Progesterone (a delta 4-3 keto-pregnane) as well as a series of pregnanes enhanced [3H]muscimol binding to rat cerebral cortex membranes. This effect was increased by preincubation in the presence of the drugs, progesterone being effective only after a 30 min preincubation. The effect of progesterone was dose-dependent from 1 nM to 100 microM reaching a maximum of 140% over control. Its 20 alpha and 20 beta reduced metabolites (20 alpha- and 20 beta-OH-4-pregnen-3-one) had no effect. Ring A reduction in either the 5 alpha or 5 beta position resulted in steroids (5 alpha- and 5 beta-pregnane-3,20-di-one) producing moderate but consistent facilitation of binding (30-40% increase at 10 microM). The presence of a hydroxyl group in C3 had variable results in the potency of pregnanes to facilitate muscimol binding. Pregnanes with a 3 beta-OH group showed weaker activity than those having a 3 alpha-OH group, 5 alpha-pregnan-3 alpha-ol-20-one being the most potent (100% stimulation). Pregnenolone, a delta 5-3 beta-OH-pregnane, was only effective at low concentrations (10 nM to 10 microM). Scatchard analysis of [3H]muscimol binding in the presence of progesterone and 5 alpha-pregnan-3 alpha-ol-20-one revealed that the observed effect was due to an increase in binding sites rather than to a change in affinity.