11,12,15-Trihydroxyeicosatrienoic acid mediates ACh-induced relaxations in rabbit aorta.
Rabbit aortic endothelium metabolizes arachidonic acid (AA) by the 15-lipoxygenase pathway to vasodilatory eicosanoids, hydroxyepoxyeicosatrienoic acids (HEETAs), and trihydroxyeicosatrienoic acids (THETAs). The present study determined the chemical identity of the vasoactive THETA and investigated its role in ACh-induced relaxation in the rabbit aorta. AA caused endothelium-dependent, concentration-related relaxations of the rabbit aorta. Increasing the extracellular KCl concentration from 4.8 to 20 mM inhibited the relaxations to AA by approximately 60%, thereby implicating K+-channel activation in the relaxations. In addition, AA caused an endothelium-dependent hyperpolarization of aortic smooth muscle from -39.6 +/- 2.7 to -56.1 +/- 3.4 mV. In rabbit aortic rings, [14C]AA was metabolized to prostaglandins, HEETAs, THETAs, and 15-hydroxyeicosatetraenoic acid. Additional purification of the THETAs by HPLC resolved the mixture into its 14C-labeled products. Gas chromatography/mass spectrometry identified the metabolites as isomers of 11,12,15-THETA and 11,14,15-THETA. The 11,12,15-THETA relaxed and hyperpolarized the rabbit aorta, whereas 11,14,15-THETA had no vasoactive effect. The relaxations to 11,12,15-THETA were blocked by 20 mM KCl. In aortic rings pretreated with inhibitors of nitric oxide and prostaglandin synthesis, ACh caused a concentration-related relaxation that was completely blocked by 20 mM KCl. Pretreatment with the phospholipase A2 inhibitors mepacrine and 7,7-dimethyl-5,8-eicosadienoic acid, the lipoxygenase inhibitors cinnamyl-3,4-dihydroxy-alpha-cyanocinnamate, nordihydroguaiaretic acid, and ebselen, or the hydroperoxide isomerase inhibitors miconazole and clotrimazole also blocked ACh-induced relaxations. ACh caused a threefold increase in THETA release. These studies indicate that AA is metabolized by endothelial cells to 11,12,15-THETA, which activates K+ channels to hyperpolarize the aortic smooth muscle membrane and induce relaxation. Additionally, this lipoxygenase pathway mediates the nonnitric oxide, nonprostaglandin relaxations to ACh in the rabbit aorta by acting as a source of an endothelium-derived hyperpolarizing factor.
Campbell
William B
WB
Department of Pharmacology and Toxicology, Medical College of Wisconsin, 8701 Watertown Plank Rd., Milwaukee, WI 53226,USA. wbcamp@mcw.edu
Spitzbarth
Nancy
N
Gauthier
Kathryn M
KM
Pfister
Sandra L
SL
D015126:8,11,14-Eicosatrienoic Acid / Q000031:analogs & derivatives* / Q000378:metabolism* / Q000493:pharmacokinetics; D000109:Acetylcholine / Q000494:pharmacology; D000818:Animals; D001011:Aorta / Q000187:drug effects / Q000502:physiology*; D016718:Arachidonic Acid / Q000493:pharmacokinetics; D002250:Carbon Radioisotopes; D004791:Enzyme Inhibitors / Q000494:pharmacology; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D009131:Muscle, Smooth, Vascular / Q000187:drug effects / Q000502:physiology; D019335:Nitroarginine / Q000494:pharmacology; D015221:Potassium Channels / Q000378:metabolism; D011817:Rabbits; D014664:Vasodilation / Q000187:drug effects / Q000502:physiology*; D014665:Vasodilator Agents / Q000494:pharmacology
D002250:Carbon Radioisotopes; D004791:Enzyme Inhibitors; D015221:Potassium Channels; D014665:Vasodilator Agents; D019335:Nitroarginine; D016718:Arachidonic Acid; C066049:11,12,15-trihydroxyeicosatrienoic acid; D015126:8,11,14-Eicosatrienoic Acid; D000109:Acetylcholine