8-(3-ISOTHIOCYANATOSTYRYL)CAFFEINE IS A SELECTIVE, IRREVERSIBLE INHIBITOR OF STRIATAL A(2)-ADENOSINE RECEPTORS

8-(3-Isothiocyanatostyryl)caffeine (ISC) was synthesized and shown to inhibit selectively the binding of [H-3]CGS 21680 (an A2a-selective ag onist) at adenosine receptors in striatal membranes. The K(i) value at A2a-receptors was found to be 110 nM (rat), with selectivity ratios f or A2a versus A1-receptors in rat, guinea pig, bovine, and rabbit stri atum of >100-fold. Preincubation of membranes with ISC caused a dose-d ependent, irreversible antagonism of the binding of [H-3]CGS 21680, wi th an IC50 value of 3 muM. The irreversibility is likely due to the pr esence of the chemically reactive isothiocyanate group, since the bind ing of the corresponding analogue in which the isothiocyanate was repl aced with a chloro group was completely reversible. The potency of ISC to irreversibly inhibit the binding of [H-3]CGS 21680 in several spec ies varied in the order rat almost-equal-to guinea pig > bovine almost -equal-to rabbit. In all four species, binding of the A1-selective ago nist [H-3]R-N6-phenylisopropyladenosine was not diminished by pre-trea tment with 2 muM ISC. The kinetics of irreversible inhibition of rat A 2a-receptors by 2 muM ISC gave a t1/2 of approximately 3 min. Followin g partial inactivation, the remaining rat A2a-binding sites retained t he same K(d) value as in control membranes for saturation by [H-3]CGS 21680. Thus, ISC appears to be a selective affinity label for A2a-vers us A1-receptors in the brain. (C) 1993 Wiley-Liss, Inc.