Sevoflurane activates PI3-K/Akt/P~(70S6K) kinase cell-survival signaling pathways and protects neuron against ischemia-reperfusion / 中国药理学通报

ABSTRACT

Aim To investigate effects of Sevoflurane on PI_3-K/Akt/P~(70S6K) cell-survival signal transduction pathways after neuron ischemia-reperfusion,and explore neuroprotection mechanisms of sevoflurane.Methods Newborn(24～48 h)Wister rats were decapitated and hippocampus tissue was dissected and cut into 1 mm?1 mm?1 mm pieces.Then digestion with 0.125% trypsin,centrifuged at 800 r?min~(-1) for 5 min at 4℃,and suspended in a medium containing DMEM supplemented to 25 mmol?L~(-1) glucose,10% fetal bovine serum,10% horse serum,and 2 mmol?L~(-1) glutamine.Cells were plated at 1.0?10~5?ml~(-1) on poly-Dlysine-treated 96-well(100 ?l/well)plates as well 6-well(2 ml/well) plates.Cultures were treated with 10 ?mol?L~(-1) cytosine arabinoside on day 4 in culture to minimize glial growth.One-half of the medium was replaced twice a week with medium containing DMEM(4.5 g?L~(-1) glucose)/F12(1 ∶1),5% fetal bovine serum and 5% horse serum.Cells were used after 7 days. For ischemia-reperfusion(oxygen glucose deprivation,OGD)experiments,cultures were washed three times in a glucose-free balanced salt solution(BSS)and placed in deoxygenated glucose-free medium and sealed under 95% N_2-5% CO_2 in an anaerobic chamber equilibrated to 37℃ and 100% humidity for 45 min.OGD was terminated by replacement of stored medium and by returning the cultures to a standard incubator maintained at 37℃ in 95% O_2-5% CO_2.Experimental group cells were respectively carried out OGD,OGD+2% Sevoflurane,OGD+2% Sevoflurane +10 ?mol?L~(-1) LY294002,OGD+2% Sevoflurane +10 ?mol?L~(-1) Triciribin,and OGD+2% Sevoflurane +10 nmol?L~(-1) Rapamycin.Control cells were cultured normally.Group Sevo was carried out OGD meanwhile anesthesized with 2% sevoflurane.Group LY,Tri and Rap cells was carried out OGD meanwhile culture medium was added 10 ?mol?L~(-1) LY294002,10 ?mol?L~(-1) Triciribin or 10 nmol?L~(-1) Rapamycin,and anesthesized with 2% sevoflurane.Compound remained present throughout the duration of the experiment until analysis 24 h later.Neuron viability and apoptosis were measured.The protein expression of PI_3-K,Akt and P~(70S6K) were detected.Results Sevoflurane enhanced expression of PI_3-K,Akt and P~(70S6K),meanwhile increased neuron viability and decreased neuron apoptosis(vs group I/R,P