【摘要】：目的:建立甲基苯丙胺(MA)依賴大鼠模型,使用天麻素進(jìn)行干預(yù),檢測谷氨酸受體(Grin2B、Glur-1)、谷氨酸轉(zhuǎn)運(yùn)體(GLT-1、EAAT-1)及凋亡因子Caspase-3在MA依賴大鼠六個(gè)相關(guān)腦區(qū)(額葉皮質(zhì)、紋狀體、伏隔核、海馬、黑質(zhì)、中腦腹側(cè)被蓋區(qū))的表達(dá),探討天麻素對(duì)甲基苯丙胺所致中樞神經(jīng)系統(tǒng)毒性損害是否具有保護(hù)作用。方法:SD大鼠80只,隨機(jī)將80只大鼠分為6周組及8周組。將每組再分為:空白對(duì)照組,天麻素前干預(yù)組,甲基苯丙胺依賴組,天麻素(10mg)干預(yù)治療組及天麻素(20mg)干預(yù)治療組。通過腹腔注射甲基苯丙胺,采用條件性位置偏愛實(shí)驗(yàn)(CPP)和刻板行為評(píng)分觀察行為學(xué)變化,建立甲基苯丙胺依賴大鼠模型。確認(rèn)大鼠產(chǎn)生甲基苯丙胺依賴后,建成不同時(shí)間段的依賴模型,注射不同劑量的天麻素(10mg或20mg)干預(yù)。應(yīng)用免疫熒光、蛋白免疫印跡技術(shù)和實(shí)時(shí)定量PCR技術(shù)檢測鼠腦不同腦區(qū)谷氨酸受體(Grin2B、Glur-1)、谷氨酸轉(zhuǎn)運(yùn)體(GLT-1、EAAT-1)及 Caspase-3 的表達(dá)變化。結(jié)果:1、與對(duì)照組相比,MA依賴組和天麻素干預(yù)各組的條件性位置偏愛實(shí)驗(yàn)結(jié)果均有顯著性差異,表明成功建立了甲基苯丙胺依賴大鼠模型。2、MA依賴組Caspase-3表達(dá)增多;4周MA依賴組NR2B表達(dá)增高;大部分腦區(qū)Glur-1表達(dá)量增高;2周MA依賴組GLT-1、EAAT-1表達(dá)量降低,4周MA依賴組GLT-1、EAAT-1表達(dá)量增高。3、與天麻素干預(yù)前比較,天麻素干預(yù)后大鼠在伴藥盒停留時(shí)間明顯縮短。刻板行為評(píng)分,3分及4分次數(shù)明顯減少。4、天麻素干預(yù)后對(duì)MA依賴組Caspase-3、谷氨酸受體(Grin2B、Glur-1)及谷氨酸轉(zhuǎn)運(yùn)體(GLT-1、EAAT-1)的增高有治療作用,且天麻素20mg干預(yù)組效果優(yōu)于10mg組。結(jié)論:1、采用多次腹腔注射甲基苯丙胺,結(jié)合條件性位置偏愛實(shí)驗(yàn)和刻板行為評(píng)分的方法成功建立甲基苯丙胺依賴大鼠模型。2、與對(duì)照組比較,除谷氨酸轉(zhuǎn)運(yùn)體(GLT-1、EAAT-1)在2周MA依賴組降低,Caspase-3、谷氨酸受體(Grin2B、Glur-1)及谷氨酸轉(zhuǎn)運(yùn)體(GLT-1、EAAT-1)表達(dá)總體呈增高趨勢,提示Caspase-3、谷氨酸受體(Grin2B、Glur-1)及谷氨酸轉(zhuǎn)運(yùn)體(GLT-1、EAAT-1)與MA神經(jīng)毒性和依賴機(jī)制密切相關(guān)。3、天麻素干預(yù)后對(duì)MA依賴組大鼠行為學(xué)指標(biāo)有改善,對(duì)Caspase-3、谷氨酸受體(Grin2B、Glur-1)及谷氨酸轉(zhuǎn)運(yùn)體(GLT-1、EAAT-1)的增高有一定改善作用,且天麻素20mg干預(yù)組效果優(yōu)于10mg組。提示天麻素對(duì)甲基苯丙胺所致中樞神經(jīng)系統(tǒng)毒性損害具有保護(hù)作用。
[Abstract]:Aim: to establish a methamphetamine (MA) dependent rat model, and to investigate the effects of Gastrodin on glutamate receptor, glutamate transporter (GLT-1EAAT-1) and apoptosis factor Caspase-3 in six brain regions (frontal cortex, striatum, nucleus accumbens) of MA-dependent rats. The expression of Gastrodin in hippocampus, substantia nigra and ventral tegmental area of the midbrain, to investigate the protective effect of Gastrodin on the central nervous system (CNS) toxicity induced by methamphetamine. Methods 80 Sprague-Dawley rats were randomly divided into 6 week group and 8 week group. Each group was divided into three groups: blank control group, Gastrodin preintervention group, methamphetamine dependent group, 10mg intervention group and 20mg intervention group. A model of methamphetamine dependent rats was established by intraperitoneal injection of methamphetamine. The behavioral changes were observed by using conditioned place preference test (CPP) and stereotypical behavior score. After methamphetamine dependence was produced in rats, different time dependent models were established and different doses of Gastrodin (10mg or 20mg) were injected. The expression of glutamate receptor (Grin2BmGlur-1), glutamate transporter (GLT-1EAAT-1) and Caspase-3 in different brain regions were detected by immunofluorescence, Western blot and real-time quantitative PCR. Results compared with the control group, there were significant differences in the experimental results of conditioned place preference between the MA dependent group and Gastrodin intervention group. The results showed that the expression of Caspase-3 was increased in the methamphetamine dependent group and the NR2B expression was increased in the MA dependent group after 4 weeks. The expression of GLT-1 + EAAT-1 decreased in the MA dependent group at 2 weeks. The expression of GLT-1 + EAAT-1 was significantly increased in the MA dependent group at 4 weeks. Compared with that before Gastrodin intervention, the residence time of Gastrodin group was significantly shorter than that before Gastrodin intervention. The scores of stereotypical behavior, scores of 3 and times of 4 scores were significantly decreased. After the intervention of Gastrodin, the increase of Caspase-3, glutamate receptor (Grin2BtGlur-1) and glutamate transporter (GLT-1EAAT-1) in MA dependent group was significantly increased, and the effect of Gastrodin 20mg intervention group was better than that of 10mg group. ConclusionThe methamphetamine dependent rat model was successfully established by multiple intraperitoneal injection of methamphetamine, combined with conditional place preference test and stereotypical behavior score, compared with the control group. The expression of glutamate transporter (GLT-1) and glutamate transporter (GLT-1EAAT-1) decreased significantly in MA-dependent group at 2 weeks, and the expression of glutamate receptor (Grin2BtGlur-1) and glutamate transporter (GLT-1EAAT-1) increased significantly in MA-dependent group. These results suggest that Caspase-3, glutamate receptor (Grin2BnGlur-1) and glutamate transporter (GLT-1) are closely related to the neurotoxicity and dependence mechanism of MA. The increase of Caspase-3, glutamate receptor (Grin2BtGlur-1) and glutamate transporter (GLT-1) were improved to some extent, and the effect of Gastrodin 20mg intervention group was better than that of 10mg group. The results suggest that Gastrodin has protective effect on central nervous system toxicity induced by methamphetamine.
【學(xué)位授予單位】：昆明醫(yī)科大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2017
【分類號(hào)】：R749.64

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