Negative regulation of cellular Ca²⁺ mobilization by ryanodine receptor type 3 in mouse mesenteric artery smooth muscle

Physiological functions of type 3 ryanodine receptors (RyR3) in smooth muscle (SM) tissues are not well understood, in spite of their wide expression. However, the short isoform of RyR3 is known to be a dominant-negative variant (DN-RyR3), which may negatively regulate functions of both RyR2 and full-length (FL) RyR3 by forming hetero-tetramers. Here, functional roles of RyR3 in the regulation of Ca²⁺ signaling in mesenteric artery SM cells (MASMCs) were examined using RyR3 homozygous knockout mice (RyR3^-/-). Quantitative PCR analyses suggested that the predominant RyR3 subtype in MASMs from wild-type mice (RyR3^+/+) was DN-RyR3. In single MASMCs freshly isolated from RyR3^-/-, the EC₅₀ of caffeine to induce Ca²⁺ release was lower than that in RyR3^+/+ myocytes. The amplitude and frequency of Ca²⁺ sparks and spontaneous transient outward currents in MASMCs from RyR3^-/- were all larger than those from RyR3^+/+. Importantly, mRNA and functional expressions of voltage-dependent Ca²⁺ channel and large-conductance Ca²⁺-activated K⁺ (BK) channel in MASMCs from RyR3^-/- were identical to those from RyR3^+/+. However, in the presence of BK channel inhibitor, paxilline, the pressure rises induced by BayK8644 in MA vascular beds of RyR3^-/- were significantly larger than in those of RyR3^+/+. This indicates that the negative feedback effects of BK channel activity on intracellular Ca²⁺ signaling was enhanced in RyR3^-/-. Thus, RyR3, and, in fact, mainly DN-RyR3, via a complex with RyR2 suppresses Ca²⁺ release and indirectly regulated membrane potential by reducing BK channel activity in MASMCs and presumably can affect the regulation of intrinsic vascular tone.