Prolonged stimulation of cultured cerebellar neurons by kainic acid (KA) leads to death of neurons first evident from the swelling of soma and neurites. Stimulation is accompanied by increases in [Ca2+]i and [Na+]i as monitored using digital imaging microfluorimetry. “Blebs” tended to form on neurites with the highest increases in [Ca2+]i. Points of Ca2+ entry into neurites via glutamate-receptor-gated channels predicted where approximately 80% of blebs would form tens of minutes later. These sites were close to neurite intersections where there was a high likelihood of synaptic contacts and were enriched in mitochondria as revealed by rhodamine 123 staining. Ca2+, but not Na+ entry, produced a loss of mitochondrial potential. Prolonged KA, but not 50K, applications could fully dissipate the neuronal Na+ gradient. Recovery of resting [Na+]i was delayed by Ca2+ loading. We propose that blebs form at certain synaptic regions due to localized ionic fluxes and local Ca2+ overloading. Increased [Ca2+]i may hamper restoration of normal [Na+]i permitting local osmotic swelling as well as activation of Ca(2+)-dependent enzymes and other processes. Na+ may slow, block, or reverse Na/Ca exchange and enhance swelling. These conditions could not be reproduced by global changes in ion concentrations produced by Ca2+ or Na+ ionophores. The earliest stages of excitotoxicity thus appear to be manifestations of localized disruptions of ionic homeostasis mediated by Ca2+ overload and Na+ influx.