帕金森小鼠模型运动技能学习障碍的突触可塑性机制

发布时间：2018-09-12 10:31

【摘要】：帕金森氏病(Parkinson's Disease, PD)是仅次于阿尔兹海默症的第二大常见的神经退行性疾病,最显著的病理变化特征是中脑多巴胺能神经元的大量耗竭。主要临床症状包括运动机能障碍和认知障碍尤其是运动技能学习障碍。目前关于PD的研究大都集中在基底神经节,表明运动机能障碍的原因主要源于基底神经节神经回路结构与功能的异常,然而对PD运动技能学习能力下降和记忆受损的原因却没有很好的解释。运动皮层是支配运动功能的高级中枢,突触可塑性的动态适应性是运动技能学习的重要基础。运动皮层中的多巴胺能信号处理可以增强相关运动的协调性并促进精细运动的完成。有研究表明在多巴胺信号明显受损的PD中运动皮层的功能发生异常,若硬脑膜外慢性刺激运动皮层可以改善PD的症状。那么,随着多巴胺能神经元的耗竭运动皮层神经环路到底发生了怎样的变化?这种变化是否是运动技能学习和记忆障碍的内在原因?为了解决这些问题,我们通过对Thy1-YFP-H line转基因小鼠的颅骨建立颅窗并结合双光子激光扫描显微镜对第五层椎体神经元的顶树突进行活体成像,研究多巴胺神经元耗竭诱导的运动皮层突触可塑性的动态变化规律,并结合PD小鼠运动技能学习训练初步在突触层面阐释其学习认知障碍的原因。主要研究内容如下：(1)建立稳定可靠的PD小鼠模型：主要应用连续腹腔注射1-甲基-4-苯基-1,2,3,6-四氢吡啶(1-Methyl-4-Phenyl-1,2,3,6-Tetrahydropyridine, MPTP)的方法建立PD小鼠模型,并通过免疫组织化学染色方法鉴定多巴胺能神经元的受损程度,检验此PD模型的可靠性和稳定性。(2)PD小鼠大脑运动皮层树突棘的动态性变化：利用双光子活体成像技术对不同PD模型小鼠的初级运动皮层反复成像,发现树突棘生成率和消亡率都显著性增加,但是这种增加只特定发生在初级运动皮层,附近的桶状皮层树突棘变化率不受影响。利用治疗PD最常用药物L-DOPA,外源补充多巴胺可以部分逆转多巴胺耗竭诱导的运动皮层树突棘可塑性变化增强的现象,显示运动皮层树突棘动态性变化的加剧应该是源于多巴胺耗竭。(3)PD小鼠初级运动皮层神经环路的重塑：随着MPTP用量的增加,多巴胺能神经元损伤加剧,PD小鼠运动皮层中树突棘的生成率、消亡率、变化率呈持续性累积增加,树突棘密度略有降低,通过反复对同一动物同一皮层区域树突棘的成像和追踪,发现原有树突棘稳定性降低,新生树突棘稳定性升高。这些数据表明随着多巴胺能神经元的耗竭,大脑运动皮层通过树突棘动态可塑性异常变化使原突触连接选择性消失、新突触连接特异性建立,致使原有神经连接发生紊乱,神经环路发生异常重塑。(4)PD小鼠运动技能学习和记忆障碍的突触机制：对PD小鼠进行运动技能学习训练,并结合双光子活体成像技术对训练过程中的小鼠不同时间点进行间隔成像,分析运动技能学习诱导的树突棘的生成率、消亡率和新生树突棘的稳定率。结果表明PD小鼠模型中随着多巴胺的耗竭,在行为上表现为运动技能的学习和记忆损伤,神经环路上表现为学习过程中大脑运动皮层树突棘不能正常增长和消亡,运动学习技能诱导的新生树突棘稳定性下降。因此,我们推测PD中运动技能学习障碍和记忆障碍的突触机制是多巴胺耗竭导致的运动皮层神经环路突触可塑性的异常。综上所述,本研究通过双光子活体成像技术首先证实了帕金森小鼠大脑运动皮层神经环路可塑性发生异常,观测到了神经信号传递基本结构单位突触的大量异常丢失和生成。结合行为学实验,初步阐明了帕金森小鼠模型运动技能学习和记忆障碍的突触机制。
[Abstract]:Parkinson's disease (PD) is the second most common neurodegenerative disease after Alzheimer's disease. The most significant pathological change is the depletion of dopaminergic neurons in the brain. The main clinical symptoms include motor dysfunction and cognitive impairment, especially motor learning impairment. Most studies focus on the basal ganglia, indicating that motor dysfunction is mainly caused by abnormalities in the structure and function of the basal ganglia neural circuits, but there is no good explanation for the decline of motor learning ability and memory impairment in PD. Adaptability is an important basis for motor learning. Dopaminergic signal processing in the motor cortex can enhance the coordination of related movements and facilitate the completion of fine movements. Symptoms. So, what happens to the motor cortical circuits as dopaminergic neurons are depleted? Is this an intrinsic cause of motor learning and memory impairment? To solve these problems, we created skull windows in the skulls of Thy1-YFP-H line transgenic mice and demonstrated them with two-photon laser scans. The apical dendrites of the fifth layer of vertebral neurons were imaged in vivo under microscope to study the dynamic changes of synaptic plasticity induced by dopamine depletion in the motor cortex. The main contents of this study were as follows: (1) Establishing stable and reliable motor cortex synaptic plasticity in PD mice. PD mouse model was established by continuous intraperitoneal injection of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). The damage degree of dopaminergic neurons was identified by immunohistochemical staining, and the reliability and stability of the PD model were tested. Dynamic changes of dendritic spines in the motor cortex of mice: Two-photon in vivo imaging of the primary motor cortex of mice with different PD models revealed a significant increase in the formation and extinction rates of dendritic spines, but this increase occurred only in the primary motor cortex, and the change rate of dendritic spines in the nearby barrel cortex remained unchanged. Effects. Using L-DOPA, the most commonly used drug in the treatment of PD, exogenous dopamine supplementation could partly reverse the increased plasticity of motor cortical dendritic spines induced by dopamine depletion, suggesting that the increased dynamic changes of motor cortical dendritic spines should be due to dopamine depletion. (3) The remodeling of primary motor cortical nerve circuits in PD mice: with the increase of dopamine depletion With the increase of MPTP dosage, the damage of dopaminergic neurons was aggravated. The formation rate, extinction rate and change rate of dendritic spines in the motor cortex of PD mice increased continuously, and the density of dendritic spines decreased slightly. These data indicate that with the depletion of dopaminergic neurons, the abnormal dynamic plasticity of the motor cortex through the dendritic spines makes the original synaptic junction selective disappear, and the new synaptic junction specificity is established, resulting in the disorder of the original neural connection, and the abnormal remodeling of the neural circuits. (4) The motor skills learning and memory of PD mice Synaptic mechanism of dysfunction: PD mice were trained in motor skill learning, and two-photon in vivo imaging was used to imaging at different intervals during training. The formation rate, extinction rate and stability rate of new dendritic spines induced by motor skill learning were analyzed. Amine depletion is manifested in behavioral learning and memory impairment of motor skills, and in neural circuits, dendritic spines in the motor cortex do not grow and die normally during learning, and the stability of the new dendritic spines induced by motor learning skills decreases. In summary, this study first confirmed the abnormality of synaptic plasticity in the motor cortex of Parkinson's mice by two-photon in vivo imaging, and observed a large number of abnormal synaptic loss and birth of the basic structural unit of neural signal transduction. The synaptic mechanism of motor learning and memory impairment in Parkinson's mouse model was elucidated by behavioral experiments.
【学位授予单位】：华中科技大学
【学位级别】：博士
【学位授予年份】：2016
【分类号】：R742.5;R-332

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