期刊
EUROPEAN JOURNAL OF PHARMACOLOGY
卷 590, 期 1-3, 页码 36-42出版社
ELSEVIER SCIENCE BV
DOI: 10.1016/j.ejphar.2008.05.036
关键词
G protein-coupled receptor; endocytosis; cyclic AMP; CREB; nucleoside; Rab5; clathrin
资金
- Swedish Brain Research Foundation
- Swedish Research Council
- Karolinska Institutet
- Alex and Eva Wallstrom Foundation
- Tore Nilsons Foundation for Medical Research
- Jeanssons Foundation
- Ake Wiberg Foundation
- Signe & Olof Wallenius Foundation
- Lakaresallskapet/Socialstyrelsen
- NIH, National Institute of Diabetes and Digestive and Kidney Diseases
G protein-coupled receptors, such as the adenosine A(2A) receptor, are dynamic proteins, which undergo agonist-dependent redistribution from the cell surface to intracellular membranous compartments, such as endosomes. In order to study the kinetics of adenosine A(2A) receptor redistribution in living cells, we synthesized a novel fluorescent agonist, Alexa488-APEC Alexa488-APEC binds to adenosine A(2A) (K-i=149 +/- 27 nM) as well as A(3) receptors (K-i=240 +/- 160 nM) but not to adenosine A, receptors. Further, we characterized the dose-dependent increase in Alexa488-APEC-induced cAMP production as well as cAMP response element binding (CREB) protein phosphorylation, verifying the ligand's functionality at adenosine A(2A) but not AN receptors. In live-cell imaging studies, Alexa488-APEC-induced adenosine A(2A) receptor internalization, which was blocked by the competitive reversible antagonist ZM 241385 and hyperosmolaric sucrose. Further, internalized adenosine A(2A) receptors co-localized with clathrin and Rab5, indicating that agonist stimulation promotes adenosine A(2A) receptor uptake through a clathrin-dependent mechanism to Rab5-positive endosomes. The basic characterization of Alexa488-APEC described here showed that it provides a useful tool for tracing adenosine A(2A) receptors in vitro. (c) 2008 Elsevier B.V. All rights reserved.
作者
我是这篇论文的作者
点击您的名字以认领此论文并将其添加到您的个人资料中。
推荐
暂无数据