Na +-independent binding of GABA to the triton X-100 treated synaptic membranes from cerebellum of rat brain

Abstract

The treatment of the membranes from cerebellum of rat brain with 0.5% Triton X-100 increases both the affinity and the density of the Na +-independent binding sites for 3H-GABA (γ-aminobutyric acid) from the values obtained from membranes of rat brain after an extensive freezing and thawing treatment (Young et al. , 1976). Upon repeated washings of the Triton-treated membranes, the binding of 3H-GABA is further increased and follows biphasic kinetics which indicates two binding components having dissociation constants of 5.9 and 27 nM and densities of 1.35 and 3.9 pmole/mg protein, respectively. GABA agonist, imidazoleacetic acid, and the GABA antagonists, bicuculline and d-tubocurarine, inhibit 50% of 3H-GABA binding at 1, 47 and 85 μM concentrations (IC 50 values), respectively. The IC 50 values for these compounds are unchanged by Na +. Thus, the Na +-independent binding of 3H-GABA to the Triton-treated membranes may represent binding to the synaptic GABA receptors.