Stimulation of the angiotensin II type I receptor on bovine adrenal glomerulosa cells activates a temperature-sensitive internalization-recycling pathway.
Angiotensin II (Ang II) is an important regulator of aldosterone production by bovine adrenal glomerulosa cells. On these cells Ang II interacts with the AT1 receptor that is coupled to a G protein controlling the activity of phospholipase C. A primary culture of bovine adrenal glomerulosa cells was used to study the internalization-recycling mechanism of the AT1 receptor after stimulation with Ang II. When cells were pretreated with 10 nM Ang II for 30 min at 37 degrees C and binding studies were performed at 12 degrees C we observed a 48% loss in [125I]Ang II binding. Scatchard analysis revealed that this loss in binding translated into a decreased affinity of the AT1 receptor without any loss in the total amount of binding sites. Under the same conditions an important internalization of [125I]Ang II was invariably observed. These observations suggest that a mechanism was at work to recycle the internalized receptors to the cell surface during the binding studies. Following internalization we indeed observed an externalization of [125I]Ang II. This phenomenon relatively rapid at 37 degrees C was much slower at 12 degrees C and completely inhibited at 4 degrees C. When cells were pretreated with 10 nM Ang II for 30 min at 37 degrees C binding assays at 4 degrees C no longer revealed a loss of binding affinity but rather a 54% reduction in the total amount of binding sites. The maximal binding capacity could be recovered during incubations at 12 degrees C. These results reveal the existence of a dynamic recycling process for the AT1 receptor. In accordance with this interpretation the phenomenon was blocked by monensin, a known inhibitor of receptor recycling. These studies suggest that the stimulation of the AT1 receptor sets in motion an internalization-recycling process that seems to be a fundamental aspect of the AT1 receptor transduction mechanism.
Richard
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DE
Department of Pharmacology, Faculty of Medicine, Université de Sherbrooke, Quebec, Canada.
Chrétien
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L
Caron
M
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Guillemette
G
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D000804:Angiotensin II / Q000378:metabolism / Q000494:pharmacology*; D000818:Animals; D002417:Cattle; D003500:Cyclization; D005455:Fluorescent Antibody Technique; D008985:Monensin / Q000494:pharmacology; D011945:Receptors, Angiotensin / Q000187:drug effects* / Q000378:metabolism; D013696:Temperature; D015384:Zona Glomerulosa / Q000187:drug effects* / Q000378:metabolism
D011945:Receptors, Angiotensin; D000804:Angiotensin II; D008985:Monensin