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 α_1A-adrenergic receptors (CHO-α_1A). 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-α_1Amicroinjected with G₁₃G225A were smaller than those in CHO-α_1A. In contrast, the magnitudes of norepinephrine-induced extracellular Ca²⁺influx in CHO-α_1Amicroinjected with G₁₂G228A were similar to those in CHO-α_1A. 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 G_q/PLC-pathway is not involved in NSCCs activation by norepinephrine, 2) G₁₃couples with CHO-α_1Aand 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-α_1A.