COMPARISON OF A(4) AND A(2A) BINDING-SITES IN STRIATUM AND COS CELLS TRANSFECTED WITH ADENOSINE A(2A) RECEPTORS

A putative A(4) adenosine receptor is characterized by a distinct structure activity profile of compounds in competition for [H-3]2-phenylaminoadenosine ([H-3]CV 1808) binding sites on rat brain membranes assayed at 4 degrees C. We now confirm that A(4) binding sites can be demonstrated on ice-cold membranes of rat striatum and demonstrate a similar binding site on COS cells transfected with rat A(2a) adenosine receptors (COS/A(2a)). The characteristic A(4) potency order is: CV 1808 > [1R-(1 alpha,2 alpha,3 beta,5 beta)]-3-(2,6-diamino-N-2-(3-carbethoxyphenyl)-9H- purin-9-yl)-5'-(N-ethylcarbamoyl)-1,2-cyclopentanediol (CGS 22988) much greater than 5'-N-ethylcarboxamidoadenosine (NECA) greater than or equal to 2-[4-(2-carboxyethyl)phenylethylamino]-5'-N-ethylcarboxamidoadenosine (CGS 21680); 9-chloro-2-(2-furyl)[1,2,4]-triaolo[1,5-c]-quinazolin-5-amine (CGS 15943) only partially inhibits binding at 1 mu M. If [H-3]CGS 21680 is used for ice-cold assays, or if either [H-3]CV 1808 or [H-3]CGS 21680 are used for assays at 21 degrees C, the potency order of competing compounds changes markedly and becomes characteristic of A(2a) adenosine receptor binding sites; CGS 15943 greater than or equal to CGS 21680 similar or equal to NECA > CGS 22988 greater than or equal to CV 1808. Binding of [H-3]CGS 21680, but not [H-3]CV 1808, is significantly enhanced by the pore-forming antibiotic, alamethicin. Guanosine 5'-O-(3-thiotriphosphate) decreases the binding of both radioligands to striatal membranes at 21 degrees C significantly more than to membranes on ice. We propose that differential effects of temperature on the binding characteristics of compounds with distinct physicochemical properties to various pools of a single A(2a) adenosine receptor can result in A(4) and A(2a) binding profiles.