Spatiotemporal Control of Vascular Ca(V)1.2 by α1(C) S1928 Phosphorylation

L-type Ca(V)1.2 channels undergo cooperative gating to regulate cell function, although mechanisms are unclear. This study tests the hypothesis that phosphorylation of the Ca(V)1.2 pore-forming subunit α1(C) at S1928 mediates vascular Ca(V)1.2 cooperativity during diabetic hyperglycemia. METHODS: A multiscale approach including patch-clamp electrophysiology, super-resolution nanoscopy, proximity ligation assay, calcium imaging‚ pressure myography, and Laser Speckle imaging was implemented to examine Ca(V)1.2 cooperativity, α1(C) clustering, myogenic tone, and blood flow in human and mouse arterial myocytes/vessels. RESULTS: Ca(V)1.2 activity and cooperative gating increase in arterial myocytes from patients with type 2 diabetes and type 1 diabetic mice, and in wild-type mouse arterial myocytes after elevating extracellular glucose. These changes were prevented in wild-type cells pre-exposed to a PKA inhibitor or cells from knock-in S1928A but not S1700A mice. In addition, α1(C) clustering at the surface membrane of wild-type, but not wild-type cells pre-exposed to PKA or P2Y(11) inhibitors and S1928A arterial myocytes, was elevated upon hyperglycemia and diabetes. Ca(V)1.2 spatial and gating remodeling correlated with enhanced arterial myocyte Ca(2+) influx and contractility and in vivo reduction in arterial diameter and blood flow upon hyperglycemia and diabetes in wild-type but not S1928A cells/mice. CONCLUSIONS: These results suggest that PKA-dependent S1928 phosphorylation promotes the spatial reorganization of vascular α1(C) into “superclusters” upon hyperglycemia and diabetes. This triggers Ca(V)1.2 activity and cooperativity, directly impacting vascular reactivity. The results may lay the foundation for developing therapeutics to correct Ca(V)1.2 and arterial function during diabetic hyperglycemia.