Selective modulation of neuronal nicotinic acetylcholine receptor channel subunits by Go-protein subunits

G-protein modulation of neuronal nicotinic acetylcholine receptor (nAChR) channels in rat intrinsic cardiac ganglia was examined using dialyzed whole-cell and excised membrane patch-recording configurations. Cell dialysis with GTP{gamma}S increased the agonist affinity of nAChRs, resulting in a potentiation of nicotine-evoked whole-cell currents at low concentrations. ACh- and nicotine-evoked current amplitudes were increased approximately twofold in the presence of GTP{gamma}S. In inside-out membrane patches, the open probability (NPo) of nAChR-mediated unitary currents was reversibly increased fourfold after bath application of 0.2 mM GTP{gamma}S relative to control but was unchanged in the presence of GDP{beta}S. The modulation of nAChR-mediated whole-cell currents was agonist specific; currents evoked by the cholinergic agonists ACh, nicotine, and 1,1-dimethyl-4-phenylpiperazinium iodide, but not cytisine or choline, were potentiated in the presence of GTP{gamma}S. The direct interaction between G-protein subunits and nAChRs was examined by bath application of either Go{alpha} or G{beta}{gamma} subunits to inside-out membrane patches and in glutathione S-transferase pull-down and coimmunoprecipitation experiments. Bath application of 50 nM G{beta}{gamma} increased the open probability of ACh-activated single-channel currents fivefold, whereas Go{alpha} (50 nM) produced no significant increase in NPo. Neuronal nAChR subunits {alpha}3-{alpha}5 and {beta}2 exhibited a positive interaction with Go{alpha} and G{beta}{gamma}, whereas {beta}4 and {alpha}7 failed to interact with either of the G-protein subunits. These results provide evidence for a direct interaction between nAChR and G-protein subunits, underlying the increased open probability of ACh-activated single-channel currents and potentiation of nAChR-mediated whole-cell currents in parasympathetic neurons of rat intrinsic cardiac ganglia.