Direct visualisation of internalization of the adenosine A3 receptor and localization with arrestin3 using a fluorescent agonist

Stoddart, Leigh A. and Vernall, Andrea J. and Briddon, Stephen J. and Kellam, Barrie and Hill, Stephen J. (2015) Direct visualisation of internalization of the adenosine A3 receptor and localization with arrestin3 using a fluorescent agonist. Neuropharmacology, 98 . pp. 68-77. ISSN 1873-7064

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Abstract

Fluorescence based probes provide a novel way to study the dynamic internalization process of G protein-coupled receptors (GPCRs). Recent advances in the rational design of fluorescent ligands for GPCRs have been used here to generate new fluorescent agonists containing tripeptide linkers for the adenosine A3 receptor. The fluorescent agonist BY630-X-(D)-A-(D)-A-G-ABEA was found to be a highly potent agonist at the adenosine A3 receptor in both reporter gene (pEC50 = 8.48 ± 0.09) and internalization assays (pEC50 = 7.47 ± 0.11). Confocal imaging studies showed that BY630-X-(D)-A-(D)-A-G-ABEA was internalized with A3 linked to yellow fluorescent protein, which was blocked by the competitive antagonist MRS1220. Internalization of untagged adenosine A3 could also be visualized with BY630-X-(D)-A-(D)-A-G-ABEA treatment. Further, BY630-X-(D)-A-(D)-A-G-ABEA stimulated the formation of receptor–arrestin3 complexes and was found to localize with these intracellular complexes. This highly potent agonist with excellent imaging properties should be a valuable tool to study receptor internalization.

Item Type: Article
Keywords: Adenosine receptors; Adenosine A3 receptor; Fluorescent agonist; Internalization; Arrestin3
Schools/Departments: University of Nottingham UK Campus > Faculty of Medicine and Health Sciences > School of Life Sciences
University of Nottingham UK Campus > Faculty of Science > School of Pharmacy
Identification Number: https://doi.org/10.1016/j.neuropharm.2015.04.013
Depositing User: Eprints, Support
Date Deposited: 18 Jul 2016 14:05
Last Modified: 19 Sep 2016 16:11
URI: http://eprints.nottingham.ac.uk/id/eprint/35144

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