Angiotensin II Upregulates CaV1.2 Protein Expression in Cultured Arteries via Endothelial H₂O₂ Production

Background: We previously reported that angiotensin II caused an endothelial-dependent increase in L-type voltage-dependent Ca²⁺ channel (CaV1.2) in cultured arteries, but the signaling pathways are not clear. Methods: Endothelial damage was generated by brief intra-arterial perfusion with 0.3% CHAPS. CaV1.2 expression, function and H₂O₂ were measured by Western blot, tension recording and Amplex Red H₂O₂ assay kit, respectively. Results: Angiotensin II dose-dependently upregulated CaV1.2 expression in endothelium-intact arteries. The angiotensin II upregulation of CaV1.2 expression in endothelium-intact arteries was blocked by NAD(P)H oxidase inhibitor diphenyleneiodonium (DPI), apocynin, a more specific NAD(P)H oxidase inhibitor gp91ds-tat and also by catalase. H₂O₂ similarly upregulated CaV1.2 expression in endothelium-intact and endothelium-damaged arteries, and the latter effect was also blocked by DPI and apocynin. Angiotensin II increased H₂O₂ production by endothelium-intact but not by endothelium-damaged arteries, and this effect was blocked by apocynin, catalase and gp91ds-tat. The upregulation of CaV1.2 by angiotensin II and H₂O₂ is accompanied by an increased tension response to KCl and the Ca²⁺ channel activator FPL 64176, and this effect was also attenuated by gp91ds-tat. Conclusion: These results suggest that angiotensin II stimulates endothelial NAD(P)H oxidase-produced H₂O₂, which may additionally act through vascular smooth muscle NAD(P)H oxidase, to upregulate vascular CaV1.2 protein.