Protease-activated receptor-2-mediated Ca2+ signaling in guinea pig tracheal epithelial cells

Abstract The protease-activated receptor-2 (PAR-2), a G protein-coupled receptor activated by trypsin, contributes to the pathogenesis of inflammatory disease including asthma. Here, we examined the mechanisms by which stimulation of PAR-2 induces an increase in intracellular Ca 2+ concentration ([Ca 2+ ]i) in guinea pig tracheal epithelial cells. Trypsin (0.01–3 units/ml) dose-dependently induced a transient increase in [Ca 2+ ]i, the increase being blocked by soybean trypsin inhibitor (SBTI 1 μM). An increase in [Ca 2+ ]i was also induced by an agonist peptide for PAR-2 (SLIGRL-NH 2 , 0.001–10 μM) but not by thrombin (3 units/ml, an activator for PAR-1, PAR-3 or PAR-4). Repeated or cross stimulation of trypsin or SLIGRL-NH 2 caused marked desensitization of the [Ca 2+ ]i response. These responses of [Ca 2+ ]i to trypsin and SLIGRL-NH 2 were attenuated by a phospholipase C inhibitor, U-73122, and a Ca 2+ -ATPase inhibitor, thapsigargin (100 nM), while removal of Ca 2+ and a L-type Ca 2+ -channel blocker, verapamil, were without significant effects. Further, trypsin was without effect on the rate of fura 2 quenching by Mn 2+ entry as an indicator of Ca 2+ influx. Thus, stimulation of PAR-2 appears to increase [Ca 2+ ]i through the mobilization of Ca 2+ from intracellular stores probably via phospholipase Cβ-linked generation of a second messenger.