Calcium-independent phospholipase A2 participates in KCl-induced calcium sensitization of vascular smooth muscle

Abstract

In vascular smooth muscle, KCl not only elevates intracellular free Ca2+ ([Ca2+]i), myosin light chain kinase activity and tension (T), but also can inhibit myosin light chain phosphatase activity by activation of rhoA kinase (ROCK), resulting in Ca2+ sensitization (increased T/[Ca2+]i ratio). Precisely how KCl causes ROCK-dependent Ca2+ sensitization remains to be determined. Using Fura-2-loaded isometric rings of rabbit artery, we found that the Ca2+-independent phospholipase A2 (iPLA2) inhibitor, bromoenol lactone (BEL), reduced the KCl-induced tonic but not early phasic phase of T and potentiated [Ca2+]i, reducing Ca2+ sensitization. The PKC inhibitor, GF-109203X (≥3μM) and the pseudo-substrate inhibitor of PKCζ produced a response similar to BEL. BEL reduced basal and KCl-stimulated myosin phosphatase phosphorylation. Whereas BEL and H-1152 produced strong inhibition of KCl-induced tonic T (∼50%), H-1152 did not induce additional inhibition of tissues already inhibited by BEL, suggesting that iPLA2 links KCl stimulation with ROCK activation. The cPLA2 inhibitor, pyrrolidine-1, inhibited KCl-induced tonic increases in [Ca2+]i but not T, whereas the inhibitor of 20-HETE production, HET0016, acted like the ROCK inhibitor H-1152 by causing Ca2+ desensitization. These data support a model in which iPLA2 activity regulates Ca2+ sensitivity.