The present study describes the binding to rat striatal A2A adenosine receptors of the new potent and selective antagonist radioligand, [3H]-5-amino-7-(2-phenylethyl)-2-(2-furyl)-pyrazolo[4,3-e]-1,2,4-triazolo[1,5-c]pyrimidine, [3H]-SCH 58261. [3H]-SCH 58261 specific binding to rat striatal membranes (>90%) was saturable, reversible and dependent upon protein concn. Satn. expts. revealed that [3H]-SCH 58261 labeled a single class of recognition sites with high affinity (Kd = 0.70 nM) and limited capacity (apparent Bmax = 971 fmol mg-1 of protein). The presence of 100 μM GTP in the incubation mixt. did not modify [3H]-SCH 58261 binding parameters. Competition expts. showed that [3H]-SCH 58261 binding is consistent with the labeling of A2A striatal receptors. Adenosine receptor agonists competed with the binding of 0.2 nM [3H]-SCH 58261 with the following order of potency: 2-hexynyl-5'-ethylcarboxamidoadenosine (2HE-NECA)>5'-N-ethylcarboxamidoadenosine (NECA)>2-[4-(2-carboxyethyl)phenethylamino]-5'-N-ethylcarboxamidoadenosine (CGS 21680)>2-phenylaminoadenosine (CV 1808)>R-N6-phenylisopropyladenosine (R-PIA)>N6-cyclohexyladenosine (CCPA)>S-N6-phenylisopropyladenosine (S-PIA). Adenosine antagonists inhibited [3H]-SCH 58261 binding in the following order: 5-amino-9-chloro-2-(furyl)-[1,2,4]-triazolo[1,5-c]quinazoline (CGS 15943)>5-amino-8-(4-fluorobenzyl)-2-(2-furyl)pyrazolo[4,3-e]-1,2,4-triazolo[1,5-c]pyrimidine (8FB-PTP)=SCH 58261>xanthine amine congener (XAC)=(E,18%-Z,82%)7-methyl-8-(3,4-dimethoxystyryl)-1,3-dipropylxanthine (KF 17837S)>8-cyclopentyl-1,3-dipropylxanthine (DPCPX)≥8-phenyltheophylline (8-PT). The Ki values for adenosine antagonists were similar to those labeled with the A2A agonist [3H]-CGS 21680. Affinities of agonists were generally lower. The A1-selective agonist, R-PIA, was found to be about 9 fold more potent than its stereoisomer, S-PIA, thus showing the stereoselectivity of [3H]-SCH 58261 binding. Except for 8-PT, the adenosine agonists and antagonists examd. inhibited [3H]-SCH 58261 binding with Hill coeffs. not significantly different from unity. The present results indicate that [3H]-SCH 58261 is the first non-xanthine adenosine antagonist radioligand which directly labels A2A striatal receptors. High receptor affinity, good selectivity and very low non-specific binding make [3H]-SCH 58261 an excellent probe for studying the A2A adenosine receptor subtype in mammalian brain.
Binding of the radioligand [3H]-SCH 58261, a new non-xanthine A2A adenosine receptor antagonist, to rat striatal membranes
BARALDI, Pier Giovanni;
1996
Abstract
The present study describes the binding to rat striatal A2A adenosine receptors of the new potent and selective antagonist radioligand, [3H]-5-amino-7-(2-phenylethyl)-2-(2-furyl)-pyrazolo[4,3-e]-1,2,4-triazolo[1,5-c]pyrimidine, [3H]-SCH 58261. [3H]-SCH 58261 specific binding to rat striatal membranes (>90%) was saturable, reversible and dependent upon protein concn. Satn. expts. revealed that [3H]-SCH 58261 labeled a single class of recognition sites with high affinity (Kd = 0.70 nM) and limited capacity (apparent Bmax = 971 fmol mg-1 of protein). The presence of 100 μM GTP in the incubation mixt. did not modify [3H]-SCH 58261 binding parameters. Competition expts. showed that [3H]-SCH 58261 binding is consistent with the labeling of A2A striatal receptors. Adenosine receptor agonists competed with the binding of 0.2 nM [3H]-SCH 58261 with the following order of potency: 2-hexynyl-5'-ethylcarboxamidoadenosine (2HE-NECA)>5'-N-ethylcarboxamidoadenosine (NECA)>2-[4-(2-carboxyethyl)phenethylamino]-5'-N-ethylcarboxamidoadenosine (CGS 21680)>2-phenylaminoadenosine (CV 1808)>R-N6-phenylisopropyladenosine (R-PIA)>N6-cyclohexyladenosine (CCPA)>S-N6-phenylisopropyladenosine (S-PIA). Adenosine antagonists inhibited [3H]-SCH 58261 binding in the following order: 5-amino-9-chloro-2-(furyl)-[1,2,4]-triazolo[1,5-c]quinazoline (CGS 15943)>5-amino-8-(4-fluorobenzyl)-2-(2-furyl)pyrazolo[4,3-e]-1,2,4-triazolo[1,5-c]pyrimidine (8FB-PTP)=SCH 58261>xanthine amine congener (XAC)=(E,18%-Z,82%)7-methyl-8-(3,4-dimethoxystyryl)-1,3-dipropylxanthine (KF 17837S)>8-cyclopentyl-1,3-dipropylxanthine (DPCPX)≥8-phenyltheophylline (8-PT). The Ki values for adenosine antagonists were similar to those labeled with the A2A agonist [3H]-CGS 21680. Affinities of agonists were generally lower. The A1-selective agonist, R-PIA, was found to be about 9 fold more potent than its stereoisomer, S-PIA, thus showing the stereoselectivity of [3H]-SCH 58261 binding. Except for 8-PT, the adenosine agonists and antagonists examd. inhibited [3H]-SCH 58261 binding with Hill coeffs. not significantly different from unity. The present results indicate that [3H]-SCH 58261 is the first non-xanthine adenosine antagonist radioligand which directly labels A2A striatal receptors. High receptor affinity, good selectivity and very low non-specific binding make [3H]-SCH 58261 an excellent probe for studying the A2A adenosine receptor subtype in mammalian brain.I documenti in SFERA sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.
