BIOASSAY OF AN ENDOTHELIUM-DERIVED HYPERPOLARIZING FACTOR FROM BOVINECORONARY-ARTERIES - ROLE OF A CYTOCHROME-P-450 METABOLITE

An endothelium-derived hyperpolarizing factor (EDHF) mediates a part o f the vasodilatory action of bradykinin. A bioassay method was develop ed to investigate the properties of EDHF on bovine coronary arterial s mooth muscle cells. Cannulated bovine coronary arteries with an intact endothelium that were treated with indomethacin and NG-nitro-L-argini ne methyl ester served as the EDHF donor. The effect of the donor vess el perfusate was examined on a 240 pS single-channel calcium (Ca2+)-ac tivated potassium (K+) current (K-Ca) and resting membrane potential i n recipient coronary arterial smooth muscle cells. The open state prob ability (NPo) of the channel averaged 0.011 +/- 0.003 during basal per fusate flow. After stimulation of the donor vessels with bradykinin (1 0-(10)-10(-6) M), the perfusate induced a 1.2- to 5-fold increase in t he NPo (n = 7, p < 0.001). This increase in channel activity was atten uated by either removing the endothelium of the donor arterial segment or upon inhibition of cytochrome P-450 in the donor arterial segment with the combination of 17-octadecynoic acid and miconazole. The resti ng cell membrane averaged -60 +/- 2 mV, and hyperpolarized to -69 +/- 1.5 mV (n = 6, p < 0.05) in response to the perfusate following stimul ation of the donor vessel with bradykinin. Addition of 14,15-epoxyeico satrienoic acid mimicked the effects of the perfusate and increased th e NPo of the K-Ca channel from 0.01 +/- 0.001 to 0.05 +/- 0.001. These findings suggest that bradykinin stimulates the release of a transfer able endothelial factor that activates K-Ca, channels and hyperpolariz es coronary arterial smooth muscle cell membranes. These findings supp ort the hypothesis that coronary arteries release an EDHF which is a c ytochrome P-450 metabolite of arachidonic acid.