Electrophysiological and biochemical techniques were used to characterize adenosine receptors in rat hippocampus. The site which mediates the inhibitory action of adenosine on excitatory synaptic transmission and on spontaneous interictal spiking had properties similar to the adenosine A1 receptor. Thus, the relative order of potency for adenosine analogs was L-PIA greater than or equal to CHA greater than NECA greater than 2CA (L-PIA = N6-phenylisopropyladenosine; CHA = N6-cyclohexyl-adenosine; NECA = adenosine 5'-ethylcarboxamide; 2CA = 2-chloroadenosine), with EC50 values for the most potent analogs between 10-30 nM. The effect of the stable adenosine analog, particularly CHA and L-PIA, was slow in onset and very slowly reversible. This is suggested to be due both to a slow dissociation of these compounds from the receptors but particularly to the slow equilibrium between the concentration of the drug in the medium surrounding the slices and the biophase within the slices. Adenosine analogs bound specifically to membrane preparations of the rat hippocampus with the order of potency 3H-CHA greater than or equal to 3H-L-PIA greater than 3H-NECA. Eadie-Hofstee plots of the binding data were curvilinear for each ligand, but only for 3H-L-PIA could the existence of two binding sites with different apparent Kd-values (0.27 and 11.8 nM) be confirmed by curve-fitting. The estimated Kd-values for CHA and NECA were 1.5 and 20 nM, respectively. The adenosine analogs also enhanced 3H-cyclic AMP accumulation in 3H-adenine-labelled hippocampal slices.(ABSTRACT TRUNCATED AT 250 WORDS)