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Objective @#To explore the effects of autophagy activation on propofol⁃induced apoptosis in hippocampal neurons.@*Methods @#Primary hippocampal neurons were extracted and isolated from fetal rats and randomly divided into control group, propofol group, rapamycin group and chloroquine group. 2,3⁃Bis⁃(2⁃methoxy⁃4⁃nitro⁃5⁃sulfo phenyl) Ⅳ2H⁃tetrazolium⁃5⁃carboxanilide ( XTT) and flow cytometry were used to detect apoptosis, and Western blot was used to detect the expression of autophagy and apoptosis⁃related proteins. Then, aged rats were randomly divided into control group, propofol group, rapamycin group and chloroquine group. 100 mg/kg propofol was used for continuous anesthesia for 1 week, during which 50 mg/kg rapamycin and 10 mg/kg chloroquine were treated. Morris water maze was used to detect cognitive function, TUNEL staining was used to detect apoptosis, Golgi staining was used to observe autophagy, and Western blot was used to detect autophagy⁃related apoptosis⁃related protein expression.@*Results @#In vitro, rapamycin obviously reversed the apoptosis caused by propofol, but chloroquine had no effect. Compared with propofol group, the expression of microtubule⁃associated protein light chain 3 Ⅱ / microtubule⁃associated protein light chain 3 I (LC3 Ⅱ/LC3 Ⅰ) and Beclin⁃1 in the rapamycin group increased, but the expression of apoptotic proteins Cleaved⁃caspase⁃3 and Bax decreased. In vivo, compared with propofol group, the water maze escape latency of the rapamycin group reduced. In addition, the number of TUNEL⁃positive cells and autophagosomes in the rapamycin group decreased. Furthermore, the expression of mammalian target of rapamycin (mTOR) in the rapamycin group decreased, and the expression of LC3 Ⅱ/LC3 Ⅰ and Beclin⁃1 increased, as well as the expression of Cleaved⁃caspase⁃3 and Bax decreased. But chloroquine had no effect on autophagy and apoptosis⁃related proteins.@*Conclusion @#Rapamycin can further activate autophagy by inhibiting the activation of mTOR signal, ameliorate the neuronal apoptosis caused by propofol, which will lead to the improvement of the cognition in rats.
ABSTRACT
General anesthetics are widely used in pregnant women,gravidas and infants.In the basic studies of rodents,mammals and non-human primate,general anesthetics can cause neurotoxicity,neuroapoptosis and damage neurodevelopment on the developing brain.Therefore,To explore protective measures and mechanism of anesthetic neurotoxicity is of great significance for formulating clinical anesthesia plan,guiding clinical obstetrics and pediatric anesthesia.This article reviewed the progress of ameliorating the neurotoxicity of general anaesthetics on developing brain including anesthetic assistants,hormone drugs,plant extracts,nutritional components and others.
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Objective To investigate the effect of propofol exposure on neuroapoptosis in pri-mary cultured cortical neurons and its mechanisms.Methods Cortical neurons were primarily cultured for seven days,then divided into two groups:control group (treated with equal volume of 20% in-tralipid),propofol-treated group (treated with 500 μmol/L propofol).The neurons were treated for 12 h.The neuron viability was determined by MTT.Neuroapoptosis was identified by Hoechest 33 258 dying.Mitochondrial membrane potential was measured by the fluorescent dye rhodamine 123 (Rh123).Western blot was performed to detect the level of cyt-c and cleaved-caspase-3.Results Neu-rons survival rate (54.4%±6.4%)in the propofol group was significantly lower than that of control group (99.8% ± 4.1%) (P < 0.05 ), the rate of neuronal apoptosis (46.5% ± 5.3%) was significantly higher than that of control group (7.2%±0.9%)(P <0.05),mitochondrial membrane potential (59.6%±4.3%)was significantly lower than that of the control group (99.9% ± 5.7%) (P <0.05 ),cyt-C protein level (0.38 ± 0.03 )was significantly higher than that of control group (0.1 5±0.02)(P < 0.05 ),level of cleaved-caspase-3 protein level (0.46 ± 0.04)was significantly higher than that of control group (0.13±0.02)(P <0.05).Conclusion Propofol induces neuroapo-tosis in primary cultured cortical neurons,which is associated with the decreased level of MPP and the increase levels of cyt-c and cleaved-caspase-3.
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Objective 17β-estradiol is known to have a neuroprotective effect.The aim of this study was to investigate the effects of 17β-estradiol on propofol-induced neuroapoptosis in primarily cultured cortical neurons. Methods Rat cortical neurons were primarily cultured for 7 days and randomly divided into groups A ( vehicle control) , B, and C, treated with equal volume of 20%intralipid, 500 μmol/L propofol, and 500 μmol/L propofol +0.1 μmol/L 17β-estradiol, respectively.At 12 hours after treatment, the morphology of the neurons was observed under the microscope, their survival rate calculated by MTT, their apoptosis was deter-mined by FCM assay, and their mitochondrial membrane potential measured by fluorescent dye rhodamine 123. Results Compared with group A, group B showed a significantly reduced number of neurons, lack of 3-dimensional appearance, unclear contour, and fractured neuron axons, but a remarkable improvement was observed in the propofol-induced morphological damage in group C.The survival rate of the neurons and the mitochondrial membrane potential were markedly decreased in group B ([52.3 ±5.2]% and [59.1 ± 5.3]%) as compared with groups A ( [99.9 ±3.6]%and [99.6 ± 5.8]%) and C ([90.1 ±7.2]%and [89.2 ±7.1]%) (both P<0.01 ) , while the rate of neuroapoptosis significantly increased in group B ([43.4 ±4.6]%) in comparison with A ([3.1 ±0.2]%) and C ([22.3 ±3.2]%) (both P<0.01). Conclusion 17β-es-tradiol can protect against propofol-induced apoptosis of primarily cul-tured neurons by inhibiting the reduction of their mitochondrial membrane potential.
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Objective To investigate the effect of minocycline on isoflurane-induced hippocampal neuroapoptosis and cognitive dysfunction in aged rats. Methods Forty-five male SD rats were randomly assigned into 3 groups ( n=15): blank control group ( group C) , 1.5% isoflurane group ( group I) and 50 mg??kg-1 minocycline+1.5% isoflurane group ( group M+I) . Minocycline was injected intraperitoneally 12 h before the start of anesthesia for group M+I.Group I and group M+I were exposed to 1.5% isoflurane for 4 h, while group C were exposed to 30% O2-70% N2.At the end of anesthesia, five rats in each group were randomized to analyse arterial blood gas. The other rats in each group were sent back to their home cage until they were fully awake.Fourteen days after anesthesia, Morris water maze was used to assess the cognitive function, and then hippocampi of rats were dissected for detection of the expression of cleaved caspase3, Bax and Bcl-2. Results No difference was found in arterial gas analysis among the 3 groups (P>0.05).Compared with group C, the rats in the group I spent more time locating the platform on the third and fourth training days and the time percentage that the rats in group I spent in the target quadrant was much less (P<0.05).However, these changes were reversed in group M+I (P<0.05).The isoflurane-induced increased level of Bax and cleaved caspase3 and decline of anti-apoptotic factor Bcl-2 were restored by minocycline pretreatment ( P<0.05) . Conclusion Minocycline could attenuate cognitive dysfunction induced by isofluranein aged rats.The mechanism is associated with inhibition of hippocampal neuroapoptosis which is increased by isoflurane.