Phospholipase C-Sx modulates sustained contraction of rat mesenteric small arteries in response to noradrenaline, but not endothelin-1

摘要

First published June 20, 2008; doi:10.1152/ajpheart.01396.2007.-Vasoconstrictors activate phospholipase C (PLC), which hydrolyzes phosphatidylino-sitol 4,5-bisphosphate (PIP2), leading to calcium mobilization, protein kinase C activation, and contraction. Our aim was to investigate whether PLC-81, a PLC isoform implicated in ai-adrenoreceptor signaling and the pathogenesis of hypertension, is involved in noradrenaline (NA) or endothelin (ET-l)-induced PIP2 hydrolysis and contraction. Rat mesenteric small arteries were studied. Contractility was measured by pressure myography, phospholipids or inositol phosphates were measured by radiolabeling with 33Pi or myo-[3H]i-nositol, and caveolae/rafts were prepared by discontinuous sucrose density centrifugation. PLC-81 was localized by immunoblot analysis and neutralized by delivery of PLC-Bi antibody. The PLC inhibitor U73122, but not the negative control U-73342, markedly inhibited NA and ET-1 contraction but had no effect on potassium or phorbol ester contraction, implicating PLC activity in receptor-mediated smooth muscle contraction. PLC-81 was present in caveolae/rafts, and NA, but not ET-1, stimulated a rapid twofold increase in PLC-(delta)1 levels in these domains. PLC-81 is calcium dependent, and removal of extracellular calcium prevented its association with caveolae/rafts in response to NA, concomitantly reducing NA-induced [33PjPIp2 hydrolysis and [3H]inositol phosphate formation but with no effect on ET-1-induced [33P]PIP2 hydrolysis. Neutralization of PLC-81 by PLC-81 antibody prevented its caveolae/raft association and attenuated the sustained contractile response to NA compared with control antibodies. In contrast, ET-1-induced contraction was not affected by PLC-Si antibody. These results indicate the novel and selective role of caveolae/raft localized PLC-Si in NA-induced PIP2 hydrolysis and sustained contraction in intact vascular tissue.