Effect of ethanol on N-methyl-D-aspartate (NMDA)-induced neurotoxicity in rat dissociated cortical cells (8-12 day cultures) was studied. Treatment of cells with NMDA (50 and 500 microM) for 15 min caused cytotoxic effects on the cells, as examined by microscopic observations and lactate dehydrogenase release from cells 18 h after the treatment. Ca2+ is essential for these effects in medium during treatment. Presence of ethanol (50-300 mM) simultaneously with NMDA protected cells from the cytotoxicity depending on the concentration of ethanol. Calcium accumulation in cells on addition of NMDA, as monitored by fluorescence ratio (F405/F485) of Indo-1-preloaded cortical cells, was also decreased depending on the concentration of added ethanol. APV (200 microM) and ketamine (100 microM) blocked both the cytotoxicity and cellular calcium accumulation due to NMDA. These results suggest that ethanol effects its protection of neurons from NMDA-induced cytotoxicity by blocking the receptor-mediated calcium influx.