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Monoamine oxidase and catechol-O-methyl transferase in tissues of rainbow trout (Salmo gairdneri rich., teleost)
Comp. Gen. Phantmc., i974, Vol. 5, 25x to 253. Pergamon Press. Printed in Great Britain
251
MONOAMINE OXIDASE AND CATECHOL-O-METHYL TRANSFERASE IN TISSUES OF RAINBOW T R O U T
(SALMO GAIP~NERI RICH., TELEOST) M A D E L E I N E M. M A Z E A U D Institut Ocdanographique, i95 rue Saint-Jacques, 75--o05 Paris, France
(Received 26 October 1973) ABSTRACT I. The catabolism of catecholamines is performed in several tissues of the rainbow trout. 2. Monoamine oxidase activity is important in the kidney and brain. 3. O-Methylation occurs mainly in the liver and seems to be the major route for catecholamine degradation. IN fish, two i m p o r t a n t mechanisms for catecholamine inactivation are deamination and 0 - m e t h y l a t i o n (Mazeaud, i969, i97i ; M a z e a u d & Mazeaud, i973). Both routes of catabolism are the same as those described in m a m m a l s , and are catalysed by monoamine oxidase ( M A O ) and catechol-0methyl transferase ( C O M T ) , as was demonstrated in m a m m a l s (Blaschko, I952 ; Axelrod, i957; Axelrod, Senoh & Witkop, i958; Axelrod & Tomchick, i958; Molinoff & Axelrod, 197t ). I n fish, according to our knowledge, no work on this subject has been reported, except for the histological evidence of M A O in the retina of a Teleost, Eugerres plumieri (Drujan & Diaz Borges, I972 ) and in the hypothalamus o f several Teleosts (Urano, I971 ). I n this preliminary work, we studied M A O and C O M T activity in several tissues of a Teleost fish, the rainbow trout (Salmo gairdneri Rich.). MATERIALS AND METHODS The rainbow trout we used came from piscicultures of the Paris region. We sampled the heart and brain in toto, the cephalic third of the kidney, pieces of liver, white or red muscle and the whole gill on the right side after eliminating bones. After sampling, every tissue was weighed, ground and maintained at + 4 ° C until it was incubated. MAO activity was determined by incubating tissue homogenates with [x'C] tryptamine ([2-x*C]tryptamine succinate (New England
Nuclear), 50 mCi./mmole), according to Wurtman & Axelrod (x963). Theamountof[l*C]indolacetic acid resulting from deamlr~ation of tryptamine was measured. A small amount of [t4C] tryptamine was extracted with the deaminated radioactive material, and a correction was made for this blank value by incubating [l'C]tryptamine with boiled homogenate. MAO activity is expressed in U/g. (IU = number ofnmoles of [14C]indolacetic acid produced per hour). C O M T activity was determined by incubating tissue extracts with epinephrine and [I'C]Sadenosyl m~thionine (Centre Energie Atomique Saclay, France, 5mCi./mmole) according to Wurtman & Axelrod (t966). The m o u n t s of []*C]metanephrine resulting from 0-methylation of epinephrine was measured. Data were corrected for blank values obtained by omitting epinephrine in the incubation medium. Activity is expressed in U/g. (IU = number of nmoles of [t4C]metanephrine produced per hour). Radioactivity was measured by liquid scintillation, and quenching corrected by the use of an external standard. At a prcllminary step, we studied the validity of mammalian methodology applied to fish (Mazeaud, unpublished results). In terms of our data, the reaction was linear with enzyme eoncentratiod over a range of xo-2oo/zg, liver and xo-5oo frg. heart in the case of MAO, and IO--5oo/zg. liver and io-2ooo/zg, heart in the case of COMT. The best temperature for incubation was shown to be 3o ° C. RESULTS AND DISCUSSION I n the rainbow trout, all tissues are able to achieve d e a m i n a t i o n a n d 0-methylation, and thus contain M A O a n d C O M T . I n the case o f C O M T , we emphasized the need
Table
Comp. Gen. Pharmac.
MAZEAUD
~52 I.--CATECHOL-0-METHYL
TRANSFERASE
(COMT) Ac-nvrrY SHOWING THE REQUIREMENT F o e MgS, IONS IN THE INCUBATION MEDIUM Tms~z
Liver Kidney Heart White muscle Red muscle Brain Gill
AewpcrrY (U/g.) Incubation Incubation medium medium with MgCls without MgCls
COMT
870.4 74"5 68.5 x8. I 32.6 52.0 3 x.8
x9"3 22"0 5"x I "9 8.4 12. i 8.6
for Mg 2+ ions in the incubation medium (Table I) as was shown for mammals (Wurtman & Axelrod, i966 ). SPECIFIC ACTIVITY OF TISstr~s
For M A O , the values vary from 31.9 U/g. in the white muscle to I28.2 U/g. in the kidney. For C O M T , values vary from 21.I U/g. inthc red muscle to Io41.2 U/g. in the liver (Table II). A comparison with mammals appears to bc difficultowing to the variety eitherof methodology or expression of the results. For instancc, inscct Egashira, Takano, Shlrnizu, Kurosawa & Kamijo (1971) use a manornctric method and express M A O activity in /zl.oz/hour/ml, of homogcnatc. For C O M T , other substrates than epinephrine may bc used, e.g., norcpincphrinc (Crout, i96I); 3, 4-dihydroxybcnzoic acid (MacCaman, 1965) or 3, 4-dihydroxypheny I acetic acid
(Assicot & Bohuon, I969; Broch & Guldberg, I97I); the end products of the enzymatic reaction may be measured fluorimetrically (Axelrod & Tomchick, 1958; Crout, 1961 ; Assicot & Bohoun, I969) , or by radiochemical methods (Axelrod Albers & Clemente, i959; MacCaman, I965). From the results obtained by all these authors, it is evident that among mammals, the specific activities of M A O and C O M T depend largely on the species studied. T h e specific activitity of M A O varies from 182 U/g. in the rabbit liver, to I2I i U/g. in the rat liver (Gripois, I97O ). In the liver of trout, the amount of M A O seems to be smaller (6~ U]g.). In the case of C O M T , the specific activity of the fiver is too U]g. in the rabbit, 4oo U]g. in man, 33oo U/g. in the cow and 5ooo U/g. in the rat. Thus, a variability factor from I to 5 ° appears. O u r results demonstrate that the value we found for C O M T activityin the trout liver (i o4o.2 U]g.) is integrated in this scale of values. DISTRIBUTION
In the rainbow trout, the most important M A O activity is in the kidney and brain. The major role of the kidney for M A O activity is also reported in several species of mammals like the rabbit, while in other species like the rat, the heart or liver possess the highest M A O activities (Gripois, I97o ). However, in the trout, the M A O ratio of the kidney to M A O of any other tissue studied is at most 6. The M A O distribution is therefore more uniform than that of COMT.
Table II.--MoNoAMnce OmDAS~. (MAO) AND CA'I"ECItOL-OMETHYL TRANSFERASZ (COMT) AcTrvrrt~s IN SOMZ TmsuEs oF Salmo gairdneri Tissue
Liver (3) Kidney (4) Heart (4) White muscle (4) Red muscle (4) Brain (4) Gill (4)
MAO AeTvCIa~ (U/g.)
COMT A~rrvrrY (U/g.)
60.0 (35"5- 11o'7) 128.2 -1-43'° 23"7 -+6"9 3I'9_+ I I. 5 35'3 + x2"6 98"3 _+5"2 52"8+ x2"4
I04X.2 (85I'O- 14o2"0 73"x _+x4"5 91.4 + 8.6 22"I + 3"0 2 x.4_+4"6 57 .8 + 5"9
37.5+8.2
I974, 5
TROUT CATECHOLAMINE METABOLISM
C O M T activity is especially high in the liver and represents 15 times the activity of the kidney and 5 ° times the activity o f the red muscle. T h e prevalence of the liver in this respect is similar to that described in mammals. (Axelrod & Tomchick, 1958; Molinoff & Axelrod, 197 i). For instance, in the rat, the C O M T activity of the liver is three times the activity of the kidney and 35 times the activity of the heart. T h e C O M T activity of the trout liver compared with the M A O activity of the kidney (where it is the most important), shows that 0-methylation plays a leading part in the catabolism of catecholamines. T h e present results bring confirmation of data we obtained by other methods in different species of fish in previous works (Mazeaud, i969; I971; Mazeaud & Mazeaud, 1973) and emphasize the role o f 0-methylation in fish as in mammals (Kopin, I96O ) for physiological inactivation of catecholamines. However, the existence of M A C in a significant a m o u n t in the trout brain m a y imply a correlation between this enzyme and the neuro-adrenergic function. In this respect, structures containing M A C were described in the hypothalamo-hypophysial system of several species o f fish (Urano, 1971 ). T h e function of these structures remains to be studied.
ACKNOWLEDGEMENTS This work was supported by the Centre National de la Recherche Scientifique, Paris (Associated Laboratory n ° 9o). I thank Miss J. Marchal for her technical assistance.
253
AXELROD,J., &: TOMCmOK,R. 0958), ' Enzymatic O-methylation of epinephrine and other catechols ', 07. Biol. Chem., 233, 7oo-7o5. BLASCa~O, H. (1952), ' Amine oxidase and amine metabolism ', Pharmac. Rev., 4, 415-458. BROCH, O. J. (Jr.), & GULDSRRO,H. C. (I971), ' On the determination of catechol-O-methyl transferase activity in tissue homogenates ', Acta Pharmac. Tox., 3o, 266-277. CRotrr, J. R. (196i), ' Inhibition of catechol-Omethyl transferase by pyrogallol in the Rat ', Biochem. Pharmac., 6~ 47-54. DRUJAU, B. D., & Dmz BoRors, J. M. (x97a), ' Adrenaline-induced changes in the metabolism of glycogen in the teleost retina ', 07. Fish. Biol., 4, 79-85. EOASHX~, T., TAKANO, K., Smmzu, K., KUROSAWA,Y., & KAMIJO, K. (I971), 'Effect ofsodium nitrite on monoamine oxidase activity in rat liver and brain ', 07ap. 07. Pharmax., ax, 274. Gmvom, D. (197o), 'Dosage de la Monoamine oxydase ', T h e e 3i~me Cycle Orsay, Paris. KoPm, I . J . (I96o), ' Technique for the study of alternate pathways; epinephrine metabolism in man ', Science, N.T., x3x, 1372. MAC~, R. E. (1965) , ' Microdetermlnation of catechol-O-methyl tramfera~e ', Life Sci., 4, 2353-a359. MAzEAtm, M. (x9.69)., ' Adrdnaline : catabolites et cxcrdtion
urmmre
chez
un
Tdleostden,
Salmo gairdner ', C. R. Hebd. S~anc. Acad. Sd., Paris, a68, IO8--I3I. MAZEAUD, M. (197I), 'Recherches star la biosynth6se, la sdcretion et le catabolisme de l'adrdnaline et de la noradrdnaline chez quelques esp6ces de Cyclostomes et de Poissons ', Th6se Doctorat 6s-Sciences, Universitd de Paris. MAZZAtm, M. & MaZ~AUD, F., (1973) , ' Excrdtion and catabolism of catecholamines in Fish. Part I--Excretion rates; Part II--Catabolites ', Corap. Gen. Pharmac., 4, I83-186. Mozasovv, P. B., & AXELROD,J. (I97t), 'Biochemistry ofcatecholamlnes', A. Rev. B/cohere., 4 0 , 465-5oo. UgANO, A. (t97Q, ' Monoamine oxidase in the hypothalamo-hypophysial region of the Brown Smooth Dogfish, Triakis sryllia ', Endocr. 07ap., t S , 37-46. WUgTM.~, ~q.. J., & AXXLROD, J . (t963), ' A sensitive and specific assay for ~he estimation of monoamine oxidase ', B/cohere. Pharmac., xa~ I439-I441. WtmTUAN, R.J., & AmZLROD,J. (1966), ' Control of enzymatic synthesis of adrenaline in the adrenal medulla by adrenal cortical steroids ', 07. Biol. Chem., suit ~23ox-23o5.
REFERENCES AssmoT, M., & BOHUON,C. (1969) , ' A simple and rapid fluorimetric determination of catechol0-methyl transferase activity ', Life Sd., 8, 93-xoo. AXXLROD, J. (1957) , ' 0-Methylation of epinephrine and other cateehols in vitro and in vivo', Science, aV.T., 126, 4oo-4ox. AX.ELROD, J., ALBEm, W., & CLEM~NTE, C. D. (1959), Distribution of eatechol-O-methyl transferase in the nervous system and other tissues ', o7..N'eurochern.,5, 68-72. AXELROD, J., SENOH, S., ~" WITKOP, B. (1958), l f ~ Word Index: Monoami~e oxidase, catechol'0-mcthylation of catecholamines /n vioo ', O-methyl transfcrase, Tcleosts, heart, liver, 07. Biol. Chem., 233 , 697-7ox. kidney, brain, muscles, gill, •a/mo gairdneri.
251
MONOAMINE OXIDASE AND CATECHOL-O-METHYL TRANSFERASE IN TISSUES OF RAINBOW T R O U T
(SALMO GAIP~NERI RICH., TELEOST) M A D E L E I N E M. M A Z E A U D Institut Ocdanographique, i95 rue Saint-Jacques, 75--o05 Paris, France
(Received 26 October 1973) ABSTRACT I. The catabolism of catecholamines is performed in several tissues of the rainbow trout. 2. Monoamine oxidase activity is important in the kidney and brain. 3. O-Methylation occurs mainly in the liver and seems to be the major route for catecholamine degradation. IN fish, two i m p o r t a n t mechanisms for catecholamine inactivation are deamination and 0 - m e t h y l a t i o n (Mazeaud, i969, i97i ; M a z e a u d & Mazeaud, i973). Both routes of catabolism are the same as those described in m a m m a l s , and are catalysed by monoamine oxidase ( M A O ) and catechol-0methyl transferase ( C O M T ) , as was demonstrated in m a m m a l s (Blaschko, I952 ; Axelrod, i957; Axelrod, Senoh & Witkop, i958; Axelrod & Tomchick, i958; Molinoff & Axelrod, 197t ). I n fish, according to our knowledge, no work on this subject has been reported, except for the histological evidence of M A O in the retina of a Teleost, Eugerres plumieri (Drujan & Diaz Borges, I972 ) and in the hypothalamus o f several Teleosts (Urano, I971 ). I n this preliminary work, we studied M A O and C O M T activity in several tissues of a Teleost fish, the rainbow trout (Salmo gairdneri Rich.). MATERIALS AND METHODS The rainbow trout we used came from piscicultures of the Paris region. We sampled the heart and brain in toto, the cephalic third of the kidney, pieces of liver, white or red muscle and the whole gill on the right side after eliminating bones. After sampling, every tissue was weighed, ground and maintained at + 4 ° C until it was incubated. MAO activity was determined by incubating tissue homogenates with [x'C] tryptamine ([2-x*C]tryptamine succinate (New England
Nuclear), 50 mCi./mmole), according to Wurtman & Axelrod (x963). Theamountof[l*C]indolacetic acid resulting from deamlr~ation of tryptamine was measured. A small amount of [t4C] tryptamine was extracted with the deaminated radioactive material, and a correction was made for this blank value by incubating [l'C]tryptamine with boiled homogenate. MAO activity is expressed in U/g. (IU = number ofnmoles of [14C]indolacetic acid produced per hour). C O M T activity was determined by incubating tissue extracts with epinephrine and [I'C]Sadenosyl m~thionine (Centre Energie Atomique Saclay, France, 5mCi./mmole) according to Wurtman & Axelrod (t966). The m o u n t s of []*C]metanephrine resulting from 0-methylation of epinephrine was measured. Data were corrected for blank values obtained by omitting epinephrine in the incubation medium. Activity is expressed in U/g. (IU = number of nmoles of [t4C]metanephrine produced per hour). Radioactivity was measured by liquid scintillation, and quenching corrected by the use of an external standard. At a prcllminary step, we studied the validity of mammalian methodology applied to fish (Mazeaud, unpublished results). In terms of our data, the reaction was linear with enzyme eoncentratiod over a range of xo-2oo/zg, liver and xo-5oo frg. heart in the case of MAO, and IO--5oo/zg. liver and io-2ooo/zg, heart in the case of COMT. The best temperature for incubation was shown to be 3o ° C. RESULTS AND DISCUSSION I n the rainbow trout, all tissues are able to achieve d e a m i n a t i o n a n d 0-methylation, and thus contain M A O a n d C O M T . I n the case o f C O M T , we emphasized the need
Table
Comp. Gen. Pharmac.
MAZEAUD
~52 I.--CATECHOL-0-METHYL
TRANSFERASE
(COMT) Ac-nvrrY SHOWING THE REQUIREMENT F o e MgS, IONS IN THE INCUBATION MEDIUM Tms~z
Liver Kidney Heart White muscle Red muscle Brain Gill
AewpcrrY (U/g.) Incubation Incubation medium medium with MgCls without MgCls
COMT
870.4 74"5 68.5 x8. I 32.6 52.0 3 x.8
x9"3 22"0 5"x I "9 8.4 12. i 8.6
for Mg 2+ ions in the incubation medium (Table I) as was shown for mammals (Wurtman & Axelrod, i966 ). SPECIFIC ACTIVITY OF TISstr~s
For M A O , the values vary from 31.9 U/g. in the white muscle to I28.2 U/g. in the kidney. For C O M T , values vary from 21.I U/g. inthc red muscle to Io41.2 U/g. in the liver (Table II). A comparison with mammals appears to bc difficultowing to the variety eitherof methodology or expression of the results. For instancc, inscct Egashira, Takano, Shlrnizu, Kurosawa & Kamijo (1971) use a manornctric method and express M A O activity in /zl.oz/hour/ml, of homogcnatc. For C O M T , other substrates than epinephrine may bc used, e.g., norcpincphrinc (Crout, i96I); 3, 4-dihydroxybcnzoic acid (MacCaman, 1965) or 3, 4-dihydroxypheny I acetic acid
(Assicot & Bohuon, I969; Broch & Guldberg, I97I); the end products of the enzymatic reaction may be measured fluorimetrically (Axelrod & Tomchick, 1958; Crout, 1961 ; Assicot & Bohoun, I969) , or by radiochemical methods (Axelrod Albers & Clemente, i959; MacCaman, I965). From the results obtained by all these authors, it is evident that among mammals, the specific activities of M A O and C O M T depend largely on the species studied. T h e specific activitity of M A O varies from 182 U/g. in the rabbit liver, to I2I i U/g. in the rat liver (Gripois, I97O ). In the liver of trout, the amount of M A O seems to be smaller (6~ U]g.). In the case of C O M T , the specific activity of the fiver is too U]g. in the rabbit, 4oo U]g. in man, 33oo U/g. in the cow and 5ooo U/g. in the rat. Thus, a variability factor from I to 5 ° appears. O u r results demonstrate that the value we found for C O M T activityin the trout liver (i o4o.2 U]g.) is integrated in this scale of values. DISTRIBUTION
In the rainbow trout, the most important M A O activity is in the kidney and brain. The major role of the kidney for M A O activity is also reported in several species of mammals like the rabbit, while in other species like the rat, the heart or liver possess the highest M A O activities (Gripois, I97o ). However, in the trout, the M A O ratio of the kidney to M A O of any other tissue studied is at most 6. The M A O distribution is therefore more uniform than that of COMT.
Table II.--MoNoAMnce OmDAS~. (MAO) AND CA'I"ECItOL-OMETHYL TRANSFERASZ (COMT) AcTrvrrt~s IN SOMZ TmsuEs oF Salmo gairdneri Tissue
Liver (3) Kidney (4) Heart (4) White muscle (4) Red muscle (4) Brain (4) Gill (4)
MAO AeTvCIa~ (U/g.)
COMT A~rrvrrY (U/g.)
60.0 (35"5- 11o'7) 128.2 -1-43'° 23"7 -+6"9 3I'9_+ I I. 5 35'3 + x2"6 98"3 _+5"2 52"8+ x2"4
I04X.2 (85I'O- 14o2"0 73"x _+x4"5 91.4 + 8.6 22"I + 3"0 2 x.4_+4"6 57 .8 + 5"9
37.5+8.2
I974, 5
TROUT CATECHOLAMINE METABOLISM
C O M T activity is especially high in the liver and represents 15 times the activity of the kidney and 5 ° times the activity o f the red muscle. T h e prevalence of the liver in this respect is similar to that described in mammals. (Axelrod & Tomchick, 1958; Molinoff & Axelrod, 197 i). For instance, in the rat, the C O M T activity of the liver is three times the activity of the kidney and 35 times the activity of the heart. T h e C O M T activity of the trout liver compared with the M A O activity of the kidney (where it is the most important), shows that 0-methylation plays a leading part in the catabolism of catecholamines. T h e present results bring confirmation of data we obtained by other methods in different species of fish in previous works (Mazeaud, i969; I971; Mazeaud & Mazeaud, 1973) and emphasize the role o f 0-methylation in fish as in mammals (Kopin, I96O ) for physiological inactivation of catecholamines. However, the existence of M A C in a significant a m o u n t in the trout brain m a y imply a correlation between this enzyme and the neuro-adrenergic function. In this respect, structures containing M A C were described in the hypothalamo-hypophysial system of several species o f fish (Urano, 1971 ). T h e function of these structures remains to be studied.
ACKNOWLEDGEMENTS This work was supported by the Centre National de la Recherche Scientifique, Paris (Associated Laboratory n ° 9o). I thank Miss J. Marchal for her technical assistance.
253
AXELROD,J., &: TOMCmOK,R. 0958), ' Enzymatic O-methylation of epinephrine and other catechols ', 07. Biol. Chem., 233, 7oo-7o5. BLASCa~O, H. (1952), ' Amine oxidase and amine metabolism ', Pharmac. Rev., 4, 415-458. BROCH, O. J. (Jr.), & GULDSRRO,H. C. (I971), ' On the determination of catechol-O-methyl transferase activity in tissue homogenates ', Acta Pharmac. Tox., 3o, 266-277. CRotrr, J. R. (196i), ' Inhibition of catechol-Omethyl transferase by pyrogallol in the Rat ', Biochem. Pharmac., 6~ 47-54. DRUJAU, B. D., & Dmz BoRors, J. M. (x97a), ' Adrenaline-induced changes in the metabolism of glycogen in the teleost retina ', 07. Fish. Biol., 4, 79-85. EOASHX~, T., TAKANO, K., Smmzu, K., KUROSAWA,Y., & KAMIJO, K. (I971), 'Effect ofsodium nitrite on monoamine oxidase activity in rat liver and brain ', 07ap. 07. Pharmax., ax, 274. Gmvom, D. (197o), 'Dosage de la Monoamine oxydase ', T h e e 3i~me Cycle Orsay, Paris. KoPm, I . J . (I96o), ' Technique for the study of alternate pathways; epinephrine metabolism in man ', Science, N.T., x3x, 1372. MAC~, R. E. (1965) , ' Microdetermlnation of catechol-O-methyl tramfera~e ', Life Sci., 4, 2353-a359. MAzEAtm, M. (x9.69)., ' Adrdnaline : catabolites et cxcrdtion
urmmre
chez
un
Tdleostden,
Salmo gairdner ', C. R. Hebd. S~anc. Acad. Sd., Paris, a68, IO8--I3I. MAZEAUD, M. (197I), 'Recherches star la biosynth6se, la sdcretion et le catabolisme de l'adrdnaline et de la noradrdnaline chez quelques esp6ces de Cyclostomes et de Poissons ', Th6se Doctorat 6s-Sciences, Universitd de Paris. MAZZAtm, M. & MaZ~AUD, F., (1973) , ' Excrdtion and catabolism of catecholamines in Fish. Part I--Excretion rates; Part II--Catabolites ', Corap. Gen. Pharmac., 4, I83-186. Mozasovv, P. B., & AXELROD,J. (I97t), 'Biochemistry ofcatecholamlnes', A. Rev. B/cohere., 4 0 , 465-5oo. UgANO, A. (t97Q, ' Monoamine oxidase in the hypothalamo-hypophysial region of the Brown Smooth Dogfish, Triakis sryllia ', Endocr. 07ap., t S , 37-46. WUgTM.~, ~q.. J., & AXXLROD, J . (t963), ' A sensitive and specific assay for ~he estimation of monoamine oxidase ', B/cohere. Pharmac., xa~ I439-I441. WtmTUAN, R.J., & AmZLROD,J. (1966), ' Control of enzymatic synthesis of adrenaline in the adrenal medulla by adrenal cortical steroids ', 07. Biol. Chem., suit ~23ox-23o5.
REFERENCES AssmoT, M., & BOHUON,C. (1969) , ' A simple and rapid fluorimetric determination of catechol0-methyl transferase activity ', Life Sd., 8, 93-xoo. AXXLROD, J. (1957) , ' 0-Methylation of epinephrine and other cateehols in vitro and in vivo', Science, aV.T., 126, 4oo-4ox. AX.ELROD, J., ALBEm, W., & CLEM~NTE, C. D. (1959), Distribution of eatechol-O-methyl transferase in the nervous system and other tissues ', o7..N'eurochern.,5, 68-72. AXELROD, J., SENOH, S., ~" WITKOP, B. (1958), l f ~ Word Index: Monoami~e oxidase, catechol'0-mcthylation of catecholamines /n vioo ', O-methyl transfcrase, Tcleosts, heart, liver, 07. Biol. Chem., 233 , 697-7ox. kidney, brain, muscles, gill, •a/mo gairdneri.