The action of 2-amino-4-phosphonobutyric acid (APB) on mossy fiber-induced excitation in CA3 neurons was studied in vitro with thin sections of guinea pig hippocampus. D- and DL-APB suppressed field potentials induced in regio CA3 by granular layer stimulation. Threshold concentration of DL-APB to induce the suppression was 2--5 microM. D-APB was about 10-fold less potent than DL-APB. Field potentials induced by fimbrial stimulation were much less affected. DL-APB was without effect on antidromic activation of granule cells. 2-Amino-phosphonovaleric acid had a similar but less potent action. Gamma-D-glutamylglycine and cis-2,3-piperidine dicarboxylic acid were almost ineffective. DL-APB suppressed excitatory postsynaptic potentials recorded intracellularly from CA3 neurons in response to granular layer stimulation but caused no marked changes in resting potentials, action potentials and membrane conductance. Single cell discharges induced by iontophoretic administration of glutamate (Glu) or aspartate (Asp) were unaffected when mossy fiber-induced excitation was suppressed by D- or DL-APB. DL-APB had no suppressing action on Glu- or Asp-induced depolarizing potentials. These results indicate that APB is relatively specific blocker of synaptic transmission between mossy fibers and CA3 neurons, and that this action does not result from blockade of receptors for Glu or Asp.