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

帕金森病（PD）是世界上第二大最常见的神经退行性疾病。其病理特征是黑质致密部多巴胺能神经元的进行性丧失和细胞浆内小包涵体（主要由突触前蛋白-突触核蛋白（α -syn）组成）的存在。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，但临床症状出现在很晚的阶段，当DA缺失约80%时。相反，最近发表的数据表明，在这些神经元开始退化之前，多巴胺能功能障碍发生在疾病的早期时间点。α -syn聚集是否可能是突触前功能障碍的中介仍未解决。本研究项目的主要目的是利用体内PET/BOLD-fMRI同时监测早期PD动物模型突触前神经传递的早期功能变化。为此，我们将利用光遗传学的方法，在体内条件下，对健康大鼠和α -syn过表达大鼠的黑质纹状体神经元进行选择性刺激和记录。腺相关病毒（AAV）载体介导的α -syn过表达提供了监测早期功能变化的机会，因为神经退行性变随着时间的推移逐渐发展，反映了早期和晚期PD。光遗传控制神经元激活将通过引入微生物光敏阳离子通道，通道视紫红质-2 （chr2）进入黑纹状体神经元来实现。为了实现hr -2和人类野生型α -syn的联合表达，我们将测试三种不同的AAV载体构建，并将最佳构建用于脑切片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.