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Excitotoxic cell death and delayed rescue in human neurons derived from NT2 cells
1Department of Pharmacology, University of Pennsylvania School of Medicine, Philadelphia 19104, USA.
Abstract:
The excitotoxic response of NT2-N cells, a clonal line of human teratocarcinoma cells that are terminally differentiated into neuron-like cells, was examined using several endpoints. A 15 min exposure to glutamate produced a dose-dependent toxicity with a maximal cell loss of 80-90% in 6 week old cells. The rapidly triggered excitotoxicity induced by glutamate was blocked by NMDA selective antagonists, was calcium dependent and pH sensitive and could be mimicked by NMDA but not by non-NMDA agonists, AMPA, kainate or quisqualate. The non-NMDA agonists however caused toxicity on prolonged exposure. The NMDA receptor modulators glycine and spermidine enhanced glutamate-mediated toxicity whereas ifenprodil potently and completely inhibited toxicity suggesting that the toxic response is mediated by the NR1/NR2B combination of NMDA subunits. These cells can be rescued from death up to 1 hr after removal of glutamate by NMDA receptor blockade, removal of extracellular Ca2+ or lowering of pH. The extent of rescue is directly related to the time elapsed before intervention. Blockage of NMDA receptor activity for 1 hr immediately after removal of glutamate is both necessary and sufficient for complete rescue. Glutamate-mediated toxicity was not prevented by nitric oxide synthase inhibitors nor was nitric oxide synthase detected in NT2-N cells indicating that nitric oxide is not required for glutamate-mediated excitotoxicity. In summary, NT2-N cells exhibit a robust excitotoxic response and represent a novel model system in which to study the molecular basis of excitotoxic cell death.
Insights
Glutamate causes significant cell death in neuron-like NT2-N cells via NMDA receptors. Interventions like NMDA receptor blockade can rescue cells, highlighting a new model for excitotoxicity research.
Area of Science:
- Neuroscience
- Cell Biology
- Toxicology
Background:
- Excitotoxicity, a neuronal cell death mechanism, is implicated in neurological disorders.
- NT2-N cells, differentiated human teratocarcinoma cells, offer a model for neuronal function.
- Understanding glutamate-induced excitotoxicity is crucial for neuroprotection strategies.
Purpose of the Study:
- To investigate the excitotoxic response of NT2-N cells to glutamate.
- To characterize the receptors and pathways involved in glutamate-mediated excitotoxicity.
- To establish NT2-N cells as a model system for studying excitotoxic cell death.
Main Methods:
- NT2-N cells were exposed to glutamate and other agonists.
- NMDA receptor antagonists, calcium dependency, and pH sensitivity were assessed.
- Glycine, spermidine, and ifenprodil were used as NMDA receptor modulators.
- Rescue experiments involved NMDA receptor blockade, calcium removal, and pH alteration.
Main Results:
- Glutamate induced dose-dependent toxicity in NT2-N cells, with maximal cell loss observed.
- Excitotoxicity was mediated by NMDA receptors, specifically the NR1/NR2B subunit combination.
- Non-NMDA agonists caused toxicity upon prolonged exposure.
- Cells could be rescued from glutamate-induced death through timely interventions.
Conclusions:
- NT2-N cells provide a robust model for studying excitotoxicity.
- The NR1/NR2B subunit is critical for glutamate-mediated excitotoxicity in these cells.
- Intervention targeting NMDA receptors can rescue neurons from excitotoxic death.