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Toxicant-induced synaptic dysfunction and neurotoxicity in Parkinson disease

帕金森病中毒物引起的突触功能障碍和神经毒性

基本信息

批准号：
9356513
负责人：
KIM TIEU
金额：
$ 23.86万
依托单位：
FLORIDA INTERNATIONAL UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2016
资助国家：
美国
起止时间：
2016-09-09 至 2019-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9356513
关键词：
3-Dimensional Acute Animal Model Animals Attenuated Cell Count Cell Culture Techniques Cell Death Complement Complex Corpus striatum structure Cytosol DNA Sequence Alteration Data Dominant-Negative Mutation Dopamine Dyes Electron Microscopy Electrophysiology (science)Environment Environmental Risk Factor Epidemiology Equilibrium Equipment Exposure to Functional disorder Gene Proteins Genes Genetic Genetic Transcription Genetic study Genus Hippocampus Goals Grant Herbicides High Pressure Liquid Chromatography Human Immunoelectron Microscopy Immunohistochemistry Impairment Individual Injectable Injection of therapeutic agent Insecticides Lesion Light Link Location Measures Membrane Potentials Meperidine Methods Microdialysis Midbrain structure Mitochondria Mitochondrial Electron Transport Complex I Modeling Monitor Morphology Motor Motor Activity Movement Mus Mutant Strains Mice Mutation Nerve Degeneration Neurons Neurotoxins OPA1 gene Organelles Organic Cation Transporter Oxidation-Reduction Oxidative Stress Oxygen Consumption Paraquat Parkinson Disease Parkinsonian Disorders Pathogenesis Patients Pesticides Pharmacology Play Process Proteins Rattus Recombinant adeno-associated virus (rAAV)Respiration Risk Rodent Model Role Rotenone Serotyping Shapes Slice Superoxides Synapses Synaptosomes Techniques Testing Time Toxic Environmental Substances United States Variant Western Blotting base cytochrome c density disease diagnosis dopaminergic neuron epidemiology study extracellular gene environment interaction genetic approach in vivo inhibitor/antagonist insight interest laser capture microdissection mitochondrial dysfunction mitochondrial membrane nervous system disorder neuropathology neurorestoration neurotoxic neurotoxicity new therapeutic target nigrostriatal pathway novel novel therapeutics protein aggregation public health relevance small molecule synaptic function therapeutic target tool toxicant trafficking vector

项目摘要

DESCRIPTION (provided by applicant): Our long term goal is to study the mechanisms of neurodegeneration induced by environmental toxicants, genetic mutations and potential gene-environment interactions to gain insights into the pathogenesis of Parkinson's disease (PD). Advances in the genetics of PD have highlighted the critical role of mitochondrial dynamics (fission / fusion / movement) in neuronal function and survival. However, because monogenic familial PD represents only a small fraction of PD cases, it is critical to determine whether perturbed mitochondrial dynamics also plays a role in the nigrostriatal damage induced by environmental neurotoxicants. Most of these exogenous toxic molecules cause mitochondrial dysfunction either directly by blocking mitochondrial respiration, or indirectly through oxidative stress. Based on our preliminary data, this proposal will utilize two complementary toxicant-based animal models of nigrostriatal neurodegeneration: A) The herbicide paraquat (PQ) induces cell death primarily through oxidative stress. B) The pesticide/insecticide rotenone directly inhibits mitochondrial function. We hypothesize that whether excessive mitochondrial fission and dysfunction is induced directly by blocking mitochondrial respiration (rotenone) or indirectly by oxidative stress (PQ), promoting mitochondrial fusion will attenuate pre-synaptic dysfunction and neurotoxicity seen in these animal models. In Aim 1, we will investigate the impact of promoting mitochondrial fusion in the PQ mouse and rotenone rat models. Because PQ does not induce striatal damage in regular mice, we will use our novel mutant mice with deletion of the organic cation transporter 3 (Oct3-/-) to create a PQ animal model with damage in both nigra and striatum, as well as to enhance relevance to human gene-environment interactions because OCT3 variants have been associated with PD. Small molecule and gene-based approaches will be used for manipulation of mitochondrial fission/fusion machinery. Both neurorestorative and neuroprotective effects of these strategies will be determined in animals with pre-existing lesions and with active neurodegeneration. Striatal mitochondrial function, evoked striatal dopamine release in freely moving animals, synaptic function using electrophysiology, motor function and the integrity of the nigrostriatal pathway will be analyzed. In Aim 2, we will investigate the mechanisms by which PQ and rotenone induce mitochondrial fission and why blocking this process is protective. Where relevant, both animal models and cell cultures will be used. A wide range of state-of-the art equipment/techniques such as 3-dimensional electron microscopy, laser capture microdissection, the Seahorse extracellular flux analyzer and amperometry will be used to quantify alterations in levels of proteins/genes of interest specifically in nigral dopaminergic neurons, mitochondrial trafficking/morphology/function and synaptic density/function. Accomplishment of these aims will provide critical information regarding how toxic insults impact nigrostriatal pathway through perturbed mitochondrial dynamics and offer insights into a potential novel therapeutic target for PD.

描述（由申请人提供）：我们的长期目标是研究由环境毒物、基因突变和潜在的基因-环境相互作用诱导的神经变性机制，以深入了解帕金森病（PD）的发病机制。PD遗传学的进展突出了线粒体动力学（分裂/融合/运动）在神经元功能和存活中的关键作用。然而，由于单基因家族性PD仅代表PD病例的一小部分，因此确定受干扰的线粒体动力学是否也在环境神经毒物诱导的黑质纹状体损伤中发挥作用至关重要。这些外源性毒性分子中的大多数直接通过阻断线粒体呼吸或间接通过氧化应激引起线粒体功能障碍。根据我们的初步数据，该提案将利用两种互补的基于毒物的黑质纹状体神经变性动物模型：A）除草剂百草枯（PQ）主要通过氧化应激诱导细胞死亡。B）杀虫剂/杀虫剂鱼藤酮直接抑制线粒体功能。我们假设，无论是通过阻断线粒体呼吸（鱼藤酮）直接诱导过度的线粒体分裂和功能障碍，还是通过氧化应激（PQ）间接诱导过度的线粒体分裂和功能障碍，促进线粒体融合都会减弱这些动物模型中观察到的突触前功能障碍和神经毒性。在目的1中，我们将研究促进PQ小鼠和鱼藤酮大鼠模型中线粒体融合的影响。由于PQ不会诱导普通小鼠的纹状体损伤，我们将使用我们的有机阳离子转运蛋白3（Oct 3-/-）缺失的新型突变小鼠来创建黑质和纹状体损伤的PQ动物模型，以及增强与人类基因-环境相互作用的相关性，因为OCT 3变体与PD相关。小分子和基因为基础的方法将用于线粒体分裂/融合机制的操作。这些策略的神经恢复和神经保护作用将在具有预先存在的病变和具有活动性神经变性的动物中确定。将分析纹状体线粒体功能、自由活动动物中诱发的纹状体多巴胺释放、使用电生理学的突触功能、运动功能和黑质纹状体通路的完整性。在目标2中，我们将研究PQ和鱼藤酮诱导线粒体分裂的机制，以及为什么阻断这一过程是保护性的。如相关，将使用动物模型和细胞培养物。将使用各种最先进的设备/技术，如三维电子显微镜、激光捕获显微切割、Seahorse细胞外通量分析仪和电流分析法来定量黑质多巴胺能神经元、线粒体运输/形态/功能和突触密度/功能中感兴趣的蛋白质/基因水平的变化。这些目标的实现将提供关于毒性损伤如何通过扰乱线粒体动力学影响黑质纹状体通路的关键信息，并提供对PD潜在新治疗靶点的见解。