Abstract Using the single-electrode voltage-clamp technique, we have examined the effects of a non-N- methyl- d-aspartate (NMDA) antagonist, Joro spider toxin (JSTX), and of an NMDA antagonist, zinc, on excitatory postsynaptic currents (EPSCs) evoked by stimulation of stratum radiatum in CA1 pyramidal cells of the guinea-pig hippocampal slice. Pressure application of a synthesized JSTX (JSTX-3) at 10–200 μM greatly reduced the EPSCs ( 14 19 cells). The block by JSTX-3 was observed in pyramidal cells where the EPSCs showed linear peak current-voltage (I–V) relations in the control. EPSCs remaining after JSTX-3 application showed non-linear peak I–V relationships ( 10 14 cells), and were blocked by puff application of the selective NMDA receptor antagonist dl-2-amino-5-phosphonovalerate (APV) at 200 μM ( 6 10 cells). In the presence of JSTX-3, the decay time constant of the EPSC was increased and was less affected by membrane potential. JSTX-3 had no detectable effects on EPSCs apparently mediated solely by NMDA receptor. These observations suggest that JSTX-3 blocks excitatory synaptic transmission mainly by suppressing non-NMDA-receptor-mediated EPSCs, and that the JSTX-3-insensitive component is mediated at least in part by NMDA receptors in the hippocampal slice. Zinc (100–200 μM) reversibly attenuated EPSCs ( 6 9 cells) and appeared to block a slower component of the EPSCs, suggesting that mainly NMDA receptor-mediated currents were affected.