Cyanide-induced alterations of cytosolic calcium levels and cytotoxicity were examined in primary cultures of rat hippocampus. Cytosolic free Ca2+ ([Ca2+]i) levels were measured in hippocampal neurons using the fluorescent probe, fura 2. A concentration-dependent rise in [Ca2+]i occurred rapidly following exposure of cells to 0.5-10 mM NaCN. In normal medium (1.3 mM Ca2+), 2 mM NaCN produced an increase in [Ca2+]i (172 +/- 27% of control), 45 sec following exposure. Ca2+ elevation produced by NaCN was blocked by removal of Ca2+ from the external medium or by pretreatment with the N-methyl-D-aspartate (NMDA) receptor antagonist, 2-amino-5-phosphonovalerate (APV). The cytotoxicity of cyanide, assessed by measuring the efflux of lactate dehydrogenase, was blocked by APV. These results indicate that in hippocampal neurons, cytosolic Ca2+ accumulation induced by cyanide originates from the extracellular compartment and the NMDA receptor ionophore is a significant route for Ca2+ entry. It is proposed that excitotoxic mechanisms may contribute to altered neuronal homeostasis and injury associated with cyanide.