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Objective:To explore the effects of acute sleep fragmentation (SF) on cognitive function and the relationship between hippocampal Homer1a and synaptic plasticity in aged rats.Methods:One hundred and eight SPF grade male SD rats aged 22 to 24 months were divided into three groups according to random number table: control group (Control group), non-sleep fragmentation group (NSF group) and sleep fragmentation group (SF group), with 36 rats in each group.A sleep fragmentation model was established by sleep deprivation rod method.Morris water maze and novel object recognition tests were used to evaluate the learning and memory function of rats.Homer1a expression in hippocampus was detected by Western blot, and its distribution in CA1 area of hippocampus was observed by immunohistochemical staining.Golgi staining was used to observe the density of dendritic spines in CA1 area of hippocampus, and in vitro electrophysiological patch clamp test was used to detect the slope of field excitatory postsynaptic potential(fEPSP) from CA3 to CA1 in hippocampus.SPSS 22.0 and GraphPad Prism 9.3 softwares were used for data statistical analysis and mapping.One-way ANOVA was used for comparison among groups, and Tukey-Kramer test was used for further pairwise comparison. Results:(1)In the behavioral tests, there were statistical differences in the times of crossing the original platform, the target quadrant residence time and the new object recognition index at 1 h and 24 h among the three groups( F=13.63, 11.34, 21.26, 16.22, all P<0.01). The times of crossing the original platform in SF group((2.00±1.27) times) was lower than that of Control group ((5.67±2.16) times) and NSF group ((6.50±2.35) times) (both P<0.05). The target quadrant residence time in SF group ((9.02±4.84) s) was shorter than that in Control group ((24.73±7.37) s) and NSF group ((27.81±8.37)s) (both P<0.05). The new object recognition index at 1 h and 24 h in SF group were lower than those in Control group and NSF group (all P<0.05). (2) In Western blot assay, the expression of Homer1a protein in hippocampus of SF group(0.91±0.13) was higher than that of Control group(0.70±0.05) and NSF group(0.74±0.04)(both P<0.05). (3) In immunohistochemical staining, the optical density value of the Homer1a protein in CA1 area of hippocampus in the SF group was higher than that in the Control group and NSF group(both P<0.05). (4) In Golgi staining, the density of dendritic spines in CA1 area of hippocampus in SF group was lower than that in Control group and NSF group (both P<0.05). (5) In vitro electrophysiological test showed that the slope of fEPSP in CA3-CA1 area of hippocampus in SF group were lower than that in Control group and NSF group (both P<0.05). Conclusion:Acute SF intervention in aged rats can cause cognitive impairment, which may be associated with the inhibition of hippocampal synaptic plasticity induced by hippocampal Homer1a overexpression.
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Objective:To evaluate the role of Homer1a/metabotropic glutamate receptor 5 (mGluR5) signaling pathway in sleep deprivation-induced cognitive dysfunction in aged rats.Methods:One hundred and four SPF healthy male Sprague-Dawley rats, aged 22-24 months, weighing 320-360 g, were divided into 4 groups ( n=26 each) using a random number table method: normal control group (group Control), sleep deprivation+ vehicle group (group SD+ Vehicle), sleep deprivation+ mGluR5 forward allosteric agent CDPPB group (group SD+ CDPPB), and sleep deprivation+ mGluR5 antagonist MPEP group (group SD+ MPEP). A 48-h sleep deprivation model was developed by sleep-deprived rod method. At the beginning of developing the model and 24 h after developing the model, CDPPB 10 mg/kg, MPEP 10 mg/kg and the equal volume of 1% Tween 80 were intraperitoneally injected in group SD+ CDPPB, group SD+ MPEP and group SD+ Vehicle, respectively.Morris water maze and novel object recognition tests were conducted to evaluate cognitive function after development of the model. The expression of Homer1a and mGluR5 in the hippocampus was detected by Western blot, the dendritic spine density in the hippocampal CA1 region was detected by Golgi staining, and the field excitatory postsynaptic potential (fEPSP) slope in the hippocampal CA1 region was detected by isolated electrophysiology. Results:Compared with Control group, the number of crossing the original platform, time of staying at the target quadrant, and novel object recognition index at 1 and 24 h after training were significantly decreased, the expression of Homer1a in the hippocampus was up-regulated, the expression of mGluR5 in the hippocampus was down-regulated, and the density of dendritic spine and fEPSP slope in the hippocampal CA1 region were decreased in group SD+ Vehicle ( P<0.05). Compared with group SD+ Vehicle, the number of crossing the original platform, time of staying at target quadrant, and novel object recognition index at 1 and 24 h after training were significantly increased, the expression of mGluR5 in hippocampus was up-regulated, and the density of dendritic spines and fEPSP slope in the hippocampal CA1 region were increased in group SD+ MPEP( P<0.05), and no statistically significant change was found in the parameters mentioned above in group SD+ CDPPB ( P>0.05). Conclusions:Sleep deprivation impairs the synaptic plasticity of hippocampal neurons by regulating Homer1a/mGluR5 signaling pathway, and thus mediating the process of cognitive dysfunction in aged rats.
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Objective:To identify the risk factors for postoperative cognitive dysfunction (POCD) and develop the prediction model in elderly patients undergoing lumbar surgery under general anesthesia.Methods:The elderly patients undergoing elective lumbar surgery under general anesthesia in our hospital from July 2021 to July 2022 were enrolled. Cognitive function was assessed at 7 days after surgery using Mini-Mental State Examination and Montreal Cognitive Assessment. When the decrease in both scales≥ 1 standard deviation, the patients were considered as having POCD. The patients were divided into POCD group and non-POCD group according to whether POCD developed. The propensity score matching was used to balance the confounding bias between two groups. The multivariate logistic regression analysis was used to identify the risk factors for POCD. The prediction model was constructed, and a nomogram was drawn for visualization of the model. The receiver operating characteristic curve, calibration plot and decision curve analysis (DCA) were drawn to evaluate the differentiation, consistency and clinical validity of the model, respectively.Results:A total of 159 patients were enrolled in this study, and the incidence of POCD was 31.4%. There were statistically significant differences in the ratio of intraoperative blood transfusion, cumulative time of hypotension, total infusion volume and operation time between two groups ( n=32 each) after propensity score matching ( P<0.05). The results of multivariate logistic regression showed that age, educational levels, diabetes mellitus, previous two or more operations under general anesthesia, APTT and cumulative time of hypotension were independent risk factors for POCD in elderly patients undergoing lumbar surgery under general anesthesia ( P<0.05). A model was developed based on the risk factors mentioned above: LogitP=-15.878+ 0.263 × Age (years) - 0.122 × Educational Level (years)+ 1.601 × Diabetes Mellitus+ 1.468 × History of General Anesthesia for 2 or more times+ 0.608 × Cumulative Time of Hypotension(min) - 0.140 × APTT (s). The area under the receiver operating characteristic curve was 0.930 (95% CI 0.887-0.973), the sensitivity was 0.920, specificity was 0.798 and Youden index was 0.718. After visualizing the model via nomogram, the model was verified by Hosmer-Lemeshow test, P=0.403, C index was 0.930, and corrected C index was 0.914. Conclusions:Age, educational levels, diabetes mellitus, previous multiple operations under general anesthesia, APTT and cumulative time of hypotension are independent risk factors for POCD in elderly patients undergoing lumbar surgery under general anesthesia, and the established risk prediction model can effectively predict the occurrence of POCD in elderly patients undergoing lumbar surgery under general anesthesia.
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Objective:To evaluate the relationship between Karyopherin β2 (Kapβ2)-mediated nuclear translocation of nuclear inhomogeneous ribonucleoprotein A2/B1 (hnRNPA2/B1) and sevoflurane-induced brain neurotoxicity in a cellular experiment.Methods:The mouse hippocampal neuronal cell line HT22 cells were inoculated in confocal culture dishes and 6-well culture plates at a density of 2×10 5 cells/well and 1×10 6 cells/well and divided into 4 groups( n=12 each) by a random number table method: control group (GFP-C group) carrying green fluorescent protein (GFP) with empty adenovirus transfection, sevoflurane group (GFP-Sev group) carrying GFP with empty adenovirus transfection, control group (GFP-Sev group) transfected with Kapβ2 gene-overexpressing adenovirus, and sevoflurane group (Kapβ2-Sev group) transfected with Kapβ2 gene-overexpressing adenovirus. After 48 h of conventional incubation, empty adenovirus-carrying GFP (GFP-C and GFP-Sev groups) and Kapβ2 gene-overexpressing adenovirus (Kapβ2-C and Kapβ2-Sev groups) were transfected. After 48 h of transfection, the cells were conventionally incubated continuously in GFP-C and Kapβ2-C groups, and the cells were incubated for 3 h with 3% sevoflurane and then were conventionally incubated for 48 h in GFP-Sev and Kapβ2-Sev groups. The expression of Kapβ2, synaptophysin (SYP), postsynaptic density protein 95 (PSD95) and hnRNPA2/B1 nucleoplasmic ratio were measured in cells by Western blot. Immunofluorescence assay was used for hnRNPA2/B1 subcellular localization. Results:Compared with GFP-C group, the expression of SYP and PSD95 was significantly down-regulated, hnRNPA2/B1 nucleoplasmic ratio was decreased, and cytoplasmic hnRNPA2/B1 expression was up-regulated in GFP-Sev group, and Kapβ2 expression was significantly up-regulated in Kapβ2-C group ( P<0.05). Compared with Kapβ2-C group, the expression of SYP and PSD95 was significantly down-regulated, hnRNPA2/B1 nucleoplasmic ratio was decreased, and cytoplasmic hnRNPA2/B1 expression was up-regulated in Kapβ2-Sev group ( P<0.05). Compared with GFP-Sev group, the expression of Kapβ2, SYP and PSD95 was significantly up-regulated, hnRNPA2/B1 nucleoplasmic ratio was increased, and cytoplasmic hnRNPA2/B1 expression was down-regulated in Kapβ2-Sev group ( P<0.05). Conclusions:Kapβ2-mediated hnRNPA2/B1 nuclear translocation may be the endogenous protective mechanism against sevoflurane-induced brain neurotoxicity.
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Objective:To investigate the relationship between autophagy and nuclear factor E2 related factor 2(Nrf2) signaling pathway during high glucose-induced damage to Schwann cells.Methods:RSC96 were cells cultured in vitro and seeded in 96-well plates (1×10 4 cells/ml, 200 μl/well) or in 6-well plates (1×10 6 cells/ml, 2 ml/well) for 48 h. The cells were divided into 3 groups ( n=25 each) using a random number table method: control group (group C), high glucose group (group H) and high glucose+ autophagy agonist rapamycin group ( group H+ RAP). The cells were cultured in the common culture medium in group C. In group H, 50 mmol/L of glucose was added to the culture medium.In group H+ RAP, 50 mmol/L of glucose and 5 μmol/L rapamycin were added to the culture medium.At 48 h of incubation, the growth of cells was observed with inverted phase contrast microscope, the cell viability was measured using MTT method, apoptosis rate was determined by flow cytometry, malondialdehyde (MDA) content was determined by thiobarbituric acid method, superoxide dismutase (SOD) activity was detected using xanthine oxidase method, and the expression of Nrf2, P62 and microtubule-associated protein 1 light chain 3 Ⅱ (LC3 Ⅱ) was determined by Western blot. Results:Compared with group C, the cell viability and SOD activity were significantly decreased, apoptotic rate and MDA content were increased, and expression of Nrf2, P62 and LC3Ⅱ was up-regulated in group H and group H+ RAP ( P<0.05). Compared with group H, the cell viability and SOD activity were significantly increased, apoptosis rate and MDA content were decreased, the expression of Nrf2 and LCII was up-regulated and P62 expression was down-regulated in group H+ RAP ( P<0.05). Conclusion:Enhanced autophagy can activate Nrf2 signaling pathway, which is the endogenous protective mechanism of Schwann cell injury induced by high glucose.
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Objective:To evaluate the effects of sevoflurane combined with propofol anesthesia on the synaptic plasticity in the hippocampus after operation in rats with mild cognitive impairment (MCI) and the relationship with potassium-chloride cotransporter-2 (KCC2)/sodium-potassium-chloride cotransporter 1 (NKCC1).Methods:Clean-grade healthy male Sprague-Dawley rats, aged 16-18 months, weighing 440-540 g, in which MCI was induced by severe bilateral common carotid artery stenosis (BCAS). Forty-eight rats with MCI were divided into 4 groups ( n=12 each) using a random number table method: sham operation group (group Sham), sevoflurane anesthesia group (group S), propofol anesthesia group (group P), and sevoflurane and propofol anesthesia group (group SP). After disappearance of eyelash reflex, open reduction and internal fixation was performed after tibial fracture was induced in S, P and SP groups.Anesthesia method was as follows: 1.7% sevoflurane was inhaled and propofol 20 mg·kg -1·h -1 was intravenously infused for 3 h in group SP, 3% sevoflurane was inhaled for 3 h in group S, and propofol was intravenously infused at rate of 40 mg·kg -1·h -1 for 3 h in group P. The novel object recognition (NOR) test was performed at 14 days after operation, and the discrimination index in NOR test was calculated.The in vivo electrophysiological experiment was performed on 19 days after operation to measure long-term potentiation and amplitude of the field excitatory postsynaptic potential (fEPSP). The expression of KCC2 and NKCC1 was determined by Western blot, and the ratio of KCC2/NKCC1 was calculated.The density of dendritic spines in the hippocampal CA1 region was determined by Golgi-COX staining performed at 30 days after operation. Results:Compared with Sham group, the discrimination index in NOR test, hippocampal KCC2/NKCC1 ratio, density of dendritic spines in hippocampal CA1 region, and amplitude of fEPSP were significantly decreased in S and P groups ( P<0.05), and no significant change was found in the parameters mentioned above in group SP ( P>0.05). Compared with group S or group P, the discrimination index in NOR test, hippocampal KCC2/NKCC1 ratio, density of dendritic spines in hippocampal CA1 region, and amplitude of fEPSP were significantly increased in group SP ( P<0.05). Conclusion:Sevoflurane combined with propofol anesthesia does not aggravate postoperative cognitive dysfunction in the rats with MCI, which may be related to maintaining the balance of hippocampal KCC2/NKCC1 and protecting the synaptic plasticity in hippocampi.
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Objective:To evaluate the relationship between dexmedetomidine-induced reduction of sevoflurane-induced neurotoxicity and cation-chloride cotransporters Na + -K + -2Cl --1 (NKCC1) /K + -2Cl --2 (KCC2) in neonatal rats. Methods:Eighty healthy male Sprague-Dawley rats at postnatal day 7 were divided into 4 groups ( n=20 each) using a random number table method: control group (group C), sevoflurane anesthesia group (group S), dexmedetomidine group (group D), and sevoflurane and dexmedetomidine group (group SD). Rats were exposed to 2.5% sevoflurane for 6 h to establish the model of sevoflurane anesthesia in group S. Dexmedetomidine 1.0 μg/kg was intraperitoneally injected, and then sevoflurane anesthesia was performed in group SD.The expression of cleaved caspase-3, NKCC1 and KCC2 was detected by Western blot at 24 h after the end of anesthesia.At 35 days after the end of anesthesia, the cognitive function was assessed using the Y maze test, and the neurons in the hippocampal CA1 area were counted using the Nissan staining method. Results:Compared with group C, the percentage of time spent in novel arm and the number of neurons in hippocampal CA1 area were significantly decreased, the expression of cleaved caspase-3 and NKCC1 was up-regulated, the expression of KCC2 was down-regulated, and the ratio of NKCC1/KCC2 was increased in S and SD groups ( P<0.05), and no change was found in the above indicators in group D ( P>0.05). Compared with group S, the percentage of time spent in novel arm and the number of neurons in hippocampal CA1 area were significantly increased, the expression of cleaved caspase-3 and NKCC1 was down-regulated, the expression of KCC2 was up-regulated, and the ratio of NKCC1/KCC2 was decreased in group SD ( P<0.05). Conclusion:The mechanism of dexmedetomidine attenuating sevoflurane-induced neurotoxicity may be related to maintaining the relatively stable expression of NKCC1/KCC2 in neonatal rats.
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Objective:To evaluate the effect of propofol combined with sevoflurane anesthesia on GABA A receptor (GABA AR)α 1/α 2 subunit homeostasis in hippocampus of rats with mild cognitive impairment (MCI). Methods:Healthy clean-grade male Sprague-Dawley rats, aged 16-18 months, weighing 450-550 g, were anesthetized.MCI was induced by ligation of bilateral common carotid arteries after anesthesia.Morris water maze test was used to select the rats with MCI at 30 days after establishment of the model.The rats with MCI were divided into 4 groups ( n=18 each) using a random number table method: sham operation group (group Sham), sevoflurane anesthesia group (group S), propofol anesthesia group (group P), and propofol combined with sevoflurane anesthesia group (group SP). The rats inhaled 3% sevoflurane for 3 h in group S. Propofol 40 mg·kg -1·h -1 was intravenously infused for 3 h in group P. The rats inhaled 1.7% sevoflurane, and propofol 20 mg·kg -1·h -1 was intravenously infused for 3 h simultaneously in group SP.Open reduction and internal fixation was performed after tibial fracture was induced in S, P and SP groups.Y-maze test was performed at 14 days after operation to assess cognitive function.The expression of potassium-chloride cotransporter-2 (KCC2), sodium-potassium-chloride cotransporter 1 (NKCC1), GABA ARα 1 and GABA ARα 2 was determined using Western blot. Results:Compared with group Sham, the percentage of time of staying at novel arm was significantly decreased, the expression of hippocampal KCC2 and GABA ARα 1 was down-regulated, and the expression of NKCC1 and GABA ARα 2 was up-regulated in S and P groups ( P<0.05), and no significant change was found in the parameters mentioned above in group SP ( P>0.05). Compared with group S or group P, the percentage of time of staying at novel arm was significantly increased, the expression of hippocampal KCC2 and GABA ARα 1was up-regulated, and the expression of NKCC1 and GABA ARα 2 was down-regulated in group SP ( P<0.05). Conclusion:The mechanism by which propofol combined with sevoflurane anesthesia does not aggravate the postoperative cognitive dysfunction may be related to maintaining GABA ARα 1/α 2 subunit homeostasis in rats with MCI.
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Objective To evaluate the effect of sevoflurane combined with propofol anesthesia on the postoperative expression of nuclear heterogeneous ribonucleoprotein A2 (hnRNPA2) in brain tissues of rats with mild cognitive impairment (MCI).Methods Male Sprague-Dawley rats,aged 16-18 months,were anesthetized with pentobarbital sodium.MCI was induced by ligation of bilateral common carotid arteries after anesthesia.Forty-eight rats with MCI were divided into 4 groups (n =12 each) using a random number table method:sham operation group (SH group),sevoflurane anesthesia group (S group),propofol anesthesia group (P group),and sevoflurane combined with propofol anesthesia group (SP group).Group S inhaled 3％ sevoflurane.Propofol was intravenously infused at a rate of 40 mg · kg-1 · h-1 in group P.In group SP,1.7％ sevoflurane was inhaled,and propofol 20 mg· kg-1 · h-1 was intravenously infused.The anesthesia time was 3 h in the three groups.After disappearance of eyelash reflex,open reduction and internal fixation was performed after tibial fracture was induced.Y-maze test was performed at 7 days after operation,and the percentage of time of staying at novel arm was calculated.The open field test was performed,and the total activity distance and time of staying at the central region were recorded.Then the rats were sacrificed,and brain tissues were obtained for determination of the expression of hnRNAP2 and γ-aminobutyric acid receptor A1 subunit (GABAA-α1) in hippocampus by immunofluorescence and Western blot,respectively.Results Compared with SH group,the percentage of time of staying at novel arm was significantly decreased,the expression of hnRNPA2 in the hippocampus was up-regulated,and the expression of GABAA-α1 was down-regulated in S and P groups,and the expression of hnRNPA2 in the hippocampus was up-regulated (P＜0.05),and no significant change was found in the percentage of time of staying at novel ann or expression of GABAA-α1 in SP group (P＞ 0.05).Compared with S group or P group,the percentage of time of staying at novel arm was significantly increased,the expression of hnRN-PA2 in the hippocampus was down-regulated,and the expression of GABAA-α1 was up-regulated in SP group (P＜0.05).There was no significant difference in the total distance or time of staying at the central region among the four groups (P＞ 0.05).Conclusion The mechanism by which sevoflurane combined with propofol anesthesia does not aggravate the postoperative cognitive dysfunction may be related to up-regulating the expression of hnRNPA2 in brain tissues and maintaining GABAA-α1 stable in rats with MCI.
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Objective To evaluate the effect of dexmedetomidine on the expression of NR1 subunit-containing NMDA receptors and GIuR2 subunit-containing AMPA receptors during hypoxic injury to rat hippocampal neurons.Methods The hippocampal neurons were isolated from Wistar rats within 24 h after birth and divided into 3 groups (n=24 each) using a random number table:control group (group C),hypoxia group (group H) and dexmedetomidine group (group D).The cells were subjected to hypoxia for 6 h to establish the model of neuronal hypoxic injury in H and D groups.In group D,0.1 μmol/L dexmedetomidine was added at 6 h of hypoxia and neurons were incubated for 3 h,and then the culture medium was replaced with a normal medium and neurons were incubated for 24 h.The neuronal viability was measured by CCK-8 assay,the leakage of LDH was detected,and the leakage rate was calculated.The expression of NR1 subunits-containing NMDA receptors and GluR2 subunits-containing AMPA receptors was detected by Western blot.The concentration of calcium ion in cytoplasm was measured using Fluo-3AM.Results Compared with group C,the neuronal viability was significantly decreased,the LDH leakage rate was increased,the expression of NR1 subunits-containing NMDA receptors in the membrane was up-regulated,the expression of GluR2 subunits-containing AMPA receptors was down-regulated,and the concentration of calcium ion in cytoplasm was increased in H and D groups (P＜0.05).Compared with group H,the neuronal viability was significantly increased,the LDH leakage rate was decreased,the expression of NR1 subunits-containing NMDA receptors in the membrane was down-regulated,the expression of GluR2 subunitscontaining AMPA receptors was up-regulated,and the concentration of calcium ion in cytoplasm was decreased in group D (P＜0.05).Conclusion The mechanism by which dexmedetomidine reduces hypoxic injury to rat hippocampal neurons may be related to inhibiting up-regulation of the expression of NR1 subunits-containing NMDA receptors in the membrane and down-regulation of the expression of GluR2 subunitscontaining AMPA receptors.
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Objective To evaluate the effect of propofol on the invasion of cerebral glioma in rats. Methods A total of 120 healthy adult male Wistar rats, weighing 250-280 g, were divided into 4 groups (n=30 each)using a random number table: sham operation group(group S), glioma group(group G) and different doses of propofol groups(group P1and group P2). The rats only underwent sphenotresia in group S. The model of brain glioma was established by injecting C6 glioma cells into the right caudate nucle-us in G, P1and P2groups. In P1and P2groups, propofol was infused at the rates of 20 mg·kg-1·h-1 and 40 mg·kg1·h-1, respectively, for 6 h through the tail vein at day 10 after establishing the model. The rats were sacrificed at day 18 after establishing the model, global brains were removed, and glioma was isolated. The weight of glioma was measured. The pathological changes were examined using hematoxylin and eosin staining. Immunohistochemical staining was used to determine the positive expression of glial fi-brillary acidic protein in glioma cells. The expression of vascular endothelial growth factor(VEGF)in both the periphery(within 2 mm diameter)and central region of glioma was detected by Western blot. Results No glioma was found in group S. The marked cavity and necrotic region in the central region of glioma and dense distribution of glioma cells and neovessels in the periphery of glioma were observed, and glial fibril-lary acidic protein was positively expressed in the majority of glioma cells in G, P1and P2groups. Com-pared with group S, the expression of VEGF was significantly up-regulated, and the number of positive cells was increased in G, P1and P2groups(P<0.05). Compared with group G, the weight of glioma was significantly decreased, the expression of VEGF was down-regulated, and the number of positive cells was decreased in P1and P2groups(P<0.05). There were no significant differences in the parameters men-tioned above between group P1and group P2(P>0.05). Conclusion Propofol can inhibit the invasion of cerebral glioma and provides anti-tumor effect in rats.