Regulation of lung epithelial ATP release

Abstract

Mucociliary clearance, the primary innate defense mechanism of the lung, is controlled by extracellular nucleotides (e.g., ATP) acting on cell surface purinergic receptors. However, the mechanisms by which ATP is released from epithelial cells are poorly understood. To identify signaling elements upstream of ATP release, we examined the effect of GPCR activation on ATP release from lung epithelial A549 cells. The protease activated receptor (PAR) agonist thrombin and the P2Y2‐R agonist UTP induced robust inositol phosphate formation and Ca2+mobilization. RT‐PCR analysis indicated P2Y2‐R and PAR3 mRNA expression. Thrombin elicited a rapid ATP release, which was inhibited by BAPTA‐AM, thapsigargin, or ML‐7 (inhibits Ca2+/CaM‐dependent MLCK). In contrast, UTP caused no ATP release. Thus, signaling elements in addition to Gq/PLCβ/Ca2+may be involved in GPCR‐induced ATP release. PAR‐induced ATP release was decreased by ROCK inhibitors (Y27632 and H1 152), suggesting involvement of Rho/ROCK downstream of G12/13. Mechanical perturbations relevant to the airways (e.g. hypotonic swelling, shear stress) cause robust epithelial ATP release. Pretreatement of cells with ML‐7, Y27632, or H1 152 impaired hypotonicity‐elicited ATP release. Our results suggest that MLCK and Rho activation participate in both receptor‐ and hypotonicity‐induced ATP release from lung epithelial cells.Supported by NIH P01HL34322