mTORC2通路在苯丙胺致大鼠纹状体损伤中的变化

发布时间：2018-04-29 05:31

  本文选题：苯丙胺 + 纹状体　； 参考：《暨南大学》2014年硕士论文

【摘要】：目的 探讨mTORC2通路在苯丙胺致大鼠纹状体损伤的变化。 方法 通过给予SD大鼠腹腔注射苯丙胺（2.5mg/kg/d）建立动物模型，将SD大鼠随机分为空白组、生理盐水组与苯丙胺处理组，苯丙胺处理组又根据给药与取材时间的不同随机分为苯丙胺1h、1d、7d和14d组（空白组未给予任何处理；生理盐水组注射等量生理盐水；苯丙胺1h组给予苯丙胺注射1次，在给药后1h取材；苯丙胺1d组、7d组和14d组分别给药1次、7次、14次，每天1次，并于末次给药后24h内取材；而空白组、生理盐水组与苯丙胺14d组一同取材）。采用Open field方法测试SD大鼠在给药前、后的自主行为活动，用刻板行为评分方法(GHF-Dodd法)评价大鼠的刻板行为。用透射电镜观察大鼠纹状体部位神经元超微结构的损伤状况；Western blot方法检测mTORC2信号在纹状体的表达；免疫组化方法定位磷酸化的mTORC2在纹状体神经元的表达分布。 结果 行为学结果自主行为活动测试发现给予苯丙胺处理后，苯丙胺1d、7d、14d组自主活动的总距离、速度、运动时间与空白组比较增加，具有显著性差异（p0.01）。此外，刻板行为评分结果显示，苯丙胺7d、14d组与空白组比较刻板行为评分增加，具有显著性差异（p0.01）。 透射电镜结果透射电镜观察发现纹状体空白组细胞结构清晰，胞核大，线粒体多呈圆形或椭圆形，，线粒体膜结构清晰、嵴排列整齐；神经纤维结构清楚、完整。生理盐水组、苯丙胺1d组与空白组比较细胞结构未见明显差异。而苯丙胺14d组纹状体细胞结构模糊，核固缩，线粒体肿胀变形、嵴排列紊乱、断裂；纹状体神经纤维结构扭曲变形、甚至可见断裂。 Western blot结果在纹状体部位，P-rictor(S1219）在苯丙胺1d组（p=0.013）、7d组（p=0.021）、14d组（p=0.031）与空白组比较表达减少有统计学差异；P-AKT(T308)在苯丙胺14d组与空白组比较表达减少，p=0.000，具有显著性差异。 免疫组化结果在纹状体部位，P-rictor(S1219）免疫阳性细胞在纹状体尾壳核（CPU）和腹侧苍白球（LGP）都有分布；阳性产物主要分布在胞浆中，胞核淡染。P-rictor(S1219）阳性细胞数在苯丙胺7d组（p=0.006）、14d组(p=0.006)与空白组比较表达减少，具有显著性差异，与Western blot结果相一致。 结论 苯丙胺对纹状体的损伤机制可能与抑制mTORC2/AKT信号的表达有关。
[Abstract]:Purpose To investigate the changes of mTORC2 pathway in rat striatum injury induced by amphetamine. Method Sprague-Dawley rats were randomly divided into blank group, saline group and amphetamine treated group by intraperitoneal injection of 2.5 mg / kg 路kg / d of amphetamine. The amphetamine treatment group was also randomly divided into two groups according to the time of administration and taking materials. The control group was given no treatment, the saline group was injected with the same amount of normal saline, and the amphetamine 1h group was given the amphetamine injection once. The samples were obtained at 1 hour after administration of amphetamine, 14 times a day, once a day, and within 24 hours after the last administration in the 1-day group and 14d group, respectively, while in the blank group, the samples were taken together with the 14d group of amphetamine. Open field method was used to measure the autonomous behavior of SD rats before and after administration, and the stereotypical behavior was evaluated by GHF-Dodd method. The ultrastructure of neurons in rat striatum was observed by transmission electron microscope. The expression of mTORC2 signal in striatum was detected by Western blot, and the expression distribution of phosphorylated mTORC2 in striatum neurons was determined by immunohistochemical method. Result The results of behavioral test showed that the total distance, speed and exercise time of the group treated with amphetamine were significantly higher than those of the blank group (p 0.01). The total distance, speed and exercise time of the group treated with amphetamine were significantly higher than those of the blank group (P < 0.01). In addition, the scores of stereotype behavior in the amphetamine group on day 14 were significantly higher than those in the control group (p 0.01). The results of transmission electron microscope showed that the structure of striatum blank group was clear, the nucleus was large, the mitochondria were round or ellipse, the membrane structure of mitochondria was clear, the ridge was arranged neatly, and the structure of nerve fibers was clear and complete. There was no significant difference in cell structure between the saline group and the 1 d group compared with the blank group. In 14 d group, the structure of striatum cells was blurred, nucleus constricted, mitochondria swollen and deformed, ridge arranged disordered and broken, and striatum nerve fiber structure distorted and even broken. Western blot results: the expression of P-rictorus S1219 in the striatum was significantly lower than that in the control group (P 0.000). The expression of P-AKTT in the control group was significantly lower than that in the control group (P 0.000) compared with the control group on the 1st day, the expression of P-AKT was significantly lower in the control group than that in the control group on the 1st day, and the expression of P-AKT was significantly lower than that in the control group on the 1st day group (P 0.013) and in the control group on the 7th day (P 0.031), there was a significant difference between the two groups. The immunoreactive cells of P-rictorus S1219 in the striatum were distributed in the caudate putamen (CPU) and the ventral globus pallidus (LGP), and the positive products were mainly distributed in the cytoplasm. The expression of P0. 006 in the P0. 006 / 14d group was significantly lower than that in the control group, which was consistent with the results of Western blot. The number of the cells in the nucleus of the control group was significantly lower than that of the control group (P < 0. 006), and the results were consistent with those of the control group (P < 0. 006), which was consistent with the results of Western blot. Conclusion The mechanism of striatum injury induced by amphetamine may be related to the inhibition of mTORC2/AKT signal expression.
【学位授予单位】：暨南大学
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：R965

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