Inhibitory effects of brefeldin A, a membrane transport blocker, on the bradykinin-induced hyperpolarization-mediated relaxation in the porcine coronary artery.

1. To elucidate the mechanism of the relaxation mediated by endothelium-derived hyperpolarizing factors (EDHFs), the effect of brefeldin A, a membrane transport blocker, on cytosolic Ca(2+) concentration ([Ca(2+)]i) and tension was determined in the porcine coronary arterial strips. We also examined the effect of brefeldin A on [Ca(2+)]i in the endothelial cells of the porcine aortic valve. 2. In the presence of 10 microM indomethacin and 30 microM N(G)-nitro-L-arginine (L-NOARG), both bradykinin and substance P induced a transient decrease in [Ca(2+)]i and tension in arterial strips contracted with 100 nM U46619 (thromboxane A2 analogue). A 6 h pre-treatment with 20 microg ml(-1) brefeldin A abolished the bradykinin-induced relaxation, while it had no effect on the substance P-induced relaxation. 3. In the absence of indomethacin and L-NOARG, brefeldin A had no effect on the bradykinin-induced relaxation during the contraction induced by U46619 or 118 mM K(+). 4. The indomethacin/L-NOARG-resistant relaxation induced by bradykinin was completely inhibited by 3 mM tetrabutylammonium (non-specific Ca(2+)-activated K(+) channel blocker), while that induced by substance P was not inhibited by 3 mM tetrabutylammonium or 1 mM 4-aminopyridine (voltage-dependent K(+) channels blocker) alone, but completely inhibited by their combination. 5. Brefeldin A had no effect on the [Ca(2+)]i elevation in endothelial cells induced by bradykinin or substance P. 6. In conclusion, bradykinin produce EDHF in a brefeldin A-sensitive mechanism in the porcine coronary artery. However, this mechanism is not active in a substance P-induced production of EDHF, which thus suggests EDHF to be more than a single entity.