The nature of the adenosine receptors involved in the enhancement of acetylcholine release in the hippocampus was studied. The A 2 A agonist, CGS 21680, increased the veratridine-evoked release of [ 3 H]acetylcholine from hippocampal synaptosomes. This presynaptic effect of CGS 21680 was greater at 3-30 nM than at 100 nM. The excitatory effect of CGS 21680 was antagonised by the A 2 antagonist, DMPX (10 μM), and by the A 2 A antagonist, CSC (200 nM), but not by the A 1 antagonist, DPCPX (20 nM). We also found co-expression of A 2 A and choline acetyltransferase mRNAs in the nucleus of the diagonal band and the medial septum, where the cholinergic cell bodies that project into the hippocampus are located. These results indicate that A 2 A adenosine receptors are present in cholinergic nerve terminals in the hippocampus and that activation of these receptors enhances acetylcholine release.