CA2-DEPENDENT AND CA2+-INDEPENDENT MECHANISM OF CYCLIC-AMP REDUCTION - MEDIATION BY BRADYKININ B-2 RECEPTORS()

1 Bradykinin caused a transient reduction of about 25% in the cyclic A MP level in forskolin prestimulated DDT1 MF-2 smooth muscle cells (IC5 0: 36.4 +/- 4.9 nM) and a pronounced, sustained inhibition (40%) of th e isoprenaline-stimulated cyclic AMP level (IC50: 37.5 +/- 1.1 nM). 2 The Ca2+ ionophore, ionomycin, mimicked both the bradykinin-induced tr ansient reduction in the forskolin-stimulated cyclic AMP level and the sustained reduction in the isoprenaline-stimulated cyclic AMP level. 3 The Ca2+-dependent effect on cyclic AMP induced by bradykinin was me diated solely by Ca2+ release from internal stores, since inhibition o f Ca2+ entry with LaCl3 did not reduce the response to bradykinin. 4 T he involvement of calmodulin-dependent enzyme activities, protein kina se C or an inhibitory GTP binding protein in the bradykinin-induced re sponses was excluded since a calmodulin inhibitor, calmidazolium, a PK C inhibitor, staurosporine and pertussis toxin, respectively did not a ffect the decline in the cyclic AMP level. 5 Bradykinin enhanced the r ate of cyclic AMP breakdown in intact cells, which effect was not mimi cked by ionomycin. This suggested a Ca2+-independent activation of pho sphodiesterase activity by bradykinin in DDT1 MF-2 cells. 6 The bradyk inin B-1 receptor agonist, desArg(9)-bradykinin, did not affect cyclic AMP formation in isoprenaline prestimulated cells, while the bradykin in Bz receptor antagonists, Hoe 140 (D-Arg[Hyp(3), Thi(5), D-Tic(7), O ic(8)]-BK) and D-Arg[Hyp(3), Thi(5,8), D-Phe(7)]-BK completely abolish ed the bradykinin response in both forskolin and isoprenaline prestimu lated cells. 7 Bradykinin caused an increase in intracellular Ca2+, wh ich was antagonized by the bradykinin B-2 receptor antagonists, Hoe 14 0 and D-Arg[Hyp(3), Thi(5,8), D-Phe(7)]-BK. The bradykinin B-2 recepto r agonist, desArg(9)-bradykinin, did not evoke a rise in cytoplasmic C a2+. 8 It is concluded, that stimulation of bradykinin B-2 receptors c auses a reduction in cellular cyclic AMP in DDT1 MF-2 cells. This decl ine in cyclic AMP is partly mediated by a Ca2+/calmodulin independent activation of phosphodiesterase activity. The increase in [Ca2+](i) me diated by bradykinin B-2 receptors inhibited forskolin- and isoprenali ne-activated adenylyl cyclase differently, most likely by interfering with different components of the adenylyl cyclase signalling pathway.