Effect of recurrent epileptiform discharges induced by magnesium-free treatment on developing cortical neurons in...

As seizures in infants and children often originate from the neocortex, neocortical epilepsy models may be appropriate for studying epileptiform activity and seizure-induced injury in the developing nervous system. However, the characterization of epileptiform activity or seizure-induced injury in cultured developing cortical neurons has seldom been reported. Therefore, We attempted to establish a cultured developing cortical neuronal epilepsy model, and to study the subsequent effect on neurons. Cultures were exposed to Mg(2+)-free media for 3 h, and then returned to regular media. Using whole-cell patch-clamp intracellular recording techniques, we found that spontaneously recurrent epileptiform discharges for at least 72 h could be induced after transient Mg(2+)-free treatment. Neuron morphology following Mg(2+)-free treatment demonstrated no prominent alterations. At different time points (6, 24 and 72 h) after Mg(2+)-free treatment, neuronal viability, identified by trypan blue staining and LDH activity, and apoptosis, measured by flow cytometry, showed modest but non-significant (P>0.05) changes compared with the age-matched control group after various culture periods (6 and 17 days) in vitro. Mitochondrial metabolic activity, measured by MTT assay, significantly decreased by 15% at 6 h after Mg(2+)-free treatment (P<0.05) in neurons cultured for 6 days, and at 24 h showed a 29% decrease in neurons cultured for 17 days (P<0.05). In conclusion, brief Mg(2+)-free treatment constitutes a cultured developing cortical neuron 'seizure' model, and can induce transient mitochondrial dysfunction without cell loss.