Fluorescent Ligands of the Bradykinin B₁ Receptors: Pharmacologic Characterization and Application to the Study of Agonist-Induced Receptor Translocation and Cell Surface Receptor Expression

Abstract

ABSTRACT

Met-Lys-bradykinin-Ser-Ser, a peptide produced by the neutrophil from kininogen, is metabolically activated by angiotensin converting enzyme in vascular tissue

“…The binding assay of 1 nM [ 3 H]Lys-des-Arg 9 -BK (Perkin Elmer Life Sciences; 88 Ci/mmol) to HEK 293a cells stably expressing B 1 R-YFP [8] was performed as described [9]. These 2 assays, based on pharmacologically functional rabbit receptors that are fluorescent (continuous stable expression judged from visual inspection) and similar to their human counterparts, were applied to construct binding competition curves for a series of unlabeled peptides.…”

N-terminal extended conjugates of the agonists and antagonists of both bradykinin receptor subtypes: Structure–activity relationship, cell imaging using ligands conjugated with fluorophores and prospect for functionally active cargoes

“…The isolated rabbit aorta is an established bioassay for the B 1 R that has been exploited to characterize novel B 1 R agonists or antagonists [4]; B 2 R ligands were assayed by the contraction of the human isolated umbilical vein [5,3,14]. Cumulative concentration effect curves were constructed for novel agonists and their potencies estimated by the halfmaximal concentration (EC 50 ) and compared to that of reference agonists of natural sequence for each receptor subtype.…”

“…FURA-2 fluorometry was applied to detached cells as described [4] to assess the effects of some novel ligands on kinin receptor signaling.…”

“…Cumulative concentration effect curves were constructed for novel agonists and their potencies estimated by the halfmaximal concentration (EC 50 ) and compared to that of reference agonists of natural sequence for each receptor subtype. Antagonists were introduced 30 min before the construction of the concentration-effect curves of the reference agonist (BK for the B 2 R, des-Arg 9 -BK for the B 1 R) and the shift to the right was used to determine their potencies using the Schild regression and pA 2 scale [3,4].…”

“…Note: Reproduced from [4] with permission from ASPET, except for data related to B-10324 (cited from [16]). …”