Characterization of chloride efflux from GT1-7 neurons: Lack of effect of ethanol on GABA(A) response

The purpose of this study of GT1-7 neurons was to partially characterize basal Cl^- transport and GABA(A) mediated Cl^- efflux and to test the effect of ethanol on a GABA(A) receptor that lacks a γ subunit. We measured GABA(A) function and Cl^- transport with ³⁶C1-. Our results show that basel ³⁶Cl^- efflux varied with temperature at 4°C, 23°C, and 37°C. At 23°C, DIDS, an inhibitorof anion exchange, reduced basel ³⁶Cl^- efflux maximally by 79.6% with an IC₅₀ of 42.1 μM, wheras burnetanide, an inhibitor of (Na-K-Cl) cotransport, had no effect on basal ³⁶Cl^- efflux at concentrations up to 150 μM. At 4°C, muscimol, a GABA(A) receptor agonist, stimulated ³⁶Cl^- efflux with an EC₅₀ of 1.47 μM. Bicuculline, a GABA(A) receptor antagonist, completely reversed the effect of 20 μM muscimol with an IC₅₀ of 6.08 μM. Ethanol, at concentrations up to 87 mM (0.4% (w/v)), had no effect on muscimol-induced ³⁶Cl^- efflux at 4°C or at 32°C. Our results indicate that stimulation of GABA(A) receptors causes an efflux of Cl^- from GT1-7 neurons. This finding is consistent with the concept that stimulation of GABA(A) receptors produces depolarization of the plasma membrane, increase in cytosolic [Ca²⁺], and GnRH release. Our results represent the first description of chloride transport in GT1-7 neurons and suggest the presence of a Cl^- exchange, but not (Na-K-Cl), transporter mechanism. Furthermore, the lack of an effect of ethanol observed in this study is consistent with the idea that a γ2L subunit may be necessary for the effects of low concentrations of ethanol at GABA(A) receptors.