NEUROPEPTIDES IN THE CNS WITH NEW MASS SPECTROMETRIC SAMPLING PROTOCOLS

描述(申请人提供)：药物滥用研究的一个重要目标是检测和描述暴露于滥用药物后神经系统生物化学的变化，并将这些变化与正常和病理条件下的高级神经元功能联系起来。在众多的信号分子中，细胞间信号肽(ISPs)是一个巨大的测量挑战；在生物系统中表达的多肽确实有数百个，其中许多只有在翻译后修饰时才具有生理活性。这项拟议的研究开发并实施了基于基质辅助激光解吸/电离质谱仪(MALDI-MS)的技术，用于发现和表征ISPs，包括确定它们在哺乳动物神经系统中的化学形态和空间定位。总体战略是开发新的技术和采样方案，以在非常小的生物样本中检测和识别新的互联网服务提供商，包括新准备或培养的单个哺乳动物细胞、小群细胞，以及从神经系统空间受限区域释放的细胞。这些技术有望发现数百个潜在的互联网服务提供商，这些服务提供商将通过研究筛选功能相关性，这些研究旨在：(A)找到具有特定翻译后修饰(例如，酰胺化)的多肽；(B)确定在神经系统中经历长途运输的多肽；(C)发现哪些多肽是以依赖活动的方式释放的。这项技术开发与尖端基础研究奖项征集是完美匹配的。通过开发强大的基于蛋白质组学的新技术来回答有关最复杂和难以捉摸的一组神经调节剂-神经肽的问题，将获得对滥用药物对中枢神经系统影响的基本洞察。

DESCRIPTION (provided by applicant): An important goal of drug abuse research is to detect and characterize changes in the biochemistry of the nervous system on upon exposure to drugs of abuse, and to relate these changes to higher neuronal functions in normal and pathological conditions. Amongst numerous signaling molecules, intercellular signaling peptides (ISPs) are a dramatic measurement challenge; there are literally hundreds of peptides expressed in biological systems, many of which are only physiologically active when posttranslationally modified. The proposed research both develops and implements matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) based technologies for the discovery and characterization of ISPs, including determination of their chemical form and spatial localization in the mammalian nervous system. The overall strategy is to develop new technologies and sampling protocols to detect and identify new ISPs in very small biological samples, including individual freshly prepared or cultured single mammalian cells, small groups of cells, and those released from spatially restricted regions of the nervous system. These technologies promise to uncover hundreds of potential ISPs that will be filtered for functional relevance by studies designed to: (A) find peptides with specific post- translational modifications (e.g., amidation); (B) determine the peptides that undergo long- distance transport in the nervous system; (C) discover which peptides are released in an activity-dependent manner. This technology development ideally matches the cutting-edge basic research award solicitation. By developing powerful new proteomics-based technologies to answer questions about the most complex and elusive set of neuromodulators, the neuropeptides, basic insight into the effects of drugs of abuse on the CNS will be gained.