In the present study, pharmacological properties of a
bradykinin B(2) receptor amplified either from guinea-pig ileum or lung and homologous to the previously reported sequence except two
amino-acid changes L(124)-->P and N(227)-->Y in the receptor
protein were characterized. Tritiated
bradykinin ([(3)H]-BK) specifically bound to the cloned guinea-pig
B(2)
bradykinin receptor stably expressed in Chinese hamster ovary cells (CHO-K1) with a K(D) value of 0.29+/-0.07 nM. In competition experiments,
bradykinin (BK) affinity constant value was 0.21+/-0.05 nM while the two specific
kinin B(1)
ligands,
des-Arg(9)-bradykinin (DBK) and des-Arg(9)-Leu(8)-bradykinin (DLBK) were unable to compete with [(3)H]-BK. As the specific
peptide antagonist D-Arg-[Hyp(3),Thi(5),D-
Tic(7),Oic(8)]-
bradykinin (
HOE140), (E)-3-(6-acetamido-3-pyridil)-N-[-N-[2,4-dichloro-3-[(2-methyl-8-quinolinyl)oxymethyl]phenyl]-N-methylaminocarbonylmethyl]acrylamide (
FR173657) and 1-[[3-[2,4-dimethylquinolin-8-yl)oxymethyl] - 2,4 - dichloro - phenyl]sulfonyl] - 2(S) - [[4-[4-(aminoiminomethyl)-phenylcarbonyl]piperazin-1-yl]carbonyl]
pyrrolidine (LF16-0335C) exhibited a high affinity for this receptor with K(i) values of 7.34+/-2.45 nM and 8.54+/-1.55 nM respectively. BK and
kallidin (KD) increased
inositol phosphates (IPs) levels with EC(50) values of 0.44+/-0.12 nM and 6.88+/-0.28 nM, respectively. Neither DLBK nor DBK (0.01 nM to 10 microM) stimulated or inhibited IPs turnover and as expected
HOE140 did not raise IPs production.
HOE140 (0.1 microM) and LF 16-0335c (1 microM) right shifted the BK response curve with pK(B) values of 9.2+/-0.4 and 8.4+/-0.3, respectively. The results indicate that this cloned guinea-pig receptor displayed typical pharmacological properties of a
bradykinin B(2) receptor and support the existence of a single
B(2) receptor in this species.