The inhaled anesthetic, isoflurane, enhances Ca2+-dependent survival signaling in cortical neurons and modulates MAP kinases, apoptosis proteins and transcription factors during hypoxia.
We tested whether the protection of hypoxic neurons by the inhaled anesthetic isoflurane is related to the Ca2+-dependent phosphorylation of MAP kinases and anti-apoptotic co-factors. In cultures of mouse cortical neurons we measured changes in the phosphorylation of Ca2+-dependent and Ca2+-independent MAP kinases, transcription factors, and apoptosis regulators after hypoxia or hypoxia combined with isoflurane (1% in gas phase). In hypoxic neurons, isoflurane reduced cell death and TUNEL staining by >80%. Isoflurane released Ca2+ from intracellular stores, increasing [Ca2+]i in oxygenated neurons by approximately 20%. Neuroprotection was associated with a smaller increase in [Ca2+]i in hypoxic neurons and required IP3 receptors and phospholipase C. In hypoxic neurons, isoflurane increased the phosphorylation of the Ca2+-dependent MAP kinases Pyk2 and p42/44 (ERK). The Ca2+-independent MAP kinase p38 pathway showed increased phosphorylation with isoflurane but not with ionomycin, a Ca2+ ionophore. JNK was phosphorylated in hypoxic neurons in the presence of isoflurane, as was the transcription factor c-Jun; JNK inhibition with SP600125 prevented both phosphorylation of c-Jun and neuroprotection. Isoflurane decreased phosphorylation of the pro-apoptotic cofactors Bad and p90RSK and increased Akt phosphorylation. However, with the exception of c-Jun, transcription factors (Elk-1, GSK-3, Forkhead, p90RSK) decreased or remained unchanged. We conclude that isoflurane's protection of hypoxic cortical neurons involves signaling that includes changes in intracellular Ca2+ regulation, several MAP kinase pathways and modulation of apoptosis regulators.
Bickler
Philip E
PE
Severinghaus-Radiometer Research Laboratories, Department of Anesthesia and Perioperative Care, University of California at San Francisco, San Francisco, California 94143-0542, USA. bicklerp@anesthesia.ucsf.edu
Fahlman
Christian S
CS
D018685:Anesthetics, Inhalation / Q000494:pharmacology*; D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D002118:Calcium / Q000378:metabolism*; D015687:Cell Hypoxia; D002478:Cells, Cultured; D002540:Cerebral Cortex / Q000187:drug effects*; D048049:Extracellular Signal-Regulated MAP Kinases / Q000502:physiology; D020287:In Situ Nick-End Labeling; D007530:Isoflurane / Q000494:pharmacology*; D020935:MAP Kinase Signaling System / Q000187:drug effects*; D051379:Mice; D018696:Neuroprotective Agents / Q000494:pharmacology*; D010766:Phosphorylation; D051057:Proto-Oncogene Proteins c-akt / Q000502:physiology; D014157:Transcription Factors / Q000378:metabolism*; D010738:Type C Phospholipases / Q000502:physiology; D048051:p38 Mitogen-Activated Protein Kinases / Q000502:physiology
D018685:Anesthetics, Inhalation; D018696:Neuroprotective Agents; D014157:Transcription Factors; D007530:Isoflurane; D051057:Proto-Oncogene Proteins c-akt; D048049:Extracellular Signal-Regulated MAP Kinases; D048051:p38 Mitogen-Activated Protein Kinases; D010738:Type C Phospholipases; D002118:Calcium