Isoflurane inhibits growth but does not cause cell death in hippocampal neural precursor cells grown in culture.
BACKGROUND
Isoflurane causes long-term hippocampal-dependent learning deficits in rats despite limited isoflurane-induced hippocampal cell death, raising questions about the causality between isoflurane-induced cell death and isoflurane-induced cognitive function. Neurogenesis in the dentate gyrus is required for hippocampal-dependent learning and thus constitutes a potential alternative mechanism by which cognition can be altered after neonatal anesthesia. The authors tested the hypothesis that isoflurane alters proliferation and differentiation of hippocampal neural progenitor cells.


METHODS
Multipotent neural progenitor cells were isolated from pooled rat hippocampi (postnatal day 2) and grown in culture. These cells were exposed to isoflurane and evaluated for cell death using lactate dehydrogenase release, caspase activity, and immunocytochemistry for nuclear localization of cleaved caspase 3. Growth was assessed by cell counting and BrdU incorporation. Expression of markers of stemness (Sox2) and cell division (Ki67) were determined by quantitative polymerase chain reaction. Cell fate selection was assessed using immunocytochemistry to stain for neuronal and glial markers.


RESULTS
Isoflurane did not change lactate dehydrogenase release, activity of caspase 3/7, or the amount of nuclear cleaved caspase 3. Isoflurane decreased caspase 9 activity, inhibited proliferation, and decreased the proportion of cells in s-phase. messenger ribonucleic acid expression of Sox2 (stem cells) and Ki67 (proliferation) were decreased. Differentiating neural progenitor cells more often select a neuronal fate after isoflurane exposure.


CONCLUSIONS
The authors conclude that isoflurane does not cause cell death, but it does act directly on neural progenitor cells independently of effects on the surrounding brain to decrease proliferation and increase neuronal fate selection. These changes could adversely affect cognition after isoflurane anesthesia.
Sall
Jeffrey W
JW
Department of Anesthesia and Perioperative Care, University of California, San Francisco, USA. sallj@anesthesia.ucsf.edu
Stratmann
Greg
G
Leong
Jason
J
McKleroy
William
W
Mason
Daniel
D
Shenoy
Shanti
S
Pleasure
Samuel J
SJ
Bickler
Phillip E
PE
D018685:Anesthetics, Inhalation / Q000009:adverse effects*; D000818:Animals; D000963:Antimetabolites / Q000378:metabolism; D001973:Bromodeoxyuridine / Q000378:metabolism; D020169:Caspases / Q000378:metabolism; D016923:Cell Death; D002454:Cell Differentiation / Q000187:drug effects*; D049109:Cell Proliferation / Q000187:drug effects*; D005904:Glial Fibrillary Acidic Protein / Q000378:metabolism; D006624:Hippocampus / Q000166:cytology / Q000187:drug effects; D007150:Immunohistochemistry; D007530:Isoflurane / Q000009:adverse effects*; D019394:Ki-67 Antigen / Q000378:metabolism; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D009474:Neurons / Q000166:cytology / Q000187:drug effects*; D039904:Pluripotent Stem Cells / Q000187:drug effects; D051381:Rats; D017207:Rats, Sprague-Dawley; D055748:SOXB1 Transcription Factors / Q000378:metabolism; D016896:Treatment Outcome
D018685:Anesthetics, Inhalation; D000963:Antimetabolites; D005904:Glial Fibrillary Acidic Protein; D019394:Ki-67 Antigen; D055748:SOXB1 Transcription Factors; C527784:Sox2 protein, rat; D007530:Isoflurane; D007770:L-Lactate Dehydrogenase; D020169:Caspases; D001973:Bromodeoxyuridine