Mapping Functional Connectivity in a Rat Model of Early Stage Parkinson´s Disease with Simultaneous Optogenetic PET/fMRI

帕金森病（PD）是全球第二大常见的神经退行性疾病。病理学标志是黑质丘脑部多巴胺能神经元的进行性损失和称为路易体的小胞浆内包涵体的存在，其主要由突触前蛋白α-突触核蛋白（α-syn）组成。黑质纹状体神经元的进行性变性导致纹状体中神经递质多巴胺（DA）的严重缺乏，但临床症状出现在非常晚的阶段，当DA缺乏约80%时。相比之下，最近发表的数据表明，多巴胺能功能障碍发生在这些神经元开始退化之前的疾病的非常早期的时间点。问题是否α-syn聚集是突触前功能障碍的一个可能的介质仍然没有解决。本研究项目的主要目的是利用PET/BOLD-fMRI技术在体内同步监测早期PD动物模型中突触前神经传递的早期功能变化。为此目的，将使用光遗传学方法建立在体内条件下选择性刺激和记录健康和α-syn过表达大鼠中黑质纹状体神经元活性的方法和方案。腺相关病毒（AAV）载体介导的α-syn的过表达提供了监测早期功能变化的机会，因为神经变性随着时间的推移逐渐发展并反映早期和晚期PD。通过将微生物光敏阳离子通道视紫红质-2（ChR-2）引入黑质纹状体神经元，将实现光遗传学控制的神经元激活。为了实现ChR-2和人野生型α-syn的组合表达，将测试三种不同的AAV载体构建体，并且最佳构建体将用于脑切片中和体内条件下的光刺激和电记录。

Parkinson´s disease (PD) is the second most common neurodegenerative disorder worldwide. The pathological hallmark is the progressive loss of dopaminergic neurons in the substantia nigra pars compacta and the presence of small intracyctoplasmatic inclusions known as Lewy Bodies, mainly composed of the presynaptic protein alpha-synuclein (alpha-syn). The progressive degeneration of nigrostriatal neurons results in a profound deficiency of the neurotransmitter dopamine (DA) in the striatum, but clinical symptoms appear at a very late stage when the DA deficit is about 80 %. In contrast, recently published data suggest that dopaminergic dysfunctions occur at a very early time point of the disease before these neurons start to degenerate. The question whether alpha-syn aggregation is a possible mediator of the presynaptic dysfunctions remains still unresolved. The main objective of this research project is to monitor early functional changes of presynaptic neurotransmission in an animal model of early stage PD using simultaneous in vivo PET/BOLD-fMRI. For this purpose methods and protocols to selectively stimulate and record the activity of nigrostriatal neurons in healthy and alpha-syn overexpressing rats under in vivo conditions will be established using the optogenetic approach. The adeno-associated viral (AAV) vector mediated over-expression of alpha-syn provides the opportunity to monitor early functional changes, since neurodegeneration develops progressively over time and reflects early and late stage PD. Optogenetically controlled neuron activation will be achieved by introduction of microbial light-sensitive cation channels, Channelrhodopsin-2 (ChR-2), into nigrostriatal neurons. To achieve combined expression of ChR-2 and human wild type alpha-syn, three different AAV vector constructs will be tested and the best construct will be used for optical stimulations and electrical recordings in brain slices and under in vivo conditions.