A Scalable N x N Electrochemical Detector Array Platform for Analysis of Quantal

描述（由申请人提供）：本项目的目标是开发一种可扩展的n x n电化学检测器阵列平台，该平台具有片上放大器，用于量子发射器释放事件的大规模并行记录。本试验将分析的神经生物学过程是胞吐和递质释放的过程，这是大脑功能及其他功能的关键过程之一。待释放的分子以高浓度储存在膜结合的细胞器中。在刺激时，这些囊泡的内容物通过将囊泡腔连接到细胞外空间的融合孔以量子事件释放。了解囊泡融合和递质释放的机制具有广泛的医学意义。在帕金森氏病的治疗中，药物左旋多巴增加了多巴胺能神经元数量减少时多巴胺的释放。另一方面，BoTox治疗通过减少递质释放发挥其作用。除了这些例子之外，许多其他药物和许多分子操纵以各种方式调节递质释放。递质释放的这种调节不仅通过改变量子释放事件的数量或频率来发生，而且还通过调节量子大小和从单个囊泡释放的动力学来发生。为了理解特定操作影响发射器释放的机制，因此有必要对单个量子释放事件进行精确测量，分析其幅度和时间过程并导出特征参数。常规地，这样的测量通过在显微镜观察下将碳纤维微电极放置在细胞（例如嗜铬细胞、多巴胺能或多巴胺能神经元或PC12细胞）附近、刺激细胞并记录一系列释放事件来进行。该项目开发的技术是从半导体行业改编而来的，涉及开发CMOS微电子芯片，用于记录可氧化的递质分子（如去甲肾上腺素、多巴胺或血清素）的单量子释放事件。该技术将允许同时记录来自数百个细胞的单个囊泡释放事件，而不需要显微镜观察和操作，从而将提供一个高通量平台来表征分子和药理学操作。该技术将加速旨在了解递质释放及其调节的分子机制的研究，以及通过调节递质释放发挥作用的治疗方法的开发和测试。以这种方式，这种新的测定平台将提供测量量子释放事件作为神经生物学终点的机会，并将提供用于靶点鉴定和药物发现的管道。 公共卫生相关性：神经递质和激素的释放受到许多蛋白质、信号分子和各种药物的调节。两个例子是药物左旋多巴，用于治疗帕金森氏病，或美容肉毒杆菌治疗。该项目的目标是开发一个可扩展的n x n电化学检测器阵列平台，该平台具有片上放大器，用于大规模并行记录单个发射器释放事件。这种新的分析平台将提供测量量子释放事件作为神经生物学终点的机会，并将为靶点识别和药物发现提供管道。

DESCRIPTION (provided by applicant): The goal of this project is the development of a scalable n x n electrochemical detector array platform with on- chip amplifiers for massively parallel recordings of quantal transmitter release events. The neurobiological process that this assay will analyze is the process of exocytosis and transmitter release, one of the key processes in brain function and beyond. The molecules to be released are stored at high concentration in membrane-bound organelles. Upon stimulation the contents of these vesicles are released in quantal events through a fusion pore that connects the vesicular lumen to the extracellular space. Understanding the mechanisms of vesicle fusion and transmitter release is of broad medical significance. In the treatment of Parkinson's disease the drug levodopa increases dopamine release from the reduced number of dopaminergic neurons. On the other hand, BoTox treatment exerts its effect through the reduction of transmitter release. In addition to these examples, many other drugs and many molecular manipulations modulate transmitter release in various ways. This regulation of transmitter release occurs not only via changing the number or frequency of quantal release events but also via modulation of quantal size and of the kinetics of release from individual vesicles. To understand the mechanism by which a specific manipulation affects transmitter release it is therefore necessary to perform precise measurements of individual quantal release events, analyze their amplitude and time course and derive characteristic parameters. Conventionally, such measurements are performed by positioning a carbon fiber microelectrode close to a cell (such as a chromaffin cell, a dopaminergic or serotonergic neuron or a PC12 cell) under microscopic observation, stimulate the cell and record a series of release events. The technology developed in this project is adapted from the semiconductor industry and involves the development of a CMOS microelectronic chip for on-chip recordings of single quantal release events of oxidizable transmitter molecules such as noradrenaline, dopamine, or serotonin. The technology will allow the simultaneous recording of single vesicle release events from hundreds of cells without the need for microscopic observation and manipulation, and will thereby provide a high-throughput platform to characterize molecular and pharmacological manipulations. The technology will accelerate the research aimed at understanding the molecular mechanisms of transmitter release and its modulation as well as the development and testing of treatments that act through the modulation of transmitter release. In this way this novel assay platform would provide opportunities to measure quantal release events as neurobiological endpoints and will provide a pipeline for target identification and drug discovery. PUBLIC HEALTH RELEVANCE: Release of neurotransmitters and hormones is regulated by many proteins, signaling molecules and various drugs. Two examples are the drug L-Dopa, used to treat Parkinson's disease, or the cosmetic BoTox treatment. The goal of this project is the development of a scalable n x n electrochemical detector array platform with on-chip amplifiers for massively parallel recordings of individual transmitter release events. This novel assay platform will provide opportunities to measure quantal release events as neurobiological endpoints and will provide a pipeline for target identification and drug discovery.