Suite of high content assays for mitochondrial dynamics in neurons

项目摘要/摘要 迫切需要开发新的治疗方法来治疗精神疾病和 神经紊乱。尽管有这种需要，但在过去两年中，在实现这一目标方面进展甚微。 几十年。对这一失败的一种解释是，基于细胞的屏幕旨在识别新的导线 传统上，化合物使用永生化的非神经元细胞系。这是可以理解的，在 回顾过去，为什么过去的屏幕未能产生人们所希望的过多的新的和重要的 与其他类型的细胞相比，化合物赋予了神经元非凡的结构和生理能力。一个 该失败的第二个可能原因是大多数基于单元的屏幕使用了基于目标的方法， 其中特定的蛋白质靶标被确定，可能为新疗法的开发带来希望。 然而，大脑以及大脑疾病如何影响其功能仍然是一个巨大的谜团。这一事实证明 表型屏幕的替代品可能会带来更多的希望。表型屏幕搜索 在不知道特定分子靶点的情况下对特定细胞生物学表型的影响 影响表型。 我们开发了一种新型的筛选平台，利用原代培养的神经元和细胞生物学， 用于识别小分子影响的表型读数。该方法灵活、可伸缩、 与传统方法相比，提供了许多优势。在这个项目中，我们建议使用以下技术开发分析方法 这个平台用于线粒体动力学过程，包括生物发生、分裂、融合、蛋白质输入、 分枝和损坏。我们将使用线粒体的荧光标记来跟踪线粒体 分析，并进行三个小分子筛选对线粒体动力学的影响 分析方法得到了优化。线粒体功能障碍与情绪等多种精神障碍有关 精神障碍、精神分裂症和自闭症；以及神经退行性疾病。因此，这个项目承诺了一个 丰富的新资源，可用于研究小分子、RNAi或基因过表达对 神经元中的线粒体动力学。

PROJECT SUMMARY / ABSTRACT There is a pressing need to develop new classes of therapeutics for the treatment of psychiatric and neurological disorders. Despite this need, little progress has been made towards this goal over the last two decades. One explanation for this failure is that the cell-based screens designed to identify new lead compounds have traditionally utilized immortalized, non-neuronal cell lines. It becomes understandable, in retrospect, why past screens have failed to yield the hoped-for plethora of new and important classes of compounds given the extraordinary architecture and physiology of neurons compared to other cell types. A second possible reason for this failure is that most cell-based screens have utilized a target-based approach, in which a specific protein target is identified that may hold promise for the development of new therapeutics. However, the brain and how brain disorders influence its function remains a large mystery. This fact argues that the alternative of phenotypic-based screens may offer more promise. Phenotypic screens search for influences on particular cell biological phenotypes without knowledge of the specific molecular targets that influence the phenotype. We have developed a novel screening platform that employs primary cultured neurons and cell biological, phenotypic readouts to identify the influences of small molecules. The methodology is flexible, scalable, and offers numerous advantages over traditional approaches. In this project, we propose to develop assays using this platform for the processes of mitochondrial dynamics, including biogenesis, fission, fusion, protein import, branching, and damage. We will use fluorescent markers for mitochondria to follow mitochondria for such assays, and conduct three screens of small molecules for influences on mitochondrial dynamics once the assays are optimized. Mitochondria dysfunction is associated with multiple psychiatric disorders like mood disorders, schizophrenia and autism; and neurodegenerative disorders as well. Thus, this project promises a rich new resource that can be used to survey small molecules, RNAi, or gene overexpression effects on mitochondrial dynamics in neurons.