本文選題：癲癇 + 經皮三叉神經電刺激　； 參考：《安徽醫(yī)科大學》2014年碩士論文

【摘要】：目的： 1.觀察經皮三叉神經電刺激（TcTNS）對匹羅卡品誘發(fā)的癲癇持續(xù)狀態(tài)（SE）所致海馬神經元損傷的保護作用。 2.觀察觀察經皮三叉神經電刺激（TcTNS）對匹羅卡品誘發(fā)的慢性癲癇模型大鼠海馬神經元GAD65/67表達的影響，以探討經皮三叉神經電刺激治療癲癇及腦保護作用的可能機制。 資料和方法： 實驗一： 選用健康成年雄性SD大鼠，采用隨機數字表法將大鼠隨機分為二組，匹羅卡品模型組和對照組，模型組用匹羅卡品誘導癲癇點燃模型，對照組注射等量的生理鹽水，經過行為學觀察兩周后，具有慢性癲癇自發(fā)性再發(fā)作的為模型制備成功大鼠，再將模型制備成功大鼠采用隨機數字表法分為兩組，治療組和模型組，分別給予三叉神經電刺激和假刺激一個月（刺激參數為頻率140Hz,電流10mA,脈寬0.5ms，正向脈沖刺激30s間歇5min，連續(xù)刺激1h，假刺激組各項參數均為0。刺激時間定為每天12:00-16:00）后終止刺激，再次注射皮羅卡品誘導一次癲癇持續(xù)狀態(tài)（SE）并于SE后24h,48h,72h處死大鼠并取材，對照組亦在相應時間點取材，采用TUNNEL和NISSEL染色觀察各組大鼠海馬神經元細胞的凋亡和壞死情況。組間比較采用配對t檢驗，以p㩳0.05定義為差別有統計學意義。 實驗二： 選用健康成年雄性SD大鼠，采用同實驗一的方法建立癲癇點燃模型，將慢性癲癇模型制備成功者用隨機數字表法分為模型組與治療組，分別給予三叉神經電刺激與假刺激一個月（刺激參數及時間同實驗一），于停止刺激后24h，72h，一周，二周，四周處死大鼠并取材，對照組亦于相應時間點取材。采用免疫組化的方法研究大鼠海馬及皮層GAD65、67的表達情況，組間比較采用配對t檢驗，組內比較采用單因素方差分析，以p㩳0.05定義為差別有統計學意義。 結果： 1．在實驗一中： 1.1TUNNEL染色：SE后各時間點治療組和模型組均可見TUNNEL陽性細胞，主要分布在海馬的CA1和CA3區(qū)，SE后24h，48h，72h治療組CA3區(qū)的TUNNEL陽性細胞較模型組顯著減少（p＜0.05），以72小時尤為明顯（p＜0.01）正常對照組可見少量TUNNEL陽性細胞。 1.2NISSEL染色：正常對照組顯示海馬CA1和CA3區(qū)神經元細胞排列整齊緊密，沒有明顯的神經元缺失，細胞核呈圓形或橢圓形，胞漿內尼氏小體豐富。模型組各時間點可見不同程度的神經元丟失，部分神經元胞體皺縮，胞漿深染，核固縮，錐體細胞存活數較正常細胞減少，尤以72小時最為明顯（p＜0.01）。經皮三叉神經刺激治療組各時間點大鼠海馬CA1，，CA3區(qū)神經元細胞尚清晰完整，部分細胞出現染色質凝集，神經元丟失情況較模型組顯著下降，即治療組Nissl染色細胞數顯著多于模型組（p＜0.05）。 2.在實驗二中： 2.1：海馬區(qū)及皮層均有GAD65的陽性細胞表達，尤以海馬的CA3區(qū)表達最為明顯，主要在胞核中表達，陽性細胞為棕紅色顆粒，經皮三叉神經刺激治療組較假刺激組相比，各時間點治療組陽性細胞表達均高于模型組（p＜0.05），治療組GAD65的表達于停止刺激后72小時-7天達到高峰，較模型組有極顯著差異（p＜0.01）。后GAD65的表達下降，至28天接近正常表達水平。 2.2：海馬區(qū)及皮層均可見GAD67陽性細胞，在海馬的CA3區(qū)及DG區(qū)表達最為明顯，主要在胞漿中表達，與模型組各時間點相比較，治療組的表達遠大于模型組（p＜0.05）。組內比較發(fā)現，GAD67無明顯的時間變化趨勢，至停止刺激后28天，治療組的GAD67的表達仍大于正常水平。 結論： 1.經皮三叉神經電刺激治療可以減少癲癇大鼠海馬神經元的損傷，從而發(fā)揮對癲癇大鼠海馬神經元的保護作用。 2.經皮三叉神經電刺激可使癲癇大鼠腦內抑制性神經遞質的標志物GAD65/67表達上調，,抑制性遞質的增強可能為三叉神經電刺激治療癲癇及發(fā)揮腦保護作用的機制之一。
[Abstract]:Purpose :

1 . To observe the protective effect of transcutaneous trigeminal nerve stimulation ( TcN ) on hippocampal neuronal injury induced by pilocarpine ( SE ) .

2 . To observe the effect of percutaneous trigeminal nerve stimulation ( TcN ) on the expression of GAD65 / 67 in rat hippocampal neurons induced by pilocarpine , and to investigate the possible mechanism of percutaneous trigeminal nerve stimulation in the treatment of epilepsy and brain protection .

Information and methods :

Experiment 1 :

Adult male SD rats were randomly divided into two groups : two groups , pilocarpine model group and control group . The model group was divided into two groups by pilocarpine . After two weeks of behavioral study , the rats were divided into two groups : treatment group and model group .

Experiment 2 :

The rats were randomly divided into three groups : model group and treatment group . The rats were sacrificed at 24 h , 72 h , one week , two weeks and four weeks after cessation of stimulation . The expression of GAD65 and 67 in hippocampus and cortex of rats was studied by immunohistochemistry .

Results :

1 . In Experiment 1 :

1 . After SE treatment group and model group , positive cells were observed , and the number of positive cells was significantly decreased at 24 h , 48 h and 72 h after SE ( p < 0 . 05 ) , and a small number of positive cells were found in the normal control group at 72 hours ( p < 0.01 ) .

1 . 2NISSEL staining : The normal control group showed that the neuronal cells in the CA1 and CA regions of the hippocampus were orderly and compact , no obvious neuronal loss was found , the nuclei were round or oval , and the Nissl in the cytoplasm was abundant .

2 . In Experiment 2 :

2.1 : The expression of GAD65 positive cells in the hippocampus and cortex was the most obvious , especially in the CA 3 region of hippocampus . The positive cells were higher than those in the model group ( p < 0.05 ) . The expression of GAD65 in the treatment group was higher than that in the model group ( p < 0.05 ) . The expression of GAD65 in the treatment group decreased from 72 hours to 7 days after cessation of stimulation , and the expression of GAD65 decreased to close to normal expression level 28 days .

2.2 : The expression of GAD67 positive cells was found in the hippocampus and in the cortex . The expression of GAD67 in hippocampus was much higher than that in model group ( p < 0.05 ) . Compared with model group ( p < 0.05 ) , the expression of GAD67 was much larger than that in model group ( p < 0.05 ) .

Conclusion :

1 . Percutaneous trigeminal nerve stimulation can reduce the damage of hippocampal neurons of epileptic rats , and play an important role in the protection of hippocampal neurons in epileptic rats .

2 . The expression of GAD65 / 67 is up - regulated by percutaneous trigeminal nerve stimulation , and the enhancement of inhibitory neurotransmitter may be one of the mechanisms of trigeminal nerve stimulation in the treatment of epilepsy and brain protection .
【學位授予單位】：安徽醫(yī)科大學
【學位級別】：碩士
【學位授予年份】：2014
【分類號】：R742.1

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