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AXON-TARGETED MICRODEVICES FOR CNS AXON TRANSPORT STUDIES

用于中枢神经系统轴突运输研究的轴突靶向微器件

基本信息

批准号：
8129436
负责人：
Shelly Elese Sakiyama-Elbert
金额：
$ 22.34万
依托单位：
WASHINGTON UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2010
资助国家：
美国
起止时间：
2010-09-01 至 2013-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8129436
关键词：
Affect Alzheimer&apos s Disease Area Axon Axonal Transport Back Biological Assay Cell Adhesion Cell Death Cells Cellular Stress Controlled Environment Data Development Devices Dimensions Disease Disease model Environment Functional disorder Genetic Models Goals Green Fluorescent Proteins Image Impairment In Vitro Injury Knowledge Label Lead Liquid substance Microfabrication Midbrain structure Mitochondria Molecular Movement Multiple Sclerosis Mus Nerve Degeneration Nervous System Trauma Neuraxis Neurodegenerative Disorders Neurons Organelles Oxidopamine Parkinson Disease Pathology Patients Pharmaceutical Preparations Physiological Play Population Process Property Public Health Research Research Personnel Risk Role Sampling Seeds Side Signal Transduction Slide Stroke Synapses System Techniques Testing Time Toxin Transfection Trauma Tyrosine 3-Monooxygenase Work axon growth axonal degeneration design disease mechanisms study dopaminergic neuron fluorophore in vivo insight mimetics nervous system disorder neuronal cell body novel novel therapeutics promoter protein aggregation prototype public health relevance research study restraint synaptic function therapeutic development tool trafficking

项目摘要

DESCRIPTION (provided by applicant): Axonal injury is a central component in many neurological disorders including trauma, stroke, and multiple sclerosis as well as neurodegenerative disorders such as Alzheimer's (AD) and Parkinson's disease (PD). The overall goal of this research is to develop compartmentalized culture chambers using microfabrication techniques that isolate axons in culture to facilitate the study of axon growth, function and pathology. Specifically, the design and fabrication of microdevices will allow the quantitative delivery of pharmacological agents selectively to axons and allow their effects on axonal transport and growth to be examined in a controlled environment. By understanding the effects of toxins or drugs on axon growth and transport, we will gain mechanistic insight into the pathophysiology of neurodegenerative disorders and injury, which could in turn guide the development of therapeutics for treatment of degeneration or injury. The focus of this project is to develop open chamber microdevices that allow axonal isolation and the targeted application of drugs/toxins. Because patient data, genetic models, and in vivo and in vitro toxin studies all support a compelling role for axonal dysfunction in PD, we will use a well characterized PD model to study toxin effects in axons at risk in this disorder. Specifically, cultures of midbrain neurons from mice expressing green fluorescent protein (GFP) under the tyrosine hydroxylase (TH) promoter in our microdevices will allow us to test in real time the hypothesis that the Parkinson mimetic 6-OHDA (6-Hydroxydopamine) triggers changes in axonal transport resulting in the loss of axonal function and ultimately cell death. The specific aims of this proposal are to: (1) determine the design constraints for an open chamber design that allows easy cell seeding and axonal assays, which can be readily scaled for medium/high throughput cultures, (2) determine the design constraints for a novel microdevice that allows the application of a drug/toxin to axons (but not cell bodies) that are synapsing onto another population of neurons, and (3) use the new microdevices to test for the first time the effects of PD-mimetic 6-OHDA on mitochondrial trafficking in dopaminergic (DA) axons. PUBLIC HEALTH RELEVANCE: If the aims of this proposal are achieved, novel microdevices will become available to a large numbers of researchers trying to understand the role that axons play in neurological disease and injury. Establishing a causal role for axon dysfunction in neurodegenerative disorders such as Parkinson's and Alzheimer's disease is a challenging task given current methodological restraints. The tools proposed here will overcome current limitations in this field and further advance our knowledge in this area by providing unique platforms to study pharmacological and molecular mechanisms of disease and to screen for novel therapeutics.

描述（由申请人提供）：轴突损伤是许多神经系统疾病（包括创伤、中风和多发性硬化症）以及神经退行性疾病（如阿尔茨海默病（AD）和帕金森病（PD））的核心组成部分。这项研究的总体目标是开发分区的培养室，使用微加工技术，隔离轴突的文化，以促进轴突的生长，功能和病理学的研究。具体而言，微器件的设计和制造将允许选择性地将药理学试剂定量递送到轴突，并允许在受控环境中检查它们对轴突运输和生长的影响。通过了解毒素或药物对轴突生长和运输的影响，我们将获得对神经退行性疾病和损伤的病理生理学的机械见解，这反过来又可以指导治疗退行性疾病或损伤的治疗方法的发展。该项目的重点是开发开放室微设备，允许轴突分离和药物/毒素的靶向应用。由于患者数据、遗传模型以及体内和体外毒素研究都支持轴突功能障碍在PD中的重要作用，因此我们将使用一个充分表征的PD模型来研究毒素对这种疾病中有风险轴突的影响。具体而言，在我们的微装置中，来自表达酪氨酸羟化酶（TH）启动子下的绿色荧光蛋白（GFP）的小鼠的中脑神经元的培养物将允许我们在真实的时间内测试帕金森模拟物6-OHDA（6-羟基多巴胺）触发轴突运输的变化导致轴突功能丧失和最终细胞死亡的假设。这项建议的具体目标是：（1）确定允许容易的细胞接种和轴突测定的开放室设计的设计约束，其可以容易地按比例用于中/高通量培养，（2）确定允许将药物/毒素应用于轴突的新型微型装置的设计约束（3）使用新的微器件首次测试PD模拟物6-OHDA对多巴胺能（DA）轴突中线粒体运输的影响。公共卫生关系：如果这项提议的目标得以实现，新型的微型设备将为大量试图了解轴突在神经疾病和损伤中所起作用的研究人员所用。在神经退行性疾病如帕金森病和阿尔茨海默病中建立轴突功能障碍的因果关系是一项具有挑战性的任务，因为目前的方法限制。本文提出的工具将克服这一领域目前的局限性，并通过提供独特的平台来研究疾病的药理学和分子机制以及筛选新的治疗方法，进一步推进我们在这一领域的知识。