鱼藤酮对小鼠运动协调和空间学习记忆能力的影响

发布时间：2018-08-17 17:46

【摘要】：帕金森病(Parkinson's disease, PD)是一种常见的神经系统退行性疾病。随着研究的深入,PD不再认为是仅仅在黑质—纹状体系统发生了病变,而是一个影响周围神经系统和中枢神经系统多个核团的疾病。PD患者的认知能力受损的发生率高达84%。迄今为止,PD的病因仍未完全阐明,尽管遗传和环境因素都起作用,但是90%以上的PD病人是散发性的。流行病学研究提示,环境因素与PD发病密切相关,长期接触农药的人群,PD的发病率较高。鱼藤酮是一种常用的除草剂和杀虫剂,能够通过血脑屏障,通过抑制线粒体呼吸链复合物I的活性,从而导致黑质多巴胺能神经元的能量合成障碍,进而引起神经元的死亡。因此,鱼藤酮被用于PD模型的制备。与其他的PD动物模型相比,鱼藤酮制备的PD模型具有明显的优势,它能够模拟PD的病理性特征变化,包括黑质多巴胺能神经元数目的降低和残存神经元内的alpha-突触核蛋白的聚集。目前,对鱼藤酮制备的PD模型的研究,多集中在黑质—纹状体系统的功能改变上,对其空间学习和记忆能力的改变尚未见报道。为了观察PD中认知功能的改变并初步探讨其机制,本实验选用12月龄的C57BL/6小鼠进行鱼藤酮灌胃处理,动物分为对照组,鱼藤酮处理3月组和处理1月组,应用免疫荧光和硫酸素T染色法观察黑质多巴胺能神经元数目的改变和神经元内alpha-突触核蛋白聚集的情况,应用转棒法观察小鼠运动协调能力的改变,应用Morris水迷宫评价小鼠空间学习与记忆能力的改变,同时通过免疫荧光双标的方法标记海马神经元的增殖细胞核抗原和微管相关蛋白,反映海马未成熟神经元的变化。研究结果如下： 1.在鱼藤酮处理3月组,小鼠黑质多巴胺能神经元数目减少24%,与对照组相比,差别有统计学意义(P0.05)；同时多巴胺能神经元内出现明显的alpha-突触核蛋白的聚集。而鱼藤酮处理1月组的小鼠未观察到明显变化。 2.在鱼藤酮处理3月组,小鼠在转棒上的停留时间降低了35%,与对照组相比,差别有统计学意义(P0.05)。而鱼藤酮处理1月组的小鼠未观察到改变。 3.鱼藤酮处理3月组的小鼠逃避潜伏期明显缩短,差别有统计学意义(P0.05)。鱼藤酮处理1月组的小鼠逃避潜伏期虽然有延长趋势,但与对照组相比,差别无统计学意义(P0.05)。 4.在水迷宫实验的第5天,鱼藤酮处理3月组小鼠到原平台位置的平均距离和在相对象限的时间分别缩短16.0%和34.5%(P0.05),穿越平台的次数和在训练象限的时间分别增加144%和33.4%(P0.05)。在水迷宫实验的第7天,到达原平台位置的时间、到原平台位置的平均距离和在相对象限的时间分别减少54.9%、22.2%和37.3%(P0.05),在训练象限的时间增加41.4%(P0.05)。这表明鱼藤酮处理3月组的小鼠对平台位置的记忆能力增强。 5.在水迷宫实验的第5天,鱼藤酮处理1月组的小鼠到原平台位置的平均距离、在相对象限的停留时间、穿越原平台位置的次数和在训练象限的时间均未检测出变化。然而,在第7天时,到原平台位置的平均距离延长19.8%(P0.05),在训练象限中停留时间百分比降低26.4%(P0.05)。这表明鱼藤酮处理1个月的小鼠记忆能力受到损害。 6.在鱼藤酮处理3月组和1月组,增殖细胞核抗原和微管相关蛋白对海马未成熟神经元的荧光双标记染色呈弱阳性,与对照组相比,差别无统计学意义。本实验证实,鱼藤酮处理3月组的小鼠出现了PD相关的病理学和行为学改变,黑质多巴胺能神经元数目减少和神经元内alpha-突触核蛋白的聚集,同时出现运动协调能力的障碍。本研究首次发现鱼藤酮处理3月组的小鼠出现了空间学习与记忆能力的升高,而鱼藤酮处理1月组的小鼠的空间记忆能力受损,这一现象并未伴随海马新生神经元的形成,其机制尚需进一步探讨。本实验为研究PD中学习与记忆功能障碍提供了一定的实验依据和思路。
[Abstract]:Parkinson's disease (PD) is a common neurodegenerative disease. With the development of research, PD is no longer considered as a disease that occurs only in the substantia nigra-striatum system, but a disease that affects multiple nuclei of the peripheral and central nervous system. The incidence of cognitive impairment in PD patients is as high as 84%. It is.
Up to now, the etiology of PD has not been fully elucidated, although genetic and environmental factors play a role, but more than 90% of PD patients are sporadic. Epidemiological studies suggest that environmental factors are closely related to the incidence of PD, long-term exposure to pesticides in the population, the incidence of PD is higher. Rotenone is a commonly used herbicide and pesticide, can pass through the blood. Brain barrier, by inhibiting the activity of mitochondrial respiratory chain complex I, leads to the dysfunction of energy synthesis of dopaminergic neurons in substantia nigra, leading to neuronal death. Therefore, rotenone is used in the preparation of PD models. Compared with other PD animal models, rotenone-based PD models have obvious advantages, which can simulate the disease of PD. Changes in rational characteristics include a decrease in the number of dopaminergic neurons in the substantia nigra and an accumulation of alpha-synuclein in the remaining neurons.
At present, the study on the PD model of rotenone mainly focuses on the functional changes of substantia nigra-striatum system, but the changes of spatial learning and memory abilities have not been reported. The changes of dopaminergic neurons in substantia nigra and the aggregation of alpha-synuclein in neurons were observed by immunofluorescence and sulfate T staining. The changes of motor coordination in mice were observed by rotating rod method. The spatial learning and memory of mice were evaluated by Morris water maze. The changes of memory and the expression of proliferating cell nuclear antigen (PCNA) and microtubule associated protein (MICP) in hippocampal neurons were labeled by immunofluorescence double labeling to reflect the changes of immature neurons in the hippocampus.
1. The number of dopaminergic neurons in substantia nigra of mice treated with rotenone for 3 months decreased by 24% compared with the control group (P 0.05), and there was an obvious accumulation of alpha-synuclein in dopaminergic neurons.
2. In rotenone treated group for 3 months, the retention time of rotator rod in mice decreased by 35%, which was statistically significant compared with the control group (P 0.05). In rotenone treated group for 1 month, no change was observed.
3. The escape latency of mice treated with rotenone for 3 months was significantly shortened, and the difference was statistically significant (P 0.05). The escape latency of mice treated with rotenone for 1 month was prolonged, but there was no significant difference compared with the control group (P 0.05).
4. On the fifth day of the water maze experiment, the average distance from the original platform and the relative quadrant time of rotenone treated mice in the three-month group were shortened by 16.0% and 34.5% (P 0.05), the times of crossing the platform and the time of training quadrant were increased by 14.4% and 33.4% (P 0.05), respectively. The average distance of platform position and the time of relative quadrant were reduced by 54.9%, 22.2% and 37.3% (P 0.05), respectively, and the time of training quadrant was increased by 41.4% (P 0.05). This indicated that rotenone could enhance the memory of platform position in mice treated with rotenone for 3 months.
5. On the fifth day of the water maze experiment, the average distance from the original platform to the original platform of mice in rotenone treatment group for one month was not changed in the time limit of residence, the times of crossing the original platform and the time of training quadrant. However, on the seventh day, the average distance to the original platform was extended by 19.8% (P 0.05) in the training quadrant. The percentage of residence time decreased by 26.4% (P 0.05). This indicated that rotenone treatment for one month impaired the memory of mice.
6. Proliferating cell nuclear antigen (PCNA) and microtubule-associated protein (MTRP) were weakly positive for double-labeled fluorescence staining of immature hippocampal neurons in rotenone treated group for 3 months and 1 month, and there was no significant difference between the two groups.
This study demonstrated that rotenone-treated mice showed PD-related pathological and behavioral changes, decreased dopaminergic neurons in substantia nigra and aggregation of alpha-synuclein in neurons, as well as impaired motor coordination in the rotenone-treated mice. The spatial memory of mice treated with rotenone for 1 month was impaired. This phenomenon did not accompany the formation of new neurons in the hippocampus. The mechanism needs to be further explored.
【学位授予单位】：青岛大学
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：R742.5

【共引文献】