Dehydroepiandrosterone sulfate (DHEAS) blocked the GABAA receptor noncompetitively in neurons grown in primary culture from the ventral midbrains of fetal rats. The apparent dissociation constant for this blockade was 4.5 microM, and one molecule of DHEAS was sufficient to block the receptor. The affinity of the blocked receptor for GABA was diminished by about one half. The findings that the DHEAS caused no rectification of chloride currents and that it did not shorten the durations of open ion channels indicated that DHEAS did not act by occluding open ion channels. Neither did it diminish their conductance. DHEAS accelerated desensitization in at least one population of receptors, diminished the amplitudes of inhibitory postsynaptic currents, and shortened their decay time constants in a concentration dependent manner.