title: Effect of recurrent epileptiform discharges induced by magnesium-free treatment on developing cortical neurons in vitro.
abstract: 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.
authors.affiliation: Division of Child Neurology, Department of Pediatrics, Peking University First Hospital, No. 1, Xi'anmen Dajie, Beijing 100034, PR China.
authors.forename: Hai-Yan
authors.initials: HY
authors.lastname: Cao
authors.forename: Yu-Wu
authors.initials: YW
authors.lastname: Jiang
authors.forename: Zhen-Wei
authors.initials: ZW
authors.lastname: Liu
authors.forename: Xi-Ru
authors.initials: XR
authors.lastname: Wu
mesh_terms: D000818:Animals; D017209:Apoptosis; D002478:Cells, Cultured; D002540:Cerebral Cortex / Q000196:embryology / Q000503:physiopathology; D003470:Culture Media; D004594:Electrophysiology; D004622:Embryo, Mammalian; D004827:Epilepsy / Q000503:physiopathology; D008274:Magnesium / Q000378:metabolism; D008564:Membrane Potentials / Q000502:physiology*; D008928:Mitochondria / Q000378:metabolism; D023421:Models, Animal; D009474:Neurons / Q000473:pathology* / Q000502:physiology*; D018408:Patch-Clamp Techniques; D051381:Rats; D017208:Rats, Wistar; D012640:Seizures / Q000503:physiopathology*
chemical_list: D003470:Culture Media; D008274:Magnesium