Autoregulation of bradykinin receptors and bradykinin-induced prostacyclin formation in human fibroblasts.

Abstract

A s bstract. The interaction of bradykinin (BK) with its specific receptors on intact cultured human fibroblasts results in production ofprostaglandins, including prostacyclin (PG12), and accumulation of cyclic AMP. Incubation of cells with 1 ,tM BK for 5 min at 37°C led to a marked reduction (75-90%) in BK-induced PGI2 release and in total number of [3H]BK-binding sites with no change in dissociation constant (6.1 and 7.6 nM for control and BK-treated cells, respectively). The decrease in receptor number did not result from BK transferred from the first incubation into the binding assay. BK-induced receptor loss was temperature dependent; exposure of cells to BK at 40C had little or no effect on receptor number. After incubation with BK for _ 15 min, further incubation in the absence of BK for 30 min at 370C almost completely restored both receptor number and BK-induced PGI2 release. With more prolonged exposure to BK (>1 h), restoration of receptors was inversely related to the length of exposure and the concentration ofBK. Recovery was unaffected by cycloheximide. During prolonged incubation without removal of BK, cells began to recover receptors by 5 h; >99% of the bradykinin initially present disappeared by 3 h. Bacitracin greatly retarded BK disappearance and totally prevented recovery. These observations provide direct evidence that the num-

Regulation of bradykinin receptor level by cholera toxin, pertussis toxin and forskolin in cultured human fibroblasts

“…The procedure used was that of Roscher et al (1983), with minor modifications. Briefly, serum-deprived cells previously (D Macmillan Press Ltd, 1991 exposed to toxins, forskolin and/or protein synthesis inhibitors were washed in chilled phosphate-buffered saline and set in an ice bath.…”

Guanosine Nucleotides Regulate B2 Kinin Receptor Affinity of Agonists But Not of Antagonists: Discussion of a Model Proposing Receptor Precoupling to G Protein

“…The role of G proteins, however, in the variety of BK signal cascades is not fully elucidated. Human foreskin fibroblasts expressing high numbers of B2 kinin receptor have already been shown to be an excellent tool to study the regulation and the properties of the human B2 kinin receptor, as well as its signal transduction pathways (Roscher et al, 1983(Roscher et al, , 1984Dengler et al, 1990;Faussner et al, 1991). Therefore, we decided to use this cell system to investigate the regulatory role of guanosine nucleotides in the binding of agonist and antagonist to B2 kinin receptors in particulate fractions of human foreskin fibroblasts.…”

“…Cultured human foreskin fibroblasts express high numbers of B2 kinin receptor, which, when occupied by agonist, are subject to strict regulatory control via internalization and down-regulation (Roscher et al, 1984). Although G proteins appear to couple these receptors to the synthesis of eicosanoids, thereby accumulating cAMP and the release of inositol phosphates (Roscher et al, 1983;Burch and Axelrod, 1987;Dengler et al, 1990), little is known regarding the exact mechanism of the interaction between the B2 kinin receptor and these G proteins.…”

“…Although G proteins appear to couple these receptors to the synthesis of eicosanoids, thereby accumulating cAMP and the release of inositol phosphates (Roscher et al, 1983;Burch and Axelrod, 1987;Dengler et al, 1990), little is known regarding the exact mechanism of the interaction between the B2 kinin receptor and these G proteins. In this study, we compared the effect of guanine nucleotides on B2 kinin receptor agonist and antagonist binding to partic-ulate fractions of human foreskin fibroblasts at 0°C and at 37°C.…”

Untitled

“…VIP and bumetanide, a loop diuretic like piretanide, were found to be active only on the basolateral face (Dharmsathaphom, 1985 (Cuthbert & Margolius, 1982), that chloride will prove to be a major current carrier, as found with VIP (Dharmsathaphorn, 1985) and forskolin. The desensitization following kinin may be due to rapid down regulation as with human fibroblasts (Roscher et al, 1984). Nevertheless desensitizing concentrations on one side of the monolayers did not compromise contralateral responsiveness.…”