Diazepam biphasically modulates [3H]TBOB binding to the convulsant site of the GABAA receptor complex

Interactions of GABA, bicuculline methochloride and diazepam with [3H]TBOB binding to rat brain membranes were evaluated in vitro. GABA displaced [3H]TBOB binding with and IC50 of 4 microM and a slope factor near unity. The competitive GABA antagonist bicuculline methochloride shifted the displacement curve of GABA parallelly to the right, indicating that the interaction of GABA with [3H]TBOB binding is of an allosteric nature. In the presence of GABA, diazepam displaced the binding of [3H]TBOB according to a two-site model: a high affinity site with an IC50 of about 50 nM and a lower affinity site with an IC50 of about 30 microM. Bicuculline methochloride abolished the nanomolar displacement by diazepam and increased the micromolar IC50 value. These results indicate that the interaction of the high affinity diazepam site with the [3H]TBOB binding site is totally GABA dependent and that the low affinity effect of diazepam on [3H]TBOB binding is at least partially GABA dependent. It is likely that the low affinity potency of diazepam to displace [3H]TBOB binding has physiological relevance.