Characterization of the GABA A receptor in the brain of the adult male bullfrog, Rana catesbeiana

Abstract

Little is known about the properties of GABA receptors in the amphibian brain. The GABA A receptor is widespread in the mammalian brain, and can be specifically labeled with the receptor agonist [ 3H]muscimol. The binding of [ 3H]muscimol to membrane preparations from the brain of the bullfrog, Rana catesbeiana, was investigated in kinetic, saturation, and inhibition experiments to determine whether this species possessed a GABA A-like receptor. Binding of 20 nM [ 3H]muscimol to membranes was specific and could be displaced by 1 mM GABA. Association binding curves showed that steady state occurred rapidly, within 2 min, and dissociation occurred within 5 min. The receptor was saturable with a single, high-affinity binding site ( K D=19.2 nM; B max=1.8 pmol/mg protein). Binding of [ 3H]muscimol was inhibited in a dose-dependent fashion by muscimol, GABA, bicuculline methiodide, and bicuculline (in order of potency). Baclofen (at doses from 10 −9 to 10 −3 M) failed to displace [ 3H]muscimol. The binding characteristics and ligand specificity of [ 3H]muscimol binding sites in the bullfrog brain support the hypothesis that this amphibian possesses a GABA A-like receptor protein similar to the GABA A receptor characterized in mammals.