Absence of direct antioxidant effects from volatile anesthetics in primarymixed neuronal-glial cultures

Background: Volatile anesthetics decrease ischemic brain injury. Mechanisms for this protection remain under investigation. The authors hypothesized t hat volatile anesthetics serve as antioxidants in a neuronal-glial cell cul ture system, Methods: Primary cortical neuronal-glial cultures were prepared from fetal rat brain. Cultures were exposed to iron, H2O2, or xanthine-xanthine oxidas e for 30 min in serum-free media containing dissolved isoflurane (0-3.2 mr) , sevoflurane (0-3.6 mr), halothane (0-4.1 mu), n-hexanol, or known antioxi dants, Cell damage was assessed by release of lactate dehydrogenase (LDH) a nd trypan blue exclusion 24 h later. Lipid peroxidation was measured by the production of thiobarbituric acid-reactive substances in a cell-free lipid system. Iron and calcium uptake and mitochondrial depolarization were meas ured after exposure to iron in the presence or absence of isoflurane. Results: Deferoxamine reduced LDH release caused by H2O2, or xanthine-xanth ine oxidase, but the volatile anesthetics had no effect. Iron-induced LDH r elease was prevented by the volatile anesthetics (maximum effect for haloth ane = 1.2 mM, isoflurane = 1.2 mM, and sevoflurane = 2.1 mM aqueous phase). When corrected for lipid solubility, the three volatile anesthetics were e quipotent against iron-induced LDH release. In the cell-free system, there was no effect of the anesthetics on thiobarbituric acid-reactive substance formation in contrast to Trolox, which provided complete inhibition. Isoflu rane (1.2 mM) reduced mean iron uptake by 46% and inhibited mitochondrial d epolarization but had no effect on calcium uptake. Conclusions: Volatile anesthetics reduced cell death induced by oxidative s tress only in the context of iron challenge. The likely reason for protecti on against iron toxicity is inhibition of iron uptake and therefore indirec t reduction of subsequent intracellular oxidative stress caused by this cha llenge. These data argue against a primary antioxidant effect of volatile a nesthetics.