Developing Electrochemical Sensors to Enable Quantitative Measure of Gliotransmitter Release from Astrocytes

开发电化学传感器以定量测量星形胶质细胞释放的胶质递质

• 批准号: 10752836
• 负责人: Ryan J. White
• 金额: $ 39.23万
• 依托单位: UNIVERSITY OF CINCINNATI
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-06-15 至 2028-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10752836
• 关键词: Affect; Affinity; Age; Architecture; Astrocytes; Binding; Biological; Brain; Brain region; Cell surface; Cells; Chemicals; Chemistry; Collagen; Communication; Corpus striatum structure; DNA; Detection; Development; Diameter; Disease; Electrochemistry; Electrodes; Environment; Exocytosis; Frequencies; Glutamates; Goals; Heterogeneity; Hippocampus; Homeostasis; Hot Spot; Immobilization; Ionomycin; Knowledge; Left; Libraries; Maps; Measurement; Measures; Molecular; Molecular Conformation; Monitor; Neurodegenerative Disorders; Neuroglia; Neurons; Nucleic Acids; Organelles; Physiologic pulse; Physiological; Population; Process; Proteins; Research; Role; Science; Serine; Signal Transduction; Specificity; Structure; Surface; System; Techniques; Technology; Testing; Therapeutic; Time; Tissues; Transcriptional Regulation; aptamer; brain dysfunction; cell type; design; developmental disease; gamma-Aminobutyric Acid; innovation; insight; meter; molecular dynamics; nano; nanomolar; nanoscale; nervous system disorder; neuronal cell body; neurovascular; real time monitoring; response; response to injury; sensor; spatiotemporal; synaptic function; technology development; temporal measurement; three dimensional cell culture; tool; transmission process

PROJECT SUMMARY Objectives. The primary objective of this proposal is to develop chemically specific electrochemical sensors to provide rapid and direct measurements of physiologically relevant chemical messenger release (technology development). The secondary objective is to apply these sensors to determine signaling heterogeneity in astrocytes from different brain regions (biological hypothesis). Significance and Knowledge Gap. Astrocytes act as integrators across many circuits and environments in the brain and how they interact with other cell types can vary in time and space. Rigorous prior research demonstrates that astrocytes are heterogenous, varying by brain region and circuit. Heterogeneity can be affected by transcriptional control related to synapse function, plasticity, molecular transmission and protein machinery and organelles that underly gliotransmission. Astrocyte heterogeneity also affects how this class of cells respond to insults and age and can potentially be predictive of disease vulnerability. Astrocytes have typically been studied as a homogenous population, thus there is a need to study functional heterogeneity of astrocytes (e.g., signaling) to determine functional roles of astrocytes in development, response to injuries, and neurodegenerative disease and how this response influences local circuit function and homeostasis. Solution and Specific Aims. A major barrier to studying gliotransmission is the lack of measurement tools that possess the combined spatiotemporal and chemical specificity to study dynamic molecular signaling from astrocytes. We aim to develop a sensor platform that overcomes this barrier by providing direct and rapid measurement of gliotransmission over broad spatial and temporal ranges. Through a collaborative proposal we propose to leverage the universal and specific chemical detection abilities of electrochemical, aptamer-based (E-AB) sensors with innovative measurement science for rapid determination of gliotransmitter dynamics and heterogeneity. With this new measurement technology, we aim to test the hypothesis that gliotransmitter signaling varies in terms of the frequency, amount, and identity of transmitters released with circuit-, and inter- and intraregional specificity. We will 1) Develop electrochemical, aptamer-based (E-AB) sensors to monitor real-time release of gliotransmitters from a cell population in 3D culture. 2) Develop recessed, microscale E-AB sensors to monitor the release of gliotransmitters from single astrocytes. 3) Develop recessed, nanoscale sensors to monitor the release of gliotransmitters from sub-cellular regions. 4) Select and characterize highly specific structure-switching nucleic acid aptamers as binding partners for glutamate, GABA, and D-serine with nM affinity for use in aims 1-3.

项目概要 目标。该提案的主要目标是开发化学特定的电化学传感器以提供 快速、直接测量生理相关化学信使的释放（技术开发）。这 第二个目标是应用这些传感器来确定来自不同大脑区域的星形胶质细胞的信号异质性 （生物学假设）。 意义和知识差距。星形胶质细胞充当大脑中许多回路和环境的整合者 它们与其他细胞类型相互作用的方式可能因时间和空间而异。先前的严格研究表明星形胶质细胞 异质的，因大脑区域和回路而异。异质性可能受到与突触相关的转录控制的影响 神经胶质细胞传递的功能、可塑性、分子传递以及蛋白质机械和细胞器。星形胶质细胞 异质性还影响此类细胞对损伤和年龄的反应，并有可能预测疾病 脆弱性。星形胶质细胞通常作为同质群体进行研究，因此有必要研究功能性星形胶质细胞 星形胶质细胞的异质性（例如信号传导）以确定星形胶质细胞在发育、对损伤的反应、 和神经退行性疾病以及这种反应如何影响局部回路功能和稳态。 解决方案和具体目标。研究胶质细胞传输的一个主要障碍是缺乏具有这种能力的测量工具 结合时空和化学特异性来研究星形胶质细胞的动态分子信号传导。我们的目标是发展 传感器平台通过提供广泛的神经胶质细胞传输的直接和快速测量来克服这一障碍 空间和时间范围。通过一项合作提案，我们建议利用通用和特定的化学物质 电化学、基于适体 (E-AB) 传感器的检测能力以及创新的测量科学可快速 神经胶质递质动力学和异质性的测定。通过这种新的测量技术，我们的目标是测试 假设胶质细胞发射器信号随释放的发射器的频率、数量和身份而变化 电路、区域间和区域内的特殊性。我们将 1) 开发电化学、基于适配体 (E-AB) 的传感器 监测 3D 培养细胞群中神经胶质递质的实时释放。 2) 开发凹进式微型 E-AB 传感器监测单个星形胶质细胞胶质递质的释放。 3) 开发嵌入式纳米级传感器来监测 从亚细胞区域释放神经胶质递质。 4) 选择并表征高度特异性的结构转换 核酸适体作为谷氨酸、GABA 和 D-丝氨酸的结合伴侣，具有 nM 亲和力，用于目标 1-3。