Study On The Neuroprotective Effect Of Nikdil-mediated Deep Hypothermia And Low Flow Mouse Model

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Effect of nicorandil with different concentrations on deep hypothermic low flow after ischemia reperfusion in spatial learning and memory ability Objective To investigate the effects of different concentrations of nicorandil on spatial learning and memory ability after deep hypothermia low flow Methods The C57BL/6 male mice(n=12)were randomly divided into the sham operation group(Sham),the operation group(DHLF),the low dose group(5mg/kg),the middle dose group(10mg/kg)and the high dose group(20mg/kg),except the sham group did not block the bilateral common carotid artery,the other groups were required to establish DHLF mouse model.The anal temperature of mice decreased to 18.5±0.5?,and the DHLF model was established.The Morris water maze test was used to evaluate the spatial learning and memory ability of mice on the 2-6 day after ischemia-reperfusion.After the end of the test,the morphological changes of hippocampal CA1 neurons in mice were assessed by Nissol staining,and the survival pyramidal neuron density was observed.Results The spatial learning and memory abilities of mice after cerebral ischemia-reperfusion injury were severely impaired(P<0.05),and the number of positive cells in the pyramidal neurons in the hippocampal CA1 region was significantly lower than that in the sham operation group(P<0.05).Compared with the DHLF group,there was no significant difference between the two groups in the number of pyramidal cells in the CA1 area and the water maze behavior in the low dose group.Middle dose of nicorandil significantly improved the decrease of spatial learning and memory ability after ischemia reperfusion compared with that in the low dose of nicorandil(P<0.05).There was a statistically significant difference in the number of pyramidal cells in the hippocampal CA1 region and the results of water maze behavior test in the two groups(P<0.05).At the same time,high dose of nicorandil preameliorated spatial learning and memory impairment after ischemia reperfusion compared with that in the DHLF group and low dose of nicorandil group(P<0.05).However,there was no significant difference in the number of pyramidal cells in the hippocampal CA1 region and the water maze test results in the two groups of high dose of nicorandil and the middle dose of nicorandil,while the high dose of nicdi increased the mortality of the mice after the reperfusion(P<0.05).Conclusion Nicorandil can inhibit the decrease of hippocampal pyramidal neurons caused by ischemia-reperfusion injury,thereby improving the ability of spatial learning and memory after ischemia-reperfusion.Part ? The mechanism of nicorandil improved spatial learning and memory ability after deep hypothermia and low flow Objective To investigate the protective effect of nicorandil improves the ability of spatial learning and memory after deep hypothermia and low flow Methods The C57BL/6 male mice(n=30)were randomly divided into the sham operation group(Sham),the operation group(DHLF),the nicorandil group(Nicorandil)and the 5-HD group,except the sham group did not block the bilateral common carotid artery,the other groups were required to establish the DHLF mouse model.The anal temperature of mice decreased to 18.5±0.5?,and the DHLF model was established.Each group was further divided into 6 subgroups: 2h,6h,12 h,24h,48 h and 72 h,and 6 mice in each subgroup.Nissol staining was used to detect pathological changes in hippocampal tissue;TUNEL staining was used to detect neuronal apoptosis in hippocampus CA1 region;ELISA was used to detect reactive oxygen species(ROS)in hippocampal tissues,and mitochondrial membrane potential(MMP)in hippocampal neurons was detected by flow cytometry.WB was used to detect the expression of Bax,Bcl-2,Cytochrome C,and cleaved caspase-3,9 of protein expression in hippocampus.The expression of p-ERK and t-ERK protein in hippocampus tissue was detected by immunofluorescence staining and Western blot.Results Histopathological found that the overall morphology of cells and tissues in the nicorandil group was improved and the apoptotic index decreased compared with that in the DHLF group(P<0.05),while the pathological damage of the 5-HD group was increased and the apoptotic index increased correspondingly(P<0.05).Nicorandil group significantly decreased the level of ROS,improve the mitochondrial membrane potential compared with that of the DHLF group.WB showed that the expression of anti-apoptotic Bcl-2 protein was up-regulated and the expression of pro-apoptotic Bax protein and Caspase protein was inhibited in the nicorandil group compared with that of the DHLF group(P<0.05),and the release of Cytochrome C from the mitochondria was inhibited in the nicorandil group compared with that of the DHLF group(P<0.05).Furthermore,immunofluorescence staining combined with confocal laser scanning microscopy showed that p-ERK1/2 was inhibited by nicorandil.WB showed that the expression level of p-ERK1/2 protein in the group of nicorandil was significantly lower than that of the DHLF group(P <0.05).The level of 2h,6h and 12 h of p-ERK protein was rising,12 h reached the peak,and 24 h,48h and 72 h decreased,but it was still higher than that of sham group,and the difference was statistically significant(P<0.05).Conclusion Nicorandil can improve the spatial learning and memory ability after DHLF by inhibiting mitochondrial apoptosis pathway,and its mechanism may be by inhibiting the ERK signaling pathway,thus playing the role of brain protection.