A Digital Feedback Clamp Instrument for Neurophysiology

用于神经生理学的数字反馈钳仪器

• 批准号: 8150427
• 负责人: Andrew A Hill
• 金额: $ 53.71万
• 依托单位: ROKHAN, LLC
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-09-30 至 2013-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8150427
• 关键词: Algorithms; Amplifiers; Animals; Automobile Driving; Back; Basic Science; Binding; Brain Stem; Cataloging; Catalogs; Cell membrane; Cells; Chills; Computer software; Computers and Advanced Instrumentation; Development; Devices; Digital Signal Processing; Electrodes; Equipment; Evaluation; Event; Exhibits; Feedback; Funding; Head; Health; Ion Channel; Ion Channel Protein; Ions; Lead; Legal patent; Libraries; Marketing; Mass Screening; Measurement; Measures; Membrane; Metabolism; Methods; Microelectrodes; Mus; Names; Neurons; Neurosciences; Noise; Outcome; Patch-Clamp Techniques; Pharmacologic Substance; Phase; Population Heterogeneity; Production; Property; Proteins; Publications; Research; Rhode Island; Role; Scheme; Signal Transduction; Small Business Innovation Research Grant; Societies; Speed; Staging; Suction; Surface; System; Techniques; Technology; Testing; The Sun; Therapeutic; Time; Tissues; Universities; Validation; Variant; Work; analog; base; conditioning; cost; design; digital; electric impedance; feeding; human disease; improved; instrument; interest; meetings; neurophysiology; novel; patch clamp; prototype; public health relevance; research study; response; seal; submicron; tool; voltage; voltage clamp; voltage gated channel; web site

DESCRIPTION (provided by applicant): The Patch Clamp technique has evolved in recent years from an esoteric method of basic electrophysiological research into properties of cell membranes, into a practical technique for mass screening compounds for potentially therapeutic pharmaceutical properties. The commercial market for such equipment is now in the region of $40M annually. Its role as a tool of basic research has also continued to expand, as a means of discovering types and functions of ion channels through cell membranes, and characterizing their response. Phase I SBIR funding has enabled us to develop a digital clamp amplifier that functions in whole cell clamping and produces waveforms identical to those from comparable analog instruments. We have demonstrated some novel and potentially useful applications, not available with analog devices, such as providing instant "feedback" to a second neuron (dynamic clamp), thus driving one neuron off another's firing rate, and voltage clamping using only a single electrode without an external switch, multiplexing the stimulation and recording stages. This project will lead to a new patch clamp amplifier based on a high-speed digital signal processor to interact with excitable tissues. The instrument integrates the traditional voltage clamp, current clamp, dynamic clamp, and patch clamp into one unit, allowing for transient-free switching among the clamping modes and interactive controls of the neurons. The focus for Phase II includes hardware improvements for patch clamping, software techniques for controls, and experimental validations with mammalian neurons. Patch clamping developed from whole cell voltage clamping, in which measured current is fed back through one of two electrodes in the cell in an amount to keep the voltage across the cell membrane constant, as measured on the second electrode. Ions cross the cell membrane through proteins on the surface which escort them through. These ion channels are frequently voltage gated, open to passing only at certain voltage ranges, or may be chemically gated, thermally gated, or other. Even different voltage gated channels for the same ion may open at different voltages. For patch clamping, the electrode tip is very small, commonly sub-micron, in contact with a part of the cell membrane surface, and a small suction is applied to seal the membrane to the pipette tip. This allows measurement and feedback over only a small bit of membrane, and recording of even single ion channel events. There is a continuing and growing need to catalog all the ion channels across cell membranes, characterize their gate controls, and their function in cell metabolism, and effects on whole animal function and health. A common application is to screen compounds for binding to ion channel proteins to open or close them. Ion channels have become a major target for development of new pharmaceuticals. Variations include voltage clamping, current clamping and conductance clamping. In present application and technology, these 3 techniques each require different analog instruments for the fast responsive feedback needed, and the different parameter measured. Single channel events occur at change rates of up 250K/sec. The high speeds of ion channel events have required analog circuitry. In previous work and publications, it has been shown that a digital throughput of at least 500 KHz would be required to allow a digital instrument to perform as well as the analog instruments in voltage clamping. Very recently, digital signal processing chips (DSP's) operating at as fast as 600 MHz have become commercially available. Digital clamping would save the added cost of A/D and signal conditioning units, and one instrument would be able to perform any of the 4 types of clamping. Other emergent properties allow new avenues of research with digital clamping amplifiers. This proposal is to develop low-noise bi-directional headstages and accessories needed to use this digital clamp amplifier in patch clamping applications, including a chilled headstage to minimize noise, and to develop a library of algorithms for adjustable feedback schemes for different applications, and test the suitability of the instrument for patch clamp experiments. The resulting product is expected to achieve a substantial share of a large market. PUBLIC HEALTH RELEVANCE: Patch clamp technique has emerged as a practical applied technique to test compounds for functional effects on ion channel proteins. Defective ion channels are implicated in several human diseases. The proposed instrument will replace 3 types of analog clamping instruments, and digitizers and signal conditioners with a single, versatile, more powerful digital clamping device capable of serving as a voltage, current or impedance clamp.

描述（由申请人提供）：近年来，膜片钳技术已经从基础电生理学研究细胞膜特性的深奥方法发展成为用于大规模筛选化合物的潜在治疗药物特性的实用技术。这种设备的商业市场现在每年约为4000万美元。它作为基础研究工具的作用也在继续扩大，作为发现细胞膜离子通道的类型和功能以及表征其反应的手段。第一阶段SBIR资金使我们能够开发一个数字钳位放大器，在整个细胞钳位功能，并产生波形相同的那些从可比的模拟仪器。我们已经展示了一些新颖的和潜在有用的应用，不可用于模拟设备，如提供即时的“反馈”到第二个神经元（动态钳位），从而驱动一个神经元关闭另一个的放电率，电压钳位只使用一个电极没有外部开关，多路复用的刺激和记录阶段。这个项目将导致一个新的膜片钳放大器的基础上高速数字信号处理器与兴奋组织相互作用。该仪器将传统的电压钳、电流钳、动态钳和膜片钳集成为一个单元，允许在钳位模式之间无瞬变切换和神经元的交互控制。第二阶段的重点包括膜片钳的硬件改进，控制的软件技术，以及哺乳动物神经元的实验验证。膜片钳是从全细胞电压钳位发展而来的，其中测量的电流通过细胞中的两个电极之一反馈，其量使细胞膜上的电压保持恒定，如在第二电极上测量的。离子通过细胞膜表面的蛋白质穿过细胞膜，蛋白质护送离子通过。这些离子通道通常是电压门控的，仅在某些电压范围内开放以通过，或者可以是化学门控的、热门控的或其他。甚至用于相同离子的不同电压门控通道可以在不同电压下打开。对于膜片钳，电极尖端非常小，通常为亚微米，与细胞膜表面的一部分接触，并且施加小的吸力以将膜密封到移液管尖端。这允许仅在一小部分膜上进行测量和反馈，并且甚至记录单个离子通道事件。有一个持续的和不断增长的需要，以目录所有的离子通道跨细胞膜，表征其门控，其功能在细胞代谢，并对整个动物的功能和健康的影响。一个常见的应用是筛选化合物与离子通道蛋白结合，以打开或关闭它们。离子通道已成为开发新药的主要目标。变化包括电压箝位、电流箝位和电导箝位。在当前的应用和技术中，这3种技术各自需要不同的模拟仪器以用于所需的快速响应反馈，并且需要测量不同的参数。单通道事件发生的变化率高达250 K/秒。离子通道事件的高速需要模拟电路。在以前的工作和出版物中，已经表明，将需要至少500 KHz的数字吞吐量，以允许数字仪器在电压箝位中执行以及模拟仪器。最近，以600 MHz的速度工作的数字信号处理芯片（DSP）已经在商业上可用。数字箝位将节省A/D和信号调理单元的额外成本，并且一台仪器将能够执行4种箝位类型中的任何一种。其他新出现的特性允许使用数字箝位放大器进行新的研究。该提案旨在开发低噪声双向云台和在膜片钳应用中使用该数字箝位放大器所需的配件，包括冷却云台以最大限度地减少噪声，并开发用于不同应用的可调反馈方案的算法库，并测试仪器对膜片钳实验的适用性。由此产生的产品预计将在一个巨大的市场中占有相当大的份额。 公共卫生关系：膜片钳技术已成为一种实用的应用技术，用于测试化合物对离子通道蛋白的功能效应。有缺陷的离子通道与几种人类疾病有关。拟议的仪器将取代3种类型的模拟钳位仪器，数字化仪和信号调理器与一个单一的，多功能的，更强大的数字钳位设备能够作为电压，电流或阻抗钳位。