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Isoflurane has either neuroprotective or neurotoxic effects. High-dose oxygen is frequently used throughout the perioperative period. We hypothesized that hyperoxia will affect cell viability of rat pheochromocytoma (PC12) cells that were exposed to isoflurane and reactive oxygen species (ROS) may be involved. PC12 cells were exposed to 1.2% or 2.4% isoflurane for 6 or 24 h respectively, and cell viability was evaluated. To investigate the effects of hyperoxia, PC12 cells were treated with 21%, 50%, or 95% oxygen and 2.4% isoflurane for 6 h, and cell viability, TUNEL staining, ROS production, and expression of B-cell lymphoma 2 (BCL-2), BCL2-associated X protein (BAX), caspase-3 and beta-site APP cleaving enzyme (BACE) were measured. ROS involvement was evaluated using the ROS scavenger 2-mercaptopropiopylglycine (MPG). The viability of cells exposed to 2.4% isoflurane was lower than that of cells exposed to 1.2% isoflurane. Prolonged exposure (6 h vs. 24 h) to 2.4% isoflurane resulted in a profound reduction in cell viability. Treatment with 95% (but not 50%) oxygen enhanced the decrease in cell viability induced by 2.4% isoflurane alone. Levels of ROS, Bax, caspase-3 and BACE were increased, whereas expression of Bcl-2 was decreased, in cells treated with 95% oxygen plus 2.4% isoflurane compared with the control and 2.4% isoflurane plus air groups. MPG attenuated the effects of oxygen and isoflurane. In conclusion, isoflurane affects cell viability in a dose- and time-dependent manner. This effect is augmented by hyperoxia and may involve ROS, the mitochondrial apoptotic signaling pathway, and β-amyloid protein. [ABSTRACT FROM AUTHOR]

• Isoflurane preconditioning protects against ischemia in vivo and in vitro model. • Applying 2-MPG just before reperfusion abrogated isoflurane preconditioning in vivo. • ROS scavenging for 10 min at the onset of reperfusion blocked isoflurane preconditioning. • ROS after OGD in an isoflurane preconditioning group was lower than in an OGD group. • ROS after reperfusion were increased by scavenging ROS immediately after OGD. In this in vivo and in vitro study, we aimed to investigate whether isoflurane preconditioning-induced neuronal protection is mediated by reactive oxygen species (ROS) signaling at the reperfusion stage. In the in vivo study, Sprague-Dawley rats were subjected to middle cerebral artery occlusion (MCAO) and in the in vitro study, rat pheochromocytoma (PC12) cells were subjected to oxygen glucose deprivation (OGD). Isoflurane preconditioning was carried out prior to MCAO or OGD and the ROS scavenger, N-2-mercaptopropiopylglycine (2-MPG), was administered at the start of reperfusion. Infarct volume, neurological severity score, and TUNEL staining were analyzed in the in vivo study and cell viability, Bcl-2/Bax ratio, cleaved caspase 3/caspase 3 ratio, and ROS fluorescence intensity were measured in the in vitro study. In the in vivo study, infarct volume, neurological severity score, and TUNEL-positive cell count were significantly decreased with preconditioning but were abrogated by administration of 2-MPG. In the in vitro study, cell viability and Bcl-2/Bax ratio were significantly increased with preconditioning, and cleaved caspase-3/caspase-3 ratio and ROS fluorescence intensity were significantly decreased. Administration of 2-MPG for 10 min abrogated this preconditioning effect, but it did not abolish the protection when administered for 60 min of reperfusion. Isoflurane preconditioning-induced protection was abolished by ROS scavengers at the start of reperfusion, indicating that ROS signaling can mediate the isoflurane preconditioning effect, which suggests that the time window can be important. [ABSTRACT FROM AUTHOR]