Pharmacological and functional characterization of the guinea‐pig B 2 bradykinin receptor stably expressed in CHO‐K1 cell line

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 n M . In competition experiments, bradykinin (BK) affinity constant value was 0.21±0.05 n M 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 n M and 8.54±1.55 n M respectively. BK and kallidin (KD) increased inositol phosphates (IPs) levels with EC 50 values of 0.44±0.12 n M and 6.88±0.28 n M , respectively. Neither DLBK nor DBK (0.01 n M to 10 μ M ) stimulated or inhibited IPs turnover and as expected HOE140 did not raise IPs production. HOE140 (0.1 μ M ) and LF 16‐0335c (1 μ M ) 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.British Journal of Pharmacology (2002) 135 , 462–468; doi: 10.1038/sj.bjp.0704494