Identification of epoxyeicosatrienoic acids as endothelium-derived hyperpolarizing factors.
Endothelial cells release several compounds, including prostacyclin, NO, and endothelium-derived hyperpolarizing factor (EDHF), that mediate the vascular effects of vasoactive hormones. The identity of EDHF remains unknown. Since arachidonic acid causes endothelium-dependent relaxations of coronary arteries through its metabolism to epoxyeicosatrienoic acids (EETs) by cytochrome P450, we wondered if the EETs represent EDHFs. Precontracted bovine coronary arteries relaxed in an endothelium-dependent manner to methacholine. The cytochrome P450 inhibitors, SKF 525A and miconazole, significantly attenuated these relaxations. They were also inhibited by tetraethylammonium (TEA),an inhibitor of Ca2+-activated K+ channels, and by high [K+]0 (20 mmol/L). Methacholine also caused hyperpolarization of coronary smooth muscle (-27 +/- 3.9 versus -40 +/- 5.1 mV), which was completely blocked by SKF 525A and miconazole. In vessels prelabeled with [3H] arachidonic acid, methacholine stimulated the release of 6-ketoprostaglandin F1alpha, 12-HETE, and the EETs. Arachidonic acid relaxed precontracted coronary arteries, which were also blocked by TEA, charybdotoxin, another Ca2+-activated K+ channel inhibitor, and high [K+]0. 14,15-EET, 11,12-EET, 8,9-EET, and 5,6-EET relaxed precontracted coronary vessels (EC50, 1 X 10(-6) mol/L). The four regioisomers were equally active. TEA, charybdotoxin, and high [K+]0 attenuated the EET relaxations. 11,12-EET hyperpolarized coronary smooth muscle cells from -37 +/- 0.2 to -59 +/- 0.3 mV. In the cell-attached mode of patch clamp, both 14,15-EET and 11,12-EET increased the open-state probability of a Ca2+-activated K+ channel in coronary smooth muscle cells. This effect was blocked by TEA and charybdotoxin. These data support the hypothesis that the EETs are EDHFs.
Campbell
W B
WB
Department of Pharmacology, Medical College of Wisconsin, Milwaukee 53226, USA.
Gebremedhin
D
D
Pratt
P F
PF
Harder
D R
DR
D015126:8,11,14-Eicosatrienoic Acid / Q000378:metabolism*; D000818:Animals; D016718:Arachidonic Acid / Q000494:pharmacology; D001685:Biological Factors / Q000378:metabolism*; D002417:Cattle; D003331:Coronary Vessels / Q000187:drug effects / Q000378:metabolism*; D000242:Cyclic AMP / Q000096:biosynthesis; D006152:Cyclic GMP / Q000096:biosynthesis; D065607:Cytochrome P-450 Enzyme Inhibitors; D004730:Endothelium, Vascular / Q000187:drug effects / Q000378:metabolism*; D066298:In Vitro Techniques; D009126:Muscle Relaxation / Q000187:drug effects; D009569:Nitric Oxide / Q000378:metabolism; D018408:Patch-Clamp Techniques
D001685:Biological Factors; D065607:Cytochrome P-450 Enzyme Inhibitors; C067410:endothelium-dependent hyperpolarization factor; D016718:Arachidonic Acid; D009569:Nitric Oxide; D000242:Cyclic AMP; D015126:8,11,14-Eicosatrienoic Acid; D006152:Cyclic GMP