1. The effects of metabotropic glutamate receptor (mGluR) agonists on excitatory transmission at mossy fibre-CA3 synapses were studied in rat hippocampal slice preparations using both extracellular and whole-cell clamp recording techniques. 2. Application of a novel and potent mGluR2/mGluR3-specific agonist (2S,1'R,2'R,3'R)-2-(2,3-dicarboxycyclopropyl)glycine (DCG-IV, 0.1 microM) reversibly suppressed field excitatory postsynaptic potentials evoked by mossy fibre stimulation. DCG-IV at the same concentration did not affect other glutamatergic excitatory transmissions at the commissural/associational input to CA3 or at the Schaffer collateral/commissural input to CA1 regions. 3. This suppressing effect of DCG-IV on mossy fibre transmission was dose dependent and partly antagonized by a competitive mGluR antagonist (+)-methyl-4-carboxylphenylglycine (1 mM). 4. The field potential changes induced by pressure application of glutamate (0.1 mM) to the stratum lucidum of the CA3 region was unaffected by 0.1 microM DCG-IV. 5. In whole-cell clamp experiments, 0.1 microM DCG-IV suppressed excitatory postsynaptic currents evoked by mossy fibre stimulation without inducing detectable inward current in CA3 neurons, and paired-pulse facilitation was enhanced by DCG-IV application. 6. These results suggest that mGluR2/mGluR3 are specifically expressed at mossy fibre synapses in the hippocampal CA3 region, and activation of the receptor suppresses synaptic transmission by an action on a presynaptic site.