Constriction of pulmonary artery by peroxide: role of Ca 2+ release and PKC

Abstract

Reactive oxygen species are implicated in pulmonary hypertension and hypoxic pulmonary vasoconstriction. We examined the effects of low concentrations of peroxide on intrapulmonary arteries (IPA). IPAs from Wistar rats were mounted on a myograph for recording tension and estimating intracellular Ca 2+ using Fura-PE3. Ca 2+ sensitization was examined in α-toxin-permeabilized IPAs, and phosphorylation of MYPT-1 and MLC 20 was assayed by Western blot. Peroxide (30 μM) induced a vasoconstriction with transient and sustained components and equivalent elevations of intracellular Ca 2+. The transient constriction was strongly suppressed by indomethacin, the TP-receptor antagonist SQ-29584, and the Rho kinase inhibitor Y-27632, whereas sustained constriction was unaffected. Neither vasoconstriction nor elevation of intracellular Ca 2+ was affected by removal of extracellular Ca 2+, whereas dantrolene suppressed the former and ryanodine abolished the latter. Peroxide-induced constriction of permeabilized IPAs was unaffected by Y-27632 but abolished by PKC inhibitors; these also suppressed constriction in intact IPAs. Peroxide caused translocation of PKCα, but had no significant effect on MYPT-1 or MLC 20 phosphorylation. We conclude that in IPAs peroxide causes transient release of vasoconstrictor prostanoids, but sustained constriction is associated with release of Ca 2+ from ryanodine-sensitive stores and a PKC-dependent but Rho kinase- and MLC 20-independent constrictor mechanism.