DETECTION OF ADENOSINE RECEPTOR ANTAGONISTS IN RAT-BRAIN USING A MODIFIED RADIORECEPTOR ASSAY

The present study describes a modified radioreceptor binding assay usi ng brain homogenate or serum from drug treated animals as the 'competi ng drug' in a conventional in vitro radioligand binding assay. Method validation involved measurement of the brain and serum concentration o f three adenosine receptor antagonists following systemic administrati on, using a [H-3]g-cyclopentyl-1, 3-dipropylxanthine ([H-3]DPCPX) bind ing assay. The intrinsic [H-3]DPCPX binding capacity of test samples w as abolished by protein denaturation (80 degrees C, 15 min) and, endog enous ligand was depleted enzymatically, prior to determination of dru g concentration. Brain and serum concentrations of the adenosine A, re ceptor antagonist, DPCPX increased in a dose related manner when measu red 20 min after intraperitoneal injection. Estimated brain concentrat ions were 13.8, 87.7 and 288 nM following injection of 0.01, 0.1 and 1 .0 mg/kg DPCPX, and serum concentrations were 26.5, 195 and 1370 nM re spectively. A time dependent decrease in both brain and serum concentr ation was noted 20-180 min following injection of 1.0 mg/kg DPCPX. The peripheral adenosine receptor antagonists, 1, 3-dipropyl-8-p-sulphoph enylxanthine (DPSPX; 5.6 mg/kg) and 8-(p-sulphophenyl)theophylline (8- PST; 20 mg/kg), were not detected in brain tissue 20 min after intrape ritoneal injection, despite serum concentrations of 56 and 52 mu M res pectively. This assay provides a useful and versatile method for deter mining the central penetration of neuroactive drugs. (C) 1997 Elsevier Science B.V.