1 The amino acid, D-aspartate, exists in the mammalian brain and is an agonist at the N-methyl-D-aspartate (NMDA) subtype of ionotropic glutamate receptors. Here, for the first time, we studied the actions of D-aspartate on alpha-amino-3-hydroxyl-5-methyl-4-isoxazolepropionate receptors (AMPARs) in acutely isolated rat hippocampal neurons. 2 In the presence of the NMDA receptor channel blocker, MK801, D-aspartate inhibited kainate-induced AMPAR current in hippocampal neurons. The inhibitory action of D-aspartate on kainate-induced AMPAR current was concentration-dependent and was voltage-independent in the tested voltage range (-80 to +60 mV). 3 The estimated EC50 of the L-glutamate-induced AMPAR current was increased in the presence of D-aspartate, while the estimated maximum L-glutamate-induced AMPAR current was not changed. D-aspartate concentration-dependently shifted the dose-response curve of kainate to the right. Schild plot analysis indicated that D-aspartate acts competitively to block AMPARs. The K(b) for D-aspartate was estimated to be 0.93 mM. 4 D-aspartate also blocked L-glutamate-induced current in Xenopus laevis oocytes that expressed recombinant homomeric AMPARs. 5 NMDA possessed similar inhibitory action on AMPARs. However, L-aspartate had little inhibitory action on AMPARs. 6 D-Aspartate, but not L-aspartate, was found to reduce the amplitude of miniature excitatory postsynaptic current in cultured hippocampal neurons. 7 Our data are consistent with a model in which D-aspartate directly competes with kainate and L-glutamate in binding to the agonist binding site of AMPARs. The prevalence of D-aspartate in the brain suggests a possible role of D-aspartate in modulating AMPAR-mediated fast excitatory synaptic transmission.