Cytospectrophotometric study of the kinetics of histochemical reaction of GABA-transaminase in cryostat sections of rat cerebellar cortex

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Abstract
A modification of the histochemical method for demonstration of GABA-transaminase is proposed. Substrate and cofactor concentrations are chosen on the basis of a kinetic investigation in cryostat sections of the rat cerebellar cortex. Enzymatic reactions were measured by quantitative microspectrophotometry. Michaelis constants for α-oxoglutarate in the Purkinje cell layer and granular layer (Km 1.7×10−3 M) and white matter (Km 3,8×10−3 M) are found. It is shown that α-oxoglutarate in concentrations higher than 5.2×10−3 M (1 mg/ml) suppresses the reaction in sections by competitive inhibition. The advisability of addition of malonate, PMS and cyanide to the incubation medium is confirmed. It is suggested that there are some isoenzymes of GABA-transaminase with predominant localization either in neurons or glia.