title: Ketamine-Induced Toxicity in Neurons Differentiated from Neural Stem Cells.
abstract: Ketamine is used as a general anesthetic, and recent data suggest that anesthetics can cause neuronal damage when exposure occurs during development. The precise mechanisms are not completely understood. To evaluate the degree of ketamine-induced neuronal toxicity, neural stem cells were isolated from gestational day 16 rat fetuses. On the eighth day in culture, proliferating neural stem cells were exposed for 24 h to ketamine at 1, 10, 100, and 500 μM. To determine the effect of ketamine on differentiated stem cells, separate cultures of neural stem cells were maintained in transition medium (DIV 6) for 1 day and kept in differentiation medium for another 3 days. Differentiated neural cells were exposed for 24 h to 10 μM ketamine. Markers of cellular proliferation and differentiation, mitochondrial health, cell death/damage, and oxidative damage were monitored to determine: (1) the effects of ketamine on neural stem cell proliferation and neural stem cell differentiation; (2) the nature and degree of ketamine-induced toxicity in proliferating neural stem cells and differentiated neural cells; and (3) to provide information regarding receptor expression and possible mechanisms underlying ketamine toxicity. After ketamine exposure at a clinically relevant concentration (10 μM), neural stem cell proliferation was not significantly affected and oxidative DNA damage was not induced. No significant effect on mitochondrial viability (3-(4,5-dimethylthiazole-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay) in neural stem cell cultures (growth medium) was observed at ketamine concentrations up to 500 μM. However, quantitative analysis shows that the number of differentiated neurons was substantially reduced in 10 μM ketamine-exposed cultures in differentiation medium, compared with the controls. No significant changes in the number of GFAP-positive astrocytes and O4-positive oligodendrocytes (in differentiation medium) were detected from ketamine-exposed cultures. The discussion focuses on: (1) the doses and time-course over which ketamine is associated with damage of neural cells; (2) how ketamine directs or signals neural stem cells/neural cells to undergo apoptosis or necrosis; (3) how functional neuronal transmitter receptors affect neurotoxicity induced by ketamine; and (4) advantages of using neural stem cell models to study critical issues related to ketamine anesthesia.
authors.affiliation: Office of the Director, National Center for Toxicological Research/Food and Drug Administration, Jefferson, AR, 72079, USA, William.slikker@fda.hhs.gov.
authors.forename: William
authors.initials: W
authors.lastname: Slikker
authors.forename: Fang
authors.initials: F
authors.lastname: Liu
authors.forename: Shuo W
authors.initials: SW
authors.lastname: Rainosek
authors.forename: Tucker A
authors.initials: TA
authors.lastname: Patterson
authors.forename: Natalya
authors.initials: N
authors.lastname: Sadovova
authors.forename: Joseph P
authors.initials: JP
authors.lastname: Hanig
authors.forename: Merle G
authors.initials: MG
authors.lastname: Paule
authors.forename: Cheng
authors.initials: C
authors.lastname: Wang
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D001253:Astrocytes / Q000166:cytology / Q000187:drug effects; D002118:Calcium / Q000378:metabolism; D002455:Cell Division / Q000187:drug effects; D002478:Cells, Cultured; D002540:Cerebral Cortex / Q000166:cytology / Q000196:embryology; D003470:Culture Media / Q000494:pharmacology; D004249:DNA Damage; D004305:Dose-Response Relationship, Drug; D045604:Extracellular Fluid / Q000378:metabolism; D005998:Glycine / Q000494:pharmacology; D036341:Intercellular Signaling Peptides and Proteins / Q000494:pharmacology; D007649:Ketamine / Q000633:toxicity*; D008928:Mitochondria / Q000187:drug effects; D016202:N-Methylaspartate / Q000494:pharmacology; D009336:Necrosis; D009410:Nerve Degeneration / Q000139:chemically induced / Q000473:pathology; D058953:Neural Stem Cells / Q000166:cytology*; D055495:Neurogenesis / Q000187:drug effects; D009474:Neurons / Q000166:cytology / Q000187:drug effects*; D009836:Oligodendroglia / Q000166:cytology / Q000187:drug effects; D051381:Rats; D017207:Rats, Sprague-Dawley
chemical_list: D003470:Culture Media; D036341:Intercellular Signaling Peptides and Proteins; D016202:N-Methylaspartate; D007649:Ketamine; D002118:Calcium; D005998:Glycine