Ca2+ signaling through the nicotinic acetylcholine receptor α5 subunit in human pulmonary arterial smooth muscle cells

In pulmonary arterial smooth muscle cells (PASMCs), elevations in cytosolic Ca 2+ concentration ([Ca 2+ ] cyt ) are essential for physiological processes such as cell contraction and viability, suggesting the importance of Ca 2+ signaling in PASMC function. In the present study, the functional expression of nicotinic acetylcholine receptors (nAChRs) in human PASMCs was investigated using quantitative real-time PCR, Western blotting, whole-cell patch-clamp recordings, cytosolic Ca 2+ imaging, and cell viability assays. Expression analyses revealed that the α5 subunit of nAChR (nAChRα5) is abundantly expressed in human PASMCs. At a holding potential of -60 mV, nicotine evoked rapidly activating and inactivating inward currents in human PASMCs, which were markedly suppressed by hexamethonium, a nonselective nAChR antagonist. Similar nicotine-induced and hexamethonium-sensitive currents were observed in HEK293 cells expressing human nAChRα5. Nicotine also induced a concentration-dependent increase in [Ca 2+ ] cyt (EC 50 = 395 µM), which was inhibited by hexamethonium. Both nicotine-induced inward currents and [Ca 2+ ] cyt elevations were attenuated by siRNA-mediated knockdown of nAChRα5. Furthermore, knockdown of nAChRα5 significantly reduced PASMC viability. Collectively, these findings demonstrate that nAChRα5 is highly expressed and functionally active in PASMCs, where it mediates nicotine-induced Ca 2+ signaling and regulates cell viability, suggesting a physiological role for this subunit in pulmonary vascular function.