β‐Adrenergic receptor stimulation of L‐type Ca2+ channels in rabbit portal vein myocytes involves both αs and βγ G protein subunits

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Abstract
Previous studies have shown that purified G protein αs and βγ subunits stimulate vascular L-type Ca2+ channels through protein kinase A and C (PKA and PKC), respectively. The present study tested whether activation of endogenous G proteins via β-adrenergic receptor binding also stimulates vascular Ca2+ channels through both Gαs and Gβγ and the subsequent activation of PKA and PKC. Peak Ba2+ current (IBa) in freshly isolated rabbit portal vein smooth muscle cells was significantly increased by bath application of 0.5 μm isoproterenol (isoprenaline; ISO) when measured using the whole-cell patch clamp method (53 ± 3 % increase, n = 15). Stimulation of IBa by ISO was partially reversed by a PKA inhibitor, KT 5720, or a PKC inhibitor, calphostin C, and completely blocked when cells were pretreated with both KT 5720 and calphostin C. Dialysis of cells with polyclonal antibody to Gαs significantly reduced but did not completely eliminate ISO-induced stimulation of IBa. The remaining stimulation was abolished by calphostin C. Dialysis of cells with a polyclonal antibody to Gβ also significantly reduced ISO-induced stimulation and the remaining stimulation was abolished by KT 5720. Dialysis of cells with both antibodies completely prevented the stimulation of IBa by ISO. ISO-induced stimulation of IBa was reversed by ICI-118,551, a specific β2-adrenoceptor antagonist, but not by CGP 20712A, a specific β1-adrenoceptor antagonist. In addition, the β2-adrenoceptor agonist zinterol significantly increased peak IBa while the β1-adrenoceptor agonist dobutamine and β3-adrenoceptor agonist BRL 37344A had little effect on peak IBa. These data suggest that β-adrenergic receptor stimulation of vascular L-type Ca2+ channels involves both αs and βγ G-protein subunits, which exert their effects through PKA and PKC, respectively.