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

Vasoconstrictors activate phospholipase C (PLC) which hydrolyses phosphatidylinositol 4,5-bisphosphate (PIP2) leading to calcium mobilisation, protein kinase C activation and contraction. Our aim was to investigate whether PLCdelta1, a PLC isoform implicated in alpha1-adrenoreceptor signalling and the pathogenesis of hypertension, is involved in noradrenaline (NA) or endothelin (ET-1) induced PIP2 hydrolysis and contraction. Rat mesenteric small arteries were studied. Contractility was measured by pressure myography, phospholipids or inositol phosphates were measured by radiolabelling with (33)Pi or [(3)H]-myo-inositol and caveolae/rafts prepared by discontinuous sucrose density centrifugation. PLCdelta1 was localised by immunoblot analysis and neutralised by delivery of PLCdelta1 antibody. The PLC inhibitor U73122 but not the negative control U73342 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. PLCdelta1 was present in caveolae/rafts and NA but not ET-1 stimulated a rapid two-fold increase in PLCdelta1 levels in these domains. PLCdelta1 is calcium dependent and removal of extracellular calcium prevented its association with caveolae/rafts in response to NA, concomitantly reducing NA-induced [(33)P]-PIP2 hydrolysis and [(3)H]-inositol phosphate formation but with no effect on ET-1-induced [(33)P]-PIP2 hydrolysis. Neutralisation of PLCdelta1 by PLCdelta1 antibody prevented its caveolae/raft association and attenuated the sustained contractile response to NA when compared to control antibodies. In contrast, ET-1-induced contraction was not affected by PLCdelta1 antibody. These results indicate the novel and selective role of caveolae/raft localised PLCdelta1 in NA-induced PIP2 hydrolysis and sustained contraction in intact vascular tissue. Key words: signal transduction , vascular smooth muscle, phosphoinositide, caveolae, lipid rafts.