ABSTRACT
Bradykinin (BK) binding sites were studied in membranes from bovine aorta. [3H]BK specifically bound to one high-affinity binding site (KD = 152 pM; Bmax = 4.6 fmol.mg-1) and was displaced by unlabeled BK (Ki = 121 pM). The B2-agonist kallidin and B2-antagonists D-Arg0[Hyp3,Leu5,8,Gly6,D-Phe7]BK, D-Arg0[Hyp3,D-Phe7]BK, [D-Phe7]BK, and [Thi5,8,D-Phe7]BK inhibited [3H]BK binding with respective Ki values of 101, 282, 678, 2000 and 6000 pM. The B1-antagonist des-Arg9[Leu8]BK had no effect. GTP, GTP gamma S, GDP, and GDP beta S but not 5'-GMP, guanosine, cyclic 3',5'-GMP, ATP, ADP, 5'-AMP, nor adenosine, inhibited [3H]BK binding with an IC50 of 1-3 microM for GTP and GDP and an IC50 of 0.1-0.3 microM for GTP gamma S and GDP beta S. GTP and GDP at 3 microM decreased the Bmax value by 30-70%. Millimolar concentrations of Ca2+ and Mg2+ ions increased [3H]BK binding and counteracted the effect of guanine nucleotides. This study demonstrates the existence of a specific high-affinity B2 BK binding site in bovine aortic endothelial cells. It suggests that this site is located on a G protein-interacting receptor.
Subject(s)
Bradykinin/metabolism , Endothelium, Vascular/metabolism , Guanine Nucleotides/pharmacology , Amino Acid Sequence , Animals , Aorta , Binding Sites , Cattle , Cells, Cultured , Molecular Sequence Data , Receptors, Bradykinin , Receptors, Neurotransmitter/antagonists & inhibitors , Receptors, Neurotransmitter/metabolismABSTRACT
The role of vascular smooth muscle cells (VSMC) in the intraparietal conversion of angiotensin I (AngI) to angiotensin II (AngII) was investigated in rat aortic tissue. The responses of rat aortic vascular smooth muscle cells to AngI and AngII were assessed by studying contraction of endothelium-denuded aortic rings and by measuring intracellular Ca++ ion concentration in primary cultures of VSMC free of endothelial cells. In both preparations, AngI and AngII induced identical responses which were inhibited by saralasin, a blocker of AngII receptors. In the presence of captopril, an inhibitor of the angiotensin converting enzyme, the increase in calcium caused by AngI was abolished in VSMC cultures and the contractile effect of this peptide in aortic rings was strongly decreased, whereas the responses to AngII remained unaffected. These results demonstrate that VSMC are able to convert AngI to AngII.
