Flow cytometry application for studies on adenosine A2A receptors expression.
Adenosine A2A receptors belong to the heptaspanning membrane receptors family A, also known as G protein-coupled receptors. In human brain they are highly expressed in striatum, where they co-exist and co-function with adenosine A1, glutamate mGlu5 and dopamine D2 receptors. As glutaminergic neurotransmission modulators in GABAergic enkephalinergic neurons, adenosine A2A receptors are attractive targets for new, alternative therapies of neurodegenerative disorders, like Parkinson's disease and Huntington's disease. The aim of the research was to obtained fluorescently tagged adenosine A2A receptors. Gene encoding human adenosine A2A receptor was inserted into plasmid pEYFP-N1, bearing enhanced yellow fluorescent protein (EYFP). The construct was expressed in HEK 293 cells. Fluorescence was observed by flow cytometry and epifluorescence microscopy. Functional ligand binding properties were investigated by saturation binding analysis of adenosine A2A receptors specific agonist [3H] CGS 21680.
Wójcik
Tomasz
T
Department of Technology and Biotechnology of Drugs, Faculty of Pharmacy, Jagiellonian University Medical College, Kraków, Poland. twojcik@cm-uj.krakow.pl
Bereta
Michał
M
Faron-Górecka
Agata
A
Dziedzicka-Wasylewska
Marta
M
Kieć-Kononowicz
Katarzyna
K
D000241:Adenosine / Q000031:analogs & derivatives / Q000494:pharmacology; D058908:Adenosine A2 Receptor Agonists; D001426:Bacterial Proteins / Q000378:metabolism; D002460:Cell Line; D005434:Flow Cytometry / Q000379:methods*; D020869:Gene Expression Profiling / Q000379:methods*; D006801:Humans; D007668:Kidney / Q000378:metabolism; D008024:Ligands; D008164:Luminescent Proteins / Q000378:metabolism; D008856:Microscopy, Fluorescence / Q000379:methods*; D010627:Phenethylamines / Q000494:pharmacology; D010957:Plasmids / Q000378:metabolism; D011485:Protein Binding; D011869:Radioligand Assay; D043705:Receptor, Adenosine A2A / Q000235:genetics*
D058908:Adenosine A2 Receptor Agonists; D001426:Bacterial Proteins; D008024:Ligands; D008164:Luminescent Proteins; D010627:Phenethylamines; D043705:Receptor, Adenosine A2A; C054562:yellow fluorescent protein, Bacteria; C061282:2-(4-(2-carboxyethyl)phenethylamino)-5'-N-ethylcarboxamidoadenosine; D000241:Adenosine