CHARACTERIZATION OF DES-ARG(9)-BRADYKININ-INDUCED CONTRACTION IN GUINEA-PIG GALLBLADDER IN-VITRO

We have reported that bradykinin induces graded contraction in guinea- pig gallbladder in vitro through activation of bradykinin B-2 receptor s and prostanoid release, while des-Arg(9)-bradykinin, a selective bra dykinin B-1 receptor agonist, causes only a weak contraction, suggesti ng the presence of badykinin B-1 receptors in this tissue. In the pres ent study, we attempted to characterise the receptor subtype and the p ossible mechanism by which des-Arg(9)-bradykinin induces contraction i n this preparation. Contractions induced by des-Arg(9)- bradykinin in guinea-pig gallbladder (1 pM to 1 mu M) increased significantly as a f unction of time elapsed after setting up of the preparation, reaching the maximum after 6 h of equilibration (EC50 16.4 pM and E-max 0.6 +/- 0.08 g). Des-Arg(9)-bradykinin-induced contraction in guinea-pig gall bladder was totally prevented by cycloheximide (70 mu M, an inhibitor of protein synthesis), indomethacin (3 mu M), ibuprofen (30 mu M), phe nidone (30 mu M) or Ca2+-free medium plus EGTA, and was partially anta gonised by MK 571 ((3-(3-(2-(7-chloro-2-quinolinyl) ethenyl) phenyl (( 3-dimethyl amino-3-oxo-propyl) thio) methyl) propanoic acid, 0.1 mu M) or by nicardipine (1 mu M), but was not affected by dazoxiben (3 mu M ), staurosporine (100 nM) or L 655,240 (240 (3-[1-(4-clorobenzil)-5-fl uoro-3-metilhyindol-2i1] 2,2-dimetilpropanoic acid, 1 mu M). Unexpecte dly, des-Arg(9)-bradykinin-induced contraction was unaffected by the s elective bradykinin B-1 receptor antagonists, des-Arg(9)-[Leu(8)]-brad ykinin and des-Arg(9)-NPC 17761 (des-Arg(0)-D-Arg [Hip(3), D-HipE (tra nstiofenil)(7), Oic(8)]-des-Arg(9)-bradykinin). However, the selective bradykinin B-2 receptor antagonists, HOE 140 (D-arg(0)-[Hyp(3), Thi(5 ), D-TiC7, Oic(8)]-bradykinin) and NPC 17731 (D-Arg(0) [Hyp(3), DHypE (transpropyl)(7), Oic(8)]-bradykinin), completely blocked des-Arg(9)- bradykinin-mediated contraction. Pre-treatment of the animals with Esc herichia coli endotoxin (lipopolysaccharide, 30 mu g/animal, i.v., 24 h) did not significantly change the response to des-Arg(9)-bradykinin induction. It is concluded that des-Arg(9)-bradykinin-induced contract ions in guinea-pig gallbladder are mediated primarily by the release o f proinflammatory eicosanoid(s) derived from the cyclo-oxygenase pathw ay. These effects are unrelated to thromboxane A(2) and do not seem to be coupled to activation of a protein kinase C-dependent mechanism. R esponse to des-Arg(9)-bradykinin increases as a function of the equili bration period of the preparation by a mechanism dependent on protein synthesis and seems to be mediated by activation of bradykinin B-2 (bu t not B-1) receptors. Finally, in contrast to that observed for bradyk inin, the contraction induced by des-Arg(9)-bradykinin in guinea-pig g allbladder is fully dependent on the influx of extracellular Ca2+, par tially through L-type Ca2+ channels. (C) 1997 Elsevier Science B.V.