Selective modulation of different GABA_A receptor isoforms by diazepam and etomidate in hippocampal neurons

Diazepam modulation of native γ2-containing GABA_A (γGABA_A) receptors increases channel conductance by facilitating protein interactions involving the γ2-subunit amphipathic (MA) region, which is found in the cytoplasmic loop between transmembrane domains 3 and 4 (Everitt et al., 2009). However, many drugs, predicted to act on different GABA_A receptor subtypes, increase channel conductance leading us to hypothesize that conductance variation in GABA_A receptors may be a general property, mediated by protein interactions involving the cytoplasmic MA stretch of amino acids. In this study we have tested this hypothesis by potentiating extrasynaptic GABA_A currents with etomidate and examining the ability of peptides mimicking either the γ2- or δ-subunit MA region to affect conductance. In inside-out hippocampal patches from newborn rats the general anesthetic etomidate potentiated GABA currents, producing either an increase in open probability and single-channel conductance or an increase in open probability, as described previously (Seymour et al., 2009). In patches displaying high conductance channels application of a δ_(392-422) MA peptide, but not a scrambled version or the equivalent γ2_(381-403) MA peptide, reduced the potentiating effects of etomidate, significantly reducing single-channel conductance. In contrast, when GABA currents were potentiated by the γ2-specific drug diazepam the δ MA peptide had no effect. These data reveal that diazepam and etomidate potentiate different extrasynaptic GABA_A receptor subtypes but both drugs modulate conductance similarly. One interpretation of the data is that these drugs elicit potentiation through protein interactions and that the MA peptides compete with these interactions to disrupt this process.