The effects of barbiturates on radioligand binding to inhibitory Ri adenosine receptors of rat brain membranes were investigated. Binding of the adenosine receptor agonist (-)N6-phenylisopropyl[3H]adenosine and the antagonist 1,3-diethyl-8-[3H]phenylxanthine was inhibited by several barbiturates. This inhibition was concentration-dependent and occurred in the range of pharmacologically effective concentrations. Pentobarbital was the most potent of the barbiturates tested with a Ki of 92 mumol/l. The (+)isomers of hexobarbital and mephobarbital were more potent than the respective (-)isomers. Barbituric acid itself did not displace either radioligand in concentrations up to 1 mmol/l. The inhibitory effect of pentobarbital was reversed by a single wash of membranes preincubated with the barbiturate. The presence of pentobarbital caused a decrease of the affinity of the receptor for the antagonist radioligand but did not alter the number of binding sites, suggesting a competitive antagonism. The effects of pentobarbital on radioligand binding to the receptor were not changed by the presence of picrotoxinin nor by the absence of chloride ions. This indicates that they are not mediated via the picrotoxinin binding site. The barbiturates could not be classified as either agonists or antagonists at the Ri adenosine receptor. The presence of GTP did not influence the inhibition of radioligand binding by pentobarbital; this is also observed for antagonists, whereas the affinity of agonists is markedly reduced by GTP. Binding of antagonists to the receptor is enthalpy-driven; the interaction of pentobarbital with the receptor was entropy-driven and the same was true for agonists.(ABSTRACT TRUNCATED AT 250 WORDS)