Role of phospholipase A2 in EDHF-Mediated relaxation of the porcine coronary artery

In porcine coronary artery, bradykinin-induced relaxation, in the presence of inhibitors of nitric oxide synthase and cyclooxygenase, is mediated by EDHF, which was proposed to be a non-cyclooxygenase metabolite of arachidonic acid. Unesterified arachidonic acid can be generated through activation of cytosolic phospholipase A(2) and/or by phospholipase C-mediated release of diacylglycerol, which is metabolized by lipases to yield arachidonic acid. To investigate the role of phospholipase A(2) and diacylglycerol lipase in bradykinin-induced EDHF production, porcine coronary rings treated with nitric oxide synthase and cyclooxygenase inhibitors were contracted with U46619. Relaxations to bradykinin or arachidonic acid were obtained before and after exposure to arachidonyl trifluoromethyl ketone or RHC 80267, inhibitors of phospholipase A(2) and diacylglycerol lipase, respectively. Arachidonyl trifluoromethyl ketone, but not RHC 80267, inhibited bradykinin-induced, EDHF-mediated relaxation. Under similar conditions, arachidonyl trifluoromethyl ketone did not inhibit the arachidonic acid-induced relaxation, suggesting that the compound did not block signaling pathways distal to arachidonic acid release. In porcine coronary arteries, arachidonyl trifluoromethyl ketone also blocked bradykinin-induced production of prostacyclin (an index of arachidonic acid release). When incubated with [H-3]arachidonic acid, porcine coronary arteries formed products which co-migrated with prostaglandins and hydroxyeicosatetraenoic acids; treatment with nitric oxide synthase and cyclooxygenase inhibitors abolished the formation of prostaglandins, but not of hydroxyeicosatetraenoic acids. These findings suggest that the production of EDHF in the porcine coronary artery is dependent upon phospholipase A(2)-mediated arachidonic acid release. Whether the arachidonic acid is metabolized to form EDHF, or whether it acts as a second messenger to stimulate EDHF production, remains to be determined.