Phenylephrine acts via IP3-dependent intracellular NO release to stimulate L-type Ca2+ current in cat atrial myocytes

This study determined the effects of alpha(1)-adrenergic receptor (alpha(1)-AR) stimulation by phenylephrine (PE) on L-type Ca (2+) current (I-Ca,I-L) in cat atrial myocytes. PE (10 mu m) reversibly increased I-Ca,I-L (51.3%; n = 40) and shifted peak I-Ca,I-L activation voltage by - 10 mV. PE-induced stimulation Of I-Ca,I-L was blocked by each of I mu m prazocin, 10 mu m (L)-NIO, 10 mu m W-7, 10 mu m ODQ, 2 mu m H-89 or 10 mu m LY294002, and was unaffected by 10 mu m chelerythrine or incubating cells in pertussis toxin (PTX). PE-induced stimulation Of I-Ca,I-L also was inhibited by each of 10 mu m ryanodine or 5 Am thapsigargin, by blocking IP3 receptors with 2 mu m 2-APB or 10 mu m xestospongin C or by intracellular dialysis of heparin. In field-stimulated cells, PE increased intracellular NO (NOi) production. PE-induced NOi release was inhibited by each of I Am prazocin, 10 AM L-NIO, 10 Am W-7, 10 Am LY294002, 2 Am H-89, 10 Am ryanodine, 5 Am thapsigargin, 2 Am 2-APB or 10 Am xestospongin C, and unchanged by PTX. PE (10 Am) increased phosphorylation of Akt, which was inhibited by LY294002. Confocal microscopy showed that PE stimulated NOi release from subsarcolemmal sites and this was prevented by 2 mm methyl-beta-cyclodextrin, an agent that disrupts caveolae formation. PE also increased local, subsarcolemmal. SR Ca2+ release via IP3-dependent signalling. Electron micrographs of atrial myocytes show peripheral SR cisternae in close proximity to clusters of caveolae. We conclude that in cat atrial myocytes PE acts via alpha(1)-ARs coupled to PTX-insensitive G-protein to release NOi, which in turn stimulates I-Ca,I-L- PE-induced NOi release requires stimulation of both PI-3K/Akt and IP3-dependent Ca2+ signalling. NO stimulates ICa,L via cGMP-mediated cAMP-dependent PKA signalling. IP3-dependent Ca2+ signalling may enhance local SR Ca2+ release required to activate Ca2+ -dependent eNOS/NOi production from subsarcolemmal caveolae sites.