Premium
Binding of the radioligand [ 3 H]‐SCH 58261, a new non‐xanthine A 2A adenosine receptor antagonist, to rat striatal membranes
Author(s) -
Zocchi Cristina,
Ongini Ennio,
Ferrara Silvia,
Baraldi Pier Giovanni,
Dionisotti Silvio
Publication year - 1996
Publication title -
british journal of pharmacology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 2.432
H-Index - 211
eISSN - 1476-5381
pISSN - 0007-1188
DOI - 10.1111/j.1476-5381.1996.tb15296.x
Subject(s) - ccpa , radioligand , xanthine , adenosine , cgs 21680 , adenosine receptor , chemistry , adenosine a2a receptor , agonist , stereochemistry , receptor , biochemistry , enzyme
1 The present study describes the binding to rat striatal A 2A adenosine receptors of the new potent and selective antagonist radioligand, [ 3 H]‐5‐amino‐7‐(2‐phenylethyl)‐2‐(2‐furyl)‐pyrazolo[4,3‐ e ]‐1,2,4‐triazolo [1,5‐ c ] pyrimidine, [ 3 H]‐SCH 58261. 2 [ 3 H]‐SCH 58261 specific binding to rat striatal membranes (> 90%) was saturable, reversible and dependent upon protein concentration. Saturation experiments revealed that [ 3 H]‐SCH 58261 labelled a single class of recognition sites with high affinity ( K d = 0.70 nM) and limited capacity (apparent B max = 971 fmol mg −1 of protein). The presence of 100 μ m GTP in the incubation mixture did not modify [ 3 H]‐SCH 58261 binding parameters. 3 Competition experiments showed that [ 3 H]‐SCH 58261 binding is consistent with the labelling of A 2A striatal receptors. Adenosine receptor agonists competed with the binding of 0.2 nM [ 3 H]‐SCH 58261 with the following order of potency: 2‐hexynyl‐5′‐N‐ethyl carboxamidoadenosine (2HE‐NECA) > 5′‐N‐ ethylcarboxamidoadenosine (NECA)>2‐[4‐(2‐carboxyethyl)‐phenethylamino]‐5′‐N‐ethylcarboxamido‐ adenosine (CGS 21680) > 2‐phenylaminoadenosine (CV 1808)> R ‐N 6 ‐phenylisopropyladenosine ( R ‐ PIA) > N 6 ‐cyclohexyladenosine (CHA) = 2‐chloro‐N 6 ‐cyclopentyladenosine (CCPA) > S‐N 6 ‐phenylisopro‐ pyladenosine (S‐PIA). 4 Adenosine antagonists inhibited [ 3 H]‐SCH 58261 binding with the following order: 5‐amino‐9‐chloro‐ 2‐(2‐furyl)‐[1,2,4]‐triazolo[1,5‐ c ] quinazoline (CGS 15943) > 5‐amino‐8‐(4‐fluorobenzyl)‐2‐(2‐furyl)‐pyra‐ zolo [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‐cyclo‐ pentyl‐1,3‐dipropylxanthine (DPCPX) ≥ 8‐phenyltheophylline (8‐PT). 5 The K i values for adenosine antagonists were similar to those labelled with the A 2A agonist [ 3 H]‐CGS 21680. Affinities of agonists were generally lower. The A i ‐selective agonist, R ‐PIA, was found to be about 9 fold more potent than its stereoisomer, S‐PIA, thus showing the stereoselectivity of [ 3 H]‐SCH 58261 binding. Except for 8‐PT, the adenosine agonists and antagonists examined inhibited [ 3 H]‐SCH 58261 binding with Hill coefficients not significantly different from unity. 6 The present results indicate that [ 3 H]‐SCH 58261 is the first non‐xanthine adenosine antagonist radioligand which directly labels A 2A striatal receptors. High receptor affinity, good selectivity and very low non‐specific binding make [ 3 H]‐SCH 58261 an excellent probe for studying the A 2A adenosine receptor subtype in mammalian brain.