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

The effect of guanine nucleotide-binding protein (G protein) modifiers on the binding of the adenosine A(2A) receptor agonist 2-[4-(2-p-carboxyethyl [H-3])phenylamino]-5'-N-ethylcarboxamidoadenosine ([H-3]CGS 21680) and of the adenosine Al receptor agonist [H-3]R-phenylisopropyladenosine ([H-3]R-PIA) to rat cortical and striatal membranes was studied. Guanosine 5'-(beta,gamma-imido)triphosphate (1-300 mu M), which uncouples all G proteins, more effectively inhibited [H-3]CGS 21680 (30 nM) binding in the cortex than [H-3]R-PIA (2 nM) binding to cortical or striatal membranes or [H-3]CGS 21680 (30 nM) binding in the striatum. N-Ethylmaleimide (1-300 mu M) or pertussis toxin (1-100 mu g/ml), which uncouple G(i)/G(o) protein-coupled receptors, more effectively inhibited [H-3]R-PIA binding to cortical or striatal membranes and [H-3]CGS 21680 binding in the cortex than [H-3]CGS 21680 binding in the striatum. Cholera toxin (2.5-250 mu g/ml), which uncouples G, protein-coupled receptors, more effectively inhibited [H-3]CGS 21680 binding in the striatum than [H-3]CGS 21680 binding in the cortex and less effectively inhibited [H-3]R-PIA binding to cortical or striatal membranes. Treatment of solubilised cortical membranes with pertussis toxin (50 mu g/ml) decreased [H-3]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-H-3]-(4-( 2-[7-amino-2-(2-furyl)(1,2,4)triazolo(2,3-a)triazin-5-ylamino]ethyl }phenol) ([H-3]ZM 241385, 1 nM) by CGS 21680 was 110 nM (95%CI: 98-122 nM) in non-treated, 230 (167-292) nM in pertussis toxin (25 mu g/ml)-treated and 222 (150-295) nM in cholera toxin (50 mu g/ml)-treated cortical membranes; in contrast, the K-i for displacement of [H-3]-5-amino-7-(2-phenylethyl)-2-(2-furyl)-pyrazolo(4,3- e)-1,2,4-triazolo(1,5-c)pyrimidine ([H-3]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 A(1) antagonist, [H-3]8-cyclopentyl-1,3-dipropylxanthine (2 nM), and the A(1) agonist, [H-3]R-PIA (2 nM), in 2 or 10 mM Mg2+-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 A(1) receptors and that limbic CGS 21680 binding sites differ from striatal-like A(2A) receptors since they couple to G(i)/G(o) proteins, as well as to G(s) proteins.