Allopregnanolone Activates GABAAReceptor/Cl−Channels in a Multiphasic Manner in Embryonic Rat Hippocampal Neurons

Although 3α-substituted metabolites of progesterone are well established to interact with GABAAreceptor/Cl−channels, the nature of the interaction(s) remains uncertain. We used patch-clamp recording to study the interaction with GABAAreceptor/Cl−channels expressed by embryonic hippocampal neurons differentiating in culture and nonneuronal cells transfected with GABAAreceptor subunits. Allopregnanolone primarily induced multiphasic current responses in neurons, which were eliminated by bicuculline, an antagonist of GABA at GABAAreceptor/Cl−channels. Similar multiphasic responses blocked by bicuculline were induced by allopregnanollone in nonneuronal cells transfected with α1and γ2subunits, indicating that the steroid activation of GABAAreceptor/Cl−channels occurred independently of GABA. Fluctuation analyses of current responses to allopregnanolone and GABA revealed underlying channel activities with similar estimated unitary properties. However, although both agonists activated Cl−channels with similar estimated short and long burst-length durations, most of those stimulated by the steroid were short, while most of those opened by GABA were long. Allopregnanolone potentiated GABA-evoked Cl−currents in nonneuronal cells transfected with α1and β2or β3subunits, which did not exhibit multiphasic responses to the steroid, indicating another, independent action of the steroid at activated receptors. Pertussis toxin treatment eliminated the low-amplitude current and attenuated the high-amplitude current induced by allopregnanolone in a reversible manner. Mastoparan, which activates G proteins directly, triggered a high-amplitude current after a delay, which was blocked by bicuculline. The results indicate that allopregnanolone interacts with GABAAreceptor/Cl−channels expressed by embryonic hippocampal neurons in multiple ways, some of which are mediated by G proteins.