Therapeutic hypothermia in experimental models of focal and global cerebral ischemia and intracerebral hemorrhage.
The commonly used inhalation anesthetic isoflurane has been reported to induce DNA damage and cytotoxicity. However, the methods to attenuate these effects remain largely to be determined. Mild hypothermia has neuroprotective effects. We therefore set out to assess whether mild hypothermia could protect the isoflurane-induced DNA damage and cytotoxicity. Moreover, we investigated the underlying mechanisms by assessing the effects of mild hypothermia on the isoflurane-induced changes in ATP levels. H4 human neuroglioma cells were treated with 2% isoflurane for 3 or 6 h with and without mild hypothermia (35°C). We assessed the cell viability by using 3-(4,5-Dimethylthiazol-2-yl)-2,5-Diphenyltetrazolium Bromide (MTT) and lactate dehydrogenase (LDH) assay. We determined DNA damage by measuring levels of phosphorylation of the histone protein H2A variant X at Ser139 (γH2A.X), the marker of DNA damage. We also measured ATP levels in the cells. Here we showed that the treatment with 2% isoflurane for 6 h induced cytotoxicity and DNA damage in the cells. Moreover, the treatment with 2% isoflurane for 3 h decreased ATP levels without inducing cytotoxicity. Mild hypothermia attenuated the isoflurane-induced cytotoxicity, DNA damage, and ATP reduction in the cells. Taken together, these data suggest that the isoflurane-induced reduction in ATP levels occurred before the isoflurane-induced cytotoxicity. Isoflurane may induce DNA damage and cause cytotoxicity through reducing ATP levels. Mild hypothermia would ameliorate isoflurane-induced DNA damage and cytotoxicity by attenuating the isoflurane-induced reduction in ATP levels. These pilot studies have established a system and will promote the future investigations of anesthesia neurotoxicity.