Although adenosine has been considered to depress release of neurotransmitter in central synapses, evidence for this notion is circumstantial. To obtain more direct and quantitative evidence, the suppressing action of adenosine on transmission through the mossy fiber synapse was examined by a quantal analysis in thin transverse sections of the guinea pig hippocampus. Adenosine suppressed excitatory postsynaptic potentials (EPSPs) induced in CA3 neurons by activation of single mossy fibers to 57 +/- 19% at 10 microM (6 neurons). The input resistance of the postsynaptic neurons was reduced to 84 +/- 5%. The suppression of the unitary EPSPs was accompanied by a reduction of the mean quantal content (m) to 51 +/- 13% with no significant decrease in mean quantal amplitude (q). These findings indicate that adenosine at 10 microM suppresses transmission through mossy fiber synapses mainly by depressing release of transmitter. This study provides direct evidence for the presynaptic action of adenosine on central synapses for the first time.