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Some effects of ultraviolet radiation on fish Bell, Gertrude Mary 1949

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(in ftf fat sc SOME EFFECTS OF ULTRAVIOLET RADIATION ON FISH by Gertrude Mary B e l l A Thesis Submitted i n P a r t i a l F u l f i l m e n t of the Requirements f o r the Degree of M A S T E R O F A R T S i n the Department of ZOOLOGY The U n i v e r s i t y of B r i t i s h Columbia APRIL, 1949 TABLE OP CONTENTS INTRODUCTION v HISTORICAL REVIEW 1 U l t r a v i o l e t l i g h t . 1 U l t r a v i o l e t i n s u n l i g h t • 1 Some b i o l o g i c a l e f f e c t s of u l t r a v i o l e t r a d i a t i o n • 2 E a r l y work u s i n g u l t r a v i o l e t 4 E f f e c t on human beings 6 E f f e c t on eggs 8 E f f e c t of X - r a d i a t i o n on f i s h 8 METHODS AND MATERIALS 10 Sources of U l t r a v i o l e t 10 S t a n d a r d i z a t i o n of lamps 10 Experimental animals 12 Methods 13 H i s t o l o g i c a l s t u d i e s 14 Comparison of lamps and s u n l i g h t 15 RESULTS 19 P r e l i m i n a r y experiments 19 Chum salmon eggs 21 Sockeye salmon e ggs 24 Sockeye salmon a l e v i n s 30 H i s t o l o g i c a l s t u d i e s 34 (A) S k i n 34 (B) Other t i s s u e s 40 i i DISCUSSION 41 U l t r a v i o l e t and r a t e of h a t c h i n g 41 M o r t a l i t i e s i n eggs and a l e v i n s 41 E f f e c t s of u l t r a v i o l e t on f i s h s k i n .... 44 (A) P r e l i m i n a r y experiments ....... 44 (B) A l e v i n sockeye salmon experiments ................... 46 E f f e c t s of u l t r a v i o l e t on other t i s s u e s . 49 SUMMARY . . . 51 ACKNOWLEDGMENTS 52 LITERATURE CITED 53 i i i LIST OF ILLUSTRATIONS TABLES Table I Table I I Table I I I Table IV Table V Table VI - Average hours of a c t u a l sunshine and of erythemal u l t r a v i o l e t energy as measured at C l e v e l a n d , Ohio. - T o t a l number of E - v i t o n hours i n c i d e n t upon a l e v l n sockeye salmon. - Percentage m o r t a l i t i e s of chum salmon eggs, - Percentage m o r t a l i t i e s of sockeye salmon eggs. - Removals, d a i l y m o r t a l i t i e s and cumulative percentage m o r t a l i t y of a l e v i n sockeye salmon. - C a l c u l a t e d t h e o r e t i c a l deaths of c o n t r o l and i r r a d i a t e d a l e v i n sockeye salmon. FIGURES F i g u r e 1 F i g u r e 2 F i g u r e 3 F i g u r e k F i g u r e 5 Figu r e 6 F i g u r e 6a F i g u r e 7 F i g u r e 8 - Some b i o l o g i c a l e f f e c t s of u l t r a v i o l e t r a d i a t i o n . - Graphs of percentage m o r t a l i t y of chum salmon eggs. - Graphs of percentage m o r t a l i t y of sockeye salmon eggs. - Sockeye salmon a l e v i n s i+6 days a f t e r i r r a d i a t i o n . - C o n t r o l sockeye salmon a l e v i n s k6 days a f t e r the experiment began. - Ski n of a l e v i n sockeye salmon 3 days a f t e r i r r a d i a t i o n o f tac. - D a i l y cumulative m o r t a l i t i e s of sockeye salmon a l e v i n s . - S k i n of c o n t r o l sockeye salmon 7 days a f t e r experiment began. - Ski n of f i s h k days a f t e r an i r r a d i a t i o n of 2x. i v P i g . 9 - S k i n of f i s h 4 days a f t e r an i r r a d i a t i o n of 8x. P i g . 10 - Ski n of f i s h 7 days a f t e r an i r r a d i a t i o n of 0.5x. P i g . 11 - S k i n of f i s h 7 days a f t e r an i r r a d i a t i o n o f 2x. P i g . 12 - S k i n o f f i s h 13 days a f t e r a dosage of 0.5x. V SOME EFFECTS OF ULTRAVIOLET RADIATION ON FISH INTRODUCTION The e f f e c t s of u l t r a v i o l e t have been e x t e n s i v e l y s t u d i e d on man and on the lower forms such as b a c t e r i a . L i t t l e or no work has been done on the lower v e r t e b r a t e s , p a r t i c u l a r l y a q u a t i c forms. Such s t u d i e s w i l l prove i n t e r e s t i n g from the comparative angle and may be of some p r a c t i c a l s i g n i f i c a n c e , s i n c e i t has been suggested t h a t young h a t c h e r y f i s h may be a d v e r s e l y a f f e c t e d by s u n l i g h t . The wavelengths used are approximately the same as those found i n the u l t r a v i o l e t of s u n l i g h t . In t h i s study the e f f e c t s of the near u l t r a -v i o l e t have been i n v e s t i g a t e d as f o l l o w s : (1) l e t h a l e f f e c t s on a d u l t g o l d f i s h (Carassius  a u r a t u s ) , coho salmon f r y (Onycorhynchus k l s u t c h ) , and d e v e l o p i n g eggs of chum salmon (Onycor-hynchus keta) and sockeye salmon (Onycorhynchus  ne r k a ) . (2) h i s t o l o g i c a l changes i n the s k i n and u n d e r l y i n g t i s s u e s of sockeye salmon a l e v i n s . (3) c o l o u r changes produced i n f i s h s k i n by u l t r a -v i o l e t r a d i a t i o n s . 1. HISTORICAL REVIEW U l t r a v i o l e t l i g h t U l t r a v i o l e t l i g h t i s commonly divided, i n t o three r e g i o n s , the near, middle and f a r or extreme u l t r a v i o l e t . The extreme u l t r a v i o l e t below 2000 A f i n d s l i t t l e b i o l o g i c and t h e r a p e u t i c use s i n c e i t i s absorbed by most substances i n c l u d i n g a i r . The middle u l t r a v i o l e t from 2000 A to 3000 A i s important i n b i o l o g i c and t h e r a p e u t i c work. I t i s not t r a n s -m i t t e d by g l a s s but i s t r a n s m i t t e d by q u a r t z and s p e c i a l g l a s s e s f r e e of i r o n . The s o l a r spectrum b a r e l y extends i n t o t h i s r e g i o n , the lower l i m i t being about 2900 A. The near u l t r a v i o l e t from 3000 A to about 4000 A produces f l u o r e s c e n c e and photographic a c t i o n . Ordinary g l a s s i s q u i t e t r a n s p a r e n t between 3900 A and 3500 A, but the tr a n s p a r e n c y r a p i d l y d i m i n i s h e s u n t i l most g l a s s e s of o r d i n a r y t h i c k n e s s are f a i r l y opaque at 3100 A. The v i s i b l e spectrum begins at a wavelength of about 3900 A, depending on the s e n s i t i v i t y of the eye (Luckiesh, 1946). U l t r a v i o l e t i n s u n l i g h t The u l t r a v i o l e t components of s u n l i g h t i n c l u d e t h a t r e g i o n of the spectrum from about 4000 A to approx-i m a t e l y 2850 A, that i s , from the lower regions of the v i s i b l e spectrum i n the blu e t o the lowest wavelengths t r a n s m i t t e d by the earth's atmosphere. The lowest l i m i t t r a n s m i t t e d to the e a r t h v a r i e s w i t h the season, the time of day, the l a t i t u d e , the a l t i t u d e , atmospheric c o n d i t i o n s and other f a c t o r s , the average r a n g i n g from 2850 A to 2950 A. Laurens (1928) gi v e s t h e d i s t r i b u t i o n of the s o l a r energy a f t e r t r a v e r s i n g the earth's atmosphere as 1 p e r c e n t i n the u l t r a v i o l e t , 40 p e r c e n t i n the v i s i b l e and 59 p e r c e n t i n the i n f r a r e d . "Wavelengths s h o r t e r than 0.32 u (3200 A) c o n s t i t u t e l e s s than 0.2$ of t o t a l s u n l i g h t and a most v a r i a b l e p a r t o f i t " (Blum, 1945). Some b i o l o g i c a l e f f e c t s of u l t r a v i o l e t r a d i a t i o n The b i o l o g i c a l e f f e c t s of the v a r i o u s s p e c t r a l r e g i o n s have been found to d i f f e r markedly. G e r m i c i d a l lamps were found to be most e f f e c t i v e i f the energy emitted was c o n c e n t r a t e d a t 2500 A to 2600 A s i n c e these r a d i a t i o n s are absorbed by b a c t e r i a . The c o a g u l a t i o n of egg albumen and the p r o d u c t i o n of c o n j u n c t i v i t i s have a maximum near 2500 A. The a c t i v a t i o n of e r g o s t e r o l i n the s k i n to form V i t a m i n D was found to have a peak between 2500 A and 3100 A. T h i s corresponds f a i r l y c l o s e l y t o the most e f f e c t i v e l i n e s which produce e r y -thema or the reddening e f f e c t of untanned human s k i n ( P i g . 1 ) . Other l i n e s i n the spectrum are found to produce the same r e s u l t s , b ut w i t h l e s s e f f i c i e n c y , s i n c e the a b s o r p t i o n of the p a r t i c u l a r substance concerned f a l l s o f f . P i g . 1 - Graph showing the r e l a t i v e e f f e c t s of u l t r a v i o l e t r a d i a t i o n i n the near and middle u l t r a v i o l e t (redrawn from L u c k i e s h , 1946). Arrows i n d i c a t e wavelengths used i n these experiments. 4. The a c t i v a t i o n of e r g o s t e r o l and the p r o d u c t i o n of erythema f a l l w i t h i n the lower l i m i t s of the u l t r a -v i o l e t t r a n s m i t t e d by s u n l i g h t , w h i l e the most e f f e c t i v e g e r m i c i d a l area i s below that of s u n l i g h t . E a r l y work u s i n g u l t r a v i o l e t Burge, F i s c h e r and N e i l (1916) found that u l t r a v i o l e t has the power to i n a c t i v a t e enzymes and hormones. These substance© are now known to have attac h e d to t h e i r p r o s t h e t i c group a p r o t e i n which governs the s p e c i f i c i t y o f the r e a c t i o n . The i n a c t i -v a t i o n of the enzyme i s p r o b a b l y due to the d e n a t u r a t i o n of the p r o t e i n moity of the. enzyme. Burge (1917) observed t h a t u l t r a v i o l e t k i l l s b a c t e r i a and paramecia by c o a g u l a t i o n of the protoplasm and not by i n a c t i v a t i o n of the enzymes. The c o a g u l a t i o n produced was s i m i l a r to t h a t produced by heat. H a r r i s and Hoyt (1917) found that t y r o s i n e and p h e n y l a l a n i n e e x h i b i t e d a b s o r p t i o n bands i n the u l t r a -v i o l e t . These substances may act as o p t i c a l s e n s i t i z e r s which render l i v i n g c e l l s s u s c e p t i b l e to the t o x i c a c t i o n of u l t r a v i o l e t l i g h t . C l a r k (1922) suggested t h a t the e f f e c t of u l t r a v i o l e t was a p h o t o l y t i c p r o c e s s . The r a d i a t i o n s when absorbed may cause s e v e r a l e f f e c t s * (1) Longer r a d i a t i o n s with r e l a t i v e l y s m a l l amounts of energy, i . e . , i n f r a r e d , v i s i b l e and near 5. u l t r a v i o l e t , when absorbed, may cause i n c r e a s e d m o l e c u l a r motion, producing a r i s e i n temper-a t u r e . (2) When r a d i a t i o n of s u f f i c i e n t energy f a l l s on a f l u o r e s c e n t substance I t has been supposed t h a t e l e c t r o n s are separated from t h e i r p a r e n t mole-c u l e s , c r e a t i n g an u n s t a b l e c o n d i t i o n . Recom-b i n a t i o n of the e l e c t r o n occurs w i t h the emission of l i g h t . The lowest wavelengths of the v i s i b l e and near u l t r a v i o l e t produce such an e f f e c t . (3) With the s h o r t u l t r a v i o l e t r e g i o n , X-rays and r a d i o a c t i v e p a r t i c l e s a photochemical e f f e c t Is produced. The e l e c t r o n s are so a c t i v a t e d by the r a d i a t i o n s that they a r e separated from t h e i r parent•molecule and are f r e e to combine w i t h some other atom or group o f atoms. C l a r k (1922) b e l i e v e d t h a t energy s h o r t e r than 3000 A a f f e c t e d l i v i n g c e l l s by i o n i z i n g the p h o t o e l e c t r i c a l c o n s t i t u e n t s thereby l e a d i n g to a chemical r e a c t i o n . L i g h t l o n g e r than 3000 A a c t e d i n the same way i n the presence of a s e n s i t i z e r , which so a f f e c t e d the s u r f a c e c o n d i t i o n s o f these c o n s t i t u e n t s t h a t t h e i r p h o t o e l e c t r i c t h r e s h o l d s h i f t e d i n t o the v i s i b l e . She b e l i e v e d that melanin pigment formed i n t h e s k i n by s u n l i g h t acted as a s e n s i t i z e r , s h i f t i n g the p h o t o e l e c t r i c p o i n t Into the 6 . near u l t r a v i o l e t . E f f e c t on human beings In human beings i t has been demonstrated t h a t s m a l l doses o f u l t r a v i o l e t can produce erythema w i t h a r e s u l t i n g i n c r e a s e i n pig m e n t a t i o n i n s k i n and h a i r , while stronger doses can cause n e c r o s i s of the t i s s u e acted upon. Wounds have been shown to h e a l more r a p i d l y , s k i n forms of lupus have been a i d e d or c l e a r e d up, and the a n t i r a c h i t i c e f f e c t i s w e l l known (Laurens, 1933). I r r a d i a t i o n s of experimental animals and human beings w i t h a r t i f i c i a l sources have shown s l i g h t e f f e c t on r e d b l o o d c e l l count and hemoglobin content of blood when these l e v e l s are normal a t the be g i n n i n g o f the i n v e s t i g a t i o n s . Anemic c o n d i t i o n s have been a i d e d , the number of e r y t h r o c y t e s , lymphocytes and e s p e c i a l l y p l a t e l e t s have been i n c r e a s e d s u b s t a n t i a l l y . Growth has not been ai d e d , b a s a l metabolism and the t h y r o i d have not been s u b s t a n t i a l l y a f f e c t e d by u l t r a v i o l e t l i g h t (Mayerson, 1935). U l t r a v i o l e t l i g h t has been c o r r e l a t e d w i t h the p r o d u c t i o n of s k i n c a n c e r s , both carcinomas and sarcomas, i n human beings and experimental animals. Blum (1945) p o i n t e d out that cancer of the s k i n occurs p r i n c i p a l l y i n p a r t s of the body exposed t o s u n l i g h t and that i t i s more common i n outdoor workers than indoor workers, e s p e c i a l l y i n those areas r e c e i v i n g more s u n l i g h t . 7. Cancer of the s k i n has been produced i n mice and r a t s by exposure to u l t r a v i o l e t r a d i a t i o n . E a r l i e r workers i n the f i e l d o f u l t r a v i o l e t therapy, b e l i e v e d t h a t pigment f o r m a t i o n p r o t e c t e d an organism a g a i n s t e x c e s s i v e r a d i a t i o n . Laurens (1928) s t a t e d t h a t , "One of the important f u n c t i o n s of pigment i s the p r o t e c t i o n which i t a f f o r d s an organism a g a i n s t e x c e s s i v e r a d i a t i o n , a r i s i n g as i t does, as a p h y s i o -l o g i c a l response t o o v e r s t i m u l a t i o n . Pinsen demonstrated the p r o t e c t i v e a c t i o n of pigment by showing t h a t on s u c c e s s i v e exposures of the arm the pigmented areas were not i n f l a m e d , w h i l e those p r e v i o u s l y covered by g l a s s , and thus p r o t e c t e d , showed an i n t e n s e r e a c t i o n . " Since 1928 the views of the a d a p t i o n o f human s k i n t o u l t r a v i o l e t r a d i a t i o n have changed. Subsequently Laurens (1933, 1935) and Blum (1945) b e l i e v e d that cutaneous r e s i s t a n c e to the u l t r a v i o l e t i n v o l v e d the s k i n o u t s i d e the l a y e r of pigment and t h a t i n c r e a s e d c o r n i f i c a t I o n was the f a c t o r t h a t determined the degree of p e n e t r a t i o n of the r a d i a n t energy. The r e a c t i o n of human s k i n Is b e l i e v e d to be a s u r f a c e one s i n c e wave-lengths l e s s than 2800 A are absorbed by the horny l a y e r s , whereas at 2500 A and 3000 A, the p e n e t r a t i o n i s g r e a t e r , more r a d i a t i o n r e a c h i n g the stratum M a l p i g h i i and the corium. Wavelengths below 2500 A are absorbed before they reach the l i v i n g l a y e r s of the s k i n , while wavelengths 8. l o n g e r than 3000 A have been found to p e n e t r a t e to c o n s i d e r a b l e d i s t a n c e s (Laurens, 1933). E f f e c t on eggs U l t r a v i o l e t has been shown to a l t e r cleavage p a t t e r n s . H i n r i c k s (1925) found that eggs of Pundulus  h e t e r o l l t l c u s i r r a d i a t e d w i t h u l t r a v i o l e t produced a b n o r m a l i t i e s . I r r a d i a t i o n s d u r i n g the e a r l y cleavage stages caused a b n o r m a l i t i e s i n the head, eyes and p e r i -c a r d i a l r e g i o n s , w h i l e i r r a d i a t i o n s d u r i n g the time when the germinal r i n g was b e i n g l a i d down produced a b n o r m a l i t i e s i n the t a i l r e g i o n . These a b n o r m a l i t i e s may be shown to be c o n s i s t e n t w i t h the m e t a b o l i c g r a d i e n t s , the most a c t i v e area m e t a b o l i c a l l y b e i n g the most a f f e c t e d when an i n h i b i t i n g i n f l u e n c e Is a c t i v e . As she p o i n t e d out, the a b n o r m a l i t i e s were not s p e c i f i c f o r u l t r a v i o l e t , but may be produced by a number of f a c t o r s such as p r e s s u r e , temperature, i n c r e a s e d s a l i n i t y , a l t e r a t i o n of chemical environment, X-rays, e t c . E f f e c t of X - r a d i a t i o n on f i s h Bonham et a l (1948) found that X - r a d i a t i o n s decreased the r e d b l o o d c e l l counts 2 to 3 weeks a f t e r i r r a d i a t i o n and the hemopoietic c e l l count i n the kidney 1 to 2 weeks a f t e r i r r a d i a t i o n i n f i n g e r l i n g Chinook salmon, Onycorhynchus tschawytscha. Weight and l e n g t h were reduced s i g n i f i c a n t l y i n the h i g h e r i r r a d i a t i o n s which they used. 9. Welander et a l (1948) showed t h a t X-rays I n f l u e n c e d the growth r a t e , development of the eye and other organs of embryo and l a r v a l chinook salmon. The l a r g e r doses which they used a r r e s t e d the develop-ment of cutaneous pigment, the v a s c u l a r system, f i n s , eyes and growth. Damage to the v a s c u l a r system appeared a f t e r 33 days. Subcutaneous e x t r a v a s a t i o n was noted a l o n g the b e l l y w a l l below the l a t e r a l l i n e near the a t t a c h -ment of the y o l k sac. At the same time white f l o c c u l e n t spots appeared on the y o l k sac. A f t e r 5 weeks l i t t l e r e d blood seemed to be c i r c u l a t i n g i n the normal c h a n n e l s . H i s t o l o g i c a l s t u d i e s r e v e a l e d l i t t l e or no damage i n the nervous, d i g e s t i v e , muscular, integumentary or s k e l e t a l systems. Hemopoietic t i s s u e of the kidney was s e v e r e l y damaged. .Studies of the pronephros o f i r r a d i a t e d f i s h r e v e a l e d c e l l counts w e l l below t h a t of the c o n t r o l s up to 23 days a f t e r i r r a d i a t i o n , a f t e r which r e c o v e r y took p l a c e . Reduction of the percentage of m i t o t i c c e l l s i n the hemopoietic t i s s u e was i n p r o p o r t i o n to the amount of dose. G l o m e r u l i were i n h i b i t e d i n development i n the mesonephros, but i n t e r r e n a l bodies were not damaged as e x t e n s i v e l y as hemopoietic t i s s u e . Gonads and s p l e e n were damaged h e a v i l y , i n both cases the c e l l s showed p y c n o s i s and subsequent d e g e n e r a t i o n . 10. METHODS AND MATERIALS . Sources of u l t r a v i o l e t P r e l i m i n a r y experiments were performed w i t h a c o l d quartz-mercury M i n e r a l i g h t lamp ( U l t r a - v i o l e t Products Inc., Los Angeles, C a l i f o r n i a ) . T h i s lamp was operated w i t h the f i l t e r a t t a c h e d and emits p r e -dominantly at the mercury resonance l i n e , 2537 A. These p r e l i m i n a r y experiments were c a r r i e d out before the method of dosage- s t a n d a r d i z a t i o n had been developed. The b u l k of the experiments were c a r r i e d out w i t h 275 watt General E l e c t r i c type RS sunlamps, o p e r a t i n g on a 110 v o l t AC c i r c u i t . The g l a s s on the bulb i s of a s p e c i a l type t r a n s m i t t i n g u l t r a v i o l e t down to 2800 A. "The lamp i s a s e l f - c o n t a i n e d u n i t w i t h a mercury d i s c h a r g e element and a t u n g s t e n - f i l a m e n t r e s i s t a n c e b a l l a s t i n c o r -p o r a t e d w i t h i n an u l t r a v i o l e t r e f l e c t o r type b u l b . The lamp takes about 3 minutes t o r e a c h f u l l l i g h t i n t e n s i t y and approximately 5 minutes f o r r e s t a r t i n g " (Weitz, 1946). S t a n d a r d i z a t i o n of lamps One of the commonest of f a u l t s i n b i o l o g i c a l r e s e a r c h u s i n g u l t r a v i o l e t r a d i a t i o n s i s the l a c k of s t a n d a r d i z a t i o n of dosage. The t o t a l energy s h o u l d be a s c e r t a i n e d by some n o n - s e l e c t i v e method. T h i s should be measured i n r e l a t i v e or absolute u n i t s by a thermopile, by the d e f l e c t i o n of a galvanometer, or by 11. some photochemical method which r e q u i r e s a d e f i n i t e amount of energy to produce a v i s i b l e change I n a p r o p e r t y of the s o l u t i o n . The method of Pohle (1926) has been used i n t h i s study. I t i s based on the f a c t that a s o l u t i o n of h y d r i o d i c a c i d i n water f r e e s i o d i n e under the Inf l u e n c e of i n t e n s e l i g h t . This f r e e i o d i n e w i l l give a blue c o l o r a t i o n w i t h the s t a r c h t e s t . D e t a i l s of the method used a r e : (1) The lamp was allowed t o warm up 15 to 20 minutes, or u n t i l the output was co n s t a n t . (2) A l a r g e amount of 5.3$ H2SO4 was made up s u f f i c i e n t f o r a l l t e s t s . (3) N/lO sodium t h i o s u l f a t e was prepared i n s u f f i c i e n t q u a n t i t y f o r a l l t e s t s by add i n g 24.83 gms. of Na2Sg O3. 5E"20 to 1000 gms. of water. The N/400 s o l u t i o n used f o r each t e s t was made up by adding 1 c c . o f N/lO Na 2 S20 3 to 39 c c . of d i s t i l l e d water. (4) A 1% s o l u t i o n of s o l u b l e s t a r c h was prepared at the beginning of each experiment by d i s -s o l v i n g 1 gm. of s o l u b l e s t a r c h i n 5 c c . of c o l d d i s t i l l e d water and adding t h i s t o 95 cc of b o i l i n g d i s t i l l e d water. (5) The potassium i o d i d e was prepared f r e s h l y f o r each t e s t by d i s s o l v i n g 250 mg. of KI i n 25 c 12. of d i s t i l l e d water. (6) In a 100 ml. beaker of 5 cm. upper diameter 25 c c . of KI p l u s 25 c c . of 5,3% H 2 S 0 4 p l u s 5 drops of 1% s t a r c h s o l u t i o n and 1 c c . of N/400 Na 2S20 3 were p l a c e d under the lamp. Blue " c l o u d s " formed and were s t i r r e d w i t h a g l a s s r o d . The time r e q u i r e d f o r the whole s o l u t i o n to t u r n blue was measured by a stopwatch. This time p e r i o d was c a l l e d 1 dos;e and i s e q u i v a l e n t t o 1 Pi n s e n . 1 Pinsen i s 10 microwatts o f erythemal f l u x or 1 E - v i t o n , i n c i d e n t upon a p r o j e c t e d a r e a of 1 centimeter (Luckiesh, 1946). A l l experiments run were c a l i b r a t e d i n terms of t h i s c o l o u r change, the time f o r the s o l u t i o n to t u r n blue b e i n g c a l l e d "1 dose" or "1 x". Test s such as the above are v a l u a b l e when u s i n g a mercury arc s i n c e t h e y g i v e an i d e a of the i n t e n s i t y decrease with age of the lamp, and a l s o give a s t a n d a r d i z e d dose which can be reproduced. Experimental animals G o l d f i s h used were obtained from a supply house i n S t o u f f v i l l e , O ntario and were 6 to 8 cm. i n l e n g t h when i r r a d i a t e d . Chum salmon eggs were the progeny of s e v e r a l parents obtained from Sweltzer creek, near Cultus Lake h a t c h e r y on the a f t e r n o o n of November 28, 1948. The 13. sockeye salmon eggs and a l e v i n s were the progeny of s e v e r a l p arents obtained a t Weaver creek on October 16, 1948. The eggs were taken i n t h e ' u s u a l way and t r a n s f e r r e d immediately to the experimental h a t c h e r y troughs a t the u n i v e r s i t y . Coho salmon f i n g e r l i n g s were o b t a i n e d from Capilano r i v e r and Nelson creek d u r i n g the l a t t e r p a r t of J u l y and e a r l y p a r t o f August. Methods The chum salmon eggs were c a r r i e d i n the h a t c h e r y troughs u n t i l a s h o r t time before the i r r a d i a t i o n , when they were removed t o aluminum coated pans o f 22.5 sq. cm. The pans were f i t t e d w i t h an i n l e t and o u t l e t a l l o w i n g running water to a depth of 4 cm. The eggs were p l a c e d on the bottom o f each pan In a s i n g l e l a y e r and were c o n f i n e d to an area of 10 sq. cm. by means of g l a s s r o d s . The d i s t a n c e from the bottom of the lamps to the bottom of the pans was 26.5 cm. A heavy g l a s s p l a t e was p l a c e d over the c o n t r o l group between the eggs and the lamp, s e r v i n g to f i l t e r out a l l the u l t r a -v i o l e t rays below about 3100 A. I n t h i s manner the experimental eggs were exposed to a band of u l t r a v i o l e t r a d i a t i o n extending from about 3100 A to 2800 A. This band i s approximately equal t o the u l t r a v i o l e t components of the s o l a r spectrum. Water allowed to c i r c u l a t e through the pans 14. kept the temperature constant w i t h i n 1° to 3° C. A f t e r the experiments had been c a r r i e d out the eggs were t r a n s -f e r r e d to an aluminum coated tub. P e r i o d i c m o r t a l i t y counts were made. The eggs of the sockeye were maintained i n the experimental h a t c h e r y troughs. These eggs were i r r a d i a t e d under e x a c t l y the same c o n d i t i o n s as the chum salmon eggs d e s c r i b e d above. The a l e v i n sockeye, on the other hand, were t r a n s f e r r e d from the t r o u g h s e v e r a l days a f t e r h a t c h i n g had ceased i n t o aluminum coated mesh baskets, 13 x 20 cm., suspended i n the trough t o a depth of 4 to 5 cm. The f i s h were i r r a d i a t e d i n these baskets and remained there u n t i l completion of the experiments. The lamp was p l a c e d s e c u r e l y i n a stand which s t r a d d l e d the trough i n such a manner t h a t the d i s t a n c e from the bottom of the lamp to the bottom of the basket was 26 cm. The lamp was allowed to warm up to f u l l i n t e n s i t y b e f o r e the i r r a d i a t i o n s commenced. D a i l y samples of each group were taken and d a i l y m o r t a l i t i e s were counted. H i s t o l o g i c a l s t u d i e s H i s t o l o g i c a l s t u d i e s were made of the i r r a d i a t e d and c o n t r o l f i s h f i x e d i n Bouin's p i c r i c a c i d - f o r m o l -a c e t i c a c i d f i x a t i v e . The Bouin's was removed w i t h 3 t o 4 washings o f 10% a l c o h o l s a t u r a t e d w i t h l i t h i u m carbonate. . Dehydration and c l e a r i n g was done i n 3 changes 15. of dioxan and the f i s h were embedded i n wax i n the u s u a l manner. S a g g i t t a l s e r i a l s e c t i o n s were made at 5 u. U n f o r t u n a t e l y the microtome used was observed to v a r y markedly i n the t h i c k n e s s of the s e c t i o n s so t h a t accurate c e l l counts c o u l d not be made. Removal of the y o l k caused some damage to the head kidney r e g i o n , but was l e s s t han the damage caused by a l l o w i n g some of the y o l k to remain. The s e c t i o n s were s t a i n e d i n H a r r i s 1 haemotoxylin and e o s i n made up i n the p r e s c r i b e d manner. Comparison of lamps and s u n l i g h t Since the band of u l t r a v i o l e t used i n these experiments centered about t h a t which Is erythemally e f f e c t i v e and s i n c e the dosage was c a l i b r a t e d In Pinsens, i t i s p o s s i b l e to make a rough comparison of the energies used i n these experiments w i t h those of the u l t r a v i o l e t i n s u n l i g h t . The H l t r a v i o l e t of s u n l i g h t i s c a l i b r a t e d I n e r y t h e m a l l y e f f e c t i v e u n i t s such as the number of E - v i t o n s (Pinsens) Incident upon a p r o j e c t e d area f o r a c e r t a i n p e r i o d of time. An exposure of 2/3 of an E - v i t o n hour p e r sq. cm. on the average untanned human s k i n r e s u l t s i n a minimum p e r c e p t i b l e erythema. L u c k i e s h (1946) has made a number of measure-ments of the u l t r a v i o l e t components of s u n l i g h t i n a suburb of C l e v e l a n d , Ohio. Although the values which 16. he gives (Table I) are s l i g h t l y h i g h e r than those which c o u l d be expected i n Vancouver, they are convenient f o r comparison with the u l t r a v i o l e t used i n these e x p e r i -ments . Table I - V a r i a t i o n of average hours of a c t u a l sunshine per month and of erythemal u l t r a v i o l e t energy i n d a y l i g h t as measured on a h o r i z o n t a l plane f o r a s i x year p e r i o d i n a suburb of C l e v e l a n d , Ohio ( a f t e r L u c k i e s h , 1946, p. 57). Month Average hours Erythemal U l t r a v i o l e t Energy of sunshine E - v i t o n hours Percent of p e r sq. cm. t o t a l f o r year January 71 51 1.8 February 98 72 2.5 March 167 182 6.3 A p r i l 221 280 9.8 May 298 430 15.0 June 305 476 16.7 J u l y 350 476 16.7 August 292 386 13.5 September 243 277 9.8 Octobe r 178 143 5.0 November 103 55 1.9 December 66 30 1.0 Seasonal v a r i a t i o n s were measured by L u c k i e s h (1946.) at C l e v e l a n d . I n A p r i l the time r e q u i r e d to produce a minimum p e r c e p t i b l e erythema a t noon was 50 minutes, while at noon i n June o n l y 17 minutes were necessary. The erythemal v a l u e s of the r a d i a t i o n s used f o r the sockeye salmon a l e v i n s ware c a l c u l a t e d . The a r e a 17. of the baskets used was 260 sq. cm. and the dosage was c a l i b r a t e d at 1 Pinsen i n 30 minutes. The number of Fi n s e n hours x the s u r f a c e a r e a i n cm. gives the dosage In E - v i t o n hours per sq. cm. Table I I - The t o t a l number of E - v i t o n hours i n c i d e n t upon a l e v i n sockeye salmon at the s u r f a c e of the water. C a l i b r a t i o n of dose T o t a l cLosage i n E - v i t o n hours on an area of 260 sq. cm. 0.5x 65 l.Ox 130 2.Ox 260 .4.Ox 520 8.Ox 1040 The a c t u a l energy r e a c h i n g each f i s h was a s m a l l f r a c t i o n o f the I n c i d e n t energy on the s u r f a c e of the water, s i n c e the s u r f a c e area of each f i s h which r e c e i v e d the r a d i a t i o n was very much s m a l l e r than the t o t a l s u r f a c e a r e a . Measurements of the a b s o r p t i o n of u l t r a v i o l e t l i g h t i n water i n d i c a t e d t h a t f o r a depth of 5 cm. 94.5$ of the i n c i d e n t i n t e n s i t y i s t r a n s m i t t e d (Landolt-B o r n s t e i n , 1923). D i s s o l v e d organic m a t e r i a l s and m i n e r a l s i n c r e a s e the a b s o r p t i o n of the u l t r a v i o l e t t o some ex t e n t . Measure-ments of Vancouver tap water, made by Dr. Crooker of the Department of P h y s i c s , i n d i c a t e d t h a t the v a l u e s f o r the 18. a b s o r p t i o n of u l t r a v i o l e t energy i n tap water d i d not d i f f e r s i g n i f i c a n t l y from those given i n L a n d o l t -B o r n s t e i n (1923). The c a l c u l a t e d values of energy i n c i d e n t upon the f i s h (Table I I ) u s i n g the RS sunlamp were presumably of the order which they could r e c e i v e i f exposed to summer s u n l i g h t . Thus the e f f e c t s observed i n these experiments are ones which c o u l d be observed i n some hat c h e r y c o n d i t i o n s . 1 9 . RESULTS P r e l i m i n a r y experiments P r e l i m i n a r y experiments u s i n g g o l d f i s h i r r a d i a t e d w i t h the c o l d quartz-mercury M i n e r a l i g h t lamp were only q u a l i t a t i v e and i n d i c a t e d t h a t an e f f e c t c o u l d be p r o -duced by u l t r a v i o l e t r a d i a t i o n . R a d i a t i o n s l a s t i n g from | t o 2 hours caused a b l i s t e r i n g of the d o r s a l r e g i o n s of the head and the t i p s of the f i n s . S u s c e p t i b i l i t y to' fungus i n c r e a s e d g r e a t l y over t h a t o f the c o n t r o l f i s h kept i n the same aquarium. In s e v e r a l cases I n f e c t i o n of the e x p e r i -ment a l s was heavy enough to cause death. Some of the i r r a d i a t e d f i s h showed i n c r e a s e d b l a c k e n i n g due to the f o r m a t i o n of melanophores i n the s k i n a f t e r a l a t e n t p e r i o d of s e v e r a l weeks. The b l a c k e n i n g began i n the b l i s t e r e d areas on the head and t i p s of the f i n s and spread g r a d u a l l y v e n t r a l l y . At the end of four weeks one f i s h had changed from the g o l d c o l o r to an almost complete b l a c k on the d o r s a l side of the l a t e r a l l i n e . S e v e r a l experiments w i t h a s t r o n g q u a r t z -mercury arc b e l o n g i n g to the Department of P h y s i c s produced s i m i l a r but more d r a s t i c e f f e c t s on the g o l d -f i s h . B l i s t e r i n g and s l u f f i n g of the s k i n wer?e;.pr--o.duce:d s h o r t l y a f t e r the i r r a d i a t i o n . Fungus appeared on the b l i s t e r e d areas and. death o c c u r r e d i n most c a s e s . F i s h which recovered from the fungus showed b l a c k e n i n g around 20. the b l i s t e r e d areas and small b l o o d c l o t s i n the f i n s . One f i s h s u r v i v e d . T h i s f i s h developed dense pigment on the d o r s a l head r e g i o n s , on the operculum, behind the eyes and on the t i p s of the f i n s . The pigment l a t e r spread to the v e n t r a l regions below the l a t e r a l l i n e s . One of the most s t r i k i n g r e s u l t s of these i r r a d i a t i o n s was the decrease i n a c t i v i t y shown. The experimental f i s h sank to the bottom of the aquarium where they remained with f i n s " f o l d e d " . In no case were these i r r a d i a t e d f i s h observed to f e e d u n t i l four days a f t e r the experiment took p l a c e . C o n t r o l f i s h s u b j e c t e d to s i m i l a r h a n d l i n g showed a c t i v e swimming motions and f e d immediately a f t e r the end of the e x p e r i -ment. Three experiments were c a r r i e d out u s i n g f i n g e r l i n g coho salmon ave r a g i n g 5 cm. i n l e n g t h . The f i s h were p l a c e d i n a b a t t e r y j a r (28 x 18 cm. i n c r o s s s e c t i o n a l area and 36 cm. i n h e i g h t ) . The b a t t e r y j a r , w i t h water to a depth of 15 cm., was p l a c e d i n a s i n k with running water c i r c u l a t i n g around the o u t s i d e , keeping the temperature c o n s t a n t . Two a i r j e t s were kept i n the j a r d u r i n g the time the f i s h remained In i t . C o n t r o l s were r u n i n a s i m i l a r manner, but w i t h a g l a s s p l a t e p l a c e d between the lamp and the b a t t e r y j a r . In each case the RS sunlamp was 40 cm. above the 21. bottom o f the b a t t e r y j a r . In one experiment i r r a d i a t i o n for 18 hours k i l l e d 29 out of 30 f i s h , i n another experiment i r r a d i a t i o n of 12 hours k i l l e d a l l of the f i s h . C o n t r o l s showed no m o r t a l i t i e s . P i s h i r r a d i a t e d i n the s i n k w i t h running water showed so"/, fatriMty a f t e r 1 week o f i r r a d i a t i o n , while the c o n t r o l s showed 2 dead out of 30. Power sources were v a r i a b l e d u r i n g t h i s p e r i o d , but r e s u l t s do show t h a t u l t r a v i o l e t r a d i a t i o n has some e f f e c t . Chum salmon eggs E i g h t experiments were c a r r i e d out u s i n g the eggs o f the chum salmon, beg i n n i n g 6 hours a f t e r f e r t i -l i z a t i o n and at the i n t e r v a l s i n d i c a t e d i n Table I I I u n t i l the supply of eggs r a n out. Percentage m o r t a l i t i e s of c o n t r o l and i r r a d i a t e d eggs at i n t e r v a l s a f t e r i r r a d i -a t i o n are i n d i c a t e d i n ^able I I I . Prom the t a b l e i t appeared that the smal l amount of u l t r a v i o l e t t r a n s m i t t e d by the g l a s s p l a t e over the c o n t r o l s was s u f f i c i e n t t o produce a l e t h a l e f f e c t at the h i g h e r doses (37x, 18.5x, and 8.14x). At l e s s e r doses the e f f e c t o f the u l t r a v i o l e t t r a n s m i t t e d by the g l a s s p l a t e was not s u f f i c i e n t t o a f f e c t the c o n t r o l s as s t r o n g l y as the I r r a d i a t e d f i s h . Date Dosage Nov. 28 37x Dec. 1 18.5x Dec. 4 8.14x Dec. 6 5.14x Dec. 6 3.4x Dec. 7 1.7x Dec. 9 1.29x Dec. 17 Table III Percentage mortalities of chum salmon eggs irradiated 6 hours after fertilization and at various periods thereafter. Total Sample Time in Days 1 2 cont. 160 86.9 irrad. 188 100 cont. 152 90.1 irrad. 159 100 dont. 218 9.6 irrad. 217 100 Cont. 125 6.9 12.5 irrad. 125 100 cont. 100 10 15 irrad. 100 100 cont. 125 14.4 18.4 irrad. 125 32 58.4 cont. 129 .77 2.27 irrad. 95 12.6 63.1 cont. 80 18.7 irrad. 80 98.7 16.5 20 19.5 68 Death expressed in percentage of total sample 4 5 6 8 9 10 11 12 14 17.1 24.3 21 25.1 73.6 35.5 80.8 7.7 80 25.5 31.05 22 45.1 86.4 13.1 87.3 Time i n Days 15 20 22 23 24 25 42.25 36 53.7 100 56.3 56.3 97.6 23.2 25.5 97.9 99 58.5 98.7 0, - c o n t r o l eggs I - i r r a d i a t e d eggs P i g . 2 Graphs o f the percentage m o r t a l i t i e s of I r r a d i a t e d and c o n t r o l chum salmon eggs over a p e r i o d of days a f t e r dosages o f 3.14x, 2x, 1.7x, and 1.29x. 24 P i g . 2 shows g r a p h i c a l l y the percentage m o r t a l i t y of the lower i r r a d i a t i o n s f o r each experiment. None of the i r r a d i a t e d eggs s u r v i v e d to h a t c h i n g . Since the eggs were s e n s i t i v e to h a n d l i n g at t h i s p e r i o d of development and s i n c e i t was not known whether the v i s i b l e l i g h t and near u l t r a v i o l e t t r a n s -m i t t e d by the g l a s s p l a t e over the c o n t r o l s a f f e c t e d the ©ggs* s e v e r a l h a n d l i n g experiments were c a r r i e d out. The eggs were t r e a t e d s i m i l a r l y to I r r a d i a t e d and c o n t r o l eggs, except that a l l l i g h t was excluded. M o r t a l i t i e s ranged from 8 t o 14 p e r c e n t over a p e r i o d of twenty days. Thus i t was concluded t h a t the v i s i b l e and near u l t r a -v i o l e t r a d i a t i o n s t r a n s m i t t e d to the c o n t r o l s had some e f f e c t on the eggs d u r i n g the e a r l y stages of development. Sockeye salmon eggs Six- s e t s of i r r a d i a t i o n s were c a r r i e d out u s i n g sockeye salmon eggs. These experiments began two days before the eggs became eyed and t h e r e a f t e r a t I n t e r v a l s as shown i n Table IV. Table IV shows the dosage, t o t a l sample and percentage m o r t a l i t y of I r r a d i a t e d and c o n t r o l eggs. Prom the 2x and 3x i r r a d i a t i o n s i t appeared that the c o n t r o l s were not a f f e c t e d w h i l e the i r r a d i a t e d eggs s u f f e r e d harmful e f f e c t s ( P i g . 3 ) . U n f o r t u n a t e l y these experiments were a c c i d e n t a l l y i n t e r r u p t e d before completion. Larger dosages (35.5x, 16.2x, l l x , and 25. 3.43x) showed more s t r i k i n g d i f f e r e n c e s , the I r r a d i a t e d f i s h being more s t r o n g l y a f f e c t e d . An i n t e r e s t i n g f e a t u r e of the i r r a d i a t i o n of December 20, 1948 was the f a c t thait as w e l l as p r o d u c i n g a l e t h a l e f f e c t on the eggs, h a t c h i n g was a c c e l e r a t e d . The u n t r e a t e d eggs i n the e x p e r i m e n t a l h a t c h e r y troughs commenced to h a t c h January 18, 1949. The i r r a d i a t e d f i s h began to h a t c h on December 21, 1948, one day a f t e r the i r r a d i a t i o n . At the end of the f i r s t week h a l f of the eggs had begun to hatch, but with g r e a t m o r t a l i t i e s . Of the s u r v i v o r s of the i r r a d i a t e d f i s h a number showed d i s t o r t i o n s of body form ( P i g . 4 ) . These malformed f i s h were allowed to develop f o r some time. F i g u r e s 4 and 5 are photographs of i r r a d i a t e d and c o n t r o l sockeye salmon a l e v i n s taken 46 days a f t e r i r r a d i a t i o n o f the eggs. The c o n t r o l f i s h ( F i g . 5) showed more advanced development i n pig m e n t a t i o n and s i l v e r i n g , a b s o r p t i o n of yo l k , development of f i n s , and growth. The i r r a d i a t e d f i s h ( F i g . 4) showed d i s t o r t e d body contours w i t h the head depressed over the y o l k sac at the l e v e l of the h e a r t . The yolk sac was absorbed l e s s r a p i d l y i n the I r r a d i a t e d animals and the f o r m a t i o n of pigment was delayed 2 weeks. U n t i l the time of death 75 days a f t e r i r r a d i a t i o n , p i g m e n t a t i o n on the d o r s a l r e g i ons of the head was l a c k i n g . The i r r a d i a t e d f i s h were a c t i v e , but c o u l d swim only i n c i r c l e s s i n c e t h e i r bodies were d i s t o r t e d . Table IV Experiments carried out using sockeye salmon eggs two days before eyeing and at intervals thereafter. Percentage mortalities for a period of days after irradiation Total Date Dosage Sample 1 2 3 4 5 6 7 8 9 10 11 12 14 15 18 20 23 26 27 32 Nov. 12 2x Nov. 13 3x Nov. 29 35.5x Dec. 2 16.2x Dec. 20 l l x Deo. 30 3.43x Control 100 0 0 0 0 0 0 0 0 irrad 100 > 0 0 0 2 10 11 12 Experiments interrupted control 40 ' 0 0 0 0 0 0 0 irrad 40 0 0 2.5 17.5 30 35 45 control 220 0. .45 2.6 2 6.8 7.7 8.6 11.3 irrad 220 22.7 73.2 91.8 100 control 220 0 0 0 .45 .90 1.8 3.15 irrad 223 59.6 86.5 86.9 89.7 9 6 . 4 98.6 100 control 200 3 4 4.5 6 irrad 200 48.5 59.5 75 87.5 89 control 200 2 8.5 31.0 34.0' irrad 200 2 29 30 44.5 34.5 97.5n 98.0 98.5 100 P i g . 3 - Graphs of the percentage m o r t a l i t i e s of i r r a d i a t e d and c o n t r o l sockeye salmon eggs over a p e r i o d of days a f t e r dosages o f 2x, 3x, 3.43x, l l x , 16.2x, and 35.5x. F i g . 4 - Sockeye salmon a l e v i n s 46 days a f t e r an i r r a d i a t i o n of 35.5x. Mag. 5.5x. 30. A s i m i l a r experiment was c a r r i e d out on December 30, 1948, u s i n g a dosage of 3.43x. Ha t c h i n g was a g a i n s t i m u l a t e d . Of the experimental f i s h , two showed curved v e r t e b r a l columns and c i r c u l a r swimming movements. P i g -mentation was d e l a y e d and a b s o r p t i o n of the y o l k sac was slower than the c o n t r o l s . Increased m o r t a l i t i e s i n the c o n t r o l s ( P i g . 3) may be accounted f o r by an a t t a c k of fungus on the eggs. The g r e a t e s t m o r t a l i t y o c c u r r e d i n these e x p e r i -ments d u r i n g the h a t c h i n g p r o c e s s . The f i s h emerged head f i r s t , body f i r s t , or y o l k sac f i r s t , as opposed t o the normally h a t c h i n g f i s h which emerged t a i l f i r s t . C o n t r o l f i s h o n l y o c c a s i o n a l l y showed s i m i l a r a b n o r m a l i t i e s i n h a t c h i n g . Sockeye salmon a l e v i n s Experiments were c a r r i e d out u s i n g sockeye salmon a l e v i n s averaging 20 cm. i n l e n g t h . These f i s h began h a t c h i n g on January 18, w i t h the peak on January 27. F i v e groups of a l e v i n s and two c o n t r o l groups were used. The 2x, 4x, and 8x I r r a d i a t i o n s were c a r r i e d out on February 4. The 0.5x aad l x i r r a d i a t i o n s were c a r r i e d out on February 11. Since the water temper-atu r e never rose above 3°C d u r i n g t h a t time, development was not v e r y ' . d i f f e r e n t and the two s e t s of r e s u l t s were co n s i d e r e d t o g e t h e r . C o n t r o l s were c a r r i e d out f o r each s e t . Table V gives the l i v e removals, R, d a i l y Table V Live removals, R, mortalities, M, and cumulative percentage mortalities, C, shown daily for a number of sockeye salmon alevins. Control (l) 5x lx 2x 4x 8x Control (2) Total Sample 366 341 303 Time R M c R M C R M c Q 10 0 5 0 0 5 0 0 1 2 1 0.28 2 0 0 2 0 0 2 2 0 2 2 0.61 2 0 0 3 2 0 2 5 2.13 2 12 4.11 4 2 0 0.29 2 28 10.73 2 13 8.62 5 2 0 2 3 11.73 2 35 20.83 6 2 0 2:: 7 13.97 2 47 37.41 7 2 0 2 13 18.13 2 35 50.0 8 2 1 0.6 2 43 31.76 2 36 63.12 9 2 0 2 35 43.67 2 35 76.07 10 2 1 0.89 2 22 50.32 2 19 83.45 11 2 0 2 23 58.07 2 16 89.85 12 2 0 0.91 2 23- 65.81 2 0 90.57 13 2 0 2 27 75.0 2 6 93.38 14 2 1 1.23 2 17 81.04 2 2 94.81 15 2 0 2 5 83.22 2 3 96.64 16 2 0 2 4 85.09 2 1 97.74 17 2 0 2 2 86.33 2 0 98.48 18 2 0 1.25 2 0 86.79 2 0 99.23 19 2 0 2 1 87.83 2 0 100.0 20 2 0 2 2 89.12 43 260 a 2 0 1.29 2 0 89.72 22 0 0 0 0 A l l Dead 23 2 0 2 4 91.72 24 0 -1 1.61 0 2 92.41 25 2 1 1.95 2 0 93.01 -26 0 0 0 1 93.40 27 2 0 1.96 2 0 94.06 28 0 0 0 0 29 2 0 2 0 30 0 0 1.97 0 0 60 6 55 269 271 245 220 291 . ;R M C R M C R M C R M C 5 0 0 5 0 0 5 0 0 10 0 0 2 4 1.52 2 7 2.94 2 80 37.56 2 2 0.71 2 23 10.34 2 30 15.67 2 69 70.61 2 1 1.08 2 61 33.97 3 106 61.37 2 52 96.17 2 2 1.82 2 62 58.36 2 42 80.08 2 5 99.51 2 0 1.83 2 70 86.27 2 42 99.12 0 1 100.0 2 0 1.84 2 18 94.07 2 0 100.0 13 207 2 0 1.86 2 12 99.60 18 227 2 5 3.74 0 2 100.0 A l l Dead 2 4 5.24 19 252 A l l Dead 24 14 A l l Dead 253 Alive 300 alive 17 alive 32. m o r t a l i t i e s , M, and cumulative percentage death, C, f o r each set o f i r r a d i a t i o n s . C o n t r o l (1) was that run w i t h the 2x, 4x, and 8x i r r a d i a t i o n s , while C o n t r o l (2) was f o r the 0.5x and l x I r r a d i a t i o n s . The experiments were terminated at the end of 30 days when a decrease i n the flow of water through the c a r r y i n g trough caused heavy m o r t a l i t i e s of a l l f i s h i n the trough. Table VI - The c a l c u l a t e d t h e o r e t i c a l number of deaths f o r each sample of c o n t r o l and i r r a d i a t e d sockeye a l e v i n s , as compared to the t o t a l sample used i n each experiment. Experiment T h e o r e t i c a l no. T o t a l sample of deaths used C o n t r o l (1) 7.17 366 0.5x dose 320.7 341 l.Ox dose 303 303 C o n t r o l (2) 18.53 291 2x dose 271 271 4x dose 245 245 8x dose 220 220 The removals were compensated f o r s i n c e there would have been a s l i g h t l y g r e a t e r number of deaths a t t r i b u t a b l e to treatment had not the removals been made (Table V I ) . The values obtained f o r the e x p e r i -mentals were s i g n i f i c a n t l y d i f f e r e n t from t h o s e of the c o n t r o l s . In a l l the i r r a d i a t i o n s c a r r i e d out w i t h the a l e v i n s , the a c t i v i t y o f the f i s h was d e f i n i t e l y a f f e c t e d , 33. D u r i n g the i r r a d i a t i o n s the a l e v i n s a c t i v e l y crowded i n t o the f a r t h e s t c o r n e r s of the h o l d i n g b a s k e t s . I n the l o n g e r exposures some of the f i s h became v i s i b l y weakened and sank to the bottom of the b a s k e t . Some made o c c a s i o n a l spasmodic movements, r i s i n g s w i f t l y from the bottom to the s u r f a c e only to f a l l back and remain q u i e s c e n t f o r l o n g p e r i o d s . These l e s s a c t i v e f i s h were observed to be the f i r s t to d i e . Subsequent o b s e r v a t i o n s of a c t i v i t y showed a g r a d a t i o n from o c c a s i o n a l movements among the f o u r hour group to great a c t i v i t y among the c o n t r o l s . When a b r i g h t l i g h t was shone on each group a s i m i l a r r e a c t i o n was noted. This l o s s o f a c t i v i t y remained i n the h i g h e r exposures u n t i l complete m o r t a l i t y o c c u r r e d / ' In the two lower i r r a d i a t i o n s , the f i s h remained l e s s a c t i v e than the c o n t r o l s f o r a p e r i o d of a p p r o x i -mately 15 days, when i t was observed that the most a f f e c t e d ' f i s h had d i e d and t h a t some r e c o v e r y had taken p l a c e among the remainder. At the c o n c l u s i o n of the experiment, the seventeen remaining f i s h of the 0.5x exposure showed the scene degree of a c t i v i t y as the c o n t r o l s . Most of the f i s h i n the h e a v i l y i r r a d i a t e d groups were unable to swim. The heartbeat became slow and spasmodic and many seemed unable to move the lower jaw i n r e s p i r a t i o n . White spots appeared on the y o l k F i g . 6a - Graph of dai ly percentage oumclative mortal!ty of sockeye salmon alevins irradiated with dosages of o.5x, lx» 2x, 4x, and 8x« 34. sac, the v i t e l l i n e v e i n s appeared prominent and d i s t e n d e d , and i n many cases an accumulation of serous f l u i d appeared below the l a t e r a l l i n e near the attachment of the y o l k sac. M o r t a l i t i e s o c c u r r e d too r a p i d l y and the water temperature was too low to o b t a i n s i g n i f i c a n t d i f f e r e n c e s i n the growth r a t e . Development of the s i l v e r y pigment was r e t a r d e d about a week i n those f i s h which s u r v i v e d the lower exposures A^ig 6a) • H i s t o l o g i c a l s t u d i e s (A) S k i n - H i s t o l o g i c a l s t u d i e s of experimental and c o n t r o l f i s h showed l i t t l e e f f e c t on the f i r s t two days a f t e r i r r a d i a t i o n . On the t h i r d day changes appeared i n the epidermis and dermis of the s k i n i n the areas d i r e c t l y exposed to the r a d i a n t energy. The n u c l e i of the s u p e r f i c i a l c e l l s rounded up i n t o one or two e o s i n o p h i l i c or b a s o p h i l i c granules and the f i b r o -e l a s t i c l a y e r of the dermis appeared s w o l l e n and d i s -tended. The goblet c e l l s were njore rounded and the pores to the e x t e r i o r were opened t o a wider extent ( P i g . 6) than those of the c o n t r o l s ( P i g . 7 ) . On the f o u r t h day a f t e r i r r a d i a t i o n the granules appeared more numerous and c o u l d be observed i n the b a s a l l a y e r s of the epidermis ( P i g . 8 ) . The goblet c e l l s were reduced i n number or completely absent i n some a r e a s . The s k i n of the more h e a v i l y i r r a d i a t e d f i s h (4x, 8x) was 35. ragged on i t s outer s u r f a c e as though i t were desquam-a t i n g ( P i g . 9 ) . Lower dosages (0.5x, Ix, 2x) showed g r a n u l a t i o n and r e d u c t i o n or l o s s of g o b l e t c e l l s , but di d not show any s i g n s of desquamation ( P i g . 10). Severe d e g e n e r a t i o n of the s k i n with the appearance of numerous b a s o p h i l i c and e o s i n b p h i l i c granules o c c u r r e d i n the h i g h e r .dosages (4x, 8x, 2x) a f t e r 5 to 8 days ( F i g . 11). The f i b r o e l a s t i c l a y e r of the dermis became s w o l l e n on the 3rd to 4th day. Only small fragments of i t could be found on those f i s h which showed severe g r a n u l a t i o n of the e p i d e r m i s . Death o c c u r r e d before s i g n s of r e c o v e r y i n the 2x, 4x and 8x dosages. Lower dosages (0.5x, l x ) showed l e s s severe g r a n u l a t i o n i n the epidermis and l e s s d i s t e n s i o n of the f i b r o e l a s t i c membrane. The most severe e f f e c t s were observed on the 8 t h day. These l a s t e d u n t i l the 13th to 15th day when r e c o v e r y s e t i n ( F i g . 12). The f i r s t appearance o f recover y was the h e i g h t e n i n g of the b a s a l c e l l s of the e p i d e r m i s . Subsequent recovery i n v o l v e d p r o l i f e r a t i o n of epidermal c e l l s and the appearance o f new gob l e t c e l l s . S e c t i o n s of s k i n from the s i d e s of the body away from the r a d i a t i o n s appeared s i m i l a r to t h a t of the con-t r o l s ( F i g . 7), w i t h g o b l e t c e l l s p r e s e n t and no gran-u l e s such as those found i n the i r r a d i a t e d a r e a s . Pig. 6 - Skin of f i s h 3 days a f t e r an i r r a d i a t i o n of 4x. Mag. 440x. Pig. 7 - Skin of control 7 day r the experiment . began. Mag. 440x. ok** F i g . 8 - Skin of f i s h 4 days a f t e r an i r r a d i a t i o n of 2x. Ma^. 440x. P i g . 11 - S k i n of f i s h 7 days a f t e r an i r r a d i a t i o n o f 2x. Mag. 440x. P i g . 12 - S k i n of f i s h 13 days a f t e r a dosage of 0.5x. Mag. 440x. 40. (B) Other t i s s u e s - T i s s u e s l y i n g deeper i n the f i s h d i d not show h i s t o l o g i c a l changes a f t e r i r r a d i a t i o n . The t h y r o i d and p i t u i t a r y glands were s i m i l a r to the c o n t r o l s b o t h i n development and h i s t o l o g i c a l s t r u c t u r e . The kidney appeared unchanged i n both the p r o n e p h r i c and mesonephric r e g i o n s . Counts of hemopoietic c e l l s were not p o s s i b l e s i n c e the t h i c k n e s s of the s l i d e s was not c o n s t a n t . However, q u a l i t a t i v e o b s e r v a t i o n s r e v e a l e d no changes i n the n u c l e i of the hemopoietic c e l l s or i n the number of r e d blo o d c e l l s i n the k i d n e y . The thymus gland, g i l l s , nervous system and d i g e s t i v e t r a c t were not a f f e c t e d . 41. DISCUSSION U l t r a v i o l e t and r a t e of h a t c h i n g H i n r i c k s (1925) found that d u r i n g p e r i o d s of low s u s c e p t i b i l i t y w i t h l e s s i n t e n s e r a d i a t i o n s there was an apparent t r a n s i t o r y s t i m u l a t i o n with regard to h a t c h i n g . More eggs i n the exposed l o t s hatched i n the e a r l y p a r t of the h a t c h i n g p e r i o d than i n the con-t r o l s . There was a s i g n i f i c a n t c o r r e l a t i o n between the percentage of hatched and normal eggs i n each l o t , s i n c e o n l y the more normal embryos were capable of h a t c h i n g . The p e r i o d of low s u s c e p t i b i l i t y was found to be that time when the embryonic a x i s was w e l l formed. These obser v a t i o n s agree w i t h t h o s e produced i n the experiments r e p o r t e d h e r e , where l a r g e doses at l a t e r stages of embryonic development produced a h a s t e n i n g of h a t c h i n g . The mechanism of the e f f e c t i s not c l e a r . I n some way the r a d i a t i o n must have weakened the c h o r i o n of the egg to such an extent that the embryo moving w i t h i n the egg d i s r u p t e d t h i s membrane. Embryonic development was not a p p a r e n t l y speeded up s i n c e the embryos which hatched were not as f u l l y d e v e l -oped as c o n t r o l s which hatched n o r m a l l y . The m o r t a l i t i e s which occurred i n the h a t c h i n g process were h i g h e r than i n the c o n t r o l s . M o r t a l i t i e s i n eggs and a l e v i n s The experiments c a r r i e d out show t h a t u l t r a v i o l e t 42. has a l e t h a l e f f e c t on b o t h salmon eggs and hatched f i s h . The e f f e c t i s p r i m a r i l y a s u r f a c e r e a c t i o n s i n c e u l t r a v i o l e t r a d i a t i o n s are absorbed by t h i n l a y e r s of s k i n and protoplasm (Laurens, 1933; Giese, 1939). The l a y e r of c a p i l l a r i e s i n the corium i s b e l i e v e d to be of g r e a t importance s i n c e the b l o o d f l o w i n g through t h i s r e g i o n absorbs the u l t r a v i o l e t and p a r t of the v i s i b l e r a d i a t i o n s . The absorbed energy can then be given to the organism as a whole (Laurens, 1935). P a r t of the energy absorbed by the s k i n and b l o o d i s transformed to heat by the a c t i v a t i o n of the molecules ( C l a r k , 1922). v Thus one of the e f f e c t s of the u l t r a v i o l e t , e s p e c i a l l y i n the r e g i o n s used i n these experiments, was the h e a t i n g of the t i s s u e s . Since the b lood .in the v e s s e l s l y i n g over tiae y o l k sac was not p r o t e c t e d by the s k i n , i t was probable t h a t the b l o o d was heated to an a p p r e c i a b l e e x t e n t . T h i s hyper-emia p r o b a b l y produced changes i n the b l o o d and other t i s s u e s which were h a r m f u l to the embryos and hatched f i s h . However c e l l i n j u r y occurs from the r a d i a t i o n s . Blum (1945) b e l i e v e s that the c e l l s , as a r e s u l t of the i n j u r y , e l a b o r a t e v a r i o u s substances which b r i n g about p h y s i o l o g i c a l responses. The l a t e n t p e r i o d between exposure and p h y s i o l o g i c a l responses may be accounted 43. f o r by assuming t h a t these p h y s i o l o g i c a l l y a c t i v e sub-stances are elaborated, by the c e l l a t a slow r a t e . Increased c o n c e n t r a t i o n of a d i l a t o r substance has a c t u a l l y been demonstrated i n the s k i n and b l o o d of animals that have been exposed t o u l t r a v i o l e t r a d i a t i o n . Lewis and Zotterman (1926) suggested t h a t i t was a h i s t a m i n e - l i k e "H" substance. More r e c e n t evidence has shown t h a t the substance r e l e a s e d i s not histamine and the term ttHM-substance has l o s t much of i t s u s e f u l n e s s . Menkin (1938) p o s t u l a t e d t h a t the substance which caused i n c r e a s e d c a p i l l a r y p e r m e a b i l i t y was not histamine but was " l e u c o t a x i n e " which i s e l a b o r a t e d as the r e s u l t of c e l l i n j u r y . "Leucotaxine" i s c r e d i t e d w i t h b r i n g i n g about the m i g r a t i o n of l e u c o c y t e s and t i s s u e edema, the l a t t e r being a d i r e c t r e s u l t of the Increased c a p i l l a r y p e r m e a b i l i t y . Edema was observed i n the a l e v i n s i r r a d i a t e d w i t h dosages 2x, 4x, and 8x, e s p e c i a l l y at the j u n c t i o n of the y o l k sac and the body w a l l . A s i m i l a r r e a c t i o n was observed by. Welander et a l (1948) who i r r a d i a t e d chinnok. salmon a few days before h a t c h i n g w i t h X-rays. The edema showed a l a t e n t p e r i o d of 30.days and was observed a l o n g the body w a l l above the y o l k sac, i n the lower jaw and i n the subocular lymph s i n u s . H i s t o -l o g i c a l examination of the t r e a t e d f i s h showed damage to the v a l v e s s e p a r a t i n g the lymph and venous systems 44 In salmon embryos, r e s u l t i n g i n e x t e n s i v e hemorrhage. Thus the m o r t a l i t y of the eggs and young salmon can be p a r t i a l l y a c c r e d i t e d to two f a c t o r s : -(1) heat hyperemia caused by the a b s o r p t i o n of the r a y s . (2) chemical changes which occur i n the b l o o d and damaged c e l l s as a r e s u l t of the absorbed r a d i a t i o n s c a u s i n g c a p i l l a r y d i l a t a t i o n and edema. E f f e c t s of u l t r a v i o l e t on f i s h s k i n (A) P r e l i m i n a r y experiments - P r e l i m i n a r y exper-iments u s i n g g o l d f i s h served to show t h a t u l t r a v i o l e t l i g h t can have an e f f e c t on f i s h . T h i s was suggested by Hart (1938) who succeeded i n p r o d u c i n g s l u f f i n g of i r r a d i a t e d s k i n and p i g m e n t a t i o n of g o l d f i s h i r r a d i a t e d w i t h a quartz-mercury lamp. The n e c r o s i s and pigment-a t i o n produced d i s a p p e a r e d i n two weeks. As was p o i n t e d out, p i g m e n t a t i o n o c c u r r e d f i r s t i n the areas of g r e a t e s t i n j u r y and was observed to s p r e a d v e n t r a l l y and l a t e r a l l y . Fukui (1927) found t h a t numerous agents induced b l a c k e n i n g , b e g i n n i n g on the d o r s a l s i d e s of the body and the t i p s of the f i n s , w i t h g r a d u a l spread-i n g to the areas below the l a t e r a l l i n e . He a l s o found t h a t r e g e n e r a t i n g t i s s u e s of the b l a c k , g o l d or white g o l d f i s h had the power of p r o d u c i n g melanophores, and t h a t the a b i l i t y to produce these melanophores appeared 45 t o be r e l a t e d to the presence of t y r o s i n e or t y r o s i n a s e i n the new s k i n . Smith (1932) found t h a t g o l d f i s h i r r a d i a t e d w i t h X-rays 7 times the dosage r e q u i r e d to produce erythema i n humans produced new pigment i n 11 days. This a r t i f i -c i a l l y induced pigment took 55 days t o d i s a p p e a r . The melanophores induced by the X-rays were u s u a l l y more d e l i c a t e and i r r e g u l a r than those found on the f i s h b efore treatment. Goodrich and Trinkhaus (1929) found t h a t c a l i c o g o l d f i s h produced pigment when i r r a d i a t e d with u l t r a v i o l e t l i g h t . They c o n s i d e r e d t h a t the c a l i c o f i s h were s p o t t e d i n v i s i b l y w i t h c o l o r l e s s chromoblasts, s i n g l y o r i n n e s t s , d u r i n g development. These m u l t i p l i e d and d i f f e r e n t i a t e d i ndependently at i r r e g u l a r i n t e r v a l s to form s p o t s or c l u s t e r s o f melanophores. U l t r a v i o l e t l i g h t i n some way s t i m u l a t e d the p r o d u c t i o n of i n c r e a s e d c l u s t e r s . M e l a n i n i s not a simple substance, but r a t h e r a mixture of pigments of I l l - d e f i n e d composition. I t has been shown t h a t the. s k i n c o n t a i n s an enzyme (dopase) capable of c o n v e r t i n g the compound 3, 4 dihydroxypheny-l a l a n i n e (dopa) i n t o m e l a n i n - l i k e m a t e r i a l , and that t h i s enzyme was absent from the s k i n of a l b i n o s . The probable p r e c u r s o r of 3, 4 d i h y d r o x y p h e n y l a l a n i n e i s t y r o s i n e (Hawk, Oser and Summerson, 1947). E a r l i e r workers t r i e d to e x p l a i n the a c t i o n of 46. u l t r a v i o l e t on s k i n , p r o d u c i n g pigment, as a photo-o x i d a t i o n in-which t y r o s i n e was d i r e c t l y o x i d i z e d t o 3, 4 d i h y d r o x y p h e n y l a l a n i n e , the o x i d a t i o n t a k i n g p l a c e i n the presence of u l t r a v i o l e t even i n the absence of t y r o s i n a s e . More r e c e n t l y the a c t i o n of the p r o d u c t i o n of melanin i n the s k i n has been e x p l a i n e d i n a s l i g h t l y d i f f e r e n t manner. The 3, 4 d i h y d r o x y p h e n y l a l a n i n e i s now b e l i e v e d t o be t r a n s p o r t e d to the s k i n from other p a r t s of the body, and i n the s k i n i s o x i d i z e d to melanin by dopase. This l a t t e r theory e x p l a i n s the l a t e n t p e r i o d between the I r r a d i a t i o n of the s k i n and the appearance of melanin (Blum, 1945). The appearance of new melanophores i n the g o l d -f i s h used i n these experiments was d e l a y e d from 2 t o 3 weeks and i n c r e a s e d i n amount f o r p e r i o d s of from 3 to 5 weeks. Fungus which a t t a c k e d the i r r a d i a t e d f i s h appeared f i r s t on the s l u f f i n g and damaged areas of the s k i n and l a t e r spread to the surrounding areas. T h i s fungus i s a s a p r o p h y t i c organism which a t t a c k s dead or damaged t i s s u e s f i r s t and l a t e r spreads to the su r r o u n d i n g t i s s u e s , causing t h e i r death. (B) A l e v i n sockeye salmon experiments - Laurens (1928) d e s c r i b e d the h i s t o l o g i c a l changes of i r r a d i a t e d s k i n : - ~ The changes occur b r i e f l y as f o l l o w s : the v e s s e l s become d i l a t e d w i t h i n f i l t r a t i o n o f the e p i t h e l i a l 47. l a y e r and an abnormal t h i c k e n i n g and d a r k e n i n g . F o l l o w i n g i n t e n s e r a d i a t i o n ther.e i s s e r o f i b r i n -ous or c e l l - r i c h , o f t e n hemorrhagic, exudation. I f the i r r a d i a t i o n i s severe enough the c o l l a g e n o u s t i s s u e becomes sw o l l e n and homogeneous, the e p i t h -e l i a n s w o l l e n and loosened, i n f i l t r a t e d and b l i s t e r e d , the l o s s of c o n t i n u i t y o c c u r r i n g a t the boundary between the granular and horny l a y e r s . I f i r r a d -i a t i o n i s q u i t e i n t e n s e thrombi form i n the v e s s e l s of the c u t i s . I r r a d i a t i o n of the ear of r a b b i t s w i t h a powerful C a r c lamp r e s u l t s i n d e g e n e r a t i o n and n e c r o s i s of the e p i t h e l i u m , b l i s t e r format i o n , s t a s i s , f i b r i n o u s exudation t o the s u r f a c e and t i s s u e spaces. In a d d i t i o n there i s a r e l a t i v e l y s l i g h t white c e l l i n f i l t r a t i o n , mainly of lympho-c y t e s , with plasma c e l l f o r m a t i o n . F i n a l l y a marked p r o d u c t i o n of f i b r o b l a s t s and the formation of new v e s s e l s and r e g e n e r a t i v e processes i n the e p i t h e l i u m . The r e s u l t s o b tained i n t h i s experiment resemble somewhat the changes i n the s k i n o f human beings and mammals. Studies of s e c t i o n s of the f i s h s k i n c l e a r l y i n d i c a t e d changes i n the epidermal and f i b r o e l a s t i c l a y e r s of the s k i n . Since f i s h do not have a horny l a y e r i n the epidermis, the r a d i a t i o n s were pro b a b l y a b l e to p e n e t r a t e t o the lower l a y e r s of the epidermis, c a u s i n g severe damage i n that r e g i o n . The damage to the g o b l e t c e l l s which o c c u r r e d 2 to 3 days a f t e r i r r a d i a t i o n p r o b a b l y had some e f f e c t on the osmotic r e g u l a t i o n of the f i s h . Loss of mucus on the outer s u r f a c e of the s k i n i s b e l i e v e d to have an e f f e c t on the osmotic r e g u l a t i o n , e s p e c i a l l y i f the u n d e r l y i n g t i s s u e i s damaged. F i s h i n f r e s h water l o s e s a l t s , e s p e c i a l l y c h l o r i r l e , t o the s u r r o u n d i n g medium when the mucus i s wiped o f f or i f the s k i n i s damaged (Krogh, 1939). This l o s s of 48. osmotic c o n t r o l was probably a c o n t r i b u t i n g f a c t o r to the death of the f i s h . The changes which o c c u r r e d i n the s k i n may be due e i t h e r t o the a b s o r p t i o n of the r a d i a n t energy by the protoplasm or by the n u c l e u s . Giese (1939) found by u s i n g monochromatic beams of u l t r a v i o l e t t h a t Paramecia were more s t r o n g l y I n h i b i t e d i n growth and k i l l e d by the wavelength 2804 A, the r e g i o n of maximal a b s o r p t i o n of the protoplasm. A wavelength of 2654 A, the r e g i o n of maxi-mal a b s o r p t i o n o f the n u c l e o p r o t e i n s , caused l e s s i n h i b i t i o n of growth i n i t i a l l y , but the recovery p e r i o d was p r o l o n g e d . In b a c t e r i a the wavelength 2654 A was found to be more e f f e c t i v e i n i n h i b i t i n g growth. Prom t h i s he concluded t h a t the immediate e f f e c t of u l t r a v i o l e t was more on the cytoplasm t h a n on the nucleus. Recovery of the cytoplasm was more r a p i d . I f the nucleus was harmed r e c o v e r y was l e s s r a p i d . In the i r r a d i a t e d s k i n the whole c e l l was a f f e c t e d by the i r r a d i a t i o n . The g r a n u l a t i o n appeared f i r s t i n the nucleus and secondly i n the cytoplasm. The h i g h e r dosages showed no r e g e n e r a t i o n of the damaged s k i n . Lower dosages ( l x , 0.5x) showed r e g e n e r a t i o n o c c u r r i n g f i r s t i n the b a s a l c e l l s . These c e l l s were probably a f f o r d e d p r o t e c t i o n by the o v e r l y i n g c e l l s so t h a t the n u c l e i were not damaged beyond r e c o v e r y . The wavelengths used I n these experiments were i n the r e g i o n which Giese (1939) found 49. most absorbed by the protoplasm. The observed e f f e c t of the u l t r a v i o l e t was l o c a l -i z e d , s i n c e areas of the s k i n p r o t e c t e d from the d i r e c t r a d i a t i o n showed no h i s t o l o g i c a l changes. The i n j u r i e s produced i n these l o c a l i z e d areas, as was p o i n t e d out e a r l i e r , produce " i n j u r y " substances which are t r a n s p o r t e d to the r e s t of the t i s s u e s by the b l o o d and a f f e c t the organism as a whole. E f f e c t of u l t r a v i o l e t on other t i s s u e s The appearance of f l o c c u l e n t spots on the y o l k sac a f t e r the i r r a d i a t i o n were pro b a b l y due to the denatur-a t i o n and c o a g u l a t i o n of the p r o t e i n s c o n t a i n e d i n .the y o l k m a t e r i a l . S i m i l a r spots were observed by Welander et a l (1948) a f t e r t r e a t i n g chinook salmon a l e v i n s w i t h X-rays, and were a t t r i b u t e d to the same cause. The f a c t t h a t t i s s u e s l y i n g deeper i n the body were not v i s i b l y a f f e c t e d i n d i c a t e s t h a t the u l t r a v i o l e t r a y s were absorbed i n the s u r f a c e l a y e r s of t i s s u e . T h i s was c o n s i s t e n t w i t h the o b s e r v a t i o n s of Bachem and Reed (1931) who measured the transparency of human s k i n to u l t r a v i o l e t . T h e i r f i n d i n g s showed that a l a y e r of s k i n 1.0 mm. t h i c k absorbed most of the u l t r a v i o l e t while v i s i b l e and i n f r a r e d r a d i a t i o n s were more p e n e t r a t i n g . X-rays because of t h e i r h i g h energy are more p e n e t r a t i n g than u l t r a v i o l e t r a y s and t h e i r primary e f f e c t i s on the organs and t i s s u e s l y i n g deeper i n the body (Bonham et a l , 1948; Welander et a l , 50. 1948). The s u p e r f i c i a l l a y e r s such as muscle and s k i n were not v i s i b l y harmed, s i n c e the r a d i a t i o n s were not a p p r e c i a b l y absorbed i n those l a y e r s . 51, SUMMARY 1. U l t r a v i o l e t l i g h t of wavelengths from 2800 A to 3100 A causes l e t h a l or harmful e f f e c t s on chum and sockeye salmon eggs, a l e v i n sockeye salmon, coho f i n g e r -l i n g s and g o l d f i s h . 2. Sockeye salmon eggs i n l a t e r stages of develop-ment were induced to hatch a month e a r l i e r than normal d e v e l o p i n g f i s h . Slowing of development and pigment-a t i o n were observed and abnormal body forms were produced. 3. Pigmentation and s l u f f i n g were produced i n g o l d -f i s h s k i n and i n c r e a s e d s u s c e p t i b i l i t y to fungus was observed. 4. H i s t o l o g i c a l s t u d i e s of the e f f e c t s o f u l t r a -v i o l e t r a d i a t i o n on a l e v i n sockeye salmon r e v e a l e d changes p r i m a r i l y i n the epidermis and f i b r o e l a s t i c - l a y e r s of the s k i n . The e f f e c t was l o c a l i z e d . Organs and t i s s u e s deeper i n the body showed no changes. 5. The a c t u a l cause of death was not determined, but a combination of f a c t o r s may have been r e s p o n s i b l e : -(a) heat hyperemia. (b) development of t o x i c substances i n the I n j u r e d t i s s u e s . (c) l o s s of osmotic r e g u l a t i o n due to s k i n i n j u r i e s . 52. ACKNOWLEDGMENTS The author wishes t o express her thanks to Dr. W. A. Clemens and the members of the s t a f f of the Department of Zoology, U n i v e r s i t y of B r i t i s h Columbia, where t h i s work was undertaken. S p e c i a l thanks are due to Dr. W. S. Hoar, under whose guidance t h i s work was c a r r i e d out and without whose h e l p f u l c r i t i c i s m s much would have been l e f t undone. Mr. J . Mason, Mr. I. B a r r e t t , Mr. R. W. R a d c l i f f e , Mr. P. Paul and Mr. H. B e l l were most h e l p f u l i n p r o v i d i n g eggs and f i s h , b u i l d i n g equipment necessary, and p r e p a r i n g i l l u s t r a t i o n s . Many thanks are due to them f o r the.ir a s s i s t a n c e . Very s p e c i a l thanks are due to my p a r e n t s , whose g e n e r o s i t y and i n t e r e s t have ever been an I n s p i r a t i o n f o r my work. 53. LITERATURE CITED Bachem A. and C. I. Reed, 1948. The p e n e t r a t i o n of l i g h t through human s k i n . Am. Journ. Phys., 97; 86-91. Blum, H.P., 1945. The p h y s i o l o g i c a l e f f e c t s of s u n l i g h t on man. Phys. Rev., 25: 483-530. Bonham, K., L. R. Donaldson, R. P. P o s t e r , A. D. Welander and A. H. Seymour, 1948. The e f f e c t of X-ray on m o r t a l i t y , weight, l e n g t h , and counts of e r y t h r o -c y t e s and hematopoietic c e l l s i n the f i n g e r l i n g Chinook salmon, Onycorhynchus tschawytscha Walbaum. Growth, 2: 107-121." Burge, W.E., 1917. The a c t i o n of u l t r a v i o l e t r a d i a t i o n i n k i l l i n g l i v i n g . c e l l s such as b a c t e r i a . Am. Journ. Phys., 43: 429-432. Burge, W. E., W. R. F i s c h e r and A. G. N e i l , 1916. Des-t r u c t i o n o f hormones, proenzymes, and enzymes by u l t r a v i o l e t r a d i a t i o n . Am. Journ. Phys., 40: 426-436.. In H i n r i c k s , M. A., 1925. M o d i f i c a t i o n of development on the b a s i s of d i f f e r e n t i a l s u s c e p t i b i l i t y t o r a d i a t i o n . I Fundulus h e t e r o l i t i c u s and u l t r a v i o l e t . r a d i a t i o n . Journ. Morph., 41: 239-268. C l a r k , J . H., 1922. The p h y s i o l o g i c a l a c t i o n of l i g h t . Phys. Rev., 2: 277-307. Fukui, Ken ' I c h i , 1927. On the c o l o r p a t t e r n produced by v a r i o u s agents i n the g o l d f i s h . F o l i a . Anat. Japon., 5: 257-302. Giese, A. C , 1939. U l t r a v i o l e t r a d i a t i o n and c e l l d i v i s i o n . 1. E f f e c t s of X 2654 and >v 2804 upon Paramecium  caudatum. Journ. C e l l . Comp. Phys., 13: 139-150. Goodrich, H. B. and J . P. Trinkhaus, 1939. The d i f f e r e n t i a l e f f e c t of r a d i a t i o n s on the Mendelian phenotypes of the g o l d f i s h , C a r a s s i u s auratus . B i o l . B u l l . , 77: 188-195. H a r r i s , F. I . and H. S. Hoyt, 1917. The p o s s i b l e o r i g i n of the t o x i c i t y of u l t r a - v i o l e t l i g h t . S c i e n c e , 46: 318-320. Hart, J . L., 1938. ' Can sunburn a f f e c t f i s h i n h a t c h e r y ponds? F i s h . Res. Bd. Can. Prog. Rept. P a c , (35): 6. 54. Hawk, P. B», B. L. Oser and Wm. H. Summerson, 1947. P r a c t i c a l p h y s i o l o g i c a l chemistry, x i v + 1-1322, 12th ed., B l a k i s t o n Co., P h i l a d e l p h i a . H i n r i c k s , M. A., 1925. M o d i f i c a t i o n of development on the b a s i s of d i f f e r e n t i a l s u s c e p t i b i l i t y to r a d i a t i o n . I Fundulus h e t e r o l i t i c u s and u l t r a v i o l e t r a d i a t i o n . Journ. Morph., 41: 239-268. Krogh, A., 1939. Osmotic r e g u l a t i o n In a q u a t i c animals, 1-242, Cambridge. L a n d o l t , H. and R. B o r n s t e i n , 1923. P h y s i k a l i s c h -chemische t a b e l e n , 2: 896. Laurens, H., 1928. The p h y s i o l o g i c a l e f f e c t s of r a d i a t i o n . Phys. Rev., 8: 1-91. Laurens, H., 1933. The p h y s i o l o g i c a l e f f e c t s of r a d i a n t energy, 1-610, Chemical Catalogue Co., New York. Laurens, H., 1935. Photochemistry i n medicine: a general o u t l i n e . Symp. Quant. B i o l . , 3: 277-295. Lewis, T. and Y. Zotterman, 1926. Heart, 13-: 203. I n Blum, H. P., 1945. The p h y s i o l o g i c a l e f f e c t s of s u n l i g h t on man. Phys. Rev., 25: 483-530. Luc k i e s h , M., 1946. The a p p l i c a t i o n of g e r m i c i d a l , erythemal and i n f r a r e d energy, v i i i + 1-463, D. Van No s t r a n d Co.., New York. Mayerson, H. S., 1935. The e f f e c t s of r a d i a t i o n on c i r c u l a t i o n , b l o o d and metabolism w i t h p a r t i c u l a r r e f e r e n c e to-growth, b a s a l metabolism and the t h y r o i d g l a nd. Symp. Quant. B i o l . , 3: 299-311. Menkin, V., 1938. The r o l e of inflammation i n immunity. Phys. Rev., 18: 366-418. Pohle, E. A., 1926. S t a n d a r d i z a t i o n of u l t r a - v i o l e t r a y s . The u l t r a - v i o l e t u n i t . Journ. Am. Med. Assoc., 86: '818-820. Smith, G. M., 1932. Melanophores induced by X-ray as compared w i t h those e x i s t i n g i n the p a t t e r n s as seen In C a r a s s i u s a u r a t u s . B i o l . B u l l . , 63: 484-491. Welander, A. D., L. R. Donaldson, R. P. P o s t e r , K. Bonham and"A. H. Seymour, 1948. The e f f e c t s of Roentgen rays on the embryos and l a r v a e of the Chinook salmon. Growth, 12: 203-242. 55. Weitz, C. E., 1946. General E l e c t r i c lamp b u l l e t i n . B u l l . LD1, Toronto. ABSTRACT U l t r a v i o l e t l i g h t o f wavelengths from 2800 A t o 3100 A, generated by a General E l e c t r i c RS sunlamp, were used to i r r a d i a t e eggs and young f i s h . The dosages were s t a n d a r d i z e d by means of a photochemical r e a c t i o n i n which i o d i n e was l i b e r a t e d from h y d r i o d i c a c i d under the I n f l u e n c e of i n t e n s e l i g h t . T h i s f r e e i o d i n e gave a blue c o l o r a t i o n w i t h the s t a r c h t e s t . P r e l i m i n a r y experiments u s i n g the g o l d f i s h (Carassius auratus) produced d e f i n i t e e f f e c t s . The s k i n was observed to b l i s t e r and s l u f f i n the exposed r e g i o n s , s u s c e p t i b i l i t y t o fungus became Increased over t h a t of the c o n t r o l s , and an i n c r e a s e i n the number of melanophores o c c u r r e d i n some of the i r r a d i a t e d f i s h a f t e r a l a t e n t p e r i o d of 2 to 3 weeks. M o r t a l i t y curves were determined f o r the eggs of the chum salmon (Onycorhynchus keta) and the sockeye salmon (Onycorhynchus n e r k a ) . Experiments u s i n g sockeye salmon eggs i n the l a t e r stages of development produced a s t i m u l a t i n g e f f e c t on the r a t e of h a t c h i n g . The a l e v i n s which hatched a month ahead of the c o n t r o l s were abnormal i n s e v e r a l r e s -p e c t s . The v e r t e b r a l column was curved downward over the r e g i o n of the h e a r t , growth was r e t a r d e d , the y o l k was not u t i l i z e d so r a p i d l y as the c o n t r o l s and p i g m e n t a t i o n was delayed. More m o r t a l i t i e s o c c u r r e d d u r i n g h a t c h i n g I n the i r r a d i a t e d than i n the c o n t r o l f i s h . Sockeye salmon a l e v i n s averaging 20 cm. i n l e n g t h were i r r a d i a t e d s h o r t l y a f t e r h a t c h i n g . Heavy l o s s e s occurred i n i r r a d i a t i o n s l a s t i n g from 15 minutes to 4 hours. H i s t o l o g i c a l s t u d i e s r e v e a l e d changes i n the epidermis and f i b r o e l a s t i c l a y e r s of the s k i n which were l o c a l i z e d t o the r e g i o n s which had been exposed. T i s s u e s under the f i b r o -e l a s t i c l a y e r d i d not show v i s i b l e changes. Larger dosages showed severe d e g e n e r a t i o n of epidermal c e l l s w i t h subse-quent s l u f f i n g and l o s s of goblet c e l l s . Lower dosages showed de g e n e r a t i o n of the outer c e l l s o f the epidermis a f t e r a l a t e n t p e r i o d of 2 to 3 days. Recovery o c c u r r e d i n a s m a l l number i r r a d i a t e d w i t h the lowest dosage about I 12 to 13 days a f t e r i r r a d i a t i o n . M o r t a l i t i e s c o u l d have been caused by:-1. heat hyperemia caused by a b s o r p t i o n of the r a y s . 2. chemical changes o c c u r r i n g i n the b l o o d as a r e s u l t of c e l l i n j u r y caused by the r a d i a t i o n s . 3. decrease i n osmotic r e g u l a t i o n caused by i n j u r i e s to the s k i n . 

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