NEUROPEPTIDES IN THE CNS WITH NEW MASS SPECTROMETRIC SAMPLING PROTOCOLS

描述（由申请人提供）：药物滥用研究的一个重要目标是检测和描述暴露于药物滥用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；在生物系统中有数百种肽表达，其中许多只有在翻译后修饰时才具有生理活性。提出的研究开发并实现了基于基质辅助激光解吸/电离质谱（MALDI-MS）的技术，用于发现和表征isp，包括确定其化学形态和哺乳动物神经系统中的空间定位。总体战略是开发新技术和采样方案，以在非常小的生物样品中检测和识别新的isp，包括单个新鲜制备或培养的单个哺乳动物细胞、小群细胞和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isp，这些isp将通过研究筛选功能相关性：(A)发现具有特定翻译后修饰（例如，酰胺）的肽；(B)测定在神经系统中进行长距离转运的肽；(C)发现哪些肽以活性依赖的方式释放。这项技术的发展理想地匹配了尖端基础研究奖的征集。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最难以捉摸的神经调节剂，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见解。

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.