Pharmacological characterization of native a7 nicotinic ACh receptors and their contribution to depolarization-elicited exocytosis in human chromaffin cells

BACKGROUND AND PURPOSE Expression of a7 nicotinic acetylcholine receptors (nAChRs) and their role in exocytosis have not yet been examined in human chromaffin cells. EXPERIMENTAL APPROACH To characterize these receptors and investigate their function, patch-clamp experiments were performed in human chromaffin cells from organ donors. KEY RESULTS The nicotinic current provoked by 300 mu M ACh in voltage-clamped cells was blocked by the nicotinic receptor antagonists a-bungarotoxin (alpha-Bgtx; 1 mM; 6 +/- 1.7%) or methyllycaconitine (MLA; 10 nM; 7 +/- 1.6%), respectively, in an irreversible and reversible manner, without affecting exocytosis. Choline (10 mM) pulses induced a biphasic current with an initial quickly activated (5.5 +/- 0.4 ms rise time) and inactivated component (8.5 +/- 0.4 ms time constant) (termed alpha 7), which was blocked by a-Bgtx or MLA, followed by a slower component (non-alpha 7). a7 nAChR currents were dissected by blocking the non-a7 nAChR current component of the ACh and choline response with the alpha 6* nAChR blocker a-conotoxin (alpha-Ctx) MII[S4A, E11A, L15A]. PNU-282987, an a7 nAChR-specific agonist, elicited rapidly activated and rapidly inactivated currents. a7 nAChR-positive allosteric modulators, such as 5-hydroxyindole (1 mM) and PNU-120596 (10 mM), potentiated responses that were blocked by a-Bgtx or MLA. Exocytosis was evoked by depolarization-elicited a7 nAChR currents, using choline in the presence of a-Ctx MII[MS4A, E11A, L15A] or PNU-282987 as agonists. CONCLUSIONS AND IMPLICATIONS Our electrophysiological recordings of pure a7 nAChR currents elicited by rapid application of agonists demonstrated that functional a7 nAChRs are expressed and contribute to depolarization-elicited exocytosis in human chromaffin cells.