γ-aminobutyric acid (GABA) receptor-mediated 36chloride (36Cl-) uptake was measured in synaptoneurosomes from rat brain. GABA and GABA agonists stimulated 36Cl- uptake in a concentration-dependent manner with the following order of potency: Muscimol>GABA> piperidine-4-sulfonic acid (P4S)>4, 5, 6, 7-tetrahydroisoxazolo-[5, 4-c]pyridin-3-ol (THIP)=3-aminopropanesulfonic acid (3APS) >taurine. Both P4S and 3APS behaved as partial agonists, while the GABAB agonist, baclofen, was ineffective. The response to muscimol was inhibited by bicuculline and picrotoxin in a mixed competitive/non-competitive manner. Other inhibitors of GABA receptor-opened channels or non-neuronal anion channels such as penicillin, picrate, furosemide and disulfonic acid stilbenes also inhibited the response to muscimol. A regional variation in muscimol-stimulated 36Cl- uptake was observed; the largest responses were observed in the cerebral cortex, cerebellum and hippocampus, moderate responses were obtained in the striatum and hypothalamus and the smallest response was observed in the pons-medulla. GABA receptor-mediated 36Cl- uptake was also dependent on the anion present in the media. The muscimol response varied in media containing the following anions: Br->Cl->N03->I->SCN->C3H500->C104->F-, consistent with the relative anion permeability through GABA receptorgated anion channels and the enhancement of convulsant binding to the GABA receptor-gated Cl- channel.