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-S1, a PLC isoform implicated in a i-adrenoreceptor signaling and the pathogenesis of hypertension, is involved in nor- adrenaline (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 radiolabeling with 33Pi or myo-[3H]i- nositol, and caveolae/rafts were prepared by discontinuous sucrose density centrifugation. PLC-61 was localized by immunoblot analysis and neutralized by delivery of PLC-~1 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-~1 was present in caveolae/rafts, and NA, but not ET-l, stimulated a rapid twofold increase in PLC-S1 levels in these domains. PLC-S1 is calcium dependent, and removal of extracellular calcium prevented its association with caveolae/rafts in response to NA, concomitantly reducing NA-induced [33P]PIP2 hy- drolysis and [3H]inositol phosphate formation but with no effect on ET-l-induced [33P]PIP2 hydrolysis. Neutralization of PLC-S1 by PLC-S1 antibody prevented its caveolae/raft association and attenu- ated the sustained contractile response to NA compared with control antibodies. In contrast, ET- I-induced contraction was not affected by PLC-S1 antibody. These results indicate the novel and selective role of caveolae/raft localized PLC-S1 in NA-induced PIP2 hydrolysis and sustained contraction in intact vascular tissue. [ABSTRACT FROM AUTHOR]