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-deri ved hyperpolarizing factors (EDHFs), the effect of brefeldin A, a membrane transport blocker, on cytosolic Ca2+ concentration ([Ca2+](i)) and tension was determined in the porcine coronary arterial strips. We also examined th e effect of brefeldin A on [Ca2+](i) in the endothelial cells of the porcin e aortic valve. 2 In the presence of 10 muM indomethacin and 30 muM N-G-nitro-L-arginine (L -NOARG), both bradykinin and substance P induced a transient decrease in [C a2+](i) and tension in arterial strips contracted with 100 nM U46619 (throm boxane A(2) analogue). A 6 h pre-treatment with 20 mug ml(-1) brefeldin A a bolished the bradykinin-induced relaxation, while it had no effect on the s ubstance 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 c ompletely inhibited by 3 mM tetrabutylammonium (non-specific Ca2+-activated K+ channel blocker), while that induced by substance P was not inhibited b y 3 mm tetrabutylammonium or 1 mM 4-aminopyridine (voltage-dependent K+ cha nnels blocker) alone, but completely inhibited by their combination. 5 Brefeldin A had no effect on the [Ca2+](i) elevation in endothelial cells induced by bradykinin or substance P. 6 In conclusion, bradykinin produce EDHF in a brefeldin A-sensitive mechani sm 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 m ore than a single entity.