r u e s i n c e t h e c e l l h e i g h t  gland a c t i v i t y at however,  at  lower temperatures.  indicated  The c o l l o i d  general discussion.  greater  colour,  conformed w i t h the r a d i o c h e m i c a l e s t i m a t e s .  h a s b e e n made t o r e s o l v e  the  An attempt  these very important anomolies i n the  - 40 -  F i g . 9. E x p e r i m e n t a l demonstration of the i n f l u e n c e of temperature on l e v e l s i n plasma and t o t a l body 4 days a f t e r a s i n g l e I 3 1 i n j e c t i o n . Each p o i n t r e p r e s e n t 4 t o 7 i n d i v i d u a l s (Covariance a n a l y s i s , T a b l e VI) . x  r PLASMA O —  Q io  O tj  O 0\  —  w  M  i 4  I  S %  BODY  cn  L  8  _ O  - 41 -  TABLE V I A n a l y s i s o f covariance f o r the r e l a t i o n s h i p s between v a r i o u s I * ! parameters and temperature (° C) 1  Parameter  Time (Days) 4  CR  I ^ l  i  PLASMA  BODY + % THYROID  Slope  df  0.130  1 and 46  1.14  1 and 46  1.08  1 and 46  0.12  8  0.172  4  -0.226  8  -0.285  4  -0.0723  8  P  -0.0818  Denotes s i g n i f i c a n c e  (p^.0.01).  Correlation coefficient  0.797** 0.710**  -0.813** -0.790**  -O.665** -0.584**  - 42 -  E i g . 10. E x p e r i m e n t a l demonstration of the i n f l u e n c e of temperature on T/S v a l u e s 4 and 8 days a f t e r a s i n g l e I 31 i n j e c t i o n . Each p o i n t r e p r e s e n t s a mean of 4 t o 7 i n d i v i d u a l s . x  - 43 -  E i g . 11. E x p e r i m e n t a l demonstration of the i n f l u e n c e of temperature on CR v a l u e s 4 and 8 days a f t e r a s i n g l e 1131 i n j e c t i o n . Each p o i n t r e p r e s e n t s a mean of 4 t o 7 i n d i v i d u a l s (Covariance a n a l y s i s , T a b l e V I ) .  r  -  IV.  44  -  EXPERIMENTAL INVESTIGATION OF TEMPERATURE AND INFLUENCE ON THYROID CELL HEIGHT AND  PHOTOPERIOD  RADIOIODINE METABOLISM  OF STEELHEAD YEARLINGS (January t o J u l y ) The p r e v i o u s seasonal o b s e r v a t i o n s and experiments t h a t temperature the t h y r o i d .  i s a major environmental  factor  suggest  influencing  T h i s does not exclude, however, the i n f l u e n c e  of other v a r i a b l e s , prominent among which c o u l d be the photoperiod.  Photoperiod i s important  i n controlling  changing certain  endocrine changes and the s p r i n g i n c r e a s e i n r a d i o i o d i n e metabolism i n y e a r l i n g and two-year-old  s t e e l h e a d c o u l d be  i n f l u e n c e d by the i n c r e a s i n g s p r i n g d a y l i g h t hours. t h i s p o s s i b i l i t y , an experiment  was  conducted  To  from l a t e  test January  to early July. F i s h were h e l d under 4 experimental regimes i n 2 m x 50  cm  x 30 cm concrete troughs w i t h a c o n t i n u a l l y r e p l e n i s h e d supply of d e c h l o r i n a t e d water. temperature  was  conditions.  In two  of the troughs the water  not c o n t r o l l e d and approximated  In the o t h e r two tanks i t was  t a i n e d between 5 and 6° G.  p r e - c o o l e d and main-  A l l 4 tanks were covered w i t h  p r o o f boxes i l l u m i n a t e d by f l u o r e s c e n t b u l b s . maintained  to the outside  on an 8-hour day and two  Two  light-  of them were  on a n a t u r a l daylength  u s i n g the c o n t r o l mechanism mentioned i n methods (Table V I I ) . F i s h were sampled r o u t i n e l y f o r h i s t o l o g i c a l and r a d i o i o d i n e determinations at approximately monthly i n t e r v a l s . The 1^-31 of temperature There was  plasma l e v e l s showed a c l e a r s e p a r a t i o n on the b a s i s as d i d the e x t r a t h y r o i d a l c l e a r a n c e ( F i g .  no n o t i c e a b l e suggestion of a p h o t o p e r i o d  12).  influence.  - 45 -  TABLE Y I I  Summary of the f o u r combined temperature and p h o t o p e r i o d c o n d i t i o n s . Symbols shown a r e as used in figures  TANK  SYMBOL  PHOTOPERIOD  TEMPERATURE  1  •  Natural  Natural  2  A  8 hours  O  Natural  A  8 hours  4  Controlled (5 - 6° C)  - 46 -  F i g . 12. E x p e r i m e n t a l demonstration of the combined i n f l u e n c e of temperature and p h o t o p e r i o d on plasma l!31 l e v e l s and percentage dose i n t o t a l body 4 days a f t e r a s i n g l e 1131 i n j e c t i o n i n 14 t o 18-month s t e e l h e a d . Each p o i n t r e p r e s e n t s a mean o f 8 t o 12 f i s h ( S t a t i s t i c s , Table V I I I ) .  - 47 The T/S  showed a s i m i l a r i n c r e a s e d response t o r i s i n g  temperature  hut showed no response t o i n c r e a s i n g p h o t o p e r i o d ( F i g . 13). comparing  By-  the f o u r c o n d i t i o n s i n June when the g r e a t e s t d i v e r -  gence between the treatment p r e v a i l e d , i t was not p o s s i b l e t o demonstrate a s i g n i f i c a n t e f f e c t of p h o t o p e r i o d , but a v e r y s i g n i f i c a n t i n f l u e n c e of temperature on I ^ ! metabolism was  shown  1  (Table V I I I ) .  In support o f the e a r l i e r f i n d i n g s , however, the ( F i g . 13)  c e l l h e i g h t d i d not agree w i t h the r a d i o i o d i n e parameters  and i n g e n e r a l the f i s h h e l d at the lower temperature had the higher c e l l height.  In f a c t , i g n o r i n g p h o t o p e r i o d which d i d not  appear t o be i n f l u e n c i n g the c e l l h e i g h t s i g n i f i c a n t l y , the means of a l l f i v e s e a s o n a l samples  of cold-adapted f i s h had a h i g h e r  c e l l h e i g h t than those s u b j e c t e d t o the i n c r e a s i n g temperature. C o l l o i d c o l o u r e s t i m a t e s have not been i n c l u d e d .  They were  extremely v a r i e d and r e v e a l e d no apparent t r e n d through the f o u r conditions.  C o n v e r s i o n r a t i o s were not a v a i l a b l e from these  experiments s i n c e a t the time they were conducted i t was r e a l i z e d how  slowly the P B I ^ 1  1  not  was produced and no 8-day r e a d i n g s  were taken. So f a r , e s t i m a t e s of t h y r o i d a c t i v i t y have been based  solely  on;;single t e r m i n a l measurements and do not i n c l u d e the CR.  In  order t o r e a c h more d e f i n i t e c o n c l u s i o n s c o n c e r n i n g the e f f e c t s of p h o t o p e r i o d and temperature on y e a r l i n g f i s h , s e r i a l  investi-  g a t i o n s were c a r r i e d out from these same c o n d i t i o n s i n e a r l y July  (high temperature and l o n g p h o t o p e r i o d under the n a t u r a l  conditions). with I ^ 1  1  F i f t y t o 55 f i s h from each c o n d i t i o n were i n j e c t e d  and 6 from each group k i l l e d a t the f o l l o w i n g  intervals:  - 48 -  F i g . 13. Experimental demonstration o f the combined i n f l u e n c e o f photoperiod and temperature on mean c e l l h e i g h t and T/S r a t i o s 4 days a f t e r a s i n g l e 1"131 i n j e c t i o n i n 14 t o 18-month s t e e l h e a d . C e l l h e i g h t - each p o i n t r e p r e s e n t s a mean o f 3 t o 4 f i s h ; T/S - each p o i n t r e p r e s e n t s a mean of 8 t o 12 f i s h ( S t a t i s t i c s , Table Y I I I ) .  - 49 -  TABLE V I I I  Summary o f s t a t i s t i c a l l y s i g n i f i c a n t d i f f e r e n c e s f o r s e v e r a l i l j l parameters between d i f f e r e n t c o n d i t i o n s of temperature and photoperiod i n y e a r l i n g s i n June. White-Wilcoxon non-parametric ranking t e s t used  HIGH TEMPERATURE Photoperiod normal  I  1  5  1  plasma  Photoperiod 8 hours  LOW TEMPERATURE Photoperiod Photoperiod normal 8 hours  L ++  fo dose body % dose t h y r o i d ++  T/S  ? + ++  Two Two Two Two  ++  groups groups groups groups  not s t a t i s t i c a l l y d i f f e r e n t (p-^0.05). almost s t a t i s t i c a l l y d i f f e r e n t (p = 0 . 0 5 ) . s i g n i f i c a n t l y d i f f e r e n t (p^.0.05). significantly different (p<0.01).  - 50 12,  36,  60,  84,  108,  132,  156,  204 and 276 h r a f t e r  Data are shown i n F i g u r e s 14 - 16 marizes the s t a t i s t i c s .  inclusive;  injection.  T a b l e IX sum-  The r e s u l t s obtained by the s i n g l e  4-day measurements were confirmed.  P h o t o p e r i o d a t t h i s time o f  year e x e r t e d no i n f l u e n c e on plasma I - ^ l c l e a r a n c e nor on t h y r o i d uptake, though temperature had a d e f i n i t e ( P i g s . 14 and 1 5 ) .  At the h i g h e r temperature, the c h a r a c t e r i s t i c  d i p h a s i c t h y r o i d uptake curve was  evident.  A similar  t a t i o n i s d e r i v e d from c o n s i d e r a t i o n o f the CR was  effect  interpre-  ( F i g . 16).  There  a d i s t i n c t s e p a r a t i o n between the e f f e c t s of the two  temperature c o n d i t i o n s but no d i f f e r e n t i a l response t o photoperiod. I t i s concluded t h e r e f o r e that i n y e a r l i n g  steelhead  ( i ) i n c r e a s i n g temperature s t i m u l a t e s r a d i o i o d i n e metabolism and yet causes a decrease i n c e l l h e i g h t ,  ( i i ) t h e r e i s no  i n f l u e n c e of i n c r e a s i n g p h o t o p e r i o d on the s t e e l h e a d t h y r o i d under the c o n d i t i o n s i n v e s t i g a t e d and  ( i i i ) although the f i s h  131 under the i n c r e a s i n g temperature show a h i g h e r r a t e o f I ^ metabolism than f i s h a t the lower temperature, the i n c r e a s i n g temperature regime induces r e l a t i v e l y l i t t l e change.  The  divergence between the e f f e c t s of the two regimes was not due only t o changes temperature.  in r^l  metabolism i n the f i s h under the v a r i a b l e  In f a c t , w i t h I - ^ l e x c r e t i o n data, the g r e a t e r  changes were found i n the f i s h h e l d under constant temperature. T h i s p o i n t s t o some change w i t h i n the f i s h not governed by temperature and p h o t o p e r i o d . used over this* p e r i o d was  What was  common t o a l l the f i s h  their rapidly increasing size.  p o s s i b i l i t y i s t h a t t h i s i s a phase of r a p i d growth  One  requiring  - 51 -  F i g . 14. E x p e r i m e n t a l d e m o n s t r a t i o n o f t h e c o m b i n e d i n f l u e n c e o f p h o t o p e r i o d a n d t e m p e r a t u r e on p l a s m a c l e a r a n c e f o l l o w i n g a s i n g l e I ? ! i n j e c t i o n i n 18-month s t e e l head. E a c h p o i n t r e p r e s e n t s a mean o f 5 t o 7 f i s h (Covariance a n a l y s i s , Table I X ) . 1  DAYS  - 52 -  F i g . 15. E x p e r i m e n t a l d e m o n s t r a t i o n o f t h e c o m b i n e d i n f l u e n c e o f p h o t o p e r i o d and t e m p e r a t u r e on T/S v a l u e s i n 18-month s t e e l h e a d f o l l o w i n g a s i n g l e I l 3 1 i n j e c t i o n . E a c h p o i n t r e p r e s e n t s a mean o f 5 t o 7 f i s h ( C o v a r i a n c e a n a l y s i s , Table EC).  - 53 -  F i g . 16. E x p e r i m e n t a l demonstration of the combined i n f l u e n c e of photoperiod and temperature on CR v a l u e s f o l l o w i n g a s i n g l e 1^31 i n j e c t i o n i n 18-month s t e e l h e a d . Each p o i n t r e p r e s e n t s a mean of 5 t o 7 i n d i v i d u a l s .  8L ST A  TABLE DC A n a l y s i s of c o v a r i a n c e performed on plasma l 5 1 and time ( h r ) , and T/S and time (hr) under f o u r d i f f e r e n t p h o t o p e r i o d and temperature regimes x  PLASMA I ? 1  Temp. ° C 5  Photoperiod  Phase  Slope  8  -  -0.013]  16  -  -0.014 J  -  -0.0171 1 \ and - 0 . 0 1 6 J 102  8  10  16  df  I  P  I and 97  ' 0.02  K  *  16  I II I II  ,  •  n  8  ± v  16  I II I II  F  0.026 7 1 \ and 0.021J 56 0.027 7 I and 0.019) 48  s  1.44  1  0.033 7 1 I and 0.014) 38 O.0367 1 > and O.OIOJ 50  l  \ and  J  99  n  1  f and  S 104 T/S df 1 • and 56  E  df  m  1 0.44 . and 105  s  6.67*  1 and - O.63 94  I.65  E  F  df  s  1 0.07 .and 55  F  s  4.48*  3.08  26.46** 26.40**  Fg F  = Variance r a t i o f o r slopes. » V a r i a n c e r a t i o f o r a d j u s t e d means.  * *  Significant difference Significant difference  m  l  0.?4  df 8  df  s  1  (p<0.05). (p<0.01).  1 1 - and 53  0.24  , 1 and 54  n  2.58  - 55 a greater is  t h y r o x i n demand, b u t a n e q u a l l y p l a u s i b l e  t h a t the changes i n I ^ 1  metabolism under constant  1  were s i m p l y a f u n c t i o n of i n c r e a s i n g V.  hypothesis conditions  size.  E F F E C T OF BODY MASS ON RADIOIODINE METABOLISM  To t e s t t h e e f f e c t  o f b o d y mass o n I ^ 1  groups of y e a r l i n g steelhead  1  metabolism,  two  o f t h e same age w e r e t a k e n i n m i d -  June from t h e f o u r h o l d i n g c o n d i t i o n s of combined temperature and p h o t o p e r i o d u s e d a b o v e .  12  consisted of  large  i n j e c t e d w i t h 0.02 of  12  w i t h 0.01  g,  av =  5.4  g)  that large  small fish  fish  after  b e t w e e n t h e s m a l l and  a l t h o u g h b o t h showed t h e e x p e c t e d  temperature and l a c k of response fact  injected  o f t h e i n j e c t e d dose r e t a i n e d i n t h e  e n t i r e b o d y was c o n s i s t e n t l y d i f f e r e n t large f i s h ,  96 h o u r s  were  consisted  and t h e s e were  A l l f i s h were k i l l e d  The p e r c e n t a g e  these  The o t h e r g r o u p  1  g,  19.0 g ) ;  av -  m l o f I ^ ! (3 JIG).  ml ( l ^ i o ) .  injection.  (11-65  fish  (3.2-8.0  small f i s h  One g r o u p f r o m e a c h c o n d i t i o n  dependence  on  t o p h o t o p e r i o d ( F i g . 17).  The  tended t o r e t a i n i n j e c t e d I ^ ! longer  than  1  e x p l a i n s why f i s h h e l d u n d e r c o n s t a n t  conditions  showed c h a n g e s i n i o d i n e m e t a b o l i s m a s t h e s e a s o n p r o g r e s s e d and t h e a v e r a g e mass i n c r e a s e d . temperature  The t e n d e n c y f o r  to increase the r a t e  of  I 3l x  increasing  l o s s was a p p r o x i m a t e l y  b a l a n c e d b y t h e t e n d e n c y f o r a g r o w i n g f i s h t o r e t a i n more i o d i n e due t o s l o w e r e x c r e t i o n . yearling fish  is  A t a low constant  Thus t h e a p p a r e n t anomoly i n t h e  e x p l a i n e d on t h e b a s i s temperature,  it  o f a change  i n body mass.  i n d u c e d a change t h a t w o u l d  o t h e r w i s e not occur and under the i n c r e a s i n g temperature the  size  - %-  F i g . 17. I n f l u e n c e o f b o d y mass on T/S v a l u e s . a n d percentage of I ^ l i t h e t o t a l b o d y 84 h o u r s a f t e r a s i n g l e 1.131 i n j e c t i o n i n 19-month s t e e l h e a d . n  5-5 °C  II°C I6HRS  8 HRS  80-  a  60-  m  40-  O  o  20-  o  80-  T/S  •  i  4020-  j^J  SMALL  o o  LARGE  16 HRS  8 HRS  effect  almost e x a c t l y  37  -  c a n c e l l e d the stimulus of  increasing  temperature. Measurement o f T/S r a t i o s i n g e n e r a l that  l l 3 1 m e t a b o l i s m was more a c t i v e  1.131  excretion,  idea  i n a smaller f i s h .  t h e i n d e p e n d e n c e o f p h o t o p e r i o d and  on t e m p e r a t u r e were e v i d e n t . tion,  supported the  As w i t h  dependence  W i t h t h e e x c e p t i o n o f one c o n d i -  s m a l l f i s h had h i g h e r T/S r a t i o s t h a n l a r g e f i s h .  no f i s h w e r e h e l d t o 8 d a y s , To v e r i f y  no CR v a l u e s w e r e  available.  some o f t h e a b o v e f i n d i n g s , a n o t h e r  was c o n d u c t e d o n 3 g r o u p s o f y e a r l i n g s t e e l h e a d 11° C (Table 2).  A l l f i s h were k i l l e d  t o o b t a i n CR v a l u e s , Again the r a t e  experiment  i n September  8 days a f t e r  injection  o f e x c r e t i o n a n d J.131 a c c u m u l a t i o n w e r e  ( F i g . 18);  the smaller s i z e  at  l a c k i n g from the previous determinations.  i n t h e same way b y b o d y mass b u t two f u r t h e r emerged  Since  (i)  1  affected  important points  t h e CR was a l s o m a r k e d l y s t i m u l a t e d i n  range and  ( i i ) w i t h each o f t h e 3 parameters  of J131 m e t a b o l i s m t h e r e was a g r e a t s i z e 16 grams a n d l i t t l e  effect  effect  between 8 and  b e t w e e n 16 a n d 30 g r a m s .  The  phenomenon may be e x p l a i n e d i n t e r m s o f a l o g b o d y mass thyroid activity relationship. cant negative  signifi-  c o r r e l a t i o n s when t h e d a t a w e r e t r a n s f o r m e d i n  t h i s manner ( F i g . 19).  Such a t r a n s f o r m a t i o n i s  c a l l y u s e d t o show d e p e n d e n c e fact  B o t h T / S a n d CR showed  log  of metabolic rate  that r a d i o i o d i n e parameters  characteristion mass and t h e  of t h y r o i d a c t i v i t y conform to  t h i s r e l a t i o n s h i p strongly suggests a general metabolic r o l e the t h y r o i d .  of  The e x c r e t i o n r a t e a l s o showed a s i m i l a r r e l a t i o n -  s h i p w i t h body w e i g h t  ( F i g . 19).  Since extrathyroidal excretion  - 58 -  TABLE X  Summary o f i n j e c t i o n d e t a i l s  for size  experiment  No. o f fish  Mass (g)  V o l . injected (cc)  15  8.17(4.40-10.50)  .02  1  0.122  4  15.87(12.72-20.00)  .04  2  0.126  9  29.28(19.00-41.20)  .08  4  0.136  I ^ 1  ijxc)  1  JLC/&  - 59 -  F i g . 18. I n f l u e n c e o f body mass (gram) on plasma T ' l e v e l s and T/S v a l u e s (4 days a f t e r i n j e c t i o n ) and CR v a l u e s (8 days a f t e r i n j e c t i o n ) . Means and standard d e v i a t i o n s a r e shown f o r l a r g e , medium and s m a l l s i z e groups. No s i g n i f i cant d i f f e r e n c e was noted between medium and l a r g e s i z e groups f o r any J.131 parameter, but both d i f f e r e d from the s m a l l group f o r a l l parameters ( p ^ . 0 1 ) .  o  to "T  7}  (0* O  o o  —T—  I' in O  31  PLASMA  o  I—I—I 2  >  cn O"  O"  - 60 -  F i g . 19. R e l a t i o n s h i p b e t w e e n t h e l o g a r i t h m o f bodymass a n d t h e l o g a r i t h m o f v a r i o u s l!31 p a r a m e t e r s . T/S ( A ) , r -0.57; CR ( • ) , r -0.53; % b o d y (O), r -O.67.  - 61 rate  of I ' 1  5 1  was  independent  s h i p t o body s i z e i n metabolism.  of t h y r o i d a c t i v i t y ,  A s i m i l a r dependence  aculeatus  The a p p a r e n t  (Wiggs,  of  r e l a t i o n s h i p whereby  ( H i c k m a n , 1959)  and i n  1962). of the double  above  f o r s i g n i f i c a n t m e t a b o l i c change  In smaller f i s h the reverse  20 g r a m s i s t h e r e f o r e  very desirable  is  for  about  logarithmic  over t h i s range a r e l a t i v e l y g r e a t  i n mass must be p r e s e n t  true.  A size  change to  be  exceeding  experimental work  one i s now w o r k i n g i n a n e s s e n t i a l l y m a s s - s t a b l e VI.  change  e x c r e t i o n o n mass  s t a b i l i z a t i o n o f t h e mass e f f e c t  16 grams was p r o b a b l y a r e s u l t  observed.  relation-  c o n f i r m s t h a t mass c a u s e s a n o v e r a l l  h a s b e e n shown i n P l a t i c h t h y s s t e l l a t u s Gasterosteus  its  as  range.  SXPERIMBNTAL INVESTIGATION OF THE INFLUENCE OF SEASONAL  CHANGE I N INCREASING PHOTOPERIOD AND INCREASING TEMPERATURE ON THE RADIOIODINE METABOLISM AND THYROID C E L L HEIGHT OF TWO-YEAR-OLD STEELHEAD By b o t h h i s t o l o g i c a l a n d r a d i o c h e m i c a l c r i t e r i a t h y r o i d activity (Fig.  i s very high i n steelhead  6) a n d a p p e a r s  on t h e b a s i s this  surge  f a r more a c t i v e  of a temperature  in thyroid activity  increasing photoperiod. thesis  by r e p e a t i n g  change  (Table V I I ) .  smoltification  t h a n w o u l d be alone.  It  i s caused by t h e  anticipated  is possible influence  A n a t t e m p t was made t o t e s t t h i s  on t w o - y e a r - o l d s  c a r r i e d out on y e a r l i n g s t u r e regimes  at the time of  the experiment  that  of hypo-  previously  under v a r i o u s p h o t o p e r i o d and temperaThe e x p e r i m e n t was p e r f o r m e d f r o m  late  January u n t i l the beginning of June, a.period covering the main  - 62 extent of i n c r e a s i n g photoperiod and a l s o the time of s m o l t i f i c a t i o n , which normally occurs i n A p r i l and May.  The  usual  1,131 and h i s t o l o g i c a l parameters were measured. 1.  jl31 Excretion T h i s has been assessed by the percentage  dose i n body (4 days) and 1^31  of the  injected  plasma c o n c e n t r a t i o n (4 d a y s ) .  From the seasonal data ( F i g . 6) i t i s seen t h a t s i g n i f i c a n t changes i n 1^31  metabolism became e v i d e n t towards the end  I f photoperiod were having any e f f e c t on 1^31  March.  i t might be expected from t h i s time onwards. March t o the end of May  of  metabolism  From the end  of  the e x c r e t i o n parameters from the 4  c o n d i t i o n s d i d conform t o a g e n e r a l scheme ( F i g . 20).  With but  one e x c e p t i o n (not s t a t i s t i c a l l y s i g n i f i c a n t ) f i s h s u b j e c t e d 131 t o p r o g r e s s i v e l y i n c r e a s i n g temperature  showed a f a s t e r I  l o s s than f i s h at the constant temperature  of 5° C.  In a d d i t i o n ,  f i s h s u b j e c t e d t o the i n c r e a s i n g photoperiod showed a 131 s i g n i f i c a n t l y f a s t e r r a t e of I  removal a t any g i v e n tempera-  t u r e than those h e l d under the constant 8-hour daylength (Table X I ) . was  The  i n f l u e n c e of the photoperiod on I 3 1 e x c r e t i o n 1  not r e a l l y as a t r i g g e r i n the s t r i c t  sense s i n c e i t d i d  not cause a sudden r i s e i n e x c r e t i o n r a t e at a p a r t i c u l a r season.  From e a r l y February onwards, the photoperiods were  q u i t e separable w i t h the l o n g e r photoperiod c a u s i n g the more rapid I ^ 1  1  elimination.  A p r i l and May  was  The  sudden r i s e i n e x c r e t i o n r a t e i n  a t t r i b u t e d t o the i n f l u e n c e of  temperature  which rose q u i t e r a p i d l y at t h i s p e r i o d and superimposed i t s i n f l u e n c e on t h a t of the photoperiod.  As f a r as the e x c r e t i o n  - 6? -  F i g . 20. E x p e r i m e n t a l demonstration of the combined i n f l u e n c e of temperature and photoperiod on plasma T.131 l e v e l s and percentage dose i n t o t a l body 4 days a f t e r 1131 i n j e c t i o n i n 26 t o 30-month s t e e l h e a d ( p o t e n t i a l m i g r a n t s ) . Each p o i n t r e p r e s e n t s a mean of 6 i n d i v i d u a l s ( S t a t i s t i c s , Table X I ) .  - 64 r a t e i s concerned  i t would appear t h a t both temperature  photoperiod combined t o induce the h i g h r a t e o f I ^ 1  c h a r a c t e r i s t i c of s m o l t i n g f i s h . a synergism between temperature  There was  and  loss  1  no i n d i c a t i o n of  and p h o t o p e r i o d .  By l a t e  May  the e f f e c t of i n c r e a s i n g photoperiod on f i s h h e l d at 5° C appeared equal t o the e f f e c t of i n c r e a s i n g temperature  on  fish  from a constant 8-hour regime. 2.  T h y r o i d Uptake of I ^ 1  T h i s was  1  measured by T/S  22 r e s p e c t i v e l y ) .  and TUF parameters ( F i g s . 21  and  For each parameter both 4-day and 8-day  v a l u e s were recorded.  S i n c e the f i s h sampled at 4 days were  d i f f e r e n t i n d i v i d u a l s from those sampled at 8 days, t h i s g i v e s a "replication". 1^31  As p o i n t e d out i n the p r e l i m i n a r y study of  metabolism the 4-day estimate i s more r e l i a b l e ,  as  131 measurements l a t e r than t h i s may  be i n f l u e n c e d by I  loss  from  the t h y r o i d as radiohormone. In every i n s t a n c e f o r a p a r t i c u l a r temperature,  the i n c r e a s -  i n g photoperiod induced a h i g h e r t h y r o i d uptake v a l u e than the constant 8-hour day.  The data a l s o showed t h a t divergence  between photoperiod i n f l u e n c e became g r e a t e r as the photoperiod d i f f e r e n t i a l increased;  i n May  h i g h (Table X I ) .  be concluded, t h e r e f o r e , t h a t i n c r e a s i n g  I t may  the T/S  and TUF  i n d i c e s were  photoperiod caused a p r o g r e s s i v e i n c r e a s e i n t h y r o i d uptake of 1^31  and t h a t r i s i n g temperature  superimposed a f u r t h e r i n c r e a s e .  131 As w i t h I ^  e x c r e t i o n data, the c o n d i t i o n of constant tempera-  t u r e and i n c r e a s i n g photoperiod brought thyroid I ^ 1  1  about a change i n  uptake comparable t o t h a t i n f i s h h e l d under a  - 65 -  F i g . 21. E x p e r i m e n t a l d e m o n s t r a t i o n o f the combined i n f l u e n c e o f t e m p e r a t u r e a n d p h o t o p e r i o d on T / S v a l u e s 4 and 8 d a y s a f t e r I ? ! i n j e c t i o n i n 26 t o 3 0 - m o n t h s t e e l head ( p o t e n t i a l m i g r a n t s ) . E a c h p o i n t r e p r e s e n t s a mean of 6 i n d i v i d u a l s ( S t a t i s t i c s , Table X I ) . 1  - 66 -  F i g . 22. E x p e r i m e n t a l demonstration of the combined i n f l u e n c e of temperature and photoperiod on TUF v a l u e s 4 and 8 days a f t e r l l 3 1 i n j e c t i o n i n 26 t o 30-month s t e e l head ( p o t e n t i a l m i g r a n t s ) . Each p o i n t r e p r e s e n t s a mean of 6 i n d i v i d u a l s ( S t a t i s t i c s , T a b l e X I ) .  - 67 constant 8-hour daylength but w i t h r i s i n g 3»  temperature.  Conversion R a t i o T h i s was  measured 8 days a f t e r i n j e c t i o n ( F i g . 23).  Although t h e r e were a few exceptions e a r l y i n the year, the  fish  under i n c r e a s i n g p h o t o p e r i o d f o r any g i v e n temperature  showed  a h i g h e r c e l l h e i g h t than those under 8 hours.  the  s e p a r a t i o n was  v e r y evident and extremely h i g h CR v a l u e s were  obtained as a r e s u l t of r i s i n g temperature photoperiod.  By May  and i n c r e a s i n g  These d i f f e r e n c e s were h i g h l y s i g n i f i c a n t  I conclude from the I ^ 1  1  (Table X I ) .  data on combined temperature  photoperiod i n f l u e n c e on two-year-old  and  steelhead, that increasing  p h o t o p e r i o d s t i m u l a t e s p r o g r e s s i v e i n c r e a s e i n r a t e of I ^ ! 1  e x c r e t i o n , I ^ ! uptake by the t h y r o i d and CR. 1  I t does not  n e c e s s a r i l y have a sudden a c t i o n but i s i n f l u e n c i n g the I ^ ! 1  metabolism over a l o n g p e r i o d .  What appears t o be a sudden  131 change i n I  metabolism i n A p r i l and May  s t i m u l a t i o n by a temperature  rise.  i s due t o the added  R a d i o i o d i n e metabolism i s  e x p o n e n t i a l l y dependent upon temperature  ( F i g s . 9 t o 11).  r e l a t i o n s h i p would tend t o emphasize the response t o r a p i d l y r i s i n g temperature,  of the t h y r o i d  and might g i v e the impression of  a synergism under the c o n d i t i o n of i n c r e a s i n g temperature photoperiod.  This  and  There i s no d e f i n i t e evidence, however, f o r a  synergism between these two  important  environmental  factors.  I t must be noted t h a t f i s h under constant 8-hour p h o t o p e r i o d and h e l d at 5° C showed no tendency  at a l l t o change i n any  aspect of -rl31 metabolism.  I t i s f i n a l l y emphasized t h a t the  i n f l u e n c e of i n c r e a s i n g photoperiod on p o t e n t i a l migrants  was  - 68 -  F i g . 23. E x p e r i m e n t a l d e m o n s t r a t i o n o f the combined i n f l u e n c e o f t e m p e r a t u r e and p h o t o p e r i o d on mean c e l l h e i g h t a n d CR (8 d a y s a f t e r I * i n j e c t i o n ) i n 26 t o 30-month s t e e l head ( p o t e n t i a l m i g r a n t s ) . C e l l h e i g h t - means ( O A ) a n d i n d i v i d u a l v a l u e s T ) shown; CR - e a c h p o i n t r e p r e s e n t s a mean o f 6 i n d i v i d u a l s . 1  O A  1  - 69 -  TABLE X I Summary of s t a t i s t i c a l l y s i g n i f i c a n t d i f f e r e n c e f o r s e v e r a l i l j l parameters between d i f f e r e n t c o n d i t i o n s of temperature and p h o t o p e r i o d i n two-year-olds ( p o t e n t i a l migrants) i n l a t e May (White-Wilcoxon t e s t )  HIGH TEMPERATURE Photoperiod increasing  LOW TEMPERATURE  Photoperiod 8 hours  Photoperiod increasing  Photoperiod 8 hours  1^31 plasma (4 days)  % dose body % dose t h y r o i d (4 days) TUF (4 days)  t_ L  • ++  •++ -++  •++  T/S (4 days)  \CR (8 days) ? + ++ +  Two Two Two Two  groups groups groups groups  .++ not s t a t i s t i c a l l y d i f f e r e n t ( p - 0 . 0 5 ) . almost s t a t i s t i c a l l y d i f f e r e n t (p*0.0j?) s i g n i f i c a n t l y d i f f e r e n t (p 4.0.05). s i g n i f i c a n t l y d i f f e r e n t (p<0.01).  - 70 completely absent 4.  i n y e a r l i n g non-migrant: f i s h .  C e l l Height Measurement o f c e l l h e i g h t ,  though samples were  small,  showed a d e f i n i t e s e p a r a t i o n due t o p h o t o p e r i o d b e t w e e n a n d May ( F i g . 2 3 ) . height  No c l e a r - c u t  effect  April  o f t e m p e r a t u r e on c e l l  c o u l d be d e m o n s t r a t e d a l t h o u g h i n 8 - h o u r p h o t o p e r i o d  f i s h t h e l o w e r t e m p e r a t u r e was g e n e r a l l y a s s o c i a t e d w i t h a taller  c e l l height than the higher temperature.  of low temperature  VII.  effect  on c e l l h e i g h t c o n f i r m s e a r l i e r f i n d i n g s .  I n terms of c o l l o i d c o l o u r , differences  This  i t was d i f f i c u l t t o r e c o g n i s e  any  a t a l l between f i s h f r o m t h e c o n d i t i o n s .  INFLUENCE OF PRECOCITY OF MALE PARR ON THYROID A C T I V I T Y I t was n o t e d i n e a r l y s p r i n g t h a t u p t o 13% o f t h e  year-old steelhead These never  sampled were p r e c o c i o u s  two-  s e x u a l l y mature m a l e s .  showed t h e t e n d e n c y t o w a r d s s i l v e r i n g a n d p r e s e r v e d  a modified parr coloration characteristic of the species.  o f s p a w n i n g members  H i s t o l o g i c a l studies of t h y r o i d s of these p a r r  r e v e a l e d t h a t c e l l h e i g h t s were g e n e r a l l y h i g h e r t h a n i n nonmature i n d i v i d u a l s ( F i g . 23). i n e a r l y March and suggested another v a r i a b l e a f f e c t i n g many i n s t a n c e s  T h i s was p a r t i c u l a r l y n o t i c e a b l e that  s e x u a l m a t u r a t i o n c o u l d be  the t h y r o i d a c t i v i t y .  i n the l i t e r a t u r e associating  a c t i v i t y with sexual maturation i n teleosts. summarized by P i c k f o r d and A t z To i n v e s t i g a t e  There  are  increased thyroid These have  been  (1957).  t h i s hypothesis further,  a n e x a m i n a t i o n was  made o f t h e TUF a n d CR d a t a a v a i l a b l e f r o m t h e s e same  sexually  - 71  -  F i g . 24. TUF (open bars) and CR (shaded bars) f o r p r e c o c i o u s two-year-old male 09ff and immature (0#) s t e e l h e a d i n e a r l y March under 4 combined c o n d i t i o n s of temperature and p h o t o p e r i o d .  mature i n d i v i d u a l s ( F i g . 24).  Although no d e f i n i t e  conclusions  c o u l d he drawn, c e r t a i n s e x u a l l y mature male p a r r d i d have t h y r o i d a c t i v i t i e s c o n s i d e r a b l y h i g h e r than the other members o f t h e i r group.  I t i s t e n t a t i v e l y suggested t h a t  sexual maturation thyroid activity.  precocious  i n male p a r r i s c o r r e l a t e d w i t h i n c r e a s i n g The  c a u s a l aspects  of t h i s c o r r e l a t i o n were  not i n v e s t i g a t e d . VIII.  INFLUENCE OF EXERCISE ON I ? 1  Mass, i n c r e a s i n g photoperiod, s t a t e of sexual maturation  1  METABOLISM OF STEELHEAD  temperature and p o s s i b l y  have been shown t o i n f l u e n c e annual  cycles i n thyroid a c t i v i t y .  One  other f a c t o r t h a t c o u l d  be  c o n t r i b u t i n g t o c y c l i c a l changes i n the n a t u r a l environment i s the water c u r r e n t .  Fluviatile fish,  p o s i t i o n by r h e o t a x i s .  i n general, maintain  In a f a s t r i v e r the energy  their  expenditure  f o r m a i n t a i n i n g p o s i t i o n i s presumably g r e a t e r i n a s l u g g i s h r i v e r and  extra^metabolic  demands c o u l d be made on the animal  and r e f l e c t e d by i n c r e a s e i n t h y r o i d a c t i v i t y .  Such an  increase  i n t h y r o i d a c t i v i t y i n f a s t c u r r e n t s has been claimed f o r Salmo g a i r d n e r i (Fontaine,  1959).  In t h i s experiment 48 f i s h were i n j e c t e d w i t h I ^ ! and 1  immediately 24 of these were p l a c e d i n a f a s t c u r r e n t and i n a slow c u r r e n t ; the f a s t c u r r e n t . At 2, 4, (Fig.  the slow c u r r e n t was  approximately  In both tanks the temperature was  24  20f» of  10- t o 10.5°  6 and 8 days, 6 f i s h were k i l l e d from each c o n d i t i o n  25). Three parameters were measured - percentage of the dose i n  C.  - 73 -  F i g . 25. I n f l u e n c e o f i n c r e a s e d swimming a c t i v i t y on v a r i o u s a s p e c t s o f I l p l m e t a b o l i s m i n 19-month s t e e l h e a d . E a c h p o i n t r e p r e s e n t s a mean o f 6 i n d i v i d u a l s ( C o v a r i a n c e a n a l y s i s , Table X I I ) .  - 74  -  t o t a l body, percentage of the dose i n the t h y r o i d and CR.  They  all  showed s i g n i f i c a n t d i f f e r e n c e s between treatments (Table X I I ) .  The  increased  swimming caused an i n c r e a s e d r a t e of e x t r a t h y r o i d a l  j l 3 1 e x c r e t i o n , a decrease i n the t o t a l l!31 t h y r o i d and an i n c r e a s e i n CR.  The  uptake by  i n c r e a s e d CR  the  i n d i c a t e s that  radiohormone p r o d u c t i o n  i s augmented, d e s p i t e a decreased  uptake by the t h y r o i d .  The  t h y r o i d i s probably due  t o l o s s of r a d i o a c t i v i t y as hormone and  would add  support t o the c o n t e n t i o n  induces an i n c r e a s e d w i t h the g e n e r a l IX.  THE  reduced accumulation of 1^31  These f i n d i n g s agree  of F o n t a i n e (1959h  INFLUENCE OF CHEMICAL CHANGES (SALINITY AND  LEVELS OF THE AND  MEDIA) ON  i n the  t h a t muscular a c t i v i t y  demand f o r t h y r o x i n .  conclusions  I ? 1  1  IODINE  METABOLISM OF STEELHEAD  CHUM SALMON (Oncorhynchus keta)  Apart from the e f f e c t s of p h y s i c a l a s p e c t s o f the ment on t h y r o i d a c t i v i t y and  i n these anadromous t e l e o s t s .  concluded that the s a l i n i t y and  were most important.  The  environ-  Hoar (1939)  surveyed the chemical f a c t o r s that p o s s i b l y i n f l u e n c e t h y r o i d and  environ-  the i n f l u e n c e of body mass and  p o s s i b l y sexual m a t u r a t i o n , t h e r e i s a l s o the chemical ment t o c o n s i d e r  1^31  the  iodine  concentration  i n f l u e n c e of s a l i n i t y was  particularly  p e r t i n e n t here, i n view of the v a r i o u s t h e o r i e s t h a t have been put forward t o e x p l a i n the r o l e of the t h y r o i d i n a p o t e n t i a l migrant.  The  previous  observations  suggested the i n f l u e n c e t h a t  v a r i o u s f a c t o r s have on the t h y r o i d , but  i t i s now  necessary t o  c o n s i d e r what t h i s i n c r e a s e d t h y r o i d a c t i v i t y means i n terms of  - 75 -  TABLE X I I A n a l y s i s of c o v a r i a n c e f o r the r e l a t i o n s h i p between v a r i o u s 1.131 parameters and time (hr) f o r p a r r h e l d i n f a s t and slow c u r r e n t s  Parameter % dose total body  CR  F  g  F  m  Condition Calm Torrent Calm Torrent  Slope  -.0031 7  df  1  , and -.0084 j 48 .0064 ] 1 , and .0083 J1 48  P  Fm x  7.5**  1.4  m  4.1*  12.8**  = Variance r a t i o f o r slopes.  = V a r i a n c e r a t i o f o r a d j u s t e d means. * S i g n i f i c a n t d i f f e r e n c e between s l o p e s ** S i g n i f i c a n t d i f f e r e n c e between s l o p e s  (p*C.05). (p<.01).  - 76 metabolism.  T h i s i n v o l v e s c o n s i d e r a t i o n of the v a r i o u s t h y r o i d  r o l e s suggested f o r a m i g r a t i n g (i)  Thyroxin  fish:  s t i m u l a t e s c e r t a i n changes i n metabolism,  such as s i l v e r i n g and  s a l i n i t y preference  s m o l t i f i c a t i o n (Fontaine, (ii)  -  1954;  t h a t occur  Baggerman, i 9 6 0 ,  1963).  A f i s h p r i o r t o seaward m i g r a t i o n tends t o become  demineralized.  Such d e m i n e r a l i z a t i o n has  several species  (reviewed  by Hoar, 1959)  been d e s c r i b e d f o r and may  be due  v a r i e t y of f a c t o r s i n c l u d i n g the i n c r e a s e d p r o d u c t i o n metabolic  at  to a  of  water or the f u n c t i o n i n g of c e r t a i n s a l t e l i m i n a t i o n  mechanisms p r i o r t o the a c t u a l entrance t o the sea. ( i 9 6 0 ) has  shown a g r e a t  Olivereau  i n c r e a s e i n a d r e n o c o r t i c a l volume at  t h i s time of s m o l t i f i c a t i o n i n Salmo s a l a r .  Hoar (1952,  1959)  suggested t h a t such changes i n the i n t e r n a l environment c o u l d promote i n c r e a s e d metabolic thyroid  demands r e s u l t i n g i n i n c r e a s e d  activity.  (iii)  The  d e s c r i b e d above.  f i n a l p o s s i b i l i t y stems from the  demineralization  Namely, t h a t under c o n d i t i o n s of e l i m i n a t i o n  of body s a l t s the a v a i l a b i l i t y of i o d i n e t o the t h y r o i d c o u l d become reduced, e s p e c i a l l y i f the surrounding deficient i n iodine. would not be  medium were  Under c o n d i t i o n s such as these,  i n iodine equilibrium.  The  the  t h y r o x i n supply  fish f o r the  body would have t o be produced d e s p i t e p r o g r e s s i v e l y lower l e v e l s of a v a i l a b l e  ,  A c t i v e t r a n s p o r t mechanisms would  be  r e l i e d upon even more f o r b u i l d i n g up the i o d i n e g r a d i e n t between t h y r o i d and plasma. low plasma I  I f the gland were compensating f o r  127 ' l e v e l s i n t h i s way  then i t c o u l d appear a c t i v e  - 77 by b o t h h i s t o l o g i c a l a n d r a d i o c h e m i c a l c r i t e r i a d e s p i t e that  the a c t u a l  increased.  output  available  o f hormone i n t o t h e p l a s m a h a d n o t  In other words,  s m o l t may be i n p a r t I  1 2  7  the  really  the h i g h t h y r o i d a c t i v i t y of  an a r t i f a o t  the  and m e r e l y a r e f l e c t i o n  i n d u c e d by a r a p i d r a t e  of I  1 2  fact  of  ^ excretion.  low This  131 w o u l d be s u p p o r t e d b y t h e d a t a s h o w i n g v e r y r a p i d I t i o n i n steelhead and F o n t a i n e ,  s m o l t s and o t h e r s p e c i e s a t m i g r a t i o n  i n (i)  (iii)  to evaluate  fresh water,  fresh water  sea water  assessed.  (ii)  t h e s e p o s s i b i l i t i e s f i s h were sea water  containing the  (36 j i g / l ) .  , f i s h the net  (Leloup  i960).  I n an attempt kept  elimina-  effect  (250/00 s a l i n e )  same amount o f  r*- 7 a s 25  sea w a t e r  and sea  1 2  ?  against  sea  the  osmotic  on t h e t h y r o i d c o u l d be i s o l a t e d  the  i o d i n e c o n c e n t r a t i o n w o u l d be t h e same i n b o t h m e d i a .  H i c k m a n (1959) h a s p o i n t e d o u t ,  the  the  level 131  same i n b o t h m e d i a s i n c e  considerably fish  the  the metabolism of  I  .  was l i v i n g  by t h e r e s p o n s e  i n iodine deficiency.  on b o t h y e a r l i n g s  (parr)  Finally,  of the These  and f i s h o f  water,  since As  l e v e l s h a v e t o be made 127 of I ' can a l t e r  i n f r e s h water w i t h f i s h i n f r e s h water  might i n d i c a t e ,  water  o n t h e t h e t h y r o i d c o u l d be  By c o m p a r i n g f r e s h w a t e r w i t h I influence  0/00  2  By c o m p a r i n g t h e f r e s h - w a t e r of  and  comparison  containing  t h y r o i d , whether  of  iodine the  t e s t s were c a r r i e d  s m o l t i n g age  (smolts  fish out  and  parr). Yearling fish, silvering, temperature  a l l w i t h v e r y p r o n o u n c e d p a r r m a r k s a n d no  were p l a c e d i n the t h r e e media i n September of 13.6°  C.  They s u r v i v e d the  transition  at  a  water  whether  - 78 the i n t r o d u c t i o n was A f t e r 10  -  d i r e c t or v i a intermediate  days they were i n j e c t e d w i t h I ^ . 1  were k i l l e d at 4 days;  1  1  i n f i s h from a l l three media.  The  1  c l e a r a n c e was  fish  1  f r e s h water d i d not d i f f e r .  apparent  i n i t i a l rates of l o s s of  plasma I ^ ! from f i s h h e l d i n i o d i n e d e p l e t e  of l o s s were a g a i n  H a l f of the  h a l f were k i l l e d a t 8 days.  A curvilinear relationship for I ^  t h i s r a t e of l o s s was  salinities.  and  iodine replete  In sea water, on the other hand,  greater.  In the second phase, the  rates  s i m i l a r i n both fresh-water media but  the  sea water adapted f i s h showed a pronounced decrease i n the of plasma I ^ l removal ( F i g . 26).  rate  A s i m i l a r d i f f e r e n c e between 131  f r e s h water and  sea water was  the whole body. that there  noted w i t h I  J  e l i m i n a t i o n from  The more d i p h a s i c curves i n sea water suggest  i s more than one  the s t e e l h e a d .  mechanism of  elimination i n  Furthermore, s i n c e the c u r v i l i n e a r form  was  more pronounced i n sea water, i t i s p o s s i b l e t h a t the  extrarenal  e x c r e t o r y pathway i s b e t t e r developed i n t h i s medium.  Possibly  t r a n s f e r t o sea water i n c r e a s e d g e n e r a l g i l l s and  s a l t l o s s through  the  reduced f u r t h e r the r o l e of the kidney mechanism.  The  127 a d d i t i o n of I of  ' i n t o the water i t s e l f does not a f f e c t the  1131. A d d i t i o n of l  (TUF)  loss  l  2  7  appeared t o reduce the t h y r o i d I 3 1 1  by a s m a l l amount and r e l a t i v e t o t h i s reduced  i n i o d i n e r e i n f o r c e d f r e s h water, the enhanced TUF the sea water suggested t h a t i n c r e a s e d stimulated  the " i o d i n e pump".  The  uptake  activity  values  in  osmotic p r e s s u r e may  conversion  showed no marked d i f f e r e n c e between the three  ratio,  have  however,  conditions.  - 19 -  F i g . 26. E f f e c t of t r a n s f e r of 20-month s t e e l h e a d t o sea water (25 o/oo s a l i n e ) and i o d i d e - r e i n f o r c e d f r e s h water on v a r i o u s aspects o f I ^ ? metabolism measured 4 and 8 days a f t e r I * ! i n j e c t i o n . Mean v a l u e s r e p r e s e n t e d (SW,X ; FW + I ' , A ; FW,0 ). 1  1  1 2  - 80 In l a t e May the experiment was r e p e a t e d a t 8 . 5 ° 0 on p o t e n t i a l migrants kept under the i n c r e a s i n g p h o t o p e r i o d and i n c r e a s i n g temperature regime.  U n f o r t u n a t e l y t h e temperature  f e l l t o a v e r y low l e v e l (1° C) i n t h e i o d i n e c o n t r o l tank (F.W. + I  1 2  7 ) and i n view o f t h e temperature i n f l u e n c e on I ^ ! 1  metabolism these d a t a were d i s c a r d e d .  Comparison c o u l d only  be made, t h e r e f o r e , between f r e s h d e c h l o r i n a t e d water and sea water  (25 o/oo s a l i n e ) ( F i g . 2 7 ) .  F o r comparison, smolts have  been t r e a t e d s e p a r t e l y from p a r r , though t h i s breakdown cons i d e r a b l y reduced the sample s i z e and made a f i n a l  critical  examination d i f f i c u l t . C e r t a i n important t r e n d s appear from t h e d a t a .  F i r s t , the  extent o f I ? l o s s , both t o t a l and e x t r a t h y r o i d a l , was reduced 1  1  i n sea water w h i l e the parameters measuring the a c t i v i t y o f g l a n d , T/S and CR were lower than i n f r e s h water.  No d i f f e r e n c e  between s i l v e r e d or s i l v e r i n g f i s h and p a r r was noted.  I t was  concluded t h a t i n t r o d u c t i o n i n t o sea water of p o t e n t i a l l y migrating f i s h  (parr or smolt) p o s s e s s i n g a h i g h t h y r o i d  activity  c o n s i d e r a b l y lowered t h y r o i d h y p e r f u n c t i o n and a l s o reduced 131 the former r a p i d r a t e o f I  excretion.  I t i s inferred  either  t h a t s t a b i l i z a t i o n o f plasma i o d i n e a t a h i g h e r l e v e l has i n h i b i t e d the t h y r o i d I ^ 1  1  accumulating e f f i c i e n c y , o r t h a t  sea water i t s e l f lowered t h y r o i d a c t i v i t y .  A third  possibility  i s t h a t h i g h e r i o d i n e l e v e l s i n themselves reduced t h y r o i d a c t i v i t y but that t h i s t r e n d was masked by the tendency f o r the i n c r e a s e d osmotic p r e s s u r e t o s t i m u l a t e a r e a l i n c r e a s e i n hormone output.  Without c o n t r o l s i n i o d i n e r e i n f o r c e d water  - 81 -  F i g . 27. E f f e c t o f t r a n s f e r o f 30-month s m o l t s and p a r r t o s e a w a t e r (23 0 / 0 0 s a l i n e ) o n v a r i o u s a s p e c t s o f l l 3 1 m e t a b o l i s m . Mean v a l u e s ( b a r ) a n d i n d i v i d u a l v a l u e s shown.  SMOLTS  PARR ••3-3  TOTAL ••3-3  t  1  f 2  «/> < _l Q. 5  -  I  1 E3  8  t9 o o  ft  ffl  o  40  8  T/S  2Cf  8  o o  sw  O  FW  - 82 T h i s i s s u e cannot be r e s o l v e d . At t h i s point i n the proceedings o f m i g r a n t age w e r e e x h a u s t e d repeated. less  the stocks  so t h e experiment  Chum s a l m o n w e r e a v a i l a b l e ,  however,  identical physiological condition.  of  steelhead  c o u l d n o t be i n a more o r  By m i d - s u m m e r u n d e r -  y e a r l i n g chum s a l m o n ( w h i c h s h o u l d h a v e m i g r a t e d i n t h e s p r i n g ) show v e r y h i g h r a t e s water.  o f I * ^ m e t a b o l i s m when r e t a i n e d i n f r e s h 1  The b i o l o g i c a l h a l f - l i f e f o r t h e l o s s  d r o p s a s l o w a s 23 h o u r s  (Eales,  o f serum I ^ ! 1  1961), w h i l e h i g h t h y r o i d u p -  t a k e a n d t h e r a p i d p r o d u c t i o n o f v e r y h i g h CR v a l u e s over  i n some c a s e s )  also  via thyroid routes. and E a l e s  indicates a very  rapid I ^ 1  I t has been suggested  (1961, 1963) t h a t  (60% o r 1  metabolism  b y H o a r (1932, 1959)  t h e s e f i s h may show i n a n  exaggerated  form the t h y r o i d h y p e r a c t i v i t y u s u a l l y e x h i b i t e d i n a smolting fish.  I t was d e c i d e d t h e r e f o r e  sea w a t e r and change  to investigate  the influence of  i n i o d i n e c o n c e n t r a t i o n o n t h e chum s a l m o n  thyroid. Twelve f i s h p o s s e s s i n g  thyroid activities  were p u t i n t o each o f t h e f o l l o w i n g : saline)  ( i ) sea water  ( i i ) f r e s h w a t e r w i t h t h e same I  sea water and ( i i i ) f r e s h water was 1 8 ° G .  1  2  (25 0 / 0 0 •.  ^ content  The w a t e r  a s 25 0 / 0 0 temperature  1  and k i l l e d  108 h o u r s  after  A t t h i s t i m e t h e e x t r a t h y r o i d a l e x c r e t i o n a n d CR  were measured Stable  itself.  1  kind  The f i s h were h e l d under t h e s e c o n d i t i o n s f o r 6  days and then i n j e c t e d w i t h I ^ injection.  of t h i s  ( F i g . 28).  iodine reinforcement,  did not a l t e r the I ? ! rate 1  as w i t h y e a r l i n g  steelhead,  o f e x c r e t i o n b u t l o w e r e d t h e CR  - 83 -  F i g . 28. E f f e c t o f t r a n s f e r o f u n d e r y e a r l i n g p o t e n t i a l m i g r a n t chum s a l m o n t o s e a w a t e r (25 o / o o s a l i n e ) a n d i o d i d e reinforced fresh water. Mean v a l u e s ( b a r ) a n d i n d i v i d u a l  v a l u e s shown X s < . 0 . 0 5 ;  XX«=<0.01.  - 84 g r e a t l y compared t o the s t r i c t l y fresh-water s i t u a t i o n .  In  a d d i t i o n , s a l i n i t y depressed 1^31 l o s s r e l a t i v e t o e i t h e r f r e s h water o r i o d i n e r e i n f o r c e d c o n t r o l s . the reduced 1^31 e x c r e t i o n  This  suggests t h a t  observed i n sea water was due t o the  o v e r a l l osmotic e f f e c t and not due t o h i g h ambient I - . 3  2 7  The  c o n v e r s i o n r a t i o i n sea water was l e s s than i n f r e s h water a l t h o u g h the i n f l u e n c e was not as pronounced as w i t h the migrant steelhead. In these data on chum, the e f f e c t s o f i o d i n e and osmotic change become much c l e a r e r . l e v e l s caused a r e d u c t i o n  I t appears t h a t  i n the CR.  increased  iodine  The h y p e r a c t i v i t y o f the  t h y r o i d as measured i n f r e s h water by r a d i o c h e m i c a l methods 127 c o u l d be a b o l i s h e d  by a h i g h ambient I  l e v e l , even though  the r a t e of i o d i n e e l i m i n a t i o n from the body had not been a l t e r e d . In sea water, however, t h y r o i d a c t i v i t y was i n c r e a s e d i o d i n e c o n t r o l s and t h i s r e v e a l e d As w i t h s t e e l h e a d  over the  response t o the osmotic change.  y e a r l i n g s , t h e r e was d e f i n i t e s t i m u l a t i o n o f  the t h y r o i d i n a medium o f h i g h s a l t I f the migrant s t e e l h e a d  content.  responded s i m i l a r l y t o the chum  i n the i o d i n e r e i n f o r c e d medium, i t would be concluded t h a t ( i ) the h i g h l e v e l of 1^31 l o s s i s reduced when the s t e e l h e a d moves i n t o the sea  ( i i ) the h i g h t h y r o i d a c t i v i t y i s p a r t l y due  t o a s t a t e o f i o d i n e d e f i c i e n c y and the  enters  sea i t s t h y r o i d may a g a i n be a c t i v a t e d due t o t h e i n c r e a s e d  salinity. an  ( i i i ) when the f i s h  Whether the change from f r e s h t o sea water produces  o v e r a l l increase  or decrease i n t h y r o i d a c t i v i t y would be due  t o s e v e r a l f a c t o r s prominent among which would be t h e blood  - 85 level  o f I * ? a t t h e t i m e o f t r a n s i t i o n and a l s o t h e  to the  salt  Thus, to  concentration. although the net r e s u l t  of t r a n s f e r  from f r e s h  s e a w a t e r may he a d r o p i n t h y r o i d a c t i v i t y ,  i o d i n e and s a l i n i t y i n f l u e n c e s t h y r o i d a c t i v i t y due t o t h e  generally  increase  a net  increase  2  to the  increased  little  of  iodine  The two g r o u p s difference  of  s a l t water  response  of  e  v  e  i  D U  t  involves  c a n he a r g u e d  that  the response  steelhead  w o u l d show,  therefore,  and t h e  however,  In the y e a r l i n g s ,  sea water  t h e r e was  thyroid activity  caused  slight  very  i s suggested.  i n the smolt  de-  stimulation.  t h e n e t r e s u l t was a d e p r e s s i o n  t h y r o i d a c t i v i t y by a l l p a r a m e t e r s by h i g h i o d i n e l e v e l s  due  some  t o e i t h e r medium h u t t h e i o d i n e s l i g h t l y  p r e s s e d T/S r a t i o s  in  a stimu-  the t r a n s i t i o n  outweighs  the  level.  i n t h i s respect.  In the smolts,  If  i n t h y r o i d a c t i v i t y then i t  stimulatory effect  analysis  water  show a d e p r e s s i o n  i n I^ ^ i  l a t i o n due t o t h e s a l i n i t y e l e v a t i o n .  the  response  1 2  of  and a marked t h y r o i d This implies that  inhibition the  high  i s p a r t l y due t o a n i o d i n e  deficiency. X.  THE R E L A T I O N S H I P BETWEEN RADIOIODINE METABOLISM AND S I L V E R I N G Thyroid hyperfunction  i n juvenile  s i l v e r i n g and s m o l t i f i c a t i o n . this belief, role  there  of t h y r o x i n .  stimulate  Though c e r t a i n e x p e r i m e n t s  i s considerable  disagreement  As noted p r e v i o u s l y ,  subepidermal guanine  salmon i s c o r r e l a t e d  regarding  with  confirm the  w h i l e t h y r o x i n may  d e p o s i t i o n i n the l a b o r a t o r y  may n o t be r e q u i r e d u n d e r n a t u r a l c o n d i t i o n s . i n d i c a t e d by c e r t a i n d a t a o b t a i n e d i n t h i s  it  T h i s seemed t o  investigation.  It  be  - 86 was  noted t h a t f i s h kept under an 8-hour daylength and  temperature of 5 "to 6° C showed no seasonal  increase  at a  i n thyroid  f u n c t i o n e i t h e r w i t h r a d i o i o d i n e or h i s t o l o g i c a l methods and  yet  c e r t a i n of them s i l v e r e d .  May  Between the p e r i o d 27 A p r i l t o 29  when i n v e s t i g a t i o n s of t h y r o i d f u n c t i o n were being made of smolting  f i s h from the f o u r photoperiod and  o u t l i n e d i n T a b l e 711,  a r e c o r d was  f i c a t i o n of each f i s h  (Table X I I I ) .  Of the 4 c a t e g o r i e s mature males) considered, one  one  made of the s t a t e of s m o l t i -  (smolts, p a r r - s m o l t s , s m o l t i f i c a t i o n was  environmental regime.  under any  temperature regimes  p a r r and  sexually  not favoured by  I t i s s t i l l p o s s i b l e t h a t the  c o n d i t i o n might tend t o have a h i g h e r  a c t i v i t y than the p a r r under the same c o n d i t i o n .  To  any  smolts  thyroid investigate  t h i s more f u l l y , two-year-olds from the i n c r e a s i n g p h o t o p e r i o d and  low temperature regime were considered  i n greater d e t a i l  t h e s e c o n s t i t u t e d a l a r g e r sample of examined f i s h . of 1^31  metabolism was  treated separately  s t a t e of s m o l t i f i c a t i o n ( P i g . 29).  as  Each aspect  i n r e l a t i o n to  I t i s concluded t h a t  the there  131 was  no c o n s i s t e n t t r e n d r e l a t i n g any  aspect of I  metabolism  t o s i l v e r i n g , and the c a u s a l r e l a t i o n s h i p between t h y r o x i n subepidermal guanine d e p o s i t i o n i s t h e r e f o r e Of f u r t h e r i n t e r e s t was  and  questioned.  the constancy between the 4 h o l d i n g  c o n d i t i o n s i n the t o t a l number of steelhead  showing any  t o s i l v e r (smolts  always between 65  75%  + parr-smolts).  T h i s was  of the t o t a l number examined i n any  one  and  c o n d i t i o n (Table X I I I ) .  A c c o r d i n g t o Maher and L a r k i n (1954), approximately 60% steelhead  tendency  of young  t r o u t from the C h i l l i w a c k R i v e r , B r i t i s h Columbia,  - 87 -  TABLE X I I I The  e f f e c t o f d i f f e r e n t temperature and p h o t o p e r i o d regimes on the percentage i n c i d e n c e o f s m o l t i f i c a t i o n i n two-year-old s t e e l h e a d between 27 A p r i l and 29 May, 1°62  HIGH TEMPERATURE Photoperiod increasing  Smolts (no p a r r marks) Parr-smolts S i l v e r , but w i t h p a r r marks) Parr (no s i l v e r Sexually mature male p a r r TOTALS  LOW TEMPERATURE  Photoperiod 8 hours  Photoperiod increasing  Photoperiod 8 hours  No.  i  24  40  7  28  39 58  7 30  18  30  9 36  13 19  11 48  10 17  7 28  10 15  3 13  8 13 6o 100  No.  2  %  8  25 100  No.  5  %  8  67 100  No.  2  %  9  23 100  - 88 -  F i g . 29. D e m o n s t r a t i o n o f t h e l a c k o f a r e l a t i o n s h i p b e t w e e n s i l v e r i n g and any a s p e c t o f 1131 m e t a b o l i s m (8 d a y s after injection). Mean v a l u e s ( b a r ) . a n d i n d i v i d u a l v a l u e s shown.  migrate as two-year-olds  89 -  a t an average l e n g t h of 16.49 cm.  Both the l e n g t h o f t h e m i g r a t i n g f i s h and the percentage showing s m o l t i f i c a t i o n agreed f a v o u r a b l y w i t h the data from the l a b o r a t o r y - h e l d f i s h c o n s i d e r e d above. XI. Any  PERIPHERAL SITES OF RADIOHORMONE CATABOLISM  attempt t o c l a r i f y the r o l e o f t h y r o x i n i n t e l e o s t s  depend on knowledge o f i t s s i t e of a c t i o n . organs o f t h y r o x i n i n f i s h are unknown.  will  So f a r t h e t a r g e t  One method would be t o  i n j e c t a sample o f r a d i o a c t i v e hormone and then f o l l o w i t s r o u t e i n t o the v a r i o u s t i s s u e s .  No pure r a d i o a c t i v e t h y r o x i n was  a v a i l a b l e i n t h e present study and so a l e s s d i r e c t was  technique  employed. When I - ^ l i  s  i n j e c t e d , the i n o r g a n i c i o n ( I ^ l ) v e r y q u i c k l y  reaches an e q u i l i b r i u m between the v a r i o u s body compartments. Thus as f a r as i n o r g a n i c r*"^ i s concerned, t i s s u e count/min/g blood count/min/g  s  i s I ^ ! i n t h e body. 1  h  o  u  l  d  r  e  m  a  i n  the r a t i o o f  constant f o r as l o n g as t h e r e  Owing t o the r e l a t i v e l y r a p i d d i f f u s i o n o f  the i o n , departure from t h i s r a t i o would o n l y be noted i n those t i s s u e s such as t h e t h y r o i d which a c t i v e l y concentrate i o d i d e . I t may a l s o be argued t h a t s i n c e 1^31 i s the predominant r a d i o a c t i v e c o n s t i t u e n t i n t h e blood t h a t the same r a t i o should h o l d between the t o t a l r a d i o a c t i v i t y i n the t i s s u e s and blood, i . e . tissue I ^ blood I 3 1 1  1  count/min/g  t o t a l t i s s u e r a d i o a c t i v i t y count/min/g  count/min/g " t o t a l blood r a d i o a c t i v i t y  count/min/g  I f t h i s r a t i o based on t o t a l r a d i o a c t i v i t y i n t i s s u e and blood should change, i t would imply a change between b l o o d and t i s s u e  - 90 of an o r g a n i c 1 ^ 1  fraction.  accumulation of o r g a n i c a l l y  A r i s e i n the bound  r a t i o would mean  i n the  sumably an accumulation of radiohormone.  tissues  - pre-  Thus by measuring  the  r a t i o of r a d i o a c t i v i t y of t i s s u e to blood at v a r i o u s times 131 after I '  i n j e c t i o n some i n d i c a t i o n might be gained of  of p e r i p h e r a l  metabolism of radiohormone.  T h i s was out  the  t h e o r e t i c a l b a s i s f o r the  below i n which 48  l i 3 1 and  fish  experiment  o n e - t h i r d of t h i s t o t a l k i l l e d at 3,  u s u a l manner and reweighed and  6 and  blown i n t o a t a r e d  counted at a d i s t a n c e of 4 cm  s c i n t i l l a t i o n counter f o r 3,000 counts. kidney, s k i n , l i v e r , gut,  brain  counted i n a s i m i l a r manner. gram t i s s u e was  planchet  and  from the  was  gill,  and  radioactivity  r a d i o a c t i v i t y per  per gram  v a l u e s f o r each r a t i o were  (IPig.  e x c e p t i o n of the g i l l 6 days.  30). t i s s u e , t h e r e was  r a t i o between 3 and  on the  absolute l e v e l s of r a d i o a c t i v i t y i n the v a r i o u s undoubtedly due  ml  end-probe  muscle were d i g e s t e d  i n the  as t h i s was  the  Weighed p i e c e s of  expressed r e l a t i v e to the  means c a l c u l a t e d  W i t h the  The  In each f i s h the  each time p e r i o d , 16  obtained and  9 days.  removed i n  planchet.  uc  the whole blood d i g e s t e d f o r 2 days w i t h one  of 2N NaOH and  At  carried  (10-40 g) were i n j e c t e d w i t h 4  From each f i s h a s m a l l whole b l o o d sample was  blood.  sites  No  significance  to g r e a t e r v a s c u l a r i t y  little  change  i s placed tissues  of the  tissue  -  i f the t i s s u e were completely v a s c u l a r i t would possess a r a t i o of e x a c t l y active  1.  Any  i n c r e a s e i n r a d i o a c t i v i t y above 1 means an  accumulation of i o d i n e  i n the g i l l t i s s u e .  For  i n g e n e r a l between 3 and  from the  other t i s s u e s ,  blood. the  T h i s i s found only  constancy of the  6 days would bear out  the  initial  ratio  - 91 -  F i g . 30. Change i n the t i s s u e : blood r a d i o a c t i v i t y i n a v a r i e t y of t i s s u e s from 19-month s t e e l h e a d . Each p o i n t r e p r e s e n t s a mean o f 14- t o 16 i n d i v i d u a l s .  - 92 assumption t h a t i A ^ l moves r a p i d l y between body compartments and p r o v i d e s  support  f o r the suggestion t h a t any  the r a t i o would be due  t o change i n d i s t r i b u t i o n of  f r a c t i o n s , which even at 6 days are o n l y present q u a n t i t i e s i n the blood The and,  l a t e r change i n  ( F i g s . 5 and  organic  i n small  16).  change i n the r a t i o i n the g i l l  tissue i s interesting  i n view of the small amount of PBI  131 u s u a l l y encountered  in  blood at 6 days, presumably i n v o l v e s an accumulation  T h i s was and  a l s o found by Hickman  i t was  (1959)  of  i n the s t a r r y f l o u n d e r  by Hickman t h a t the h i g h I - ^ l c o n c e n t r a t i o n  concluded  there could be r e c o n c i l e d w i t h the known f u n c t i o n of the in  salt excretion.  gills  However, w h i l e the data of Hickman showed  a steady r a t i o between g i l l  and blood 24 hours a f t e r  injection,  these data on s t e e l h e a d i n d i c a t e a p r o g r e s s i v e accumulation r a d i o a c t i v i t y i n the g i l l with time. for  No  satisfactory  explanation  t h i s phenomenon can be g i v e n , u n l e s s the l a t e r h i g h  were due  to accumulation  of  i  na  f  o  m  of  values  other than the  freely  d i f f u s i b l e ion. The all  t i s s u e : b l o o d r ^ l r a t i o changed between 6 and  t i s s u e s , but was  9 days i n  c o n s i s t e n t l y most pronounced i n the  very  m e t a b o l i c a l l y a c t i v e t i s s u e s , i . e . b r a i n , gut, l i v e r and  kidney.  S k i n showed only a s l i g h t percentage i n c r e a s e and muscle o n l y a moderate percentage change from 6 t o 9 days ( F i g s . 30 and That the r i s e i n the t i s s u e : b l o o d r a t i o between 6 and due  t o an accumulation  9 days i s  of r a d i o t h y r o x i n i s borne out by  o b s e r v a t i o n t h a t over t h i s p e r i o d , d e t e c t a b l e amounts of are r e l e a s e d from the gland  ( F i g s . 5 and  16).  31).  the  - 93 -  F i g . 31. P e r c e n t a g e change days a f t e r i n j e c t i o n ) of t i s s u e : d a y s a f t e r i n j e c t i o n i n 19-month  ( r e l a t i v e to the value 3 b l o o d r a t i o s 6 and 9 steelhead.  6  DAYS  -  It i s therefore  94 -  concluded that i f change i n  t o t a l t i s s u e count/min/g blood  count/min/g  i s i n d i c a t i v e of b u i l d up o f r a d i o a c t i v e hormone i n the t i s s u e , then the h i g h e s t and  gut.  accumulation was observed i n b r a i n , l i v e r ,  kidney  In a l l these t i s s u e s energy demands are probably h i g h .  In l e s s a c t i v e t i s s u e s such as muscle and s k i n , the P B I ^ 1  accumulation was l e s s .  T h i s suggests t h a t the t h y r o i d hormone  i s not u t i l i z e d i n s p e c i f i c areas but probably r e q u i r e d metabolically  a c t i v e system.  I t would argue a g a i n s t  by any  a speci-  f i c a l l y l o c a l i z e d r o l e of the t h y r o i d i n t e l e o s t s and i m p l i e s a g e n e r a l metabolic r o l e .  - 95 DISCUSSION I.  COMPARISON OF PARAMETERS FOR MEASURING THYROID A C T I V I T Y AMD THE EFFECT OF TEMPERATURE ON THESE PARAMETERS S e v e r a l d i s c r e p a n c i e s between t e c h n i q u e s  t h y r o i d a c t i v i t y may be c i t e d .  Swift  (1959),  f o r measuring simultaneously  u s i n g h i s t o l o g y and r a d i o c h e m i s t r y t o measure s e a s o n a l i n t h y r o i d a c t i v i t y i n brown t r o u t , t e c h n i q u e s were n o t n e c e s s a r i l y showed t h a t h i g h t e m p e r a t u r e  demonstrated that  correlated.  (1955)  c a u s e d no h i s t o l o g i c a l c h a n g e i n shown a v e r y  s t i m u l a t i o n o f t h e t h y r o i d by t e m p e r a t u r e w i t h I131  techniques.  Eales  (1963)  o b t a i n e d some c o n f o r m i t y  between  h i s t o l o g i c a l and r a d i o c h e m i c a l methods i n two s p e c i e s Oncorhynchus.  coho and s o c k e y e ,  disagreement  i n general these discrepancies r e l a t e  It is  temperatures  or to  seasonal That  between h i s t o l o g i c a l and r a d i o i o d i n e measurements  important f o r the explanation that In the present  further  of  emphasized  may o n l y o c c u r when t e m p e r a t u r e i s t h e m a i n v a r i a b l e i s  on p e r c e n t a g e  pink  e i t h e r t o comparisons  changes p r o b a b l y i n d u c e d by temperature d i f f e r e n c e s . disagreement  juvenile  Other i n s t a n c e s  have been c i t e d by M a t t y ( i 9 6 0 ) .  of thyroid a c t i v i t y at different  of  but i n two o t h e r s p e c i e s ,  a n d chum t h e r e was a c o m p l e t e d i s a g r e e m e n t .  that  t h e two  Olivereau  t h e e e l , b u t L e l o u p and F o n t a i n e (i960) have significant  change  study,  very  follows.  estimation of thyroid a c t i v i t y  based  l ! 3 1 a c c u m u l a t i o n , c a n be e n t i r e l y e l i m i n a t e d f r o m  c o n s i d e r a t i o n as i t does n o t a l l o w f o r change i n e x t r a -  thyroidal I 31 loss. 1  This disadvantage  a n d T / S w h i c h show c l o s e  i s e l i m i n a t e d b y TUF  correspondence both i n seasonal  data  - 96 and i n e x p e r i m e n t a l temperature t r e a t m e n t s .  T h e same i s  o f t h e CR w h i c h c o n f o r m s w i t h t h e TUF and T / S i n d e x . behaviour of r a d i o i o d i n e at various l e v e l s  observations.  Thus  c o r r e l a t e w i t h the  This i s s t r i k i n g l y evident  t h e e x p e r i m e n t on t e m p e r a t u r e  i s concerned  (Figs.  where  8 to  11).  The CR was c o r r e l a t e d p o s i t i v e l y w i t h t e m p e r a t u r e w h i l e  the  c e l l h e i g h t a s s e s s m e n t s showed a n e g a t i v e  The  correlation.  c o l l o i d c o l o u r showed a p o s i t i v e c o r r e l a t i o n s i m i l a r t o 1^31  data.  the  Comparable d i s c r e p a n c i e s were d e m o n s t r a t e d i n t h e  seasonal data,  especially  in late  summer.  The m a i n q u e s t i o n  b e r e s o l v e d i s why t h e r a d i o c h e m i c a l d a t a a n d c o l l o i d s h o u l d show i n c r e a s e i n t h y r o i d a c t i v i t y w i t h r i s i n g and t h e c e l l h e i g h t d e c r e a s i n g a c t i v i t y w i t h r i s i n g I s i t p o s s i b l e t o r e c o n c i l e t h e s e two c o n f l i c t i n g b e a r i n g i n mind t h a t w i t h r e s p e c t influences  the  of t h y r o i d a c t i v i t y  seems c o n s i s t e n t b u t i t d o e s n o t n e c e s s a r i l y histological  true  to  data temperature  temperature.  observations,  to other environmental  , c e l l h e i g h t and r a d i o i o d i n e assessments  generally  agree? The f o l l o w i n g a t t e m p t t o do t h i s , r e s t s (i)  c e l l height  i s c o n t r o l l e d s o l e l y b y TSH  c o n t r o l from h i g h e r nervous c e n t r e s ,  on 3 a s s u m p t i o n s ( i i ) a s s u m i n g no  t h e r e l e a s e o f TSH i s  c o n t r o l l e d by a f e e d b a c k m e c h a n s i m t h a t  i s dependent upon the  b l o o d hormone ( t h y r o x i n a n d p r o b a b l y t r i i o d o t h y r o n i n e l e v e l s ) and  ( i i i ) that w i t h i n c r e a s i n g temperature there  need by t h e t i s s u e s reasonable aspect  for thyroxin.  is a  The l a t t e r i s n o t a n u n -  a s s u m p t i o n i f t h y r o x i n i s i n v o l v e d i n some  o f c e l l u l a r m e t a b o l i s m whose r a t e  temperature.  greater  increases  basic  with  -  - 97 Consider a f i s h acclimated to a low temperature. p o i k i l o t h e r m i c a n i m a l and t h e r a t e reactions so-is  w o u l d t e n d t o be l o w .  and t h e  output  the blood l e v e l s  o f TSH augmented  The c e l l h e i g h t w o u l d become g r e a t e r  v i a the  must assume  that  be i n c r e a s e d  rate  reactions  will  constants  determining the r a t e  a l s o be s t i m u l a t e d .  I f the  raised increased  feedback.  However,  constants  a  w o u l d be  a n d so w o u l d t h e  o f t h e g l a n d as measured r a d i o c h e m i c a l l y . lothermone  is  Due t o t h e p o s t u l a t e d  p e r i p h e r a l m e t a b o l i s m o f hormone,  is  f o r many b i o c h e m i c a l  As the temperature  the t h y r o x i n requirement.  depleted  constants  It  activity  in a  f o r a l l the  by r i s i n g t e m p e r a t u r e  poikibody  and,  rate  of o p e r a t i o n of the t h y r o i d  stimulatory effect  on t h e  will  peripheral  c a t a b o l i s m of t h y r o x i n were s i m u l t a n e o u s l y b a l a n c e d by an i d e n t i c a l s t i m u l a t i o n of fall  i n blood levels  pituitary.  I f the  e n t i r e temperature  synthesis,  a n d no i n c r e a s e d r e l e a s e o f TSH f r o m t h e  same r e l a t i o n s h i p w e r e t o h o l d o v e r range  then the  t h y r o x i n w o u l d be c o m p l e t e l y its  synthesis.  w i t h temperature techniques  t h e n t h e r e w o u l d be no  increasing tissue  compensated  demands  but a marked s t i m u l a t i o n where  the f o l l o w i n g species r e v e a l  As O l i v e r e a u  histology  radiochemical  (1955a,b,c)  has  no h i s t o l o g i c a l c h a n g e t o  shown  tempera-  ture - Cyprinus carpio, Tinea v u l g a r i s , A n g u i l l a a n g u i l l a , auratus  and S c y I l i u m c a n i c u l a .  A n g u i l l a a n g u i l l a , however, shown b y I - ^ l m e t h o d s r i s i n g temperature.  In at  for  by t h e s t i m u l a t i o n o f  Thus one w o u l d e x p e c t no c h a n g e i n  were concerned.  the  least  one o f t h e s e  L e l o u p and F o n t a i n e  Mugil species,  (i960) h a v e  an e x t r e m e l y marked p o s i t i v e  response  A l l t h e s e examples w o u l d conform,  to  therefore,  - 98 w i t h t h e above t h e o r y t h a t respond t o temperature  t h y r o x i n p r o d u c t i o n and c o n s u m p t i o n  t o the  same  degree.  Though s u c h an i d e a l and e c o n o m i c a l t e m p e r a t u r e i o d i n e m e t a b o l i s m may o p e r a t e , temperature  range,  evidence  suggest that  to  t h y r o x i n output at H o a r and S a l e s ,  in goldfish  low temperatures that  drop,  r e d u c t i o n than those  at  TSH w o u l d be r e l e a s e d  since,  represented  degrees.  b u i l d up i n t h e p l a s m a ,  ( F i g . 52).  alone,  the  l e s s than i t s use.  level  A t 10°  C,  increased,  (proportional to  c e l l height  t h y r o x i n producing system i s  at  lower  XIQ).  temperatures,  i n h i b i t e d more t h a n  the  The e n s u i n g l o w b l o o d t h y r o x i n  w o u l d c a u s e a p r o d u c t i o n o f TSH a n d i n c r e a s e d species i t  be  a n d TSH p r o d u c t i o n w o u l d be i n h i b i t e d  t h y r o x i n u t i l i z a t i o n system.  In other  figure,  Under these c o n d i t i o n s t h y r o x i n would  T h u s one w o u l d e x p e c t a g r e a t e r because the  This  From t h i s  of temperature  with a r e s u l t i n g reduction i n c e l l height  levels  stimulated.  b o t h o u t p u t and p e r i p h e r a l u t i l i z a t i o n have  but to d i f f e r e n t  cata-  of t h y r o x i n would  ( p r o p o r t i o n a l t o x^) m i g h t  due t o t h e e f f e c t  is  the  its peripheral  serum l e v e l  o f hormone p r o d u c t i o n i s c o n s i d e r a b l y however,  1959;  the  i n h i b i t e d more b y  and c e l l h e i g h t s  5° C, a h i g h c e l l h e i g h t  expected  than  ( H o a r , 1958,  controlling  governing  Under these c o n d i t i o n s the  has been h y p o t h e t i c a l l y  is  when t h e c e l l h e i g h t  t h e enzymes  t h y r o x i n p r o d u c t i o n and s e c r e t i o n a r e  bolism.  there  A l s o t h e d a t a on Salmo g a i r d n e r i i m p l y  This indicates  temperature  For instance,  of  entire  t h y r o x i n demands may be g r e a t e r  low temperature  more TSH p r o d u c t i o n a t greatest.  i t need n o t h o l d over t h e  nor for a l l species.  1965).  control  is possible  that  cell  blood thyroxin levels  height. might  - 99 -  F i g . 32. Diagrammatic r e p r e s e n t a t i o n of the h y p o t h e s i s t h a t t h y r o x i n u t i l i z a t i o n and p r o d u c t i o n respond d i f f e r e n t i a l l y t o temperature. X 5 and X I Q r e p r e s e n t the d i f f e r e n c e s between u t i l i z a t i o n and p r o d u c t i o n at 5 and 10° C. Since Xcj i s g r e a t e r than X Q i t would imply g r e a t e r r e l e a s e of TSH at low temperatures and a l s o a g r e a t e r c e l l h e i g h t . ±  - 100 -  drop at h i g h e r temperatures.  These  s p e c i e s w o u l d show  c e l l height w i t h i n c r e a s i n g temperature Phoxinus phoxinus  as demonstrated  ( B a r r i n g t o n and M a t t y ,  The a b o v e h y p o t h e s i s  (1965).  by  1954).  may a l s o a c c o u n t  r e p o r t e d by H o a r and E a l e s  increasing  f o r t h e anomoly  T h e y showed t h a t  although  t h y r o i d a c t i v i t y m e a s u r e d r a d i o c h e m i c a l l y was much l o w e r a t l o w temperatures,  i n preventing  cold  death i n g o l d f i s h .  T h i s c o u l d be e x p l a i n e d b y a s s u m i n g  that  at  ( i ) t h y r o x i n i s needed  and  TSH was s t i l l  low temperatures  ( i i ) due t o a d i f f e r e n t  of importance  effect  d u c t i o n and p e r i p h e r a l m e t a b o l i c  replenished. Hochachka  t h y r o x i n was b e i n g  rate  t h a n i t was b e i n g  I n a d d i t i o n , Hochachka and Hayes  (1962) h a v e shown t h a t  amounts  o f c o l d on t h y r o x i n p r o -  sites,  removed f r o m t h e b l o o d a t a g r e a t e r  i n adequate  (19&2) a n d  t h y r o x i n treatment  directs  carbohydrate  metabolism v i a the pentose  shunt  fontinalis.  T h e y h a v e shown a l s o t h a t ,  at low temperatures,  the pentose  phosphate  i n Salvelinus  s h u n t becomes more p r o m i n e n t .  This  s u g g e s t s t h e r e may a l s o be e x t r a t h y r o x i n demands a t l o w t e m p e r a tures  i n goldfish  anything that enhance  (Hoar and E a l e s ,  1963).  stimulated the a c t i v i t y  Under these  conditions  of the t h y r o i d would  s u r v i v a l a n d t h e a c t i o n o f TSH i s i m m e d i a t e l y  under-  standable. Both c e l l height  and r a d i o i o d i n e a r e i m p o r t a n t i n e s t i m a t i n g  t h y r o i d a c t i v i t y under d i f f e r e n t jl31  consistently  the c e l l h e i g h t  temperature  a l l o w s a dynamic a n a l y s i s  i s dependent  regimes.  While the  of thyroid  activity,  o n TSH s e c r e t i o n .  T h i s TSH s e c r e t i o n  may be s t i m u l a t e d v i a t h e h y p o t h a l a m i c o - h y p o p h y s e a l the plasma feedback  mechanism.  I n the l a t t e r  axis  case t h i s  o r by can give  - 101 r i s e t o d i s c r e p a n c y between h i s t o l o g i c a l and r a d i o i o d i n e c r i t e r i a . A l t h o u g h t h e h i s t o l o g i c a l c h a n g e may m e a s u r e  relative  b e t w e e n hormone p r o d u c t i o n a n d c o n s u m p t i o n i t for  assessing  the general a c t i v i t y of the  c a n be  differences unreliable  gland.  Why t h e c o l l o i d s h o u l d show c h a n g e s p a r a l l e l i n g t h e  radio-  chemical assessments r a t h e r than c e l l heigh i s not c l e a r . what apart  i s now known o f t h y r o x i n b i o s y n t h e s i s , from the i n i t i a l  uptake of  i t appears  Bern,  The r o l e o f t h e c e l l s  1962).  t o be t h e m a i n c e n t r e  release  o f t h e hormone,  it  for  the  (Gorbman a n d  i s u n c e r t a i n but  p r o b a b l y produce the c o l l o i d t h y r o g l o b u l i n s . appears  that  i o d i n e the remainder of  m e t a b o l i c pathway i s pursued i n the c o l l o i d i t s e l f  they  Since the  colloid  i o d i n a t i o n of t y r o s i n e  i s perhaps  I emphasize  that the effect  of temperature  i o d i n e m e t a b o l i s m i s p r o b a b l y u n i q u e among a l l t h e considered. directly  the t h y r o i d t i s s u e .  its estimates. on  factors  I t may be t h e o n l y e n v i r o n m e n t a l f a c t o r  affects  and  not s u r p r i s i n g t h a t  s t a t e s h o u l d c o n f o r m more c l o s e l y w i t h t h e r a d i o c h e m i c a l Finally,  Prom  that  A l l the remaining factors  s e x u a l m a t u r i t y , p h o t o p e r i o d a n d s a l i n i t y m i g h t be e x p e c t e d a c t more i n d i r e c t l y , p r e s u m a b l y b y c h a n g i n g TSH l e v e l s v i a the feedback Thus,  or v i a the hypothalamico-hypophyseal  i n a l l instances  except temperature  the l i t e r a t u r e reveals  Before  t h i s t o be  that  i n brown t r o u t  (Salmo t r u t t a )  (1955, (i)  axis.  criteria.  In  true.  c o n c l u d i n g the d i s c u s s i o n on temperature,  m u s t be made t o t h e w o r k o f S w i f t  either  s t i m u l a t i o n , one w o u l d  e x p e c t a g r e e m e n t b e t w e e n h i s t o l o g i c a l and I ^ l general  to  1959) who  reference  concluded  r a d i o i o d i n e assessments  -  -  102  -  of t h y r o i d a c t i v i t y p a r a l l e l e d the c e l l h e i g h t measurements and ( i i ) t h y r o i d a c t i v i t y was  i n v e r s e l y r e l a t e d to temperature.  These c o n c l u s i o n s d i r e c t l y oppose my  own  and  s i n c e they  d e r i v e d from-:a s p e c i e s v e r y c l o s e l y r e l a t e d t o the  are  steelhead  they r e q u i r e c l o s e r examination. S w i f t ' s c o n c l u s i o n s based on c e l l h e i g h t c o u l d be u n r e l i a b l e i n d i c a t i o n s of t h y r o i d a c t i v i t y i f the t h e s i s developed here temperature i n f l u e n c e i s c o r r e c t . 131 I '  Furthermore, S w i f t based h i s  131 on the r a t e of l o s s o f I ' from the head r e g i o n  estimates  between 6 and  72 hours.  Yet,  from my  s t u d i e s on the  t h y r o i d , maximal t h y r o i d uptake of I ^ ^ l ± 6 days. p 131 f i B I  on  3  steelhead  only achieved  at  T h i s i s a l s o the time at which d e t e c t a b l e amount of r  s  t  appear i n the b l o o d .  It i s possible that Swift 131  was  measuring predominantly r a t e of e x t r a t h y r o i d a l I '  loss,  from the v a s c u l a r head r e g i o n and f o r t h i s reason not measuring t h y r o i d a c t i v i t y at a l l . II.  FACTORS CONTRIBUTING TO SEASONAL CHANGES IH THYROID ACTIVITY IN JUVENILE STEELHEAD S e a s o n a l change i n t h y r o i d a c t i v i t y has been i n t e r p r e t e d  s o l e l y i n terms of r a d i o c h e m i c a l  data.  The  important  influence  of temperature has been d i s c u s s e d at c o n s i d e r a b l e l e n g t h ,  but  size,  environ-  sexual maturation,  ment and  photoperiod,  i o d i n e l e v e l of the  s a l i n i t y a l s o modify t h y r o i d a c t i v i t y .  Size:  The  e f f e c t of s i z e on I - ^ l metabolism agrees w i t h  the work of Hickman (1959) and Wiggs (1°62) and  i n d i c a t e s a more  r a p i d r a t e of r a d i o i o d i n e metabolism i n s m a l l e r f i s h .  That  - 103 t h e i n c r e a s e d m e t a b o l i s m i s due t o t h e s m a l l s i z e n o t due t o a p h a s e factors.  i t s e l f and  o f i n c r e a s e d growth i s i n d i c a t e d by  several  P r o m i n e n t among t h e s e i s t h e q u a l i t a t i v e c o n f o r m i t y o f  t h e b o d y mass a n d t h y r o i d d a t a t o t h e d o u b l e l o g a r i t h m i c r e l a t i o n s h i p o f b o d y mass a n d t i s s u e m e t a b o l i s m . that  t h e t h y r o i d may h a v e a v e r y g e n e r a l  T h i s would suggest  r o l e i n metabolism and  be r e l a t e d t o t h e h i g h e r 0£ demands o f s m a l l e r f i s h . been demonstrated Further evidence  i n Platichthys stellatus against  b y Hickman (1959).  the higher thyroid a c t i v i t y of small  f i s h b e i n g a s s o c i a t e d w i t h p e r i o d s o f more a c t i v e the  s i m i l a r age o f s m a l l a n d l a r g e  T h i s w o u l d mean t h a t large  T h i s has  growth  f i s h used i n these  is  studies.  t h e s m a l l f i s h were s t u n t e d r e l a t i v e  ones and h a d a s l o w e r g r o w t h r a t e .  to the  I f s t u n t e d f i s h had a  h i g h e r t h y r o i d a c t i v i t y t h a n more r a p i d l y g r o w i n g f i s h ,  this  w o u l d c o n t r a d i c t t h e f i n d i n g s o f B a r r i n g t o n (19&3) t h a t  thyroxin  s t i m u l a t e s growth i n Salmo g a i r d n e r i .  support  to the idea that alone that  i t i s a c h a n g e i n b o d y mass a n d n o t g r o w t h  i s influencing thyroid activity  The i n f l u e n c e  o f body m a s s ,  could explain the seasonal of  I t lends further  i n this  investigation.  p a r t i c u l a r l y on s m a l l  changes observed  i n certain  j u v e n i l e O n c o r h y n c h u s w h e r e t h e r e was c o n s i d e r a b l e  from a temperature  correlation (Eales,  1963).  age  Sexual P r e c o c i t y i n Male P a r r :  species departure  I t could  h a v e some b e a r i n g o n t h e d i s c r e p a n c y n o t e d b y S w i f t between brown t r o u t o f d i f f e r e n t  fish,  also  (1955,  1959)  classes. The r e l a t i o n s h i p  s e x u a l m a t u r a t i o n a n d t h y r o i d a c t i v i t y i s one o f t h e c o n t r o v e r s i a l aspects of thyroid function.  between least  Thyroid a c t i v i t y ,  - 104 w i t h few e x c e p t i o n s ,  increases  e i t h e r by h i s t o l o g i c a l o r r a d i o -  c h e m i c a l c r i t e r i a over the p e r i o d of sexual m a t u r a t i o n . recent  o f many a p p r a i s a l s  by Matty  The most  of t h i s aspect of t h y r o i d f u n c t i o n  is  (i960).  T h i s study supports the c o n c l u s i o n t h a t the gonadal a c t i v i t y are r e l a t e d . respective  In two-year-old  of the immediate temperature  t h y r o i d and  steelhead,  ir-  or photoperiod c o n d i t i o n ,  c e r t a i n extremely h i g h h i s t o l o g i c a l and r a d i o c h e m i c a l assessments of t h y r o i d a c t i v i t y were noted i n s e x u a l l y mature male i n d i v i d u a l s . This additional influence recognised,  but only at  on t h e t h y r o i d a c t i v i t y must  c e r t a i n times  of the year.  be  Metabolic  demands d u r i n g s e x u a l m a t u r a t i o n w o u l d t e n d t o be h i g h e r , it  is possible  creased.  and  t h a t t h e demands f o r t h y r o x i n w o u l d a l s o be i n -  There  i s no e v i d e n c e  thyroxin stimulates  the onset  favouring the theory of  that  sexual maturity i n  precocious  males. Photperiod: study that  It  seems c o n c l u s i v e l y d e m o n s t r a t e d  in this  i n c r e a s i n g p h o t o p e r i o d ( J a n u a r y t o Jiune) i n d u c e s  change i n t h e t h y r o i d p h y s i o l o g y o f y e a r l i n g s t e e l h e a d , a very d e f i n i t e precisely age  is  effect  o n l a r g e r members o f t h e  one y e a r o l d e r  (potential migrants).  important i n p o t e n t i a t i n g response  n o t been r e v e a l e d  by t h i s s t u d y .  i n c r e a s i n g photoperiod i s of changes w i t h i n the status  but  Whether s i z e  to photoperiod  importance i n causing  f i s h prior to migration.  One p o s s i b i l i t y i s t h a t t h e  has  species or  has  This would i n d i c a t e that  of the t h y r o i d i n the metabolic  resolved.  same  no  the  metabolic  However,  the  sequence has not  exact  been  hypothalamico-hypophyseal  systems r e c e i v i n g  centres  release  t h y r o x i n p r o d u c t i o n which i n t u r n causes  smolt.  equally r e a l i s t i c  -  impulses from p i n e a l or o p t i c  TSH w h i c h s t i m u l a t e s changes i n the  105  Evidence for t h i s  explanation is  induces  increased metabolic  general  activity)  that  that  demands  is negligible  increasing  (perhaps  t h y r o i d a c t i v i t y v i a the feedback.  photoperiod  due t o  s e c o n d a r i l y b r i n g about  a n d an  increased  increase  in  A third possibility is  that  p h o t o p e r i o d a c t i n g v i a t h e p i t u i t a r y and a d r e n a l c o r t e x  (shown  b y O l i v e r e a u i n 1?60:  affects  t o be a c t i v e  i n migrating smolts)  m i n e r a l water metabolism to the extent  that  faster  the blood i o d i n e  than they are r e p l a c e d ,  available considered  to the t h y r o i d i s reduced. in detail  This p o s s i b i l i t y  v a l i d comparisons  A s H i c k m a n (1962) h a s c a n be made b e t w e e n  t h e f i s h must be i n i o d i n e b a l a n c e w i t h i t s amount o f I ^  7  lost level is  below.  Iodine A v a i l a b i l i t y : before  so t h a t  iodides are  emphasized,  I-*-31 d e t e r m i n a t i o n s , environment,  e n t e r i n g t h e body and t h y r o i d i s  equal to  i.e. that  l e a v i n g t h e body a n d t h y r o i d .  One ; s i t u a t i o n i n w h i c h t h i s  d i t i o n may n o t be s a t i s f i e d  i n an i o d i n e d e f i c i e n t  ment, where p l a s m a I-*A s L e l o u p and F o n t a i n e generally This i s  27  is  levels  c o u l d be c o n s i d e r a b l y  (i960) h a v e s h o w n , m a r i n e  h a v e a h i g h e r p l a s m a 1^27  a t t r i b u t e d to the  i n t h e two m e d i a .  different  amounts  R o b e r t s o n and C h a n e y  Salmo g a i r d n e r i t h a t  termined the plasma i o d i n e  environreduced.  of i o d i n e  forms. present  (1953) d e m o n s t r a t e d  t h e amount o f a m b i e n t  con-  teleosts  than fresh-water  iodine directly  in de-  levels.  S i n c e the t h y r o i d uptake  of I  1  2  7  the  i s d i r e c t l y dependent  on  - 106 the plasma s u p p l y o f I  127  l e v e l s becomes l i m i t i n g .  , any  -  c o n d i t i o n tending  t o l o w e r plasma  I f a c e r t a i n m i n i m a l uptake of l 7  is  1 2  r e q u i r e d by the t h y r o i d t o produce an adequate hormone s u p p l y , 127 i n o r d e r t o m a i n t a i n t h i s l e v e l i n the g l a n d , the I  ' gradient  between t h y r o i d and plasma becomes g r e a t e r and more demands are placed  on a c t i v e t r a n s p o r t mechanisms.  T h i s i s the t h e o r e t i c a l  b a s i s of the i o d i n e d e f i c i e n c y g o i t r e d e s c r i b e d  for Salvelinus  f o n t i n a l i s (Marine and L e n h a r t , 1910), Salmo g a i r d n e r i ( R o b e r t s o n and Ghaney, 1953)  and  f o r Salmo s a l a r (La Roche, 1952).  Under t h e s e c o n d i t i o n s o f low plasma i o d i n e the g l a n d  increases  i t s iodine concentrating  hyper-  plasia.  e f f i c i e n c y by h y p e r t r o p h y and  As the above a u t h o r s demonstrated, t h i s  hyperplasia  and h y p e r t r o p h y can be c o m p l e t e l y removed by p r o v i d i n g the  fish  w i t h adequate i o d i n e . The  same response t o changing i o d i n e l e v e l s c o u l d  c e i v a b l y occur i n l e s s s t r i n g e n t circumstances.  The  con-  higher  the  l e v e l of a v a i l a b l e i o d i n e t h e l e s s e f f i c i e n t the " i o d i n e pump" has  t o be.  None o f the methods c u r r e n t l y used i n d e t e r m i n i n g  t h y r o i d a c t i v i t y w i l l d i s t i n g u i s h t r u e and in thyroid activity.  apparent  increases  I n the f o r m e r c o n d i t i o n , the net  of hormone i s i n c r e a s e d w h i l e under c o n d i t i o n s of low  output environ-  m e n t a l i o d i n e , the i o d i n e a c c u m u l a t i o n i s more e f f i c i e n t but net hormone p r o d u c t i o n  the  i s not augmented.  There i s e v i d e n c e t o suggest t h a t many f r e s h - w a t e r f i s h compensating f o r some measure o f i o d i n e d e f i c i e n c y . e a s i l y be demonstrated by o b s e r v i n g  This  are  can  the h i s t o l o g i c a l or r a d i o -  c h e m i c a l response by the t h y r o i d when the a v a i l a b l e s t a b l e  - 107 iodine i s increased.  I f these parameters  then adequate i o d i n e i s a v a i l a b l e ; this  i s i n d i c a t i v e that part  i f they are reduced  o f what was o r i g i n a l l y  m e a s u r e d a s t h y r o i d a c t i v i t y was r e a l l y low i o d i n e l e v e l s . of  iodine levels  generally the  reducing thyroid a c t i v i t y  c o n s i d e r e d as g o i t r o u s .  decreased  that  (1959) h a v e d e m o n s t r a t e d  i n c o n d i t i o n s not  i n the iodine level  in  of t h e  a n d B e r g , G-orbman a n d K o b a y a s h i  this for several  h i s t o l o g i c a l or r a d i o c h e m i c a l a c t i v i t y study,  to  i n the l i t e r a t u r e  fresh-water  I n no i n s t a n c e has a d d i t i o n o f i o d i n e i n c r e a s e d  In the present  being  H i c k m a n (1959) o b s e r v e d ,  increase  t h y r o i d uptake  then  only an adaptation  T h e r e a r e many i n s t a n c e s  starry flounder,  water  do n o t c h a n g e a t a l l  species.  either the  of the g l a n d .  underyearling  steelhead  showed  only  a m i l d r e d u c t i o n i n t h y r o i d uptake on t h e a d d i t i o n of h i g h levels at t h i s  o f i o d i n e t o t h e medium. s t a t e of development  or l e s s adequate. smoltification  I t i s therefore  t h e ambient  significant  a c t i v i t y when i n t r o d u c e d i n t o s e a w a t e r t h i s treatment It  generally  i s concluded,  depression despite  on t h e chum s a l m o n w o u l d s u p p o r t  this.  of t h y r o i d  the fact  that  on the t h y r o i d .  t h a t the smolt has very  problem i n o b t a i n i n g adequate i o d i n e .  able  at the time of  makes h i g h e r demands  therefore,  that  i o d i n e l e v e l was more  The same s p e c i e s t e s t e d  showed a v e r y  concluded  definite  The e x p e r i m e n t a l  work  Since the water  avail-  t o b o t h n o n - m i g r a n t a n d s m o l t i n g f i s h came f r o m t h e same  reservoir  ( C l e v e l a n d Dam) a n d t h e e x t e r n a l a v a i l a b i l i t y o f  i o d i n e was p r o b a b l y u n c h a n g e d endogenous  one must t h e r e f o r e  p h y s i o l o g i c a l changes.  investigate  A change i n e f f i c i e n c y  of  - 108 1127 hut  a s s i m i l a t i o n from the food c o u l d a l s o have some e f f e c t , t h i s has  not been considered  t o emerge was i n steelhead any  -  further.  A s i g n i f i c a n t point  the extremely r a p i d e l i m i n a t i o n of i n j e c t e d at the time of m i g r a t i o n .  other time of the year.  T h i s was  h i g h e r than at  Such a r a p i d l o s s of I 3 1 x  W  i n s e v e r a l species  considered general  demineralization  t h a t has  sa  consequence of  medium i . e .  the  been observed i n many d i a d r o -  A f i s h l i v i n g i n waters a l r e a d y  show a g r e a t e r  ocean (reviewed low i n i o d i n e  l o s s from the body than uptake from the  plasma i o d i n e l e v e l s would be low  and  thyroid  compensation f o r t h i s would occur as o u t l i n e d above. and F o n t a i n e ( i 9 6 0 ) contend t h a t the r a p i d a r e f l e c t i o n of a more a c t i v e 1^31 This  1  1  i  o  s  t  metabolism v i a the  routes. 2  i n the blood of a m i g r a t i n g  salmon.  F o n t a i n e ( i 9 6 0 ) have made a comprehensive study of  d i s t r i b u t i o n of s t a b l e i o d i n e i n the v a r i o u s o f the A t l a n t i c salmon smolt and  parr.  bulk  l e v e l s of l l 7  On t h i s b a s i s one would a n t i c i p a t e low  and  thyroid.  t h a t the  from the body i s v i a e x t r a t h y r o i d a l  i n the t h y r o i d and  Leloup  l o s s i s merely  study shows t h a t t h i s i s not the case and  of the I ^  also  It i s  Q  immediately p r i o r t o descent t o the  by Hoar, 1939). could  of P a c i f i c salmon ( E a l e s , l 6 l ) .  t h a t t h i s l o s s of I - ^ l i  mous species  as  (l°60)  found i n A t l a n t i c salmon smolts by Leloup and F o n t a i n e and  1.131  both  Leloup the  body compartments  While the t h y r o i d  iodine  l e v e l s are much lower i n smolts than i n p a r r , the blood i o d i n e <, i n the smolt i s a c t u a l l y h i g h e r . against  At f i r s t  s i g h t t h i s argues  the above h y p o t h e s i s , but Leloup and F o n t a i n e have a l s o  shown t h a t most of t h i s I 7 1 2  i s l o o s e l y bound t o p r o t e i n .  (This  - 109 i s a "binding d i f f e r e n t from t h a t formed i n P B I s p l i t by t r i c h l o r o a c e t i c a c i d ) . molecule  1  ^ sinoe i t i s  T h i s e f f e c t i v e l y makes t h e I  1  2  7  much l a r g e r and l e s s d i f f u s i b l e and would e x p l a i n why  a l t h o u g h the i n j e c t e d non-bound r ^ l ±  r a p i d l y l o s t , the smolt  3  127 s t i l l has h i g h plasma I  ' levels.  A c c o r d i n g t o Leloup and  Fontaine t h i s b i n d i n g mechanism, a l s o present i n s t e e l h e a d , permits a s t o r e of i o d i n e t o be b u i l t up i n the b l o o d a t s m o l t i fication.  T h i s i s claimed t o be o f s e l e c t i v e advantage due t o  the h i g h t h y r o i d a c t i v i t y a t t h i s time when the demands f o r i o d i n e would be c o n s i d e r a b l e . a b i n d i n g o f I-*-  27  I t i s argued here, however, t h a t  t o plasma p r o t e i n s does not n e c e s s a r i l y i n c r e a s e  127 the I  ' plasma l e v e l s a v a i l a b l e f o r t h y r o i d uptake.  The very  mechanism t h a t renders I ^ 7 bound t o p r o t e i n s and t h e r e f o r e 2  reduces  i t s r a t e o f e x t r a t h y r o i d a l e l i m i n a t i o n may a l s o render  i t u n a v a i l a b l e f o r uptake by the t h y r o i d .  I n other words the  b i n d i n g o f p r o t e i n s t o i o d i d e may decrease the f r e e i o d i d e a v a i l a b l e f o r t h y r o i d uptake and may accentuate any tendency towards g o i t r e . One experiment t o t e s t t h i s h y p o t h e s i s would be t o i n v e s t i gate the uptake o f J.131 from a p o o l o f protein-bound blood. then  j!31 i n the  I f the h y p o t h e s i s o f Leloup and F o n t a i n e were c o r r e c t ( i ) t h e r e would be a reduced  of the p r o t e i n - t a g g e d I 3 1 1  rate of extrathyroidal loss because t h i s l 3 1 1  a  n  d  w  a  s  being  l o s t so s l o w l y from the blood i t would be taken up i n i n c r e a s e d amounts by the t h y r o i d . Chavin  (1956) on C a r a s s i u s a u r a t u s .  g o l d f i s h with I The  Such a experiment was performed by  1  5  1  He i n j e c t e d one group o f  and one group w i t h I 3 1  bound t o albumin.  1  e x c r e t i o n r a t e o f the p r o t e i n - t a g g e d I 3 1 was v e r y slow but 1  - 110 the t h y r o i d accumulation of r a d i o a c t i v i t y was with free I ^ . 1  of I 7  T h i s supports the present t h e o r y t h a t  1  i o d i n e a v a i l a b l e t o the t h y r o i d . t h a t the smolt may  be  2  }  can t h e r e f o r e  i n i t s body.  conclude despite levels  e i t h e r o r g a n i c a l l y bound or f r e e , i n the t h y r o i d  itself  would support t h i s .  The  binding  The  of  v e r y low  2  of I- - 7  One  i n an i o d i n e d e f i c i e n t c o n d i t i o n  the apparent abundance of 1"17  of i o d i n e to p r o t e i n i s p o s s i b l y  a f o r t u i t o u s change occasioned by an i n c r e a s e of a blood p r o t e i n f r a c t i o n t h a t has for  binding  to p r o t e i n s does not n e c e s s a r i l y augment the p o o l  1 2  1  no h i g h e r than  at s m o l t i f i c a t i o n  an e x c e p t i o n a l  affinity  iodides. I t i s concluded, t h e r e f o r e ,  t h y r o i d a c t i v i t y can be l e v e l , and  t h a t the fresh-water t e l e o s t  i n f l u e n c e d markedly by the plasma i o d i n e  t h a t t h i s l e v e l i n t u r n i s dependent on the  r e s u l t of both the uptake of I 7 1 2  or the water and  net  from the medium v i a the  food  on i t s t o t a l r a t e o f l o s s from the body.  i s suggested t h a t the a l t e r e d p h y s i o l o g y  of the  l e s s i o d i n e a v a i l a b l e t o the t h y r o i d and  t h a t this, t o some  yet u n a s c e r t a i n e d  degree, c o n t r i b u t e s  t o the  It  smolt renders  high  thyroid  " a c t i v i t y " at s m o l t i f i c a t i o n . Salinity: was  The  p r i n c i p l e object  of the  entire investigation  an a n a l y s i s of f a c t o r s c o n t r i b u t i n g t o s e a s o n a l change i n  l l 3 1 metabolism and o f development.  t h y r o i d f u n c t i o n during  the f r e s h - w a t e r phase  On t h i s b a s i s i n v e s t i g a t i o n of the e f f e c t of  s a l i n i t y appears i n c o n s i s t e n t , as f i s h i n f r e s h water  (by  d e f i n i t i o n ) would never be exposed t o such a s i t u a t i o n . of response t o s a l i n i t y , however, could h e l p t o e l u c i d a t e  A  study the  - I l l change  i n iodine metabolism at s m o l t i f i c a t i o n .  been i n d i c a t e d t h a t  this  c a n be e x p l a i n e d on t h e b a s i s  i o d i n e d e f i c i e n c y but t h e .extent and o t h e r t h e o r i e s be e x c l u d e d  I t has already-  of t h i s effect  of  i s n o t known  put forward to explain t h i s increase  cannot  immediately.  One s u g g e s t i o n ,  referred  to earlier,  i s that  a smolting  fish  p o s s i b l y due t o i n c r e a s e d amounts o f m e t a b o l i c w a t e r and t o c h a n g e s i n i t s i o n r e t a i n i n g m e c h a n i s m s p r i o r t o movement t o t h e sea,  has problems a s s o c i a t e d  1 9 5 2 , 1959; E a l e s , should a l l e v i a t e water  smolt.  1961).  with salt  and w a t e r balance  Consequently,  transfer  the osmotic problems encountered  (Hoar,  t o sea water by t h e f r e s h -  T h u s t h y r o i d a c t i v i t y w o u l d be h i g h i n t h e s m o l t  i n f r e s h w a t e r w h e r e t h e m e t a b o l i c demands a r e p r e s u m a b l y but reduced i n sea water. and p o s t - m i g r a n t fresh water, water.  The d a t a f r o m b o t h y e a r l i n g  steelhead  chum w h e r e m e t a b o l i c demands m i g h t be h i g h i n  however,  show i n c r e a s e d t h y r o i d a c t i v i t y  i n sea  T h i s a g r e e s w i t h t h e f i n d i n g s o f H i c k m a n (1959)  euryhaline flounder,  Platichthys stellatus.  where  on t h e  salinity  i n c r e a s e d b o t h oxygen consumption and t h y r o i d a c t i v i t y . not support the theory that greater  great,  i n f r e s h water  t h y r o x i n demands o f t h e s m o l t a r e  due t o o s m o t i c  I n c o n c l u s i o n , many f a c t o r s contribute to the seasonal  I t does  cycle  affect  stress. t h y r o i d a c t i v i t y and  of the steelhead.  Universal  among t h e s e i s t e m p e r a t u r e a n d a t a l l s e a s o n s t h e t h y r o i d to this  important v a r i a b l e .  t o temperature size  Superimposed on t h e b a s i c  are the effects  i s p r o b a b l y always  of other f a c t o r s .  exerting  some e f f e c t .  responds  response  Of t h e s e ,  body  Thyroid a c t i v i t y  - 112 i s l o g a r i t h m i c a l l y r e l a t e d t o body mass. r e l a t i o n s h i p , appreciable  As a r e s u l t of t h i s  e f f e c t s on the t h y r o i d are noted "only  over the lower s i z e range and become n e g l i g i b l e above 20 grams. At a r e s t r i c t e d time o f year, the s t a t e of sexual m a t u r a t i o n (as shown here i n p r e c o c i o u s demand on the t h y r o i d .  male p a r r ) can p l a c e an e x t r a  In general,  the t h y r o i d appears r e f r a c t o r y  t o changes i n p h o t o p e r i o d but p o t e n t i a l migrants show a very d e f i n i t e response t h a t i s superimposed on a p o s i t i v e response, t o temperature i n t h e s p r i n g .  The p h y s i o l o g i c a l route by which  the i n c r e a s i n g photoperiod p r o g r e s s i v e l y e x e r t s i t s e f f e c t i s unknown.  F i n a l l y , c e r t a i n o f these changes i n t h y r o i d may be  apparent r a t h e r than r e a l . f a c t o r s on plasma I  l 2  Due t o t h e e f f e c t s o f d i f f e r e n t  7 l e v e l s , the a v a i l a b i l i t y of i o d i n e t o  the t h y r o i d may be a l t e r e d .  I t i s considered  that t h i s e f f e c t  may be p a r t i c u l a r l y important i n the smolt but n e g l i g i b l e i n the  parr. III. Any  THE ROLE OF THE THYROID IN THE STEELHEAD  comprehensive theory  concerning  the t h y r o i d r o l e i n  s t e e l h e a d must take i n t o account t h a t r i s i n g temperature, s i z e , sexual m a t u r a t i o n and exposure t o h i g h e r g r e a t e r demands on t h e t h y r o i d .  in poikilotherms.  s a l i n i t y a l l exert  Without e x c e p t i o n  f a c t o r s have been a s s o c i a t e d w i t h h i g h e r  small  a l l these  r a t e s o f metabolism  T h i s suggests t h a t the t h y r o i d i s i n v o l v e d  i n some b a s i c metabolic r o l e .  There i s no evidence from t h i s  study t o i m p l i c a t e a s p e c i f i c r o l e of the t h y r o i d i n a diadromous t e l e o s t although s e v e r a l such r o l e s have been suggested i n the  - 113 literature.  These r e l a t e t o the r o l e of t h y r o x i n i n i n d u c i n g  s a l i n i t y preference 1949) 1950)  and .  -  (Baggerman, i 9 6 0 ,  1963), s i l v e r i n g  changes i n glycogen metabolism (Fontaine  Data presented  (Robertson,  and Hatey,  e a r l i e r showed the independence of  s i l v e r i n g on i n c r e a s e d t h y r o x i n output and  I concluded t h a t  s t i m u l a t i o n of guanine d e p o s i t i o n i s a pharmacological  the  effect  of t h y r o x i n , s i n c e i n c r e a s e d t h y r o x i n i s not r e q u i r e d i n  nature.  Furthermore, t h y r o x i n a d m i n i s t r a t i o n does not always produce s i l v e r i n g and  t h y r o i d e x t r a c t s c o n t a i n i n g mainly i o d i n e  t h y r o g l o b u l i n are o f t e n more potent.  The  and  same i s t r u e where  experiments w i t h t h y r o x i n and t h y r o i d e x t r a c t s have been used t o s t i m u l a t e m o b i l i z a t i o n of glycogen c h a r a c t e r i s t i c of the smolt.  In a d d i t i o n , though Baggerman (1963) obtained p o s i t i v e  r e s u l t s with the e f f e c t o f TSH  on s a l i n i t y p r e f e r e n c e  i n h i b i t i o n of s a l i n i t y p r e f e r e n c e  and  an  with t h y r o i d i n h i b i t o r s i n  j u v e n i l e Oncorhvnchus, she could o b t a i n no change i n s a l i n i t y response w i t h t h y r o x i n alone (Baggerman, i 9 6 0 ) . Although such s p e c i f i c r o l e s of t h y r o x i n cannot be e l i m i n a t e d , c o n s i d e r a t i o n of the b e t t e r e s t a b l i s h e d metabolic  general  r o l e s of t h y r o x i n might be more p r o f i t a b l e .  g e n e r a l metabolic  entirely  Such a  r o l e i s i n f a c t suggested by the data on  of p e r i p h e r a l c a t a b o l i s m  of r a d i o a c t i v e hormone.  sites  A l l the  m e t a b o l i c a l l y a c t i v e t i s s u e s showed a s i m i l a r r i s e i n hormone accumulation suggesting tissue-specific action.  a systemic  r a t h e r than an organ or  Muscle w i t h a lower metabolism than  kidney, l i v e r , b r a i n or probably gut had  a much lower accumu-  l a t i o n of presumed hormonal r a d i o a c t i v i t y , w h i l e  s k i n the most  - 114  metabolically  inactive  concluded therefore cells will  of the  o f a l l showed t h e  that  quantitative  to t h e i r rate  of  r o l e of t h y r o x i n i n metabolism, mammals, i s t h e a l m o s t c o m p l e t e  It  and the p o s i t i v e  requirements  f o r not accepting  similar to that inability  oxygen consumption i n p o i k i l o t h e r m s .  i n the g o l d f i s h a r e  rise.  of  results  b e s t known.  a  general  described  a few  However,  n o t mean t h a t  i t h a s no m e t a b o l i c * r o l e .  merely  (1953)  rate  does  i s possible  t h y r o x i n p l a y s a s i m i l a r r o l e at the b i o c h e m i c a l l e v e l warm a n d c o l d - b l o o d e d v e r t e b r a t e s .  factor  c o n t r o l s them.  i n the r e a c t i o n but i t  to certain reactions In the l a t t e r ,  serve e x a c t l y  the  sole  factor.  limiting  increase  same s u b c e l l u l a r r o l e , It  is  former not  speculated  i n both o n l y does a  reason  t h o u g h t h y r o x i n may i t may be r a r e l y that  i n t h y r o x i n l e v e l m i g h t n o t be a b l e substances  i n such a  were a l s o  If  the  situation  t o cause an  under c e r t a i n r a t h e r r a r e experimental  increase  lacking.  conditions  t h y r o x i n m i g h t be t h e s o l e  limiting factor.  case  be t r u e w i t h M u l l e r ' s w o r k )  (as c o u l d c o n c e i v a b l y  that  i s presumably  and f o r t h i s  however,  i n metabolism because other v i t a l However,  I n the  stimulate  because  s t i m u l a t e a change i n m e t a b o l i c It  in  notable  o b t a i n e d by M u l l e r  not  limiting  all  each  of t h y r o x i n to  There are  thyroxin w i l l  t h y r o x i n take part  is  metabolism.  One o f t h e p r i n c i p l e r e a s o n s  exceptions  least  t h y r o x i n i s probably important to  body but t h a t  be g e a r e d  -  t h i s were  the  then  a d m i n i s t r a t i o n of t h y r o x i n would a l l o w metabolism to proceed a faster  rate  and oxygen c o n s u m p t i o n w o u l d  Such a passive  role  at  increase.  o f t h y r o x i n w o u l d p r o b a b l y be  accompanied  - 115  -  by an e q u a l l y p a s s i v e form of c o n t r o l back method p o s t u l a t e d e a r l i e r .  i . e . v i a the plasma f e e d -  I t i s proposed t h a t as a  r e s u l t of a l l the thyroxin-demanding processes temperature, small s i z e , sexual maturation,  ( r i s e i n body  increased  salinity  and p o s s i b l y i n c r e a s e d e x e r c i s e ) plasma l e v e l s of hormone would be d e p l e t e d and system.  then r e i n s t a t e d v i a the hypophyseal feedback  Where the e f f e c t of photoperiod  i s concerned i t i s  tempting to suggest t h a t t h i s s t i m u l a t e s the t h y r o i d d i r e c t l y v i a the hypothalamico-hypophyseal system. d i r e c t evidence e i t h e r induce t h y r o i d due  f o r t h i s and  But there i s no  i t i s equally l i k e l y that i t could  d e m i n e r a l i z a t i o n with secondary e f f e c t s on  to iodine d e f i c i e n c y .  l e n g t h e n i n g photoperiod  the  A t h i r d p o s s i b i l i t y i s that  s t i m u l a t e s the smolt t o h i g h e r  levels  of g e n e r a l a c t i v i t y t h a t s e c o n d a r i l y cause i n c r e a s e d t h y r o x i n production. The  g e n e r a l s p e c u l a t i v e c o n c l u s i o n i s that t h y r o x i n  subserve a very s i m i l a r b i o c h e m i c a l and homiotherms.  may  r o l e i n both p o i k i l o t h e r m s  While i n the l a t t e r , however, there  are  d i s t i n c t advantages t o i t s c o n t r o l l i n g metabolism by being  a  l i m i t i n g f a c t o r i n c e r t a i n reactions, i n poikilotherms t h i s i s not the case.  Instead, the p r o d u c t i o n of t h y r o x i n may  governed s o l e l y by the demands of the t i s s u e s and c o n t r o l l e d t o a l a r g e extent, feedback mechanism.  be  i t s production  or e n t i r e l y by the hypophyseal  - 116 SUMMARY AND CONCLUSIONS I.  R e l i a b l e estimates of t h y r o i d a c t i v i t y can be made u s i n g s i n g l e t e r m i n a l assessments  of TUF, T/S or CR a t 4 or 8  days a f t e r i n j e c t i o n i n j u v e n i l e s t e e l h e a d . II.  Seasonal changes i n r a d i o i o d i n e metabolism  revealed  some c o r r e l a t i o n w i t h temperature but smolts departed from t h i s r u l e i n having h i g h e r than a n t i c i p a t e d v a l u e s . Smolts had a h i s t o l o g i c a l l y a c t i v e g l a n d but, i n g e n e r a l , c e l l h e i g h t showed a n e g a t i v e c o r r e l a t i o n w i t h temperature. III.  E x p e r i m e n t a l i n v e s t i g a t i o n of temperature on t h y r o i d a c t i v i t y r e v e a l e d a d i r e c t dependence of a l l aspects o f J.131 metabolism on temperature from 4 t o 18° C. H i s t o l o g i c a l l y , however, the g l a n d was more a c t i v e a t low temperatures.  IV.  P h o t o p e r i o d e x e r t e d no e f f e c t on c e l l h e i g h t or r a d i o i o d i n e metabolism of s t e e l h e a d y e a r l i n g s from January to  V.  July.  Body mass had an important e f f e c t on r a d i o i o d i n e metab o l i s m w i t h s m a l l f i s h having the h i g h e r a c t i v i t y .  The  l o g a r i t h m i c dependence of t h y r o i d a c t i v i t y on mass p e r m i t t e d n e g l e c t of t h i s f a c t o r i n f i s h over 2 0 grams. VI.  P o t e n t i a l migrant s t e e l h e a d examined from January t o June showed a marked p o s i t i v e response t o both i n c r e a s i n g temperature and i n c r e a s i n g p h o t o p e r i o d .  The  h i g h smolt v a l u e was due t o summation o f the photop e r i o d and temperature response.  There was no evidence  - 117  -  f o r a synergism between temperature and  photoperiod  influences. VII.  Precocious had  s e x u a l l y mature male p a r r i n March f r e q u e n t l y  a higher  radiochemical VIII. IX.  Exercise  t h y r o i d a c t i v i t y both by h i s t o l o g i c a l  and  techniques (than immature forms).  s l i g h t l y i n c r e a s e d the c o n v e r s i o n  S a l i n i t y g e n e r a l l y caused an i n c r e a s e  ratio.  i n thyroid activity.  P a r r i n mid-summer were only s l i g h t l y s e n s i t i v e t o increased  i o d i n e l e v e l s i n the medium but  smolts  and  chum salmon showed a great r e d u c t i o n i n t h y r o i d  activity  v/hen the i o d i n e c o n c e n t r a t i o n  raised.  of the medium was  T h i s i n d i c a t e s that p a r t of the h i g h a c t i v i t y of smolt i s due X.  t o an i o d i n e d e f i c i e n c y .  P o t e n t i a l migrant steelhead temperature and activity. and  the  h e l d under c o n d i t i o n s of  8-hour daylength had  low  unchanging t h y r o i d  A normal percentage of these f i s h s i l v e r e d  the r o l e of t h y r o x i n i n s m o l t i f i c a t i o n i s t h e r e f o r e  questioned. XI.  "Radiohormone" was but  shown t o b u i l d up i n s e v e r a l t i s s u e s  e s p e c i a l l y i n those that were m e t a b o l i c a l l y most  active.  Gill  showed an accumulation of  !-131  that  could  be i n d i c a t i v e of a r o l e i n i o n exchange w i t h the surrounding medium. I n t e g r a t i o n of these f i n d i n g s suggests t h a t  environmental  temperature and mass of f i s h always p l a y a r o l e i n determining thyroid activity. can be discounted  However, i n f i s h - over 20 grams the as an important v a r i a b l e .  latter  It i s believed  that  - 118 temperature  i s u n i q u e amongst  thyroid i n that  affecting  the  i t may a l t e r t h y r o i d f u n c t i o n w i t h o u t  the  m e d i a t i o n of TSH.  the v a r i a b l e s  This could explain discrepancies  h i s t o l o g i c a l and r a d i o c h e m i c a l d a t a , and i n t h e  be t h y r o i d r e s p o n s e s  It  is  size  and t e m p e r a t u r e  t o gonad m a t u r a t i o n ,  suggested that  gonad m a t u r a t i o n ,  a l l require metabolic  t h y r o i d a c t i v i t y w o u l d be a n t i c i p a t e d . iodine concentration is  really  often  is,  however,  considered  more e f f i c i e n t  at  exercise  develop  increase  i t s general  It  increased  The r e s p o n s e  to  In the  increased  of the gland  of is  smolt such a  i s postulated  that  f i s h and t h a t  role  t h y r o x i n subserves a very  by t h e demands  in  photolow  an  activity.  l a c k of a s p e c i f i c  b i o c h e m i c a l r o l e and t h a t  mechanism.  water.  i n s i l v e r i n g and  accumulation i n a c t i v e l y metabolizing tissues,  concluded that  governed  and  s m o l t i f i c a t i o n which lead to  in thyroid  I n view of the  of the  t h e t h y r o i d " i o d i n e pump" becomes  apparent.  at  exercise,  work and an i n c r e a s e  p e r i o d c o u l d cause i o n i c imbalance i n the iodine levels  may  an i n d i c a t i o n t h a t p a r t  low i o d i n e l e v e l s .  c o m p e n s a t i o n seems v e r y  is  increased  t o be t h e a c t i v i t y  a compensation whereby  apparent  study  influences  p h o t o p e r i o d and i o d i n e c o n c e n t r a t i o n  osmotic pressure  what  in this  literature.  S u p e r i m p o s e d on t h e  salinity,  observed  between  fundamental  i t s production i s probably  of the t i s s u e s  it  acting via a  largely feedback  - 119  -  BIBLIOGRAPHY Baggerman, B. i 9 6 0 . S a l i n i t y p r e f e r e n c e , t h y r o i d a c t i v i t y and the seaward m i g r a t i o n of f o u r s p e c i e s of P a c i f i c salmon (Oncorhvnohus). J . F i s h . Res. Bd. Canada, 17: 295-322. Baggerman, B. 1963. The e f f e c t of TSH and a n t i t h y r o i d substances on s a l i n i t y p r e f e r e n c e and t h y r o i d a c t i v i t y i n j u v e n i l e P a c i f i c salmon. Can. J . Z o o l . , 41: 307-319. B a r r i n g t o n , E. J . W. 1961. Metamorphic processes i n f i s h e s and lampreys. Am. Z o o l o g i s t , 1: 9 7 - 1 0 6 . B a r r i n g t o n , E. J . W. 1963. An i n t r o d u c t i o n t o g e n e r a l and comparative e n d o c r i n o l o g y . Clarendon P r e s s , Oxford. B a r r i n g t o n , E. J . W. and Matty, A. J . 1954. Seasonal v a r i a t i o n i n the t h y r o i d gland of the minnow, Phoxinus phoxinus L., w i t h some o b s e r v a t i o n s on the e f f e c t of temperature. Proc. Z o o l . Soc. London, 124: 8 9 - 9 5 . Berg, 0 . , Gorbman, A. and Kobayashi, H. 1959. The t h y r o i d hormones i n i n v e r t e b r a t e s and lower v e r t e b r a t e s . Symposium on Comparative E n d o c r i n o l o g y , e d i t e d by A. Gorbman. John Wiley and Sons, New York, pp. 302-319. Chavin, W. 1956. T h y r o i d d i s t r i b u t i o n and f u n c t i o n i n the g o l d f i s h , C a r a s s i u s auratus L. J . E x p t l . Z o o l . , 133: 259-279.  .>.  • .• ———•  Comar, C. L. 1955. R a d i o i s o t o p e s i n b i o l o g y and McGraw-Hill Book Co., Toronto.  agriculture.  E a l e s , J . G. 1961. A comparative study of i o d i n e metabolism i n j u v e n i l e Oncorhynchus. M.Sc. T h e s i s , Univ. of B r i t i s h Columbia. E a l e s , J . G. 1963. A comparative study of t h y r o i d f u n c t i o n i n migrant j u v e n i l e salmon. Can. J . Z o o l . , 41: ( i n p r e s s ) . Fontaine, M. 1954. B i o l . Rev.,  Du determinisme p h y s i o l o g i q u e des m i g r a t i o n s . 29: 390-418.  F o n t a i n e , M. 1959. I n f l u e n c e de l a nage a contre-courant sur l e metabolisme de l ' i o d e et l e fonctionnement t h y r o i d i e n chez l a T r u i t e a r c - e n - c i e l (Salmo g a i r d n e r i i R i c h ) . C. R. Soc. B i o l . , P a r i s , 249: 343-347. G a y l o r d , H. R. and Marsh, M. C. 1912. Carcinoma of the t h y r o i d i n salmonid f i s h e s . B u l l . U. S. Bur. F i s h . , 32: 367-524. Gorbman, A. and Bern, H. A. 1962. A textbook of comparative endocrinology. John W i l e y and Sons, New York.  - 120 Gurr, E . 1953. A p r a c t i c a l manual o f m e d i c a l and b i o l o g i c a l s t a i n i n g techniques. Leonard H i l l L i m i t e d , London. Hagen, G, 1936. 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T h y r o i d f u n c t i o n i n some anadromous and l a n d locked t e l e o s t s . T r a n s . Roy. Soc. Canada, 4 6 : 4 9 - 5 3 . Hoar, W. S. 1959. Endocrine f a c t o r s i n the e c o l o g i c a l a d a p t a t i o n of f i s h e s . Symposium on Comparative E n d o c r i n o l o g y , e d i t e d by A. Gorbman, John W i l e y and Sons, New York, pp. 1 - 2 3 . Hoar, W. S. and B e l l , C. M. 1950. The t h y r o i d gland i n r e l a t i o n t o the-seaward m i g r a t i o n of P a c i f i c salmon. Can. J . Res.,  28:  126-136.  Hoar, W. S. and E a l e s , J . G. 1963. The t h y r o i d gland and low temperature r e s i s t a n c e of g o l d f i s h . Can. J . Z o o l . , 41: ( i n p r e s s ) . Hochachka, P. W. and Hayes, F. R. 1962. The e f f e c t o f temperat u r e a c c l i m a t i o n on pathways of g l u c o s e metabolism i n the t r o u t . Can. J". Z o o l . , 40: 261-270. Hochachka, P. W. 1962. T h y r o i d a l e f f e c t s on pathways f o r carbohydrate metabolism i n a t e l e o s t . Gen. and Comp. E n d o c r i n o l . , 2: 4 9 9 - 5 0 5 . Honma, Y. 1959. S t u d i e s on the endocrine glands o f a salmonid f i s h , ayu, P l e c o g l o s s u s a l t i v e l i s . 1. Seasonal v a r i a t i o n i n t h e endocrines o f t h e annual f i s h . J . Fac. S c i . N i i g a t a Univ. (Ser. I I ) , 2: 2 2 5 - 2 3 3 . La Roche, G. 1952. E f f e t s de p r e p a r a t i o n s t h y r o i d i e n n e s e t d'iodures s u r l e g o i t r e ("pseudo-cancer) des salmonides. Revue Canadienne de B i o l o g i e , 11: 4 3 9 - 4 4 5 .  - 121 Leloup,  J . and F o n t a i n e , M . i 9 6 0 . Iodine metabolism i n lower vertebrates. A n n . N . Y . A c a d . S c i . , 86: 316-353.  L i e b e r , A . 1936. fossilis.  Jahreszyklus der Schildruse von Misgurnus Z . w i s s . Z o o l . , 148: 364-400.  M a h e r , F . P . a n d L a r k i n , P . A . 1954. L i f e h i s t o r y o f t h e s t e e l head t r o u t o f t h e C h i l l i w a c k R i v e r , B r i t i s h C o l u m b i a . T r a n s . Am. F i s h . S o c , 84: 27-38. M a r i n e , D. 1914. The r a p i d i t y o f t h e i n v o l u t i o n o f a c t i v e t h y r o i d h y p e r p l a s i a s of brook t r o u t f o l l o w i n g the use of f r e s h sea f i s h as f o o d . J . E x p t l . M e d . , 19: 376-382. M a r i n e , D. a n d L e n h a r t , C . H . 1910. O b s e r v a t i o n s a n d e x p e r i m e n t s on t h e s o - c a l l e d c a r c i n o m a o f S a l m o f o n t i n a l i s a n d i t s relation to ordinary goitre. J . E x p t l . M e d . , 12: 311-337. Matty, A . J . i960. Thyroid cycles i n f i s h . L o n d o n , 2: 1-15. Muller,  Symp. Z o o l . S o c .  J . 1953. U b e r d i e W i r k u n g v o n T h y r o x i n u n d T h y r e o t r o p e m Hormon a u f d e n S t o f f w e c h s e l u n d d i e F a r b u n g d e r G o l d f i s c h e s . Z . 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P r o d u c t i o n of the s i l v e r y smolt stage i n rainbow t r o u t by i n t r a m u s c u l a r i n j e c t i o n o f mammalian t h y r o i d e x t r a c t and t h y r o t r o p i c hormone. J". E x p t l . Z o o l . , 110: 3 3 7 - 3 5 5 . Robertson, 0 . H. and Chaney, A. L. 1953. T h y r o i d hyperplasia and t i s s u e i o d i d e content i n spawning rainbow t r o u t : a comparative study of Lake Michigan and C a l i f o r n i a searun t r o u t . P h y s i o l . Z o o l . , 26: 328-340. Snedecor, G. W. 1956. S t a t i s t i c a l methods. C o l l e g e P r e s s , Ames, Iowa.  The Iowa S t a t e  S t e e l , R. G. D. and T o r r i e , J . H. i 9 6 0 . P r i n c i p l e s and procedures of s t a t i s t i c s . McGraw-Hill Book Co., New York. S w i f t , D. R. 1955. Seasonal v a r i a t i o n s i n the growth r a t e , t h y r o i d gland a c t i v i t y and food r e s e r v e s o f brown t r o u t (Salmo t r u t t a L . ) . J . E x p t l . B i o l . , 3 2 : 751-764. S w i f t , D. R. 1959. Seasonal v a r i a t i o n i n t h e a c t i v i t y o f the t h y r o i d gland o f y e a r l i n g brown t r o u t , Salmo t r u t t a L. J . E x p t l . B i o l . , 36: 120-125. S w i f t , D. R. i 9 6 0 . C y c l i c a l a c t i v i t y o f the t h y r o i d gland o f f i s h i n r e l a t i o n t o environmental changes. Symp. Z o o l . Soc. London, 2: 17-27. Wiggs, A. J . 1962. Some f a c t o r s a f f e c t i n g r a d i o i o d i d e metabolism i n the threespine stickleback. M.Sc. T h e s i s , U n i v . of B r i t i s h Columbia. Z a i t z e v , A. V. 1955. A h i s t o l o g i c a l i n v e s t i g a t i o n of the annual changes o f the t h y r o i d g l a n d of the p i k e , and the neuros e c r e t o r y a c t i v i t y of the hypothalamic n u c l e i i n the seasonal change o f the t h y r e o t r o p i c f u n c t i o n o f the hypophysis. D o k l . Akad. Nauk, U. S. S. R., 104: 315-318 (In Russian, quoted from P i c k f o r d and A t z , 1 9 5 7 ) .  

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