ENDOTHELIUM-DEPENDENT RELAXATIONS MEDIATED BY INDUCIBLE B-1 AND CONSTITUTIVE B-2 KININ RECEPTORS IN THE BOVINE ISOLATED CORONARY-ARTERY

1 Rings of bovine left anterior descending coronary artery (LAD) were contracted with the thromboxane A(2)-mimetic, U46619 (1-30 nM), to approximately 40% of their maximum contraction to 125 mM KCl Krebs solution (KPSSmax) for comparison of responses to the B-1 and B-2 kinin receptor agonists, des-Arg(9)-bradykinin (des-Arg(9)-BK) and bradykinin (BK), respectively. Relaxation responses were normalized as percentages of the initial U46619-induced contraction level, while contractile responses were expressed as percentages of KPSSmax. 2 After 6 h of in vitro incubation in Krebs solution at 37 degrees C, des-Arg(9)-BK ((p)EC(50), 8.00 +/- 0.08; maximum response (R(max)) 93.9 +/- 1.9%) and BK ((p)EC(50), 9.75 +/- 0.07; R(max), 100.1 +/- 0.7%) caused endothelium-dependent relaxations in precontracted rings of bovine LAD which were competitively and selectively antagonized by the B-1 receptor antagonist, des-Arg(9)-[Leu(8)]-BK (pA(2), 6.27 +/- 0.11) and the B-2 receptor antagonist Hoe-140 (pA(2), 9.63 +/- 0.14), respectively. 3 At 3 h of in vitro incubation, the sensitivity ((p)EC(50), 7.45 +/- 0.10) and R(max) (84.6 +/- 3.3%) to des-Arg(9)-BK were significantly less than those obtained in the same tissues at 6 h ((p)EC(5)0, 7.94 +/- 0.06; R(max), 91.4 +/- 2.5%), whereas endothelium-dependent relaxations to BK and ACh were unaffected by incubation time. 4 Relaxation responses to des-Arg(9)-BK, but not BK, at both 3 h and 6 h were significantly attenuated by the protein synthesis inhibitors, cycloheximide (30 and 100 mu M) and actinomycin D (2 mu M). 5 At 6 h, the nitric oxide (NO) synthase inhibitor, N-G-nitro-L-arginine (L-NOARG, 100 mu M), caused a significant 2 fold decrease in (p)EC(50) (9.58 +/- 0.03) but had no effect on R(max) for BK. For des-Arg(9)-BK, L-NOARG (100 mu M) caused a marked and significant decrease in both the (p)EC(50) and R(max) and revealed contractions to low concentrations of des-Arg(9)-BK. In both cases, L-NOARG inhibition was reversed in the presence of L-arginine (10 mM). 6 At 6 h, removal of the endothelium abolished relaxation responses to des-Arg(9)-BK and BK, and for des-Arg(9)-BK, but not BK, unmasked concentration-dependent contractions ((p)EC(50), 7.57 +/- 0.09; R(max), 83.4 +/- 9.1%). The sensitivity of contractions to des-Arg(9)-BK increased slightly from 3 h ((p)EC(50), 7.37 +/- 0.08) to 6 h ((p)EC(50), 7.62 +/- 0.12) of in vitro incubation; however, there was a small but significant depression in the maximum response over this time (R(max), 126.8 +/- 8.5% and 103.3 +/- 8.6% for 3 h and 6 h of incubation respectively). 7 In conclusion, the bovine LAD contains inducible B-1 and constitutive B-2 endothelial cell kinin receptors, both of which mediate endothelium-dependent relaxation partly via the release of NO. B-1 receptors were also present on the smooth muscle layer of the bovine LAD.