Allopregnanolone Activates GABAA Receptor/Cl- Channels in a Multiphasic Manner in Embryonic Rat Hippocampal Neurons

Laboratory of Neurophysiology, National Institute of Neurological Disorders and Stroke, National Institutes of Health, Bethesda, Maryland 20892 Liu, Qi-Ying, Yoong H. Chang, Anne E. Schaffner, Susan V. Smith, and Jeffery L. Barker. Allopregnanolone Activates GABA A Receptor/Cl Channels in a Multiphasic Manner in Embryonic Rat Hippocampal Neurons. J. Neurophysiol. 88: 1147-1158, 2002. Although 3 -substituted metabolites of progesterone are well established to interact with GABA A receptor/Cl channels, the nature of the interaction(s) remains uncertain. We used patch-clamp recording to study the interaction with GABA A receptor/Cl channels expressed by embryonic hippocampal neurons differentiating in culture and nonneuronal cells transfected with GABA A receptor subunits. Allopregnanolone primarily induced multiphasic current responses in neurons, which were eliminated by bicuculline, an antagonist of GABA at GABA A receptor/Cl channels. Similar multiphasic responses blocked by bicuculline were induced by allopregnanollone in nonneuronal cells transfected with 1 and 2 subunits, indicating that the steroid activation of GABA A receptor/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 1 and 2 or 3 subunits, 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 GABA A receptor/Cl channels expressed by embryonic hippocampal neurons in multiple ways, some of which are mediated by G proteins.