Abstract 322: A Crucial Role of Rho/Rho-kinase Pathway in Constriction and Closure of Human and Rabbit Ductus Arteriosus: Redox Activation Mediates Calcium Sensitization
Background - At birth, functional closure of the ductus arteriosus (DA) is initiated within minutes by inhibition of oxygen-sensitive, voltage-gated K+ channels. The resulting membrane depolarization and activation of L-type calcium channels initiates vasoconstriction. The subsequent maintenance of vasoconstriction results in part from increases in endothelin and decreases vasodilator prostaglandins. We hypothesized that Rho-kinase pathway is crucial to maintaining elevated tension in response to O2 and Endothelin-1 (ET-1) in human and rabbit DA after birth. Methods and Results - O2 constriction persists in the absence of extracellular calcium, albeit at a significantly reduced level, consistent with the occurrence of calcium sensitization. Two different Rho-kinase inhibitors (Y-27632 and fasudil) rapidly lower basal tone in hypoxic human DAs and reverse O2-constriction. Likewise, in isolated DA from term fetal rabbits, Rho-kinase inhibitors cause dose-dependent relaxation of oxygen-constricted DA and attenuate constriction to ET-1, phenylephrine and potassium chloride. Rho kinase inhibitors induced greater relaxation in ET-1- constricted DA and pulmonary artery than aortic rings. Exposure to O2 for 1-hour increased expression of Rho/Rho-kinase proteins in Human DA smooth muscle cells and vasa vasorum. The mechanism by which O2 activates Rho kinase appears to be independent of GTP-RhoA form and involves activation RhoB expression. The effects of O2 are mimicked in part by H2O2, suggesting redox activation of Rho kinase. Conclusions - In the human ductus activation of the Rho kinase pathway is crucial to maintaining basal tone and constriction to oxygen. Oxygen activates the Rho kinase pathway through a redox mechanism causing calcium sensitization. Activation of Rho kinase by O2 causes a secondary upregulation of RhoB and ROCK-1 expression and activity by a RhoA-independent mechanism. Activation of Rho kinase is an appealing strategy to close DA whereas Rho kinase inhibition has the potential to maintain DA patency as a bridge to cardiac surgery in patients with DA-dependent congenital heart disease.