IP3 constricts cerebral arteries via IP3 receptor-mediated TRPC3 channel activation and independently of sarcoplasmic reticulum Ca2+ release

Vasoconstrictors that bind to phospholipase C-coupled receptors elevate inositol-1,4,5-trisphosphate (IP3). IP3 is generally considered to elevate intracellular Ca2+ concentration ([Ca2+](i)) in arterial myocytes and induce vasoconstriction via a single mechanism: by activating sarcoplasmic reticulum (SR)-localized IP3 receptors, leading to intracellular Ca2+ release. We show that IP3 also stimulates vasoconstriction via a SR Ca2+ release-independent mechanism. In isolated cerebral artery myocytes and arteries in which SR Ca2+ was depleted to abolish Ca2+ release (measured using D1ER, a fluorescence resonance energy transfer-based SR Ca2+ indicator), IP3 activated 15 pS sarcolemmal cation channels, generated a whole-cell cation current (I-Cat) caused by Na+ influx, induced membrane depolarization, elevated [Ca2+](i), and stimulated vasoconstriction. The IP3-induced I-Cat and [Ca2+](i) elevation were attenuated by cation channel (Gd3+, 2-APB) and IP3 receptor (xestospongin C, heparin, 2-APB) blockers. TRPC3 (canonical transient receptor potential 3) channel knockdown with short hairpin RNA and diltiazem and nimodipine, voltage-dependent Ca2+ channel blockers, reduced the SR Ca2+ release-independent, IP3-induced [Ca2+](i) elevation and vasoconstriction. In pressurized arteries, SR Ca2+ depletion did not alter IP3-induced constriction at 20 mm Hg but reduced IP3-induced constriction by approximate to 39% at 60 mm Hg. [Ca2+](i) elevations and constrictions induced by endothelin-1, a phospholipase C-coupled receptor agonist, were both attenuated by TRPC3 knockdown and xestospongin C in SR Ca2+-depleted arteries. In summary, we describe a novel mechanism of IP3-induced vasoconstriction that does not occur as a result of SR Ca2+ release but because of IP3 receptor-dependent I-Cat activation that requires TRPC3 channels. The resulting membrane depolarization activates voltage-dependent Ca2+ channels, leading to a myocyte [Ca2+](i) elevation, and vasoconstriction.