Microfluidic System for Monitoring Gliotransmitter Release

用于监测神经胶质递质释放的微流体系统

• 批准号: 9788379
• 负责人: Richard Bertram
• 金额: $ 16.24万
• 依托单位: FLORIDA STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-09-30 至 2021-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9788379
• 关键词: Acetylcholine; Acute; Adenosine; Adenosine Triphosphate; Affect; Amines; Astrocytes; Back; Behavior; Biological Assay; Biology; Brain; Brain Diseases; Cells; Chronic; Coculture Techniques; Data; Detection; Development; Devices; Dopamine; Drug Addiction; Drug Exposure; Fluorescence; Fluorescent Probes; Frequencies; Functional disorder; Future; Glutamates; Goals; Health; Human; Image; Ion Channel; Knowledge; Lasers; Lead; Luciferases; Measurement; Measures; Methods; Microfluidic Microchips; Microfluidics; Mission; Monitor; Naphthalene; Neuraxis; Neuroglia; Neurons; Neurosciences; Neurotransmitters; Norepinephrine; Perfusion; Pharmaceutical Preparations; Physiological; Play; Public Health; Reaction; Research; Research Personnel; Resolution; Rewards; Role; Sampling; Serine; Serotonin; Stimulus; Structure; Synapses; Synaptic Transmission; System; Techniques; Time; United States National Institutes of Health; assay development; cell type; dopaminergic neuron; drug of abuse; innovation; luciferin; neurotransmitter release; novel; postsynaptic neurons; presynaptic neurons; release factor; response; small molecule; temporal measurement; tool

The precise mechanisms through which astrocytes modulate synaptic transmission are not clear. It is also not clear how dysfunction in these cells, including those caused by drugs of abuse, can lead to brain disorders. The objective of this proposal is to develop a method that will allow the culture of astrocytes within a microfluidic platform to enable real-time measurements of intracellular Ca2+ concentration ([Ca2+]i) via fluorescent probes simultaneously with secretion of neuroactive substances in response to various stimuli. Guided by our strong preliminary data, we will obtain the objective of this proposal by pursuing the following specific aims: 1) monitoring D-serine and glutamate secretion simultaneous with [Ca2+]i imaging within astrocyte cultures, and 2) quantitation of ATP release from astrocyte cultures. In the first aim, we will develop a microfluidic system to monitor, with high time resolution, the release of primary amines from astrocytes simultaneous with intracellular [Ca2+]i imaging. In the second aim, we will integrate an enzymatic assay for adenosine triphosphate to allow monitoring of all the major gliotransmitters from astrocytes. The research proposed in this application is innovative because it will utilize a broad approach to the measurement of multiple factors released from astrocytes combined with simultaneous [Ca2+]i imaging, providing a highly dynamic view of gliotransmission. The results of this system will be significant because it will provide the first tool capable of investigating release of all major gliotransmitters simultaneously in an automated fashion in response to a number of stimulants. Ultimately, this device will produce a novel tool for future studies regarding astrocyte biology and the roles these cells have in neuroprotective behavior, tripartite synaptic transmission, and the effects of drugs of abuse.

星形胶质细胞调节突触传递的确切机制尚不清楚。也不是 明确这些细胞的功能障碍（包括由滥用药物引起的细胞功能障碍）如何导致大脑疾病。 该提案的目的是开发一种方法，允许在一个特定的环境中培养星形胶质细胞。 微流控平台可通过以下方式实时测量细胞内 Ca2+ 浓度 ([Ca2+]i) 荧光探针与响应各种刺激的神经活性物质的分泌同时发生。 在我们强有力的初步数据的指导下，我们将通过追求以下目标来实现本提案的目标 具体目标：1) 通过 [Ca2+]i 成像同时监测 D-丝氨酸和谷氨酸分泌 星形胶质细胞培养物，以及 2) 星形胶质细胞培养物中 ATP 释放的定量。在第一个目标中，我们将开发 微流体系统以高时间分辨率监测星形胶质细胞释放伯胺 与细胞内 [Ca2+]i 成像同时进行。在第二个目标中，我们将整合酶测定 三磷酸腺苷可监测星形胶质细胞的所有主要神经胶质递质。研究 本申请中提出的方案具有创新性，因为它将利用广泛的方法来测量 星形胶质细胞释放的多种因子与同步 [Ca2+]i 成像相结合，提供了高度 胶质细胞传输的动态视图。该系统的结果将是重大的，因为它将提供第一个 能够以自动方式同时调查所有主要胶质递质释放的工具 对多种兴奋剂的反应。最终，该设备将为未来的研究提供一种新颖的工具 星形胶质细胞生物学以及这些细胞在神经保护行为、三方突触传递、 以及滥用药物的影响。