The Mechanism Study Of Electrical Stimulation Of Cerebellar Fastigial Nucleus Resisting The Inflammatory Injury Following Focal Cerebral Ischemia And Reperfusion In Rats

Background: More attention has been paid to the roles of the mechanism of inflammatory injury in the cerebral ischemia and reperfusion recently. Nuclear factor-κB activation has been taken as the key aspect in the inflammatory responses for cerebral ischemia and reperfusion. However, which roles do the NF-κB play in the cerebral ischemia and reperfusion, the neuroprotection or the neurodegeration? Further studies on this aspect would be necessary due to the disagreement on this problem.Recently studies showed that FNS as a non-drug therapeutic tool has the exact neuroprotection and anti-inflammatory effects. Up to date, no studies were showed whether FNS had the direct regulation of NF-κB activation post-ischemia or not, nor did the studies on the mechanism of FNS neuroprotection at the transcription level.Brain circulation therapy machine based on the mechanisms of FNS showed the satisfied clinical effects on several CNS diseases by percutaneously electrical stimulation of cerebellar fastigial nucleus(similar to electroacupuncture"wangu" point of Traditional Chinese Medicine). However, few studies on this aspect whether the clinic effects of percutaneously electrical stimulation of cerebellar fastigial nucleus were produced by cerebellar fastigial nucleus or not. There may be a better understanding for the mechanism of FNS clinical effects by studying the relationship between electroacupuncture "wangu" point and cerebellar fastigial nucleus. Objective: (1) To investigate the expression and the activation of NF-κB and the change of NF-κB DNA binding activity following focal cerebral ischemia and reperfusion in rats. The inhibitor of NF-κB activation PDTC was applied to block its activation in order to further confirm which roles it played in brain inflammatory damage after focal cerebral ischemia and reperfusion in rats, neurodegeneration or neuroprotection? (2) To further explore the mechanisms of electrical stimulation of cerebellar fastigial nucleus (FNS) resisting the inflammatory injury following focal ischemia and reperfusion on the transcriptional level.(3) To explore the effect of electroacupuncture "wangu" point (EA) on the inflammatory injury following focal cerebral ischemia and reperfusion transcriptionally and further investigate whether the effect was dependent on intact cerebellar fastigial nucleus or not. And address the cause-effect relationship between the clinical effects of percutaneous FNS and the cerebellar fastigial nucleus, by which we can further understand for the mechanism of percuaneous FNS clinical effects.Methods: The models of male adult Wistar rats with or withoutbilateral FN neurons lesioned by the microinjection of ibotenic acid 5 days before FNS or EA and transient right middle cerebral artery occlusion (TMCAO) were used. The different interventional therapeutic methods including FNS, FN sham stimulation, EA and the administration of PDTC intravenously were applied to our study. The injuried brain tissue was determined for cresyl violet staining (Nissl's staining) and hematoxylin and eosin (HE) staining to observe the ischemic pathological changes of brain tissue and estimate the survival neurons in ischemic cortex and for TTC staining to observe the area of infarctions, for immunohistochemical staining of the active form of NF-κB p65 protein subunit, in order to observe activation and nuclear translocation of NF-κB, for hybridization in situ to detect expression of NF-κB p65 mRNA, for RT-PCR to detect expression of IL-1β mRNA, for Western blot analysis to detect the content of NF-κB p65 protein subunit in nuclear extracts from ischemic cortex as well as for electrophoretic mobility shift assay (EMSA) to detect the NF-κB DNA binding activity in nuclear extracts isolated from ischemic cortex. In addition, the neurological symptom score of ischemic rats must be estimated. Our experiment was designed to fully understand the changes of NF-κB p65 biological performances in the course of focal cerebral ischemia and reperfusion...