Metabotropic Regulation of RhoA/Rho-Associated Kinase by L-type Ca2+ Channels New Mechanism for Depolarization-Evoked Mammalian Arterial Contraction

Abstract

Background: Sustained vascular smooth muscle contraction is mediated by extracellular Ca2 influx through L-type voltage-gated Ca2 channels (VGCC) and RhoA/Rho-associated kinase (ROCK)-dependent Ca2 sensitization of the contractile machinery. VGCC activation can also trigger an ion-independent metabotropic pathway that involves G-protein/phospholipase C activation, inositol 1,4,5-trisphosphate synthesis, and Ca2 release from the sarcoplasmic reticulum (calcium channel-induced Ca2 release). We have studied the functional role of calcium channel-induced Ca2 release and the inter-relations between Ca2 channel and RhoA/ROCK activation. Methods and Results: We have used normal and genetically modified animals to study single myocyte electrophys iology and fluorimetry as well as cytosolic Ca2 and diameter in intact arteries. These analyses were complemented with measurement of tension and RhoA activity in normal and reversibly permeabilized arterial rings. We have found that, unexpectedly, L-type Ca2 channel activation and subsequent metabotropic Ca2 release from sarcoplasmic reticulum participate in depolarization-evoked RhoA/ROCK activity and sustained arterial contraction. We show that these phenomena do not depend on the change in the membrane potential itself, or the mere release of Ca2 from the sarcoplasmic reticulum, but they require the simultaneous activation of VGCC and the downstream metabotropic pathway with concomitant Ca2 release. During protracted depolarizations, refilling of the stores by a residual extracellular Ca2 influx through VGCC helps maintaining RhoA activity and sustained arterial contraction. Conclusions: These findings reveal that calcium channel-induced Ca2 release has a major role in tonic vascular smooth muscle contractility because it links membrane depolarization and Ca2 channel activation with metabotropic Ca2 release and sensitization (RhoA/ROCK stimulation)