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Role of Blood Glucose Concentration Fluctuation Control in Neuronal Damage of Rat Hippocampus after Transient Global Brain Ischemia

Author: LiuLiPing
Tutor: MiWeiDong
School: PLA Postgraduate Medical School
Course: Anesthesiology
Keywords: 2-VO EICT animal model Sprague-Dawley rats ischemia/reperfusion glucostasis shuttle box apoptosis Ischemia/reperfusion Glucostasis PET GLUT1 MAPK insulin
CLC: R743.3
Type: PhD thesis
Year: 2008
Downloads: 209
Quote: 0
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Abstract


Part 1 Establishment of the rat global brain ischemia model with blood glucose concentration fluctuation controlObjective: To establish a simple, reliable and minor trauma model of global cerebral ischemia with blood glucose concentration fluctuation control in Sprague-Dawley rats. Methods: The transient global cerebral ischemia and reperfusion models in Sprague-Dawley rats were made by bilateral common carotid artery occlusion combined with hemorrhagic hypotension (2-VO model). Blood glucose concentration fluctuation control was made by modified method of Euglycemic hyperinsulinemic clamp technique (EICT). Then the reliability of this model was evaluated based on brain auditory evoked potential (BAEP) and 2,3,5-triphenyltetrazolium chloride staining. Results: Blood glucose concentration can be controled subtly during rat global cerebral ischemia/reperfusion. BAEP dispear when rat undergo transient global brain ischemia 5 minute and come back after 5 minute reperfusion, and 2,3,5-triphenyltetrazolium chloride staining clearly indicated the broad brain damage but not infarct. Conclusion: The model is simple, reliable, minor trauma and stable. It is dependable to use this experimental model to study global cerebral ischemia and blood glucose control. Part 2 Role of blood glucose concentration fluctuation control in neuronal damage of rat hippocampus after transient global brain ischemiaObjective: To observe the role of glucostasis in Neuronal damage of rat hippocampus after transient global ischemia/reperfusion. Methods: Sixty-four normal SD rats randomized 4 groups, pseudooperation group(CON), global brain ischemia/reperfusion group(I/P), blood glucose fluctuation group(BGF) and blood glucose stabliliation group(BGS). The global cerebral ischemia rat models were made by bilateral common carotid artery occlusion combined with hemorrhagic hypotension. Perioperative blood glucose concentration fluctuation were controlled by EICT. After 15 minutes ischemia, rats were reperfused and eight of each group sacrificed at 150 minutes after that to obtain brain. Each brain was cut in the middle, hippocampus of a half brain was separated and frozen in liquid nitrogen for western blot assay, another half brain was fixed in 4% paraformaldehyde for histopathology and immunohistochemistry test. The rest eight rats of each group were performed shuttle box at 1 to 5 days after operation and sacrificed to obtain brain for pathology test. Shuttle box, HE staining and TUNEL immunostaining were used to observe animal memory ability, neuronal damage and neuronal apoptosis, respectively. Results: neuronal damage in hippocampus rise at 3 hour and 5 day after transient global ischemia, blood glucose stabilize at±0.5mmol·L-1 range about preoperative level can alleviate neuronal damage and apoptosis, while damage and apoptosis of BGF group that blood glucose fluctuate at±1.2 mmol·L-1 range about preoperative level have no significant differences compare to which of ischemia/reperfusion group. Result of shuttle box is consistent with neuronal apoptosis, blood glucose stabliliation can improve animal conditioned reflex behavior building speed damaged by transient global ischemia/reperfusion. Conclusion: perioperative glucostasis can improve neuronal damage and apoptosis of rat hippocampus after global brain ischemia. Part 3 Role of blood glucose concentration fluctuation control in glucose metabolism of rat brain after transient global brain ischemiaObjective: To observe the role of glucostasis in energy metabolism of rat hippocampus after transient global ischemia/reperfusion. Methods: Sixty-four normal SD rats randomized 4 groups, pseudooperation group(CON), global brain ischemia/reperfusion group(I/P), blood glucose fluctuation group(BGF) and blood glucose stabliliation group(BGS). Positron emission tomography (PET) and immunostaining were used to observe animal energy metabolism and GLUT1 expression. Results: Expressional levels of GLUT1 in hippocampus decrease at 5 day after transient global ischemia, blood glucose stabilizing at±0.5mmol·L-1 range about preoperative level can increase GLUT1 expression levels, while blood glucose fluctuating at±1.2mmol·L-1 range about preoperative level has no significant difference compare to which of ischemia/reperfusion groups. Results of PET are consistent with GLUT1 expression. Conclusion: Perioperative glucostasis may regulate energy metabolism of rat hippocampus on 5days after transient global brain ischemia/reperfusion through GLUT1 expression.Part 4 Role of blood glucose concentration fluctuation control in MAPK apoptosis pathway of rat brain after transient global brain ischemiaObjective: To investigate the effect of glucostasis in MAPK pathway after transient global ischemia in Sprague-Dawley rats. Methods: The hippocampus tissue obtained in 150min and 5d after ischemia/reperfusion were observed by western blot to analysis the expression of pERK, JNK and p38MAPK pathway protein, respectively. Results: pERK protein expression level increased at 150 minutes after rat global ischemia/reperfusion and maintained high level at 5 days. Glucostasis can upregulate pERK expression further, while glucose fluctuation has no significant difference effect compared to ischemia/reperfusion. JNK and p38 levels have no obvious change at 150 minutes, but upregulated at 5 days after rat global ischemia/reperfusion. Insulin infusion can decrease the expression of JNK and p38 caused by ischemia/reperfusion. There have no significant difference of JNK and p38 levels between glucose influctuation group and glucose stabilization group. Conclusion: Glucostasis change pERK pathway of MAPK, while the effects to JNK and p38MAPK pathway may achieved by insulin infusion.

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