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Brain epinephrine and norepinephrine in the neonate chick: Phylogenetic implications
Comp. gen. Pharmac., x97i , 2. [Scientechnica (Publishers) Ltd.]
Ix8
BRAIN
EPINEPHRINE
AND
NOREPINEPHRINE
IN
THE
N E O N A T E CHICK: P H Y L O G E N E T I C I M P L I C A T I O N S JOSEPH
P. H A N I G *
Department of Pharmacology, New York Medical College, io5th Street and 5th Avenue, New York, New York IOO29, U.S.A. ABSTRACT x. Average norepinephrine and epinephrine levels in whole brain of i 12 neonate chicks, Gallus domesticus, were found to be o'522 and o'277 lag. per g. respectively, a ratio of 2 : I. 2. In contrast, norepinephrine greatly predominates over epinephrine in fish and mammalian brain, whereas the reverse is true for amphibians. 3. The ratio of catecholamines in chick brain most resembles that of reptiles and this is consistent with the established phylogenetie position of Aves. T h E vast m a j o r i t y o f research u n d e r t a k e n to e l u c i d a t e the role o f the c a t e c h o l a m i n e s in the c e n t r a l nervous system has been perf o r m e d in rats a n d o t h e r m a m m a l s . This class o f v e r t e b r a t e s has a r a t h e r c h a r a c t e r istic whole b r a i n c a t e c h o l a m i n e profile in w h i c h n o r e p i n e p h r i n e (NE) p r e d o m i n a t e s (0"4-0"6 lag. p e r g.) a n d e p i n e p h r i n e (E) is b a r e l y d e t e c t a b l e (von Euler, x946 , I956; Vogt, 1954; Bogdanski, Bonomi, a n d Brodie, z963). I n contrast, the relative concentrations of these two amines in b r a i n a r e considera b l y different in lower classes of v e r t e b r a t e s T h e reptiles, a c o m m o n ancestor of b o t h m a m m a l s a n d Ayes ( R o m e r , i967) , have a r a t i o of b r a i n N E to E of 3 : I, whereas the a m p h i b i a n s , only one step lower phylogenetically, h a v e a v e r y large p r e d o m i n a n c e of E in the b r a i n w i t h little or no N E present (Brodie a n d Bogdanski, I964). I n the course of o u r studies of the role of biogenic amines in the C.N.S. of the n e o n a t e chick, Callus domesticus, the question arose as to w h e t h e r N E a n d E levels in b r a i n were consistent w i t h the g e n e r a l l y a c c e p t e d phylogenetic position of Ayes, i.e., a n offshoot from reptiles t h a t p a r a l l e l e d chronologically the d e v e l o p m e n t of m a m m a l s ( R o m e r , i967). I n o r d e r to investigate this question, whole b r a i n N E a n d E levels were e s t i m a t e d fluorim e t r i c a l l y in n e o n a t e chicks. * Present address: Food and Drug Administration, Bureau of Drugs, Division of Drug Biology, Washington D.C. 2o2o4, U.S.A.
M A T E R I A L S AND M E T H O D S One-day-old sex-linked hybrid cockerels were housed in a temperature-controlled brooder and given free access to food and water. All animals were used for experimentation within I week of hatching. NE and E were estimated in whole brain by a slight modification of the differential fluorometric procedure of Chang ([964). The catecholamines contained in the supernatant of perchloric acid/brain homogenates were adsorbed on to alumina activated by the procedure of Anton and Sayre (I962) and eluted, after two successive water washings, with 0-2 N acetic acid. These extracts were oxidized with o. I ./V iodine followed by 25 per cent alkaline sulphite and 0"5 jV acetic acid and the fluorescent trihydroxyindoles formed from E and NE were read in an Aminco-Bowman spectrophotofluorometer at 500 mla and 485 mla respectively following activation at 4io mla and 385 mla. Simultaneous equations were used to calculate the concentration of E and NE in each sample. RESULTS AND DISCUSSION Analysis of N E in whole b r a i n of I I 2 chicks resulted in an a v e r a g e v a l u e Of 0.522 lag. p e r g. E in whole b r a i n was found to have a n a v e r a g e value o f 0.277 lag. p e r g. T h e endogenous levels of b o t h c a t e c h o l a m i n e s in chick b r a i n were of the s a m e o r d e r o f m a g n i t u d e with a r a t i o of 2 : i respectively. This ratio is v e r y different from t h a t r e p o r t e d for fish (I o : [) b y von Euler (I 96 I), a m p h i bians (i : I O), or m a m m a l s (x o : x), b u t does closely resemble t h a t o f reptiles (3 : I). T h e levels o f b o t h catecholamines in chick brain, r e p o r t e d above, a r e in general agreem e n t with those r e p o r t e d b y J u o r i o a n d V o g t
197I, 2, I I 8 - I I 9
BRAIN EPINEPHRINE AND NOREPINEPHRINE
II9
In conclusion, we have attempted to (I967) for mature chickens. A direct comparison could not be made, however, since demonstrate, in chicks, a consistency bewe did not estimate these amines in specific tween the catecholamine pattern in the brain brain areas. O u r norepinephrine value for and the phylogenetic position or relative whole brain is in close agreement with that of degree of evolutionary development of Aves Watts, Mendez, Reilly, and Krop (1969) , in comparison to other vertebrate classes. O n although they either did not look for or did both counts, Ayes most resemble reptiles but not detect epinephrine in whole brain of do have certain mammalian tendencies with chick. It is interesting that Watts and his co- onset of maturity. workers were able to demonstrate a significant increase in brain norepinephrine with REFERENCES onset of maturity in chickens. This may ANTON,A. H., and SAYRE,D. F. (I962), ' A study of the factors affecting the aluminium oxidereflect the onset of development changes, trihydroxyindole procedure for the analysis of such as conversion from near poikilothermy catecholamines ', 07. Pharmac. exp. Ther., I38~ to homeothermy and others, which make the 36o-375. mature chicken, in certain respects, more BOGDANSKI,D. F., BONOMX,L., and BRODm, B. B. similar to mammals than to reptiles. (x963), ' Occurrence of serotonin and catecholamines in brain and peripheral organs of As one ascends the vertebrate phylogenetic various vertebrate classes ', Life Sci., ~, 8 o ~ 4. scale, a general pattern of reversal of cateBRODIE, B. B., and BOGDANSKI, D. F. (x964), cholamine predominance seems to occur 'Biogenic amines and drug action in the until one reaches the reptiles. Beyond this nervous system of various vertebrates ', Prog. branch point reversal does not occur, but Brain Res., 9, 234-242rather alteration becomes a matter of degree; CHANG, C. C. (I964), ' A sensitive method for spectrophotofluorometric assay of catecholAyes going in one direction by showing a amines ', Int. 07../Veuropharmac., 3, 643"649 • relative increase in brain epinephrine and VON EULER, U. S. (I946), ' A specific sympathomammals in the opposite direction with a mimetic ergone in adrenergic nerve fibres (sympathin) and its relation to adrenaline and very large relative increase of brain norepinenor-adrenaline ', Acta physiol, scand., I2, 73-97phrine. It would appear, in this case, that Noradrenaline, pp. 133-i54. Springmorphological evolution of the C.N.S. does - - -field,(I956), Ill. : Charles C. Thomas. have chemical sequelae with regard to - - - - ( x 9 6 1 ) , 'Occurrence and distribution of catecholamine predominance. This variacatecholamines in the fish brain ', Acta physiol. scand., 52, 62-64. tion may represent a form of neurochemical JuoRxo, A. V., and VOGT, M. (I967), 'Monoevolution with a phylogenetic component amines and their metabolites in avian brain ', between successive vertebrate classes and an J. Physiol., Lond., x~, 489-518. ontogenetic component within each of these ROMER, A. S. (1967), ' Major steps in vertebrate evolution ', Science, N.Y., x58, 1629-1637. classes. It is worthy of consideration that fish, one VOGT, M. (I954), ' T h e concentration of sympathin in different parts of the central nervous of the most primitive of vertebrates, and system under normal conditions and after the mammals, the most advanced, show the administration of drugs ', 07. Physiol., Lond., x23, greatest similarity in brain catecholamine 451-48I. ratios (von Euler, 196I). This may serve to WATTS,J. S., MENDEZ, H. C., RmLLV,J. F., and KROP, S. (I969), 'Brain amine content and illustrate the need for further investigation of functional maturity in the fowl: The effects of the nature and significance of evolutionary pargyline ', Archs int. Pharmacodyn., x78, x53processes that have given rise to different 159. neurotransmitter-enzyme systems in various Key Word Index: Neonate chick, whole brain phyla of the animal kingdom, particularly catecholamines, epinephrine, norepinephrine, with regard to neurohumeral modulation of neurochemical-phylogenetic correlates, developseemingly similar homeostatic functions. mental pharmacology.
Ix8
BRAIN
EPINEPHRINE
AND
NOREPINEPHRINE
IN
THE
N E O N A T E CHICK: P H Y L O G E N E T I C I M P L I C A T I O N S JOSEPH
P. H A N I G *
Department of Pharmacology, New York Medical College, io5th Street and 5th Avenue, New York, New York IOO29, U.S.A. ABSTRACT x. Average norepinephrine and epinephrine levels in whole brain of i 12 neonate chicks, Gallus domesticus, were found to be o'522 and o'277 lag. per g. respectively, a ratio of 2 : I. 2. In contrast, norepinephrine greatly predominates over epinephrine in fish and mammalian brain, whereas the reverse is true for amphibians. 3. The ratio of catecholamines in chick brain most resembles that of reptiles and this is consistent with the established phylogenetie position of Aves. T h E vast m a j o r i t y o f research u n d e r t a k e n to e l u c i d a t e the role o f the c a t e c h o l a m i n e s in the c e n t r a l nervous system has been perf o r m e d in rats a n d o t h e r m a m m a l s . This class o f v e r t e b r a t e s has a r a t h e r c h a r a c t e r istic whole b r a i n c a t e c h o l a m i n e profile in w h i c h n o r e p i n e p h r i n e (NE) p r e d o m i n a t e s (0"4-0"6 lag. p e r g.) a n d e p i n e p h r i n e (E) is b a r e l y d e t e c t a b l e (von Euler, x946 , I956; Vogt, 1954; Bogdanski, Bonomi, a n d Brodie, z963). I n contrast, the relative concentrations of these two amines in b r a i n a r e considera b l y different in lower classes of v e r t e b r a t e s T h e reptiles, a c o m m o n ancestor of b o t h m a m m a l s a n d Ayes ( R o m e r , i967) , have a r a t i o of b r a i n N E to E of 3 : I, whereas the a m p h i b i a n s , only one step lower phylogenetically, h a v e a v e r y large p r e d o m i n a n c e of E in the b r a i n w i t h little or no N E present (Brodie a n d Bogdanski, I964). I n the course of o u r studies of the role of biogenic amines in the C.N.S. of the n e o n a t e chick, Callus domesticus, the question arose as to w h e t h e r N E a n d E levels in b r a i n were consistent w i t h the g e n e r a l l y a c c e p t e d phylogenetic position of Ayes, i.e., a n offshoot from reptiles t h a t p a r a l l e l e d chronologically the d e v e l o p m e n t of m a m m a l s ( R o m e r , i967). I n o r d e r to investigate this question, whole b r a i n N E a n d E levels were e s t i m a t e d fluorim e t r i c a l l y in n e o n a t e chicks. * Present address: Food and Drug Administration, Bureau of Drugs, Division of Drug Biology, Washington D.C. 2o2o4, U.S.A.
M A T E R I A L S AND M E T H O D S One-day-old sex-linked hybrid cockerels were housed in a temperature-controlled brooder and given free access to food and water. All animals were used for experimentation within I week of hatching. NE and E were estimated in whole brain by a slight modification of the differential fluorometric procedure of Chang ([964). The catecholamines contained in the supernatant of perchloric acid/brain homogenates were adsorbed on to alumina activated by the procedure of Anton and Sayre (I962) and eluted, after two successive water washings, with 0-2 N acetic acid. These extracts were oxidized with o. I ./V iodine followed by 25 per cent alkaline sulphite and 0"5 jV acetic acid and the fluorescent trihydroxyindoles formed from E and NE were read in an Aminco-Bowman spectrophotofluorometer at 500 mla and 485 mla respectively following activation at 4io mla and 385 mla. Simultaneous equations were used to calculate the concentration of E and NE in each sample. RESULTS AND DISCUSSION Analysis of N E in whole b r a i n of I I 2 chicks resulted in an a v e r a g e v a l u e Of 0.522 lag. p e r g. E in whole b r a i n was found to have a n a v e r a g e value o f 0.277 lag. p e r g. T h e endogenous levels of b o t h c a t e c h o l a m i n e s in chick b r a i n were of the s a m e o r d e r o f m a g n i t u d e with a r a t i o of 2 : i respectively. This ratio is v e r y different from t h a t r e p o r t e d for fish (I o : [) b y von Euler (I 96 I), a m p h i bians (i : I O), or m a m m a l s (x o : x), b u t does closely resemble t h a t o f reptiles (3 : I). T h e levels o f b o t h catecholamines in chick brain, r e p o r t e d above, a r e in general agreem e n t with those r e p o r t e d b y J u o r i o a n d V o g t
197I, 2, I I 8 - I I 9
BRAIN EPINEPHRINE AND NOREPINEPHRINE
II9
In conclusion, we have attempted to (I967) for mature chickens. A direct comparison could not be made, however, since demonstrate, in chicks, a consistency bewe did not estimate these amines in specific tween the catecholamine pattern in the brain brain areas. O u r norepinephrine value for and the phylogenetic position or relative whole brain is in close agreement with that of degree of evolutionary development of Aves Watts, Mendez, Reilly, and Krop (1969) , in comparison to other vertebrate classes. O n although they either did not look for or did both counts, Ayes most resemble reptiles but not detect epinephrine in whole brain of do have certain mammalian tendencies with chick. It is interesting that Watts and his co- onset of maturity. workers were able to demonstrate a significant increase in brain norepinephrine with REFERENCES onset of maturity in chickens. This may ANTON,A. H., and SAYRE,D. F. (I962), ' A study of the factors affecting the aluminium oxidereflect the onset of development changes, trihydroxyindole procedure for the analysis of such as conversion from near poikilothermy catecholamines ', 07. Pharmac. exp. Ther., I38~ to homeothermy and others, which make the 36o-375. mature chicken, in certain respects, more BOGDANSKI,D. F., BONOMX,L., and BRODm, B. B. similar to mammals than to reptiles. (x963), ' Occurrence of serotonin and catecholamines in brain and peripheral organs of As one ascends the vertebrate phylogenetic various vertebrate classes ', Life Sci., ~, 8 o ~ 4. scale, a general pattern of reversal of cateBRODIE, B. B., and BOGDANSKI, D. F. (x964), cholamine predominance seems to occur 'Biogenic amines and drug action in the until one reaches the reptiles. Beyond this nervous system of various vertebrates ', Prog. branch point reversal does not occur, but Brain Res., 9, 234-242rather alteration becomes a matter of degree; CHANG, C. C. (I964), ' A sensitive method for spectrophotofluorometric assay of catecholAyes going in one direction by showing a amines ', Int. 07../Veuropharmac., 3, 643"649 • relative increase in brain epinephrine and VON EULER, U. S. (I946), ' A specific sympathomammals in the opposite direction with a mimetic ergone in adrenergic nerve fibres (sympathin) and its relation to adrenaline and very large relative increase of brain norepinenor-adrenaline ', Acta physiol, scand., I2, 73-97phrine. It would appear, in this case, that Noradrenaline, pp. 133-i54. Springmorphological evolution of the C.N.S. does - - -field,(I956), Ill. : Charles C. Thomas. have chemical sequelae with regard to - - - - ( x 9 6 1 ) , 'Occurrence and distribution of catecholamine predominance. This variacatecholamines in the fish brain ', Acta physiol. scand., 52, 62-64. tion may represent a form of neurochemical JuoRxo, A. V., and VOGT, M. (I967), 'Monoevolution with a phylogenetic component amines and their metabolites in avian brain ', between successive vertebrate classes and an J. Physiol., Lond., x~, 489-518. ontogenetic component within each of these ROMER, A. S. (1967), ' Major steps in vertebrate evolution ', Science, N.Y., x58, 1629-1637. classes. It is worthy of consideration that fish, one VOGT, M. (I954), ' T h e concentration of sympathin in different parts of the central nervous of the most primitive of vertebrates, and system under normal conditions and after the mammals, the most advanced, show the administration of drugs ', 07. Physiol., Lond., x23, greatest similarity in brain catecholamine 451-48I. ratios (von Euler, 196I). This may serve to WATTS,J. S., MENDEZ, H. C., RmLLV,J. F., and KROP, S. (I969), 'Brain amine content and illustrate the need for further investigation of functional maturity in the fowl: The effects of the nature and significance of evolutionary pargyline ', Archs int. Pharmacodyn., x78, x53processes that have given rise to different 159. neurotransmitter-enzyme systems in various Key Word Index: Neonate chick, whole brain phyla of the animal kingdom, particularly catecholamines, epinephrine, norepinephrine, with regard to neurohumeral modulation of neurochemical-phylogenetic correlates, developseemingly similar homeostatic functions. mental pharmacology.