Abstract We identified the possible endogenous factor effective to modulate the binding of [ 3H]-labeled excitatory amino acid agonists and antagonists in the 100,000 × g supernatant of Triton X-100 (0.01%)-treated cell membranes from frog spinal cords. The factor inhibited the binding of [ 3H]glutamate to Triton X-100-treated cell membranes, to which the binding capacity of [ 3H]glutamate increased much more than that to intact cell membranes. The binding capacities of [ 3H]AMPA (an AMPA type agonist) and [ 3H]CPP (an NMDA type antagonist) to cell membranes remained low by Triton treatment, but they were enhanced significantly by the addition of the factor. The effect of the factor on the [ 3H]kainate binding was hardly observable. The factor may provide key information on receptor structures and the classification of receptor types concerning excitatory amino acids in the mammalian central nervous system.