Hydroxyt­ryptophan-decarboxylase activity in nerve endings of the rat brain.

G. RODRÍGUEZ DE LORES ARNAIZ; E. DE ROBERTIS

JOURNAL OF NEUROCHEMISTRY

WILEY-BLACKWELL PUBLISHING, INC

Año: 1964 vol. 11 p. 213 - 219

0022-3042

Journal of Neurochemistry, 1964. Vol. 11, pp. 213 to 219. Pergamon Press Ltd. Printed in Northern Ireland 5-HYDROXYTRYPTOPHAN DECARBOXYLASE ACTIVITY IN NERVE ENDINGS OF THE RAT BRAIN* GEORGINA RODRIGUEZ DE LORES ARNAIZ and E. DE ROBERTIS Instituto de Anatomia General y Embriologfa, Facultad de Ciencias Mkdicas, Buenos Aires, Argentina (Received 14 August 1963) IN RECENT years the localization in nerve endings and synaptic vesicles, in addition to the physiological action, has become one of the main criteria for identifying a substance as a ?transmitter? in brain. In a study of the ACh-system in rat brain the three main components ACh, AChE (DE ROBERTIS, PELLEGRINO DE IRALDI, RODRIGUEZ DE LORES ARNAIZ and SALGANICOFF, 1962u), and ChAc (DE ROBERTIS, RODRIGUEZ DE LORES ARNAIZ, SALGANICOFF, PELLEGRINO DE IRALDI and ZIEHER, 1963), were found to be concentrated in a special population of nerve endings contained in a submitochondrial fraction. Under the electron microscope a synaptic complex was recognized which comprises the nerve ending with the two synaptic membranes attached by the intersynaptic filaments and the subsynaptic web (DE ROBERTIS et ul., 1961). The finer localization of the ACh-system within the synaptic complex showed that while both ACh and ChAc were mainly related to the synaptic vesicles, AChE appeared to be mostly concentrated in the synaptic membranes. Similar studies on two components of the 5-HT-system, MA0 and 5-HT, showed that the inactivating enzyme is exclusively localized in mitochondria, following exactly the distribution of SDH (RODRIGUEZ DE LORES ARNAIZ and DE ROBERTIS, 1962), and 5-HT is concentrated in microsomes and in the same population of nerve endings containing the ACh-system (ZIEHER and DE ROBERTIS, 1963). Upon disintegration of the synaptic complex by osmotic shock (DE ROBERTIS et al., 19623) MA0 remained within the mitochondria and 5-HT was evenly distributed among the resulting fractions. A study of the localization of 5-HTP-D, the enzyme that forms 5-HT by decarboxylation of 5-HTP (UDENFRIEND, CLARK and TITUS, 1953), first seemed hopeless in view of the reports in the literature indicating that this is a completely soluble enzyme (CLARK, WEISSBACH and UDENFRIEND, 1954; BOGDANSKI, WEISSBACH and UDENFRIEND, 1957; UDENFRIEND, BOGDANSKI and WEISSBACH, 1957; PAGE, 1958; ROSENGREN, 1960). But in preliminary experiments on rat brain, in which we used the homogenization technique currently employed in our laboratory, we * This paper was supported by the Consejo Nacional de Investigaciones Cientificas y Tecnicas, Buenos Aim, Argentina and Grant NB 03991 of the National Institute of Health, U.S.A. Abbreviations used: AChE, acetylcholinesterase; ChAc, cholineacetylase; MAO, monoamineoxidase ; SDH, succinate dehydrogenase; SHTP-D, 5-hydroxytryptophan decarboxylase ; 5-HTP, 5-hydroxytryptophan; Hp, total particulate fraction; Hs, total soluble supernatant ; N, nuclear fraction; Mit, crude mitochondria1 fraction; Mic, rnicrosomal fraction; Sup, final supernatant; NA, noradrenaline; RSA, relative specific activity. 1 213 214 GEORGINA RODRIGUEZ DE LORES ARNAIZ and E. DE ROBERTIS found that only about 50 per cent of the 5-HTP-D was soluble and the rest remained in the particulate fraction. In view of this finding it seemed of interest to follow the content of this ´bound´ 5-HTP-D in the different subcellular fractions of brain.