Calcium-independent phospholipase A2-derived arachidonic acid is essential for endothelium-dependent relaxation by acetylcholine

The role of calcium-independent phospholipase A(2) (iPLA(2))produced arachidonic acid (AA) in acetylcholine (ACh)-mediated, endothelium-dependent vascular relaxation was investigated. ACh-induced relaxation of phenylephrine-constricted isolated rat mesenteric resistance arteries was attenuated following pretreatment with (E)-6-(bromomethylene) tetrahydro-3(1-naphthalenyl)- 2H-pyran-2-one (BEL; 1 muM; p < 0.01), a highly selective suicide substrate inhibitor of iPLA(2). Following BEL, the ACh relaxation could be completely restored following pretreatment with picomolar quantities of the cell-permeant methyl ester analog of AA (arachidonic acid methyl ester, AAMe). Higher amounts of AA-Me (1 μM) had a direct endothelium-dependent relaxing action, which was inhibited by the nitric-oxide synthase inhibitor (N-ω-nitro-L-arginine; 100 μM), independent of ACh, and unaffected by BEL. Neither the ACh relaxation restoring action nor the direct relaxing action of AA-Me was affected by preincubation with inhibitors of the lipoxygenase (esculetin, 10 μM) or cytochrome P450 monooxygenase (17-octadecynoic acid; 10 μM) pathways; and both actions of AA-Me were enhanced following preincubation with the cyclooxygenase inhibitor indomethacin (10 μM; p < 0.05). The results of the present study indicate that iPLA(2)-produced AA plays an essential role in ACh-mediated endothelium-dependent relaxation in rat mesenteric resistance arteries.