Differential targeting and signalling of voltage-gated T-type Cav3.2 and L-type Cav1.2 channels to ryanodine receptors in mesenteric arteries

Abstract

Key points: In arterial smooth muscle, Ca2+ sparks are elementary Ca2+-release events generated by ryanodine receptors (RyRs) to cause vasodilatation by opening maxi Ca2+-sensitive K+ (BKCa) channels. This study elucidated the contribution of T-type Cav3.2 channels in caveolae and their functional interaction with L-type Cav1.2 channels to trigger Ca2+ sparks in vascular smooth muscle cells (VSMCs). Our data demonstrate that L-type Cav1.2 channels provide the predominant Ca2+ pathway for the generation of Ca2+ sparks in murine arterial VSMCs. T-type Cav3.2 channels represent an additional source for generation of VSMC Ca2+ sparks. They are located in pit structures of caveolae to provide locally restricted, tight coupling between T-type Cav3.2 channels and RyRs to ignite Ca2+ sparks. Abstract: Recent data suggest that T-type Cav3.2 channels in arterial vascular smooth muscle cells (VSMCs) and pits structure of caveolae could contribute to elementary Ca2+ signalling (Ca2+ sparks) via ryanodine receptors (RyRs) to cause vasodilatation. While plausible, their precise involvement in igniting Ca2+ sparks remains largely unexplored. The goal of this study was to elucidate the contribution of caveolar Cav3.2 channels and their functional interaction with Cav1.2 channels to trigger Ca2+ sparks in VSMCs from mesenteric, tibial and cerebral arteries. We used tamoxifen-inducible smooth muscle-specific Cav1.2−/− (SMAKO) mice and laser scanning confocal microscopy to assess Ca2+ spark generation in VSMCs. Ni2+, Cd2+ and methyl-β-cyclodextrin were used to inhibit Cav3.2 channels, Cav1.2 channels and caveolae, respectively. Ni2+ (50 μmol L−1) and methyl-β-cyclodextrin (10 mmol L−1) decreased Ca2+ spark frequency by ∼20–30% in mesenteric VSMCs in a non-additive manner, but failed to inhibit Ca2+ sparks in tibial and cerebral artery VSMCs. Cd2+ (200 μmol L−1) suppressed Ca2+ sparks in mesenteric arteries by ∼70–80%. A similar suppression of Ca2+ sparks was seen in mesenteric artery VSMCs of SMAKO mice. The remaining Ca2+ sparks were fully abolished by Ni2+ or methyl-β-cyclodextrin. Our data demonstrate that Ca2+ influx through CaV1.2 channels is the primary means of triggering Ca2+ sparks in murine arterial VSMCs. CaV3.2 channels, localized to caveolae and tightly coupled to RyR, provide an additional Ca2+ source for Ca2+ spark generation in mesenteric, but not tibial and cerebral, arteries.