Endothelium-derived hyperpolarizing factor, but not nitric oxide, is reversibly inhibited by brefeldin A.
The subcellular localization of the enzymes synthesizing endothelium-derived vasodilator autacoids has been proposed to play a role in determining the ability of endothelial cells to enhance autacoid production in response to stimulation. We therefore investigated the effects of brefeldin A-induced disruption of the Golgi apparatus and Golgi-plasma membrane trafficking on the production of nitric oxide (NO), prostacyclin, and the endothelium-derived hyperpolarizing factor (EDHF) by native and cultured endothelial cells. In porcine coronary artery segments, brefeldin A (35 micromol/L, 90 minutes) did not affect relaxations to sodium nitroprusside or the K+ channel opener cromakalim but elicited a rightward shift in the concentration-response curve to bradykinin without altering the maximum vasodilator response (Rmax). Brefeldin A failed to attenuate the bradykinin-induced, NO-mediated relaxation under depolarizing conditions but inhibited the bradykinin response under conditions of combined cyclooxygenase/NO synthase blockade, suggesting that this agent selectively interferes with the production of EDHF. Indeed, incubation of porcine coronary arteries with brefeldin A, which did not affect the bradykinin-induced accumulation of either cyclic GMP or 6-keto-prostaglandin F1alpha, markedly and reversibly attenuated the EDHF-mediated hyperpolarization of detector smooth muscle cells in a patch-clamp bioassay system. The microtubule destabilizer nocodazole also affected both the EC50 and Rmax to bradykinin in porcine coronary arteries. Since EDHF is thought to be a cytochrome P450-derived metabolite of arachidonic acid and both brefeldin A and nocodazole are known to interfere with the targeting of cytochrome P450 from the Golgi apparatus to the plasma membrane, it is conceivable that brefeldin A inhibits EDHF formation by preventing the targeting of the EDHF-synthesizing enzymes to the plasma membrane.
Bauersachs
J
J
Institut für Kardiovaskuläre Physiologie, Zentrum der Physiologie, Klinikum der J.W. Goethe-Universität, Frankfurt am Main, Germany. r.busse@em.uni-frankfurt.de
Fleming
I
I
Scholz
D
D
Popp
R
R
Busse
R
R
D015121:6-Ketoprostaglandin F1 alpha / Q000378:metabolism; D000818:Animals; D001011:Aorta / Q000502:physiology; D001685:Biological Factors / Q000096:biosynthesis* / Q000494:pharmacology*; D001920:Bradykinin / Q000494:pharmacology; D020126:Brefeldin A; D002478:Cells, Cultured; D003331:Coronary Vessels / Q000502:physiology; D006152:Cyclic GMP / Q000378:metabolism; D003517:Cyclopentanes / Q000494:pharmacology*; D004357:Drug Synergism; D004730:Endothelium, Vascular / Q000502:physiology*; D006056:Golgi Apparatus / Q000187:drug effects / Q000502:physiology; D006801:Humans; D066298:In Vitro Techniques; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D009119:Muscle Contraction / Q000187:drug effects / Q000502:physiology; D009131:Muscle, Smooth, Vascular / Q000187:drug effects / Q000502:physiology*; D009569:Nitric Oxide / Q000502:physiology*; D019001:Nitric Oxide Synthase / Q000378:metabolism; D009599:Nitroprusside / Q000494:pharmacology*; D011817:Rabbits; D013552:Swine; D014471:Umbilical Veins; D014664:Vasodilation
D001685:Biological Factors; D003517:Cyclopentanes; C067410:endothelium-dependent hyperpolarization factor; D009599:Nitroprusside; D020126:Brefeldin A; D009569:Nitric Oxide; D015121:6-Ketoprostaglandin F1 alpha; D019001:Nitric Oxide Synthase; D006152:Cyclic GMP; D001920:Bradykinin