Abstract

Pannexin-1 (Panx1) forms plasma membrane channels that allow the exchange of small molecules between the intracellular and extracellular compartments, and are involved in diverse physiological and pathological responses in the nervous system. However, the signaling mechanisms that induce their opening still remain elusive. Here, we propose a new mechanism for Panx1 channel activation through a functional crosstalk with the highly Ca(2+)permeable alpha 7 nicotinic acetylcholine receptor (nAChR). Consistent with this hypothesis, we found that activation of alpha 7 nAChRs induces Panx1-mediated dye uptake and ATP release in the neuroblastoma cell line SH-SY5Y-alpha 7. Using membrane permeant Ca(2+)chelators, total internal reflection fluorescence microscopy in SH-SY5Y-alpha 7 cells expressing a membrane-tethered GCAMP3, and Src kinase inhibitors, we further demonstrated that Panx1 channel opening depends on Ca(2+)signals localized in submembrane areas, as well as on Src kinases. In turn, Panx1 channels amplify cytosolic Ca(2+)signals induced by the activation of alpha 7 nAChRs, by a mechanism that seems to involve ATP release and P2X7 receptor activation, as hydrolysis of extracellular ATP with apyrase or blockage of P2X7 receptors with oxidized ATP significantly reduces the alpha 7 nAChR-Ca(2+)signal. The physiological relevance of this crosstalk was also demonstrated in neuroendocrine chromaffin cells, wherein Panx1 channels and P2X7 receptors contribute to the exocytotic release of catecholamines triggered by alpha 7 nAChRs, as measured by amperometry. Together these findings point to a functional coupling between alpha 7 nAChRs, Panx1 channels and P2X7 receptors with physiological relevance in neurosecretion.