Voltage-sensitive dye imaging from the nervous system

神经系统电压敏感染料成像

• 批准号: 8097413
• 负责人: DEJAN P ZECEVIC
• 金额: $ 35.05万
• 依托单位: YALE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-07-15 至 2013-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8097413
• 关键词: Action Potentials; Affect; Area; Autistic Disorder; Behavior Control; Brain; Brain Diseases; Brain region; Cell membrane; Data; Dendritic Spines; Development; Diffuse; Disease; Dyes; Epilepsy; Excitatory Synapse; Fluorescence Microscopy; Frequencies; Image; Individual; Label; Laser Scanning Microscopy; Lasers; Light; Maps; Membrane Potentials; Microscopy; Monitor; Mood Disorders; Nervous system structure; Neurodevelopmental Disorder; Neurons; Noise; Optics; Plant Roots; Preparation; Process; Protocols documentation; Pump; Research; Resolution; Schizophrenia; Signal Transduction; Slice; Source; Speed; Synaptic plasticity; System; Testing; Time; Vertebral column; charge coupled device camera; improved; information processing; insight; neural circuit; prevent; public health relevance; research study; solid state; success; synaptic function; tool; voltage

DESCRIPTION (provided by applicant): New or improved tools to gain a better understanding of how neurons process information and modulate activity are critical. Because the principal function of a neuron is to process electrical signals, one tool that is most in need for improvement is membrane potential imaging with voltage-sensitive probes (Vm imaging). This application seeks to build a high speed imaging system that will have the sensitivity to cover the full range of spatial and temporal resolutions required in monitoring neuronal activity in brain slices. Presently, such a system is not in existence. The spatial resolution will range from recording Vm-signals from individual dendritic spines to monitoring simultaneously activity of large ensembles of neurons. Subcellular, single spine resolution in Vm-imaging has been attempted (Nuriya et al, 2006) but never satisfactory achieved. Single spine resolution is, however, essential because excitatory synapses on individual dendritic spines are elementary units underlying mechanisms of short and long term plasticity. At the same time, cellular mechanisms of plasticity must be integrated in the context of organizational and integrative aspects of persistent synaptic plasticity in different brain region at the macroscopic level. Monitoring large neuronal ensembles is needed to provide spatial maps of plasticity expression over large areas of the brain circuitry. In order to accurately monitor action potential (AP) signals, high-temporal resolution of 2,000 - 10,000 frames per second will be provided by utilizing a high-speed CCD camera. The basic principles governing the sensitivity of high-speed, high-spatial resolution Vm- imaging indicate how proposed improvements can be implemented. Also, all necessary components for the proposed system are available and have been tested and demonstrated to be functional (preliminary experiments). The imaging system will use a frequency-doubled diode-pumped Nd:YVO4 at 532 nm as an excitation source in wide-field epi-fluorescence microscopy and a high-speed CCD camera for acquisition. An additional aim of this proposal is to further develop and test new voltage-sensitive probes and protocols for selectively labeling the plasma membrane of individual neurons with minimal intracellular labeling in order to achieve the maximum signal-to-noise (S/N). PUBLIC HEALTH RELEVANCE: The research on normal synaptic function and plasticity in the nervous system seeks fundamental insights into the mechanisms by which neural circuits control behavior. Through these insights, it may become possible to clarify the root causes of diseases that affect millions, including schizophrenia, mood disorders, degenerative brain disorders, epilepsy, and neurodevelopmental disorders such as autism.

描述（由申请人提供）：新的或改进的工具，以获得更好地了解神经元如何处理信息和调节活动是至关重要的。由于神经元的主要功能是处理电信号，因此最需要改进的一种工具是使用电压敏感探针的膜电位成像（Vm成像）。本申请旨在构建一种高速成像系统，该系统将具有覆盖监测脑切片中的神经元活动所需的全部空间和时间分辨率范围的灵敏度。目前，这样的制度还不存在。空间分辨率的范围将从记录来自单个树突棘的Vm信号到同时监测大型神经元集合的活动。已经尝试了Vm成像中的亚细胞、单棘分辨率（Nuriya等人，2006），但从未达到令人满意的效果。然而，单棘分辨率是必不可少的，因为单个树突棘上的兴奋性突触是短期和长期可塑性机制的基本单位。同时，可塑性的细胞机制必须在宏观水平上整合在不同脑区持续突触可塑性的组织和整合方面的背景下。需要监测大的神经元系综，以提供大脑回路大区域的可塑性表达的空间图。为了准确监测动作电位（AP）信号，将利用高速CCD相机提供每秒2，000 - 10，000帧的高时间分辨率。控制高速、高空间分辨率Vm成像的灵敏度的基本原理指示可以如何实施所提出的改进。此外，拟议系统的所有必要组件都是可用的，并已测试和证明是功能性的（初步实验）。成像系统将使用倍频二极管泵浦Nd：YVO 4在532 nm作为激发源，在宽场落射荧光显微镜和高速CCD相机的采集。该提案的另一个目的是进一步开发和测试新的电压敏感探针和方案，用于以最小的细胞内标记选择性地标记单个神经元的质膜，以实现最大的信噪比（S/N）。公共卫生关系：对神经系统中正常突触功能和可塑性的研究旨在从根本上了解神经回路控制行为的机制。通过这些见解，有可能澄清影响数百万人的疾病的根本原因，包括精神分裂症、情绪障碍、退行性脑部疾病、癫痫和自闭症等神经发育障碍。