MT Function and Dysfunction in Single Neurons in Vivo

描述（由申请人提供）：该申请涉及广泛的挑战领域（06）：使能技术，以及特定的挑战主题，01-OD-101：开发新的工具和技术来询问体内的人类线粒体功能。线粒体功能障碍被认为是衰老和神经退行性疾病的主要原因。虽然MT功能可以通过氧化磷酸化和ATP产生的特性来测量，但这些定义的测量并没有直接导致对病理生理学的明显理解。阿尔茨海默氏症（AD）帕金森氏症（PD）亨廷顿氏症的共同特点是MT不能满足细胞的能量需求。然而，AD、PD和HD显示特定区域的细胞死亡，即使在这些区域中，也只有选定的细胞类型被靶向。因此，区域特异性意味着MT功能障碍的发生是对特定细胞随年龄或疾病进展而发生的细胞代谢变化的反应。由于系统的异质性，存在实质性的障碍，使得很难确定主要缺陷是特定MT无法将代谢前体转化为ATP，还是细胞途径无法产生足够的代谢前体来合成ATP。为了克服这些限制，我们开发了一种全新的超高灵敏度质谱技术，称为纳米结构引发质谱（NIMS），它能够在150nm分辨率下进行质谱分析。因此，NIMS具有前所未有的在空间上跟踪单个细胞或单个MT中的质量组成的能力。NIMS旨在创建分子特征，即使在相邻的细胞类型之间也能独特区分，并且不会破坏大脑的三维结构。我们已经将NIMS应用于全脑切片内任何区域的单个细胞的代谢物组成成像和定位，并直接测量单个细胞的mt特异性变化。碎片化、灵敏度和低空间分辨率等问题限制了其他质谱技术在实现NIMS分辨率和成像能力方面的应用。然而，我们的方法提供了，也许是唯一现有的技术进步，有效和精确地解决，区域，细胞类型和mt特异性异质性。NIMS可以应用于任何疾病，任何组织切片，它的想法是整理大脑中发生的复杂变化。

DESCRIPTION (provided by applicant): This application addresses broad Challenge Area (06): Enabling Technologies, and specific Challenge Topic, 01-OD-101: Development of new tools and technologies to interrogate human mitochondrial function in vivo. Dysfunction of Mitochondria (MT) dysfunction is thought to be a primary contributor to aging and neurodegenerative disease. Although MT function is measurable by well-established properties of oxidative phosphorylation and ATP production, these defined measures have not directly led to an obvious understanding of pathophysiology. Alzheimer's (AD) Parkinson's (PD) Huntington's disease shares the property that MT is not keeping up with the energy demands of the cell. However, AD, PD, and HD display region-specific cell death, and even within those regions, only select cell types are targeted. Regional specificity, therefore, implies that MT dysfunction develops in response to the changing cellular metabolism in only specific cells with age or as disease progresses. Due to system heterogeneity, there have been substantial barriers that have made it difficult to determine whether the primary defect is the inability of particular MT to convert metabolic precursors to ATP, or a failure of cellular pathways to generate sufficient metabolic precursors for ATP synthesis. To overcome these limitations, we have developed a completely new, ultra-high sensitivity mass spectrometry technique called Nanostructure-Initiator Mass Spectrometry (NIMS), which is capable of mass analysis at 150nm resolution. Thus, NIMS has the unprecedented ability to spatially track mass composition in a single cell or in a single MT. NIMS is designed to create molecule signature that uniquely distinguish among even adjacent cell types, and does not destroy the 3-dimenational architecture of the brain. We have adapted NIMS to image and localize individual cells by their metabolite composition in any region within whole brain slices, and to directly measure MT-specific variations in single cells in situ. Issues with fragmentation, sensitivity, and low spatial resolution have limited the use of other mass spectroscopy techniques to achieve the resolution and imaging capabilities NIMS. However, our approach provides, perhaps the only existing technological advance that efficiently and precisely resolve, region, cell type, and MT-specific heterogeneity. NIMS can be applied to any disease, to any tissue section, and is idea to sort out complex changes that occur in the brain. PUBLIC HEALTH RELEVANCE: At least 2% of Americans will be afflicted with some form of Alzheimer's disease (AD) (4,000,000), Parkinson's disease (PD) (1,500,000), or Huntington's disease (HD) (200,000), among others. Each of these disorders can affect patients for decades, yet no effective long-term approaches to therapy are currently available. Because the number of affected individuals will grow exponentially, the gap between the problem's size and our capabilities for treatment will widen. Thus, an understanding of the pivotal role of MT dysfunction promises to increase therapeutic options.