title: The role of the N-methyl-D-aspartate receptor in ketamine-induced apoptosis in rat forebrain culture.
abstract: Recent data suggest that anesthetic drugs may cause widespread and dose-dependent apoptotic neurodegeneration during development. The window of vulnerability to this neurotoxic effect, particularly with N-methyl-D-aspartate (NMDA) antagonists such as ketamine, is restricted to the period of synaptogenesis. The purposes of this study are to determine whether treatment of forebrain cultures with ketamine results in a dose-related increase in neurotoxicity and whether upregulation of NMDA receptor subunit NR1 promotes ketamine-induced apoptosis. Forebrain cultures were treated for 12 h with 0.1, 1, 10 and 20 microM ketamine or co-incubated with NR1 antisense oligonucleotide (2 microM). After washout of the ketamine, cultures were kept in serum-containing medium (in presence of glutamate) for 24 h. Application of ketamine (10 and 20 microM) resulted in a substantial increase in DNA fragmentation as measured by cell death enzyme-linked immunosorbent assay, increased number of terminal dUTP nick-end labeling positive cells, and a reduction in mitochondrial metabolism of the dye 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide. No significant effect was seen in the release of lactate dehydrogenase, indicating that cell death presumably occurred via an apoptotic mechanism. Co-incubation of ketamine with NR1 antisense significantly reduced ketamine-induced apoptosis. Western analysis showed that neurotoxic concentrations of ketamine increased Bax and NR1 protein levels. NR1 antisense prevented this increase caused by ketamine, suggesting that ketamine-induced cell death is associated with a compensatory upregulation of the NMDA receptor. These data suggest that NR1 antisense offers neuroprotection from apoptosis in vitro, and that upregulation of the NR1 following ketamine administration is, at least, partially responsible for the observed apoptosis.
authors.affiliation: Division of Neurotoxicology, National Center for Toxicological Research, Food and Drug Administration, Jefferson, AR 72079-0502, USA. cwang@nctr.fda.gov
authors.forename: C
authors.initials: C
authors.lastname: Wang
authors.forename: N
authors.initials: N
authors.lastname: Sadovova
authors.forename: X
authors.initials: X
authors.lastname: Fu
authors.forename: L
authors.initials: L
authors.lastname: Schmued
authors.forename: A
authors.initials: A
authors.lastname: Scallet
authors.forename: J
authors.initials: J
authors.lastname: Hanig
authors.forename: W
authors.initials: W
authors.lastname: Slikker
mesh_terms: D000778:Anesthetics, Dissociative / Q000633:toxicity*; D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D015153:Blotting, Western; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D004797:Enzyme-Linked Immunosorbent Assay; D020287:In Situ Nick-End Labeling; D007649:Ketamine / Q000633:toxicity*; D016376:Oligonucleotides, Antisense / Q000494:pharmacology; D016548:Prosencephalon / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology*; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000187:drug effects / Q000378:metabolism; D051381:Rats; D016194:Receptors, N-Methyl-D-Aspartate / Q000378:metabolism*; D013873:Thionucleotides / Q000494:pharmacology; D051028:bcl-2-Associated X Protein
chemical_list: D000778:Anesthetics, Dissociative; C494828:Bax protein, rat; D016376:Oligonucleotides, Antisense; D019253:Proto-Oncogene Proteins c-bcl-2; D016194:Receptors, N-Methyl-D-Aspartate; D013873:Thionucleotides; D051028:bcl-2-Associated X Protein; D007649:Ketamine