Mechanisms Of Isoflurane Induced Neuronal PC12Cell Apoptosis: Roles Of Inositol1,4,5-trisphosphate Receptors

Objective: Although inhalation anesthetics are safe and effective inclinical anesthesia, but a growing number of experimental studies found thatinhalation anesthetics can induce cytotoxicity, leading to neurodegeneration orpostoperative cognitive dysfunction. Studies show that isoflurane can inducetoxic effects on various types of cells and tissues, but the mechanism ofisoflurane on neuron is still not clear, through the induction of endoplasmicreticulum calcium release lead to intracellular calcium imbalance is animportant mechanism.The regulation of intracellular calcium homeostasis isthe interaction of the three types of receptor from endoplasmic reticulum,including Ca²⁺ATPase, inositol-1,4,5-triphosphate receptor （IP₃R） andryanodine receptor（RyR）. Our purpose is to investigate the role of IP₃R inisoflurane-induced cytotoxicity.Methods: The PC12cells were incubated with50ng/ml2.5s nervegrowth factor （NGF） for7d which possess the properties of neurons, can beused as a neuronal model and widely used in the study of neural physiology,pathology and pharmacology. The neuronal PC12cells were randomly dividedinto four groups （n=6）: control group （group C）, IP₃R antagonist group（group X）, isoflurane group （group I）, isoflurane+IP₃R antagonist group（group I+X）. Group C is without any treatment, Groups I and I+X weretreated with1.2%isoflurane for12h, Groups X and I+X were pretreated withIP₃R antagonist-Xestospongia C100nM for30min. After12h collectedcells respectively, anne-xin V/propidium iodide apoptosis assay andTdT-mediated dUTP Nick-End Labeling （TUNEL） assay were performed toevaluate isoflurane-induced cell apoptosis. Isoflurane-evoked changes of thecalcium concentration ([Ca²⁺]i) in the cytoplasm were measured by flowcytometry. Reverse transcription-polymerase chain reaction （RT-PCR） wasperformed to evaluate IP₃R mRNA expression. Results: Compared with group C, the cell apoptosis rate and [Ca²⁺]iwasno significant difference in group X （P>0.05）, IP₃R mRNA expression waslower in group X（P<0.05）, cell apoptosis rate and [Ca²⁺]iincreased in groups Iand I+X（P<0.05）, IP₃R mRNA expression was higher in group I （P <0.05）,but there was no significant difference in group I+X （P>0.05）. Compared togroup X, the cell apoptosis rate,[Ca²⁺]iand IP₃R mRNA expression increasedin groups I and I+X （P <0.05）. Compared to group I, the cell apoptosis rate,[Ca²⁺]iand the IP₃R mRNA expression decreased in group I+X （P <0.05）.Conclusion: These results suggest that exposure to1MAC isoflurane for12h causes cell apoptosis, the mechanism is intracellular calcium regulationdisorder-calcium overload, may partly by direct activation of IP₃R on the ERmembrane, IP₃R inhibitor could inhibit this effect partly, I-IP₃R is one of theimportant targets.