Effects of drugs on γ-aminobutyric acid receptors, uptake, release and synthesis in vitro

Abstract

The effects of various convulsant and anticonvulsant drugs have been studied using in vitro assays for the postsynaptic action of the neurotransmitter γ-aminobutyric acid (GABA). GABA caused a receptor-ionophore mediated increase in chloride permeability in crayfish muscle. At 100 μ M concentrations, benzyl penicillin, bicuculline, diethyl barbiturate, diazepam, imipramine, and haloperidol partially inhibited this response while picrotoxinin inhibited it 100% (I 50 = 3 μ M). Muscimol (potently) andβ-( p-chlorphenyl) GABA (weakly) mimic GABA action in this assay. Muscimol and bicuculline (potent), and benzyl penicillin andβ-( p-chlorphenyl) GABA (weak), but not the other drugs, probably exerted their effects at the GABA receptor level, because only these four drugs among those tested were inhibitors of GABA receptor binding sites in mouse brain homogenates (sodium-independent sites having the specificity expected of receptor sites). Pentylenetetrazol weakly inhibited GABA receptor binding and GABA-induced chloride flux. Several other convulsants suspected of GABA antagonist activity, such as trimethyl caprolactam, isopropyl bicyclophosphate, tetramethylene disulfotetramine, and D-tubocurarine, showed no direct inhibition of either GABA receptor sites in mouse brain or of GABA-stimulated chloride flux in crayfish muscle. The drugs were also examined for effects on in vitro assays of GABA uptake, glutamate decar☐ylase activity (GAD), and Ca 2+-stimulated GABA release with mouse brain homogenates. GABA uptake by mouse brain particulate fractions was inhibited 50% by approximately 100μ M haloperidol, imipramine, chlorpromazine, diazepam, benzyl penicillin, bicuculline, andβ-( p-chlorophenyl) GABA, but by muscimol only at concentrations near1m M. Diazepam and benzyl penicillin also stimulated Ca 2+-independent efflux of GABA from mouse brain fractions enriched in synaptosomes. Diazepam, chlorpromazine, imipramine, and haloperidol inhibited Ca 2+-dependent release of GABA from brain particles enriched in synaptosomes. No drugs tested were found to inhibit L-glutamate decar☐ylase (GAD) activity at concentrations under100 μ M. The drugs diphenylhydantoin and diethyl barbiturate showed no effects on these GABA neurochemical assays which could be related to anticonvulsant action. Convulsant activity of benzyl penicillin appears to be related to competitive antogonism of GABA receptors, although the drugs is not very potent or specific. Diazepam at concentrations of50–100 μ M affected GABA transport in a manner consistent with anti-convulsant activity, although at similar concentrations it also could inhibit GABA release and postsynaptic action. Lioresal seems to be a definite GABA agonist, although weak and non-specific, while muscimol is a very potent and specific GABA agonist. Analogues of both these compounds are promising candidates for treatment of neurological disorders involving GABA dysfunction.