ARACHIDONIC-ACID RELEASE FROM AORTIC SMOOTH-MUSCLE CELLS INDUCED BY [ARG(8)]VASOPRESSIN IS LARGELY MEDIATED BY CALCIUM-INDEPENDENT PHOSPHOLIPASE-A2

To identify the phospholipase mediating the majority of [Arg8]vasopres sin (AVP)-induced release of arachidonic acid in A-10 smooth muscle ce lls, we exploited the specificity inherent in the mechanism-based inhi bitor, hylene)tetrahydro-3-(1-naphthalenyl)2H-pyran-2-one (HELSS), whi ch possesses a 1,000-fold selectivity for inhibition of calcium-indepe ndent versus calcium-dependent phospholipases A2. Utilizing [H-3]arach idonic acid-labeled A-10 smooth muscle cells, one-half of AVP-inducibl e [H-3]arachidonic acid release was inhibited by pretreatment with onl y 1 muM HELSS and two-thirds of AVP-stimulated [H-3]arachidonic acid r elease was inhibited by 5 muM HELSS. The inhibition of [H-3]arachidoni c acid release by HELSS was saturable (i.e. no additional inhibition o f [H-3]arachidonic acid release was present at 10 muM HELSS), specific (i.e. the activities of six intracellular enzymes, as well as the rat e of glucose oxidation, were not altered by HELSS treatment), and nont oxic (i.e. HELSS-treated cells excluded trypan blue dye and did not le ak intracellular enzymes into the medium). Collectively, these results demonstrate that HELSS blocks AVP-induced arachidonic acid release by specific and irreversible inhibition of calcium-independent phospholi pase A2 and underscore the importance of calcium-independent phospholi pase A2 in agonist-induced arachidonic acid release in at least some c ell types.