Abstract

Endogenous neurosteroids have rapid actions on ion channels, particularly GABA_A receptors, which are potentiated by nanomolar concentrations of 3α-hydroxypregnane neurosteroids. Previous evidence suggests that 3β-hydroxypregnane steroids may competitively antagonize potentiation induced by their 3α diastereomers. Because of the potential importance of antagonists as experimental and clinical tools, we characterized the functional effect of 3β-hydroxysteroids. Although 3β-hydroxysteroids reduced the potentiation induced by 3α-hydroxysteroids, 3β-hydroxysteroids acted noncompetitively with respect to potentiating steroids and inhibited the largest degrees of potentiation most effectively. Potentiation by high concentrations of barbiturates was also reduced by 3β-hydroxysteroids. 3β-Hydroxysteroids are also direct, noncompetitive GABA_A receptor antagonists. 3β-Hydroxysteroids coapplied with GABA significantly inhibited responses to ≥15 μm GABA. The profile of block was similar to that exhibited by sulfated steroids, known blockers of GABA_A receptors. This direct, noncompetitive effect of 3β-hydroxysteroids was sufficient to account for the apparent antagonism of potentiating steroids. Mutated receptors exhibiting decreased sensitivity to sulfated steroid block were insensitive to both the direct effects of 3β-hydroxysteroids on GABA_Aresponses and the reduction of potentiating steroid effects. At concentrations that had little effect on GABAergic synaptic currents, 3β-hydroxysteroids and low concentrations of sulfated steroids significantly reversed the potentiation of synaptic currents induced by 3α-hydroxysteroids. We conclude that 3β-hydroxypregnane steroids are not direct antagonists of potentiating steroids but rather are noncompetitive, likely state-dependent, blockers of GABA_Areceptors. Nevertheless, these steroids may be useful functional blockers of potentiating steroids when used at concentrations that do not affect baseline neurotransmission.