title: G protein coupling of CGS 21680 binding sites in the rat hippocampus and cortex is different from that of adenosine A1 and striatal A2A receptors.
abstract: The effect of guanine nucleotide-binding protein (G protein) modifiers on the binding of the adenosine A2A receptor agonist 2-[4-(2-p-carboxyethyl[3H])phenyl-amino]-5'-N-ethylcarboxamidoadenosine ([3H]CGS 21680) and of the adenosine A1 receptor agonist [3H]R-phenylisopropyladenosine ([3H]R-PIA) to rat cortical and striatal membranes was studied. Guanosine 5'-(beta,gamma-imido)triphosphate (1-300 microM), which uncouples all G proteins, more effectively inhibited [3H]CGS 21680 (30 nM) binding in the cortex than [3H]R-PIA (2 nM) binding to cortical or striatal membranes or [3H]CGS 21680 (30 nM) binding in the striatum. N-Ethylmaleimide (1-300 microM) or pertussis toxin (1-100 microg/ml), which uncouple G(i)/G(o) protein-coupled receptors, more effectively inhibited [3H]R-PIA binding to cortical or striatal membranes and [3H]CGS 21680 binding in the cortex than [3H]CGS 21680 binding in the striatum. Cholera toxin (2.5-250 microg/ml), which uncouples G(S) protein-coupled receptors, more effectively inhibited [3H]CGS 21680 binding in the striatum than [3H]CGS 21680 binding in the cortex and less effectively inhibited [3H]R-PIA binding to cortical or striatal membranes. Treatment of solubilised cortical membranes with pertussis toxin (50 microg/ml) decreased [3H]CGS 21680 (30-100 nM) binding which almost fully recovered after reconstitution with G(i)/G(o) proteins. The K(i) for displacement of [2-3H]-(4{2-[7-amino-2-(2-furyl)(1,2,4)triazolo(2,3-a)(1,3,5)triazin+ ++-5-ylamino]ethyl}phenol) ([3H]ZM 241385, 1nM) by CGS 21680 was 110 nM (95%CI: 98-122 nM) in non-treated, 230 (167-292) nM in pertussis toxin (25 microg/ml)-treated and 222 (150-295) nM in cholera toxin (50 microg/ml)-treated cortical membranes; in contrast, the K(i) for displacement of [3H]-5-amino-7-(2-phenylethyl)-2-(2-furyl)-pyrazolo(4,3-e)-1,2,4-triazol o(1,5-c)pyrimidine ([3H]SCH 58261, 1 nM) by CGS 21680 was 74 (57-91) nM in non-treated, 71 (44-100) nM in pertussis toxin-treated and 147 (100-193) nM in cholera toxin-treated cortical membranes. Finally, CGS 21680 displaced monophasically the binding of the A1 antagonist, [3H]8-cyclopentyl-1,3-dipropylxanthine (2 nM), and the A1 agonist, [3H]R-PIA (2 nM), in 2 or 10 mM Mg(2+)-medium, either at 25 degrees C or 37 degrees C, in cortical or striatal membranes. These results indicate that CGS 21680 does not bind to A1 receptors and that limbic CGS 21680 binding sites differ from striatal-like A2A receptors since they couple to G(i)/G(o) proteins, as well as to G(s) proteins.
authors.affiliation: Laboratory of Neurosciences, Faculty of Medicine, University of Lisbon, Portugal. racunha@neurociencias.pt
authors.forename: R A
authors.initials: RA
authors.lastname: Cunha
authors.forename: M D
authors.initials: MD
authors.lastname: Constantino
authors.forename: J A
authors.initials: JA
authors.lastname: Ribeiro
mesh_terms: D000241:Adenosine / Q000031:analogs & derivatives* / Q000378:metabolism / Q000494:pharmacology; D000818:Animals; D001665:Binding Sites; D001667:Binding, Competitive / Q000187:drug effects; D001921:Brain / Q000187:drug effects / Q000378:metabolism*; D002540:Cerebral Cortex / Q000187:drug effects / Q000378:metabolism; D002772:Cholera Toxin / Q000494:pharmacology; D003342:Corpus Striatum / Q000187:drug effects / Q000378:metabolism; D005033:Ethylmaleimide / Q000494:pharmacology; D019204:GTP-Binding Proteins / Q000378:metabolism*; D006165:Guanylyl Imidodiphosphate / Q000494:pharmacology; D006624:Hippocampus / Q000187:drug effects / Q000378:metabolism; D008297:Male; D008566:Membranes / Q000187:drug effects / Q000378:metabolism; D037342:Pertussis Toxin; D010627:Phenethylamines / Q000378:metabolism* / Q000494:pharmacology; D058906:Purinergic P1 Receptor Agonists; D011743:Pyrimidines / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D018047:Receptors, Purinergic P1 / Q000378:metabolism*; D014227:Triazines / Q000378:metabolism; D014230:Triazoles / Q000378:metabolism; D014316:Tritium; D010566:Virulence Factors, Bordetella / Q000494:pharmacology
chemical_list: C098657:5-amino-7-(2-phenylethyl)-2-(2-furyl)pyrazolo(4,3-e)-1,2,4-triazolo(1,5-c)pyrimidine; D010627:Phenethylamines; D058906:Purinergic P1 Receptor Agonists; D011743:Pyrimidines; D018047:Receptors, Purinergic P1; D014227:Triazines; D014230:Triazoles; D010566:Virulence Factors, Bordetella; C097270:ZM 241385; D014316:Tritium; C061282:2-(4-(2-carboxyethyl)phenethylamino)-5'-N-ethylcarboxamidoadenosine; D006165:Guanylyl Imidodiphosphate; D002772:Cholera Toxin; D037342:Pertussis Toxin; D019204:GTP-Binding Proteins; D000241:Adenosine; D005033:Ethylmaleimide