CORE--ANALYTIC NEUROCHEMISTRY AND SPECTROSCOPY CORE

在过去的二十年里，我们对大脑功能的了解有了巨大的增长，特别是对那些 构成突触传递的基础的事件。数百个实验室辛勤地工作着 描述神经递质和突触的代谢、药理、运输和发育 仪器。这项事业的幸运结果是对信息错综复杂的详细理解 神经系统中的交换。我们可以确定神经传递快和慢的分子基础。我们 认识到神经胶质细胞在维持神经纤维的离子内稳态、终止 神经传递和向神经元提供补充神经递质池的前体。 我们已经开始意识到，突触--就像大脑本身一样--不是一个成品，而是一个 一种永续进化的状态，这种状态的发展是按照精心制定的时间表进行的 促进新信息的获取和适应不断变化的外部环境。 对突触生物化学的科学兴趣既反映了该学科的基本重要性，也反映了 认识到神经递质失衡是许多人类疾病的核心，其中包括缺氧性脑疾病 损伤、先天代谢障碍、癫痫、药物滥用、抑郁等。智力低下及其他 发育障碍通常是由突触生化和/或突触发生的紊乱引起的。 对这些疾病的有效治疗前提是了解相关的病理生物学。 我们构建了分析神经化学和光谱核心，为我们的用户提供了一系列 适应突触全面检查的服务--来自个体的体外分析 代谢物到离子通量的原位表征到体内神经递质的测量和 神经递质受体和转运体。我们已经能够提供尖端技术- 质谱学、正电子发射断层扫描、光学成像-这些都超出了分析能力 个别实验室。此外，我们通过让每个用户熟悉我们的分析来支持他们 并通过帮助每个用户解释实验结果来实现。 在MRDDRC最初提交时，该核心赞助了三项服务：高效液相色谱(HPLC) 神经递质及其代谢物的分析、GC-MS技术和神经细胞培养设备。在……里面 1995年，当前的细胞神经科学核心(核心A)被“梳理”为一个单独的核心，以改善我们的 细胞和病理服务的曲目。现代分析神经化学与光谱学核心 (核心D)然后采用其目前的配置。我们仍然提供用于氨基酸定量的hplc资源。 酸和生物胺具有很高的灵敏度，我们提供质谱分析服务。 用于体内和体外动力学研究的稳定同位素浓缩。在2000年，我们增加了一种基于细胞的分析 一种测量由细胞死亡和/或凋亡激活的分子的设备，包括反应性的钙离子 氧物种、钙激活的蛋白水解酶和半胱氨酸天冬氨酸蛋白酶-3。我们还测量了DNA片段化(一种较晚的标记 细胞凋亡)、乳酸脱氢酶(细胞死亡的细胞质标记物)和福尔马赞3-(4，5-二甲基噻唑- (2-基)-2，5-二苯基四氮唑(MTT)，活细胞的线粒体标记物。我们加了一个正电子 用于阐明活体分子和细胞突触过程的发射断层扫描(PET)设备 有机体。一项新的倡议是目前拟议的蛋白质组学服务，该服务将使蛋白质识别在 复杂混合物以及翻译后修饰位点的表征。 这一核心的意义体现在许多出版物和赠款申请中，即核心服务 已成功启用。由于丰富的分析基础设施，几项新的科学倡议是可行的 核心所能承受的。一个例子是罕见疾病研究联盟，一个研究尿素的企业 这是其他几个MRDDRCs1之间协作的一个显著例子。我们 随着我们为核心增加新的服务，包括蛋白质组学设施，预计这一趋势将继续下去。 该核心包含几个不同的分析组件：高效液相色谱、质谱学、细胞分析、正电子发射计算机断层扫描和 蛋白质组学。我们承认这个曲目范围很广，但我们希望通过创建一个 促进生化研究的单一单元，从单个分析物(高效液相色谱，质谱仪)延伸而来 从光谱学和蛋白质组学)到完整细胞(基于细胞的分析)到整个生物体(PET)。这不仅是因为 安排使组织统一，也允许增加经济，这是我们必须权衡的一个考虑因素 在解除武装、复员和重返社会方案的总体预算受到严格限制的时候，必须谨慎行事。创建更多的核心将 使这种节约化变得不可能。相反，当前的结构使我们能够利用制度性 支持，这在为我们的用户提供实际成本中所占的比例要大得多，因为这些状态- 艺术服务。

The past two decades have seen enormous growth in our understanding of brain function, especially of those events that form the basis of synaptic transmission. Hundreds of laboratories have labored diligently to characterize the metabolism, pharmacology, transport and development of the neurotransmitters and the synaptic apparatus. The fortunate outcome of this enterprise is a detailed understanding of the intricacies of information exchange in the nervous system. We can identify the molecular basis for fast and slow neural transmission. We appreciate the importance of glial cells in maintaining ionic homesostasis in the neuropil, in terminating neurotransmission and in furnishing to neurons the precursors for replenishment of the neurotransmitter pool. We have begun to appreciate that the synapse - like the brain itself- is not a finished product, but a structure in a state of perpetual evolution, the development of which proceeds according to a carefully wrought schedule that facilitates the acquisition of new information and the adaptation to an ever changing external environment. Scientific interest in synaptic biochemistry reflects both the fundamental importance of the subject and the recognition that neurotransmitter imbalance is at the heart of many human diseases, among them hypoxic brain injury, inborn errors of metabolism, epilepsy, substance abuse, depression, etc. Mental retardation and other developmental disabilities commonly result from derangements of synaptic biochemistry and/or synaptogenesis. Effective treatments for these disorders presuppose an understanding of the relevant pathobiology. We structured the Analytical Neurochemistry and Spectroscopy Core to provide our users with an array of services that accommodates comprehensive scrutiny of the synapse - from the in vitro analysis of individual metabolites to in situ characterization of ionic flux to in vivo measurement of neurotransmitters and neurotransmitter receptors and transporters. We have been able to make available sophisticated technologies - mass spectrometry, positron emitting tomography, optical imaging - that exceed the analytic capability of an individual laboratory. Furthermore, we support our users by familiarizing each of them with our analytical potential and by helping each user to interpret experimental results. At the initial submission of the MRDDRC, this core sponsored three services: liquid chromatographic (HPLC) analysis of neurotransmitters and their metabolites, GC-MS technology, and a neuronal cell culture facility. In 1995 the current Cellular Neuroscience Core (Core A) was "teased" out as a separate core to improve our repertoire of cellular and pathology services. The current Analytical Neurochemistry and Spectroscopy Core (Core D) then assumed its present configuration. We still provide HPLC resources for the quantitation of amino acids and biogenic amines with high sensitivity and we furnish mass spectrometry services for the determination of stable isotope enrichment for kinetic studies, both in vivo and in vitro. In 2000 we added a Cell Based Assay Facility that measures molecules that are activated by cell death and/or apoptosis, including Ca2+, reactive oxygen species, Ca2+-activated proteases, and caspase-3. We also measure DNA fragmentation (a late marker of apoptosis), lactate dehydrogenase (a cytoplasmic marker of cell death) and the formazan 3-(4,5-dimethylthiazol- 2-yl)-2,5-diphenyltetrazoliumbromide (MTT), a mitochondrial marker for living cells. We added a Positron Emission Tomography (PET) Facility to elucidate molecular and cellular synaptic processes in the living organism. A novel initiative is the current proposed Proteomics service that will enable protein identification in complex mixtures as well as the characterization of sites of post-translational modification. The significance of this Core is reflected in the many publications and grant applications that the core services successfully enabled. Several new scientific initiatives were feasible because of the rich analytic infrastructure that the Core affords. One example is the Rare Disease Research Consortium, a venture that studies the urea cycle defects and that represents a notable example of collaboration among several other MRDDRCs1. We anticipate this trend to continue as we add novel services to the Core, including the Proteomics Facility. This Core contains several different analytic components: HPLC, mass spectrometry, cell-based assays, PET and proteomics. We acknowledge that this repertoire is broad, but we wanted to accommodate our users by creating a single unit that facilitates biochemical study along a continuum extending from individual analytes (HPLC, mass spectrometry and proteomics) to intact cells (cell-based assays) to the whole organism (PET). Not only does this arrangement confer organizational unity, it also allows increased economy, a consideration that we must weigh carefully at a time of strict limits on the overall budget of the MRDDRC. The creation of additional cores would have made impossible such economization. Instead, the current structure allows us to leverage institutional support, which comprises by far the greater proportion of the actual costs of providing our users,with these stateof- the-art services.