Modulation of recombinant α₆β₂γ₂ GABA_A receptors by neuroactive steroids

The sensitivity of the recombinant α₆β₂γ₂ GABA_A receptor expressed in HEK 293 cells to neuroactive steroids was studied. The steroids 3α-hydroxy-5α-pregnan-20-one (3α-OH-DHP), pregnenolone sulfate and 3α-OH-DHP sulfate have different modulatory effects on [³H]muscimol or [³H]Ro15-4513 binding to the α₆β₂γ₂ than to the α₁β₂γ₂ receptor. Binding of both radioactive ligands to the α₆β₂γ₂ receptor was maximally potentiated with each steroid used (10 nM) and decreased with further increases in steroid concentration. Using whole-cell recording, the GABA response of clusters of transfected HEK 293 cells was strongly potentiated by 3 or 10 nM 3α-OH-DHP. In contrast, this response was reduced by 100 nM 3α-OH-DHP. This latter effect appears to be related to the acceleration of the GABA response desensitization, observed in isolated cells. 3α-OH-DHP (10 or 100 nM) was able to activate a response in the absence of GABA. It is proposed that the interaction of neuroactive steroids with the α₆β₂γ₂ receptor involves at least two distinct binding sites. One of them might be located close to the GABA binding site.