Phenylephrine acts via IP 3 ‐dependent intracellular NO release to stimulate L‐type Ca 2+ current in cat atrial myocytes

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