In vivo chemical monitoring using capillary separations

使用毛细管分离进行体内化学监测

• 批准号: 7759356
• 负责人: ROBERT T KENNEDY
• 金额: $ 5.53万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-05-05 至 2013-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7759356
• 关键词: Acute; Amino Acids; Amphetamines; Animal Welfare; Arts; Behavior; Behavioral; Bibliography; Biological Assay; Blood capillaries; Brain; Brain Chemistry; Brain region; Cells; Chemicals; Circadian Rhythms; Communities; Complex; Country; Coupled; Coupling; Cues; Detection; Development; Dexamethasone; Disease; Dopamine; Drug Addiction; Electrophoresis; Endorphins; Enkephalins; Environment; Environmental Impact; Equipment; FK506; Feeding behaviors; Generations; Gliosis; Goals; Hour; Human; IACUC; Immunoassay; Immunosuppressive Agents; Injection of therapeutic agent; International; Intervention; Learning; Leptin; Mass Fragmentography; Measurement; Methods; Microdialysis; Microfluidics; Modeling; Monitor; Names; Nerve Tissue; Neurodegenerative Disorders; Neuroglia; Neurons; Neuropeptides; Neurosciences; Neurosciences Research; Neurotensin; Neurotransmitters; Nicotine; Oils; Opioid Peptide; Peptides; Phenotype; Physiologic pulse; Positron-Emission Tomography; Principal Investigator; Problem Solving; Rattus; Reaction; Recovery; Research; Research Ethics Committees; Resolution; Resources; Rewards; Role; Sampling; Scheme; Signal Transduction; Stream; System; Systems Analysis; Technology; Testing; Time; Tissues; Vertebrates; Work; abstracting; addiction; aqueous; base; capillary; capillary liquid chromatography; drug of abuse; expiration; human subject; improved; in vivo; interest; leptin receptor; miniaturize; neurochemistry; neurotransmitter release; neurotrophic factor; novel; prevent; programs; psychostimulant; receptor; relating to nervous system; research study; tool

Our overall objective is to develop new bioanalytical methods for exploring brain chemistry dynamics in vivo. Neurons and glia in the brain communicate by releasing neurotransmitters that interact with receptors on neighboring cells. Monitoring the concentration dynamics of neurochemicals and metabolites in vivo is a vital tool in the effort to understand brain function, diseases, and treatments. A versatile and effective approach for in vivo monitoring of chemical messages is to couple sampling methods, such as microdialysis, to analytical measurements. Although this approach has proven invaluable, its utility is limited by poor temporal resolution, poor spatial resolution, poor results for neuropeptide monitoring, and application to only acute measurements. In this project, we will develop technology and methods to solve these problems. Temporal resolution is important because concentrations of transmitters can change rapidly during behavior and experimental maneuvers. Temporal resolution is often limited by dispersion of concentration pulses as they are transported to the analytical system. We will develop a microfluidic sampling system whereby the aqueous sample stream is segmented into droplets within a stream of oil and the droplets subsequently analyzed by rapid chip-based electrophoresis assays. Sample stream segmentation will prevent dispersion during mass transport and allow temporal resolution of 10 s or better for many neurotransmitters. This system will be coupled to miniaturized sampling probes to improve spatial resolution and allow access to small brain regions. Neuropeptides regulate many brain functions; however, monitoring them in vivo is limited by the sensitivity of current methods so that samples must be collected for ~30 min resulting in poor temporal resolution. We will develop high sensitivity neuropeptide assays based on capillary liquid chromatography and microfluidic immunoassays. The assays will have detection limits of 1 pM for 1 µL samples allowing an unprecedented 10-fold improvement in temporal resolution for neuropeptide monitoring. In vivo chemical measurements are nearly always performed acutely; however, it would be extremely useful to be able to monitor neurochemistry over a period of weeks to monitor progressive changes associated with diseases, like addiction, or normal function, like learning. Long term monitoring is typically prevented by reactive gliosis, a tissue reaction that results in encapsulation of the probe and prevents sampling from active neural tissue. We will explore the use of pharmacological interventions with compounds known to suppress reactive gliosis and support neuroregeneration to prolong in vivo monitoring. Finally, we will perform fundamental neuroscience studies as a means of testing the methods and demonstrating their utility to the broader neuroscience community. These applications include determining: 1) the role of leptin receptors in regulating dopamine and feeding behavior; 2) the effect of psychostimulants on opioid peptides, and 3) neurochemical differences underlying distinct behavioral phenotypes that are a model for vulnerability to drug addiction.

我们的总体目标是开发新的生物分析方法，探索脑化学动力学， vivo.大脑中的神经元和神经胶质细胞通过释放神经递质与受体相互作用来进行交流。 相邻的细胞。监测体内神经化学物质和代谢物的浓度动态至关重要 这是一种了解大脑功能、疾病和治疗方法的工具。一种通用而有效的方法， 化学信息的体内监测是将取样方法（例如微透析）与分析方法（例如微透析）结合起来， 测量.虽然这种方法已被证明是非常宝贵的，但其实用性受到时间分辨率差的限制， 空间分辨率差，神经肽监测结果差，仅适用于急性测量。 在本项目中，我们将开发解决这些问题的技术和方法。 时间分辨率是重要的，因为发射器的浓度可以在发射期间迅速变化。 行为和实验演习。时间分辨率通常受到浓度分散的限制 脉冲，因为它们被传输到分析系统。我们将开发一个微流控采样系统 由此将含水样品流分割成油流内的液滴，并且液滴 随后通过基于芯片的快速电泳测定进行分析。样本流分割将防止 分散在质量运输和允许10秒或更好的时间分辨率为许多神经递质。 该系统将与小型化取样探头连接，以提高空间分辨率， 小的大脑区域。神经肽调节许多脑功能;然而，在体内监测它们是有限的 由于当前方法的灵敏度，因此必须收集样本约30分钟，导致时间较差 分辨率我们将开发基于毛细管液相色谱法的高灵敏度神经肽检测方法， 微流控免疫测定。对于1 µL样品，检测限为1 pM， 神经肽监测的时间分辨率前所未有地提高了10倍。体内化学品 测量几乎总是敏锐地进行;然而，能够 在几周内监测神经化学，以监测与疾病相关的进展性变化，如 成瘾，或正常功能，如学习。长期监测通常会被反应性神经胶质增生所阻止， 组织反应，导致探针封装并阻止从活性神经组织取样。我们 将探索使用已知抑制反应性神经胶质增生的化合物进行药物干预， 支持神经再生以延长体内监测。最后，我们将进行基础神经科学 研究作为测试方法的手段，并证明它们对更广泛的神经科学的实用性 社区这些应用包括确定：1）瘦素受体在调节多巴胺中的作用， 摄食行为; 2）精神兴奋剂对阿片肽的影响; 3）神经化学差异 潜在的不同的行为表型是一个模型的脆弱性药物成瘾。