Short-term desensitization of the angiotensin II receptor of bovine adrenal glomerulosa cells corresponds to a shift from a high to a low affinity state.
Angiotensin II (Ang II) is an important regulator of aldosterone production by bovine adrenal glomerulosa (BAG) cells. Ang II interacts with a specific receptor coupled to a guanyl nucleotide-binding protein (G protein) that controls the activity of phospholipase C. A primary culture of BAG cells was used to study short-term desensitization of the Ang II receptor. After short exposures to Ang II, BAG cells lost some [125I]Ang II binding capacity. This loss was dependent on the duration of the pretreatment and on the concentration of Ang II used. A maximal loss of [125I]Ang II binding of 55 +/- 10% was observed after a pretreatment of 30 min with 30 nM Ang II. The EC50 was 1.3 +/- 0.6 nM (mean +/- SD of three experiments). The desensitization was readily reversible, since most of the binding capacity (higher than 90%) was recovered after a 60-min incubation, at 37 C, in the absence of Ang II. Scatchard studies revealed that the Ang II receptor of BAG cells exists under two affinity states with one dissociation constant of 0.2 nM and another dissociation constant of 1.5 nM. After a 30-min exposure of BAG cells to 10 nM Ang II, an important decrease of high affinity binding sites was observed. The maximal amount of binding sites was similar on control and desensitized cells (around 52,000 receptors per cell). GTP gamma S, a potent activator of G proteins, decreased [125I]Ang II binding to permeabilized BAG cells. This GTP gamma S effect was not observed on permeabilized BAG cells that had previously been desensitized with 10 nM Ang II. These results suggested that, similarly to GTP gamma S, short exposure to 10 nM Ang II caused the uncoupling of Ang II receptor from its G protein. DuP-753 (a selective AT1 angiotensin II type 1 receptor antagonist) markedly unhibited, whereas PD-123319 (a selective AT2 angioten II type 2 receptor antagonist) had no effect on Ang II receptor desensitization, indicating that the AT1 receptor subtype was responsible for the observed phenomenon. Pretreatment of BAG cells with staurosporine (a protein kinase C inhibitor) and R24571 (a calmodulin inhibitor) did not modify Ang II-induced desensitization of AT1 receptor.(ABSTRACT TRUNCATED AT 400 WORDS)
Department of Pharmacology, Faculty of Medicine, University of Sherbrooke, Quebec, Canada.
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Boulay
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Chrétien
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DE
Richard
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Guillemette
D000450:Aldosterone / Q000378:metabolism; D000470:Alkaloids / Q000494:pharmacology; D000804:Angiotensin II / Q000378:metabolism / Q000494:pharmacology; D000818:Animals; D001713:Biphenyl Compounds / Q000494:pharmacology; D002417:Cattle; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D019204:GTP-Binding Proteins / Q000502:physiology; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000494:pharmacology; D007093:Imidazoles / Q000494:pharmacology; D019808:Losartan; D011485:Protein Binding; D047428:Protein Kinase Inhibitors; D011494:Protein Kinases / Q000502:physiology; D011945:Receptors, Angiotensin / Q000819:agonists / Q000032:analysis* / Q000502:physiology*; D019311:Staurosporine; D013777:Tetrazoles / Q000494:pharmacology; D013997:Time Factors; D015384:Zona Glomerulosa / Q000737:chemistry* / Q000166:cytology* / Q000378:metabolism
D000470:Alkaloids; D001713:Biphenyl Compounds; D007093:Imidazoles; D047428:Protein Kinase Inhibitors; D011945:Receptors, Angiotensin; D013777:Tetrazoles; D000804:Angiotensin II; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D000450:Aldosterone; D011494:Protein Kinases; D019204:GTP-Binding Proteins; D019311:Staurosporine; D019808:Losartan