Parallel Solid-State Electrodes for Turn-Key Intracellular Electrophysiology

描述(申请人提供)：大脑中相互连接的细胞网络称为神经元，构成了所有认知功能的基础。在过去的几十年里，我们对大脑功能的理解取得了关键进展，这是允许监测神经元电活动的技术的结果。这项技术，广泛地被称为电生理学，允许研究负责感觉、运动、思维、学习和记忆的连接神经元的电路。这些技术还揭示了神经元之间的异常电信号如何导致功能障碍，如自闭症或阿尔茨海默病以及中风或创伤性损伤造成的损伤。最灵敏的电生理记录形式监测单个细胞中非常微小的电流，在神经元内部放置玻璃吸管。这些“细胞内”膜片钳记录是探索神经元如何工作--以及不工作--的强大工具。尽管有这些巨大的进步，目前的细胞内记录技术仍然有很大的局限性：穿刺细胞会破坏细胞，导致短时间的记录和异常的生物物理和生化特性；需要熟练的科学家，减少可以使用这种技术的实验室数量；同时从一个或两个以上的神经元进行记录的可能性很小，这给研究神经元通信带来了挑战。隐形生物科学的建立是为了克服现有细胞内技术的根本局限性。我们团队发明了一种新技术，我们称之为“隐形”或仿生电极。这些电极能够融合到细胞膜上，提供与细胞之间最小的损害和电气上的紧密连接。我们的初步测量表明，高质量的长期细胞内记录可以与传统的膜片钳相媲美。仿生探针基于标准的硅微加工工艺，能够在廉价的芯片上实现大阵列的电极。利用第一阶段SBIR赠款的支持，我们将改进设备，并对这种改变游戏规则、廉价且易于使用的细胞内记录平台进行可行性研究。我们的长期目标是开发这项技术，在研究人员中进行广泛的商业分发，以推进基本发现，加快药物开发，并改善那些患有脑部疾病的人的健康和福祉。为了实现这一雄心勃勃的计划，我们概述了第一阶段的两个具体目标：目标1：优化仿生电极的性能和生产。在这个目标中，我们将评估不同的仿生电极几何和结构设计的性能。将评估设计的电学特性以及培养神经元的电生理性能。制造过程将得到简化和结构化，目标是最终实现大规模制造。具体的里程碑目标包括&lt；2 mV噪声的电气性能、优于0.1ms的时间分辨率和&lt；200 mW的输入阻抗。时间：第二季度和第三季度。目的2：利用培养的细胞进行仿生探针的功能表征第二个目的是表征用于记录大鼠海马神经元的仿生装置的性能。这种对细胞内记录能力的严格测试将允许直接与黄金标准的基于吸管的膜片钳进行比较。AIM 1中不同探头设计的细胞记录将用于优化制造技术和探头设计。长达数天甚至数周的长期记录将被用来展示探针的寿命和时间能力，远远超过传统膜片钳的能力。时间：第三季度和第四季度。我们聚集了一支在电生理学、微/纳米制造、细胞间通信和业务方面拥有专业知识的强大团队，以支持隐形生物科学公司的这项技术开发。在这个节目的最后，我们将有一个经过实验审查的系统，用于“交钥匙”的细胞内记录。这些将提供简单的细胞准备，每个芯片16个单独可寻址的电极，高质量的记录，并与现有的电生理软件和记录硬件兼容。我们相信，这些设备将在神经科学界引起广泛的兴趣，无论是作为基础研究工具，还是在药物发现和个性化医学中的高级应用。

DESCRIPTION (provided by applicant): Interconnected networks of cells in the brain called neurons underlie all cognitive functions. Key advances in our understanding of brain function during the last several decades have resulted from technologies that permit monitoring of electrical activity in neurons. This technique, broadly referred to as electrophysiology, permits the study of circuits of connected neurons responsible for sensation, movement, thought, learning, and memory. These techniques have also revealed how abnormal electrical signaling between neurons can lead to dysfunction as occurs in disorders such as autism or Alzheimer's disease as well as due to damage from a stroke or traumatic injury. The most sensitive form of electrophysiological recording monitors very small electrical currents in single cells with glass pipettes placed inside the neuron. These 'intracellular' patch-clamp recordings are a powerful tool for exploring how neurons work-and don't work. Despite these great advances, current intracellular recording technology has significant limitations: puncturing the cell damages it, leading to short recordings and abnormal biophysical and biochemical properties; skilled scientists are required, reducing the number of labs that can use this technique; and simultaneous recording from more than one or two neurons is rarely possible, making it challenging to study neuronal communication. Stealth Biosciences was established to overcome fundamental limitations of existing intracellular techniques. Our group invented a new technology which we call "Stealth" or biomimetic electrodes. These electrodes are able to fuse into the cellular membrane, providing a minimally damaging, electrically tight junction with the cell. Our initial measurements have demonstrated high-quality, long-term intracellular recordings that rival that of traditional patch-clamps. Biomimetic probes are based on standard silicon microfabrication processing, enabling large arrays of electrodes on inexpensive chips. Using support from a Phase I SBIR grant, we will refine the device and perform feasibility studies of this game changing, inexpensive, and easy-to-use intracellular recording platform. Our long-term goals are to develop this technology for wide commercial distribution among researchers to advance basic discoveries, accelerate drug development, and improve the health and well-being of those suffering from disorders of the brain. To achieve this ambitious program we outline two Phase I Specific Aims: Aim 1: Optimize Biomimetic Electrode Performance and Production In this Aim, we will assess the performance of different geometric and architectural designs for biomimetic electrodes. Designs will be evaluated for electronic characteristics as well as for electrophysiological performance with cultured neurons. The fabrication process will be streamlined and structured with the goal of eventual large-scale fabrication. Specific milestone goals include electrical performance of < 2mV noise, better than 0.1 ms time resolution, and <200MW input impedance. Timing: Q2 and Q3. Aim 2: Functional Characterization of Biomimetic Probes with Cells in Culture The second Aim will characterize the biomimetic device performance for recording from rat hippocampal neurons. This stringent test of intracellular recording capabilities will allow direct comparison to the gold- standard pipette-based patch-clamps. Cell recordings from the different probe designs in Aim 1 will be used to optimize fabrication techniques and probe design. Long-term recordings extending for days and possibly weeks will be used to demonstrate lifetime and temporal capabilities of the probes far exceeding what is possible with conventional patch-clamps. Timing: Q3 and Q4. We have brought together a strong team with expertise in electrophysiology, micro/nanofabrication, cell-to-cell communication, and business to support this technology development at Stealth Biosciences. At the end of this program, we will have an experimentally vetted system for 'turnkey' intracellular recordings. These will provide simple cell-preparation, >16 individually addressable electrodes per chip, high-quality recordings, and compatibility with existing electrophysiological software and recording hardware. We believe these devices will find broad interest within the neuroscience community, both as a basic research tool, and for advanced applications in drug discovery and personalized medicine.