| Postoperative cognitive dysfunction (POCD) is a central nervous system complications which seriously affecting the life quality of patients and a heavy burden on the family and society caused by surgery, anesthesia and other factors. In the ageing brain, most region-specific changes in dendritic morphology, cellular connectivity, Ca2+ dysregulation, gene expression or other factors affect plasticity and ultimately alter the network dynamics of neural ensembles that support cognition. These age-related alterations may make the elderly more susceptible to the anesthetics and more vulnerable to developing functional impairment (e. g. cognitive decline).25.8% 1 week after surgery and 9.9% 3 months after surgery of elderly patients suffer from POCD. The mechanism of cognitive dysfunction is not well understood but only age was a risk factor for late postoperative cognitive dysfunction. Since sevoflurane has low blood-gas solubility coefficients and rapid recovery, it is used widely in clinical practice. However, studies showed that it induces apoptosis and increases beta-amyloid protein levels and increases tau phosphorylation through specific kinases activation and spatial memory deficits. Findings show that mitogen-activated protein kinases/extracellular regulated protein kinases(MAPK/ERK), CREB and the transcription factor Egrl are essential components of a signaling cascade required for the expression of long-term recognition memory. Therefore, this pathway may be involved in cognitive decline process caused by sevoflurane and play an important role.Section 1 Retrieval of Consolidated Spatial Memory in the Water Maze Is Correlated with Expression of pCREB and Egrl in the Hippocampus of Aged MiceObjectiveThe objective of this research is to study the relationship of the expression of phosphorylated cyclic AMP response element-binding protein (pCREB) and early growth response protein 1 (Egr1) in the hippocampus of aged mice with retrieval of consolidated spatial memory in a water maze.MethodsTwenty-four aged mice were allocated into four groups:â‘ no training or probe test (naive);â‘¡ no training but exposed to the same probe test (NTPRT); â‘¢received training and probe test (PRT);â‘£received training but no probe test (NPRT). Three days prior to the acquisition phase, each mouse received a pretraining session to adapt to the water temperature and the surrounding environment. Place navigation test lasted for 14 days. The time limit was set to 60 seconds. A trial terminated when the animal reached the platform, where it remained for 60 sec. Mice that failed to find the platform within this time limit were placed onto the platform by the experimenter and had to stay there for 60 sec to observe hints. On the fourth, seventh, eleventh and fourteenth day, the animals were generally given probe trials. Animal movements were recorded using the videotrack system. This processing allowed us to calculate the escape latency (time required to find the platform, in seconds) and the percentage of time spent in the target quadrant. In percentages, more than 30% meant the acquisition of the spatial reference memory. This processing allowed us to calculate the escape latency (time required to find the platform, in seconds) and the percentage of the time (time spent in the target quadrant divided by the total time,%). The percentages more than 30% meant the acquisition of the spatial reference memory. On day 14, all 12 aged mice were familiar with the maze and they were randomized into the no probe test (NPRT) group and the probe test (PRT) group. On day 15, the PRT group and NTPRT group were subjected to a probe test while the NPRT group and naive group were not. On day 15, mice were deeply anesthetized at 15 min after the probe trial and the brains were removed. pCREB immunoreactivity (pCREB-ir) and Egrl immunoreactivity (Egrl-ir) in the hippocampal CA1 and CA3 areas were examined.Results1. Behavioral ResultsThe training protocol led to a good consolidated spatial memory of the platform location. The performance of the mice from day 1 to day 14 shows that they progressively learned the task, as indicated by decreasing latencies from day 1 to day 13 and increasing percentages from day 4 to day 14. This was confirmed by analyses of variance performed on data from the whole training period (from day 1 to day 14), yielding the main effect time (F 9,126=22.08, p<0.0001), and the main effect was on day 12 (F=152.41, p<0.0001). When analyzing performance from day 4 to day 14, statistical analyses showed significant effect times (F 3,39=11.32, p<0.0001), preceding the asymptotic level of performance reached at day 11 (data not shown). Memory of the platform location was assessed with the probe trial on days 4,7,11, and 14. Percentages of the time spent in the target quadrant relative to the opposite quadrant (t=229.448, p<0.0001) revealed a great retention of the platform location. There was no difference in the performance of day 14 between the NPRT and PRT groups (t =0.075, p=0.789). There was no thigmotaxis and floating behavior during training, and statistical analyses of the mean swim speed during training did not reveal any significant group effect (F<1,p>0.3).2. Immunohistochemical ResultsAll mice were sacrificed at 15 min after the final probe trial on day 15. The results showed: â‘ NPRT group vs. PRT group:The levels of pCREB immunoreactivity (pCREB-ir) in the CA1 area of the NPRT group were significantly lower than those of the PRT group (p value<0.05). The levels of pCREB immunoreactivity (pCREB-ir) in the CA3 area of the NPRT group were significantly lower than those of the PRT group (p value<0.05). The levels of Egrl immunore-activity (Egr1-ir) in the CA1 area of the NPRT group were significantly lower than those of the PRT group (p value<0.05). The levels of Egrl immunore-activity (Egr1-ir) in the CA3 area of the NPRT group were significantly lower than those of the PRT group (p value<0.05).â‘¡NPRT group and PRT group vs. NTPRT group and Naive group:The pCREB-ir levels in the CA1 area of the NPRT and PRT groups were significantly higher than those of the Naive and NTPRT controls (p value<0.05). The pCREB-ir levels in the CA3 area of the NPRT and PRT groups were significantly higher than those of the Naive and NTPRT controls (p value<0.05). The Egrl-ir levels in the CA1 area of the NPRT and PRT groups were significantly higher than those of the Naive and NTPRT controls (p value<0.05). The Egrl-ir levels in the CA3 areas of the NPRT and PRT groups were significantly higher than those of the Naive and NTPRT controls (p value<0.05).â‘¢ NTPRT group vs. Naive group:The pCREB-ir levels in the CA1 area were no difference between the NTPRT group and Naive group (p value>0.05). The pCREB-ir levels in the CA3 area were no difference between the NTPRT group and Naive group (p value>0.05). The Egr1-ir levels in the CA1 area were no difference between the NTPRT group and Naive group (p value>0.05). The Egrl-ir levels in the CA3 area were no difference between the NTPRT group and Naive group (p value>0.05).â‘£ CA1 area vs. CA3 area:In all groups, the level of pCREB-ir was significantly higher in the CA3 area compared to the CA1 area (all p values<0.05), while the level of Egr1-ir was significantly higher in the CA1 area compared to the CA3 area (all p values<0.05).Conclusion1. Retrieval of consolidated spatial memory in the water maze is corre-lated with expression of pCREB and Egrl in the hippocampus of aged mice.2. The level of pCREB-ir was significantly higher in the CA3 area compared to the CA1 area, while the level of Egr1-ir was significantly higher in the CA1 area compared to the CA3.Section 2 Impaired Spatial Learning and Memory after Sevoflurane Anesthesia in Aged Mice Is Associated with Down-regulated pCREB and Egrl in the HippocampusObjectiveAnesthetics and postoperative cognitive deficits are recently debated, while the precise mechanism is not well understood. Early growth response protein 1(Egr1) and the phosphorylation of cyclic AMP response element-binding protein (pCREB) in the hippocampus are critical for the generation of late LTP or for the formation of long-term memory. To better understand the neural effects of inhalational anesthetics, we studied the behavioral and pCREB and Egrl changes in hippocampus of aged mice that were exposed to sevoflurane for 4 h.MethodsEighteen-month-old mice were randomly assigned to receive 3% sevoflurane or 100% oxygen for 4 h. Spatial learning and memory were tested with the Morris water maze 48 h after exposure. Place navigation test lasted for 6 days. On day 6, mice were subjected to a probe test after navigation test and were deeply anesthetized at 15 min after the probe trial. The brains were removed and pCREB and Egrl in the hippocampus were examined by western blot and RT-PCR.Results1. Behavioral resultsThe performance of the mice from day 1 to day 6 showed that escape latency of Sevo group was significantly shorter than Control group on day 4, day 5 and day 6(all p value<0.05). Percentages of the time spent in the target quadrant showed that Sevo group was significantly lower than Control group(p value<0.01).2. Western blot resultsâ‘ The pCREB protein levels in the hippocampus of Sevo group were significantly lower than that in Control group (p value< 0.01).â‘¡ The Egrl protein levels in the hippocampus of Sevo group were significantly lower than that in the Control group (p value< 0.05).3. RT-PCR resultsâ‘ The CREB mRNA levels in the hippocampus were no difference between Sevo group and Control group (p value>0.05).â‘¡ The Egrl mRNA levels in the hippocampus of Sevo group were significantly lower than that in the Control group (p value< 0.05).Conclusion1. Aged mice occurred spatial learning and memory deficits after sevoflurane anesthesia..2. Impaired spatial learning and memory after sevoflurane anesthesia in aged mice is associated with down-regulated pCREB and Egrl in the hippocampus. |
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