Characterization of G proteins involved in activation of nonselective cation channels and arachidonic acid release by norepinephrine/α1A-adrenergic receptors

Abstract

We demonstrated recently that norepinephrine activates Ca2+-permeable nonselective cation channels (NSCCs) in Chinese hamster ovary cells stably expressing α1A-adrenergic receptors (CHO-α1A). Moreover, extracellular Ca2+through NSCCs plays essential roles in norepinephrine-induced arachidonic acid release. The purpose of the present study was to identify the G proteins involved in the activation of NSCCs and arachidonic acid release by norepinephrine. For these purposes, we used U73122, an inhibitor of phospholipase C (PLC), and dominant negative mutants of G12and G13(G12G228A and G13G225A, respectively). U73122 failed to inhibit NSCCs activation by norepinephrine. The magnitudes of norepinephrine-induced extracellular Ca2+influx in CHO-α1Amicroinjected with G13G225A were smaller than those in CHO-α1A. In contrast, the magnitudes of norepinephrine-induced extracellular Ca2+influx in CHO-α1Amicroinjected with G12G228A were similar to those in CHO-α1A. In addition, neither a Rho-associated kinase (ROCK) inhibitor nor a phosphoinositide 3-kinase inhibitor affected norepinephrine-induced extracellular Ca2+influx. G13G225A, but not G12G228A, also inhibited arachidonic acid release partially. These results demonstrate that 1) the Gq/PLC-pathway is not involved in NSCCs activation by norepinephrine, 2) G13couples with CHO-α1Aand plays important roles for norepinephrine-induced NSCCs activation, 3) neither ROCK- nor PI3K-dependent cascade is involved in NSCCs activation, and 4) G13is involved in norepinephrine-induced arachidonic acid release in CHO-α1A.