To examine the effect of microglial conditioning medium (MCM) on damaged neurons, cultured neurons exposed to glutamate were incubated with MCM. More neurons survived in these groups, particularly in the case of glutamate stimulation of microglia. Therefore cultured neurons exposed to glutamate were incubated with MCM at predetermined intervals after glutamate stimulation. Neuronal cultures using MCM at 1, 2 and 6 h after glutamate stimulation had more surviving cells than those using unstimulated MCM. Neuronal cultures using MCM at 12 and 24 h had as much surviving cells as those using unstimulated MCM. These results indicate that the neuroprotective effect of microglia appears only in the early phase after treatment of glutamate and disappears in the late phase.