Sparse, Strong and Large Area Targeting of Genetically Encoded Indicators

遗传编码指标的稀疏、强和大面积目标

• 批准号: 9037189
• 负责人: SRDJAN D ANTIC
• 金额: $ 54.98万
• 依托单位: UNIVERSITY OF CONNECTICUT SCH OF MED/DNT
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-09-18 至 2018-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9037189
• 关键词: Adverse effects; Algorithms; Animals; Antibiotics; Area; BRAIN initiative; Blindness; Brain; Calcium; Cells; Cognitive; Craniotomy; Data; Data Analyses; Development; Disease; Dyes; Electrodes; Emotional; Event; Funding; Genetic; Genetic Recombination; Glass; Goals; Golgi Apparatus; Health; Image; Imaging Device; Imaging problem; Individual; Label; Life; Maps; Measurement; Mediating; Mental disorders; Metals; Methodology; Methods; Microscopy; Molecular; Monitor; Motor; Mus; Nerve Tissue; Neuraxis; Neurons; Neurosciences; Noise; Optics; Pattern; Population; Population Heterogeneity; Probability; Procedures; Process; Protocols documentation; Pyramidal Cells; Reporting; Resolution; Scanning; Signal Transduction; Slice; Solutions; Somatosensory Cortex; Staining method; Stains; Symptoms; Synaptic Potentials; System; Techniques; Technology; Time; Tissues; Transgenic Organisms; Translating; Trimethoprim; Work; base; cellular targeting; data acquisition; design; in vivo; information processing; interest; neuronal cell body; neuronal circuitry; neurophysiology; neurotechnology; novel; preempt; protein expression; public health relevance; recombinase; reconstruction; response; segregation; sensory stimulus; small molecule; spatiotemporal; two-photon; voltage

 DESCRIPTION (provided by applicant): Electrical (voltage) signal is the primary substrate of information processing in the brain. Detecting and recording voltage changes from neurons in living animals remains the ultimate goal of experimental neuroscience to the present day. Standard glass and metal electrodes are hugely invasive and their use suffers from poor spatial resolution, limited coverage, and blindness to cellular identity. The popular calcium-sensitive indicators generate signals contaminated with changes in intracellular calcium that are unrelated to neuronal electrical signals, and only indirectly report the electrical signals with distorted tiing and highly distorted waveform. A conceptually ideal principle to achieve monitoring of neuronal electrical activity is provided by optical voltage imaging using genetically-encoded voltage indicators (GEVIs). However, even the best performing GEVIs currently available are suitable only for in vivo monitoring of compound synaptic potentials (the summated voltage signal from unidentified number of neurons), but fail to resolve simultaneously signals from many individual cells in intact nervous tissue. While better performing GEVIs with higher sensitivity are expected to emerge from the on-going BRAIN Initiative-funded activities, this alone does not resolve the single-cell resolution voltage imaging problem: signals from individual cells are not spatially segregated. The solution to this issue requires other technologies including refined genetic targeting and data analysis methods. We plan to develop protocols for sparse GEVI targeting of central nervous system neurons. Sparse cellular targeting will allow imaging of neuronal activity with little spatial overlap. We also plan to develop data analysis routines for isolating single cel responses from data obtained in sparsely labelled tissue and, building on this, in densely targeted tissue. In summary, we propose to develop a novel genetically-directed voltage imaging tool that is qualitatively different than those currently available, along with data analyss methods to facilitate the use of electrical signals from large number of neurons embedded in functioning neuronal networks. The novel GEVI labelling protocols and data acquisition and analysis algorithms developed here should be immediately useful and impactful for studying brain function in health and disease.

 描述（由申请人提供）：电（电压）信号是大脑中信息处理的主要基质。检测和记录活体动物神经元的电压变化仍然是当今实验神经科学的最终目标。标准玻璃和金属电极具有巨大的侵入性，其使用存在空间分辨率差、覆盖范围有限以及对细胞身份无视等问题。流行的钙敏感指示器产生的信号受到细胞内钙变化的污染，与神经元电信号无关，并且仅间接报告具有扭曲的连接和高度扭曲的波形的电信号。使用基因编码电压指示器（GEVI）的光学电压成像提供了实现神经元电活动监测的概念上理想的原理。然而，即使是目前性能最好的 GEVI，也仅适用于复合突触电位（来自未识别数量的神经元的电压信号总和）的体内监测，但无法同时解析来自完整神经组织中许多单个细胞的信号。虽然正在进行的 BRAIN Initiative 资助的活动预计会出现性能更好、灵敏度更高的 GEVI，但这本身并不能解决单细胞分辨率电压成像问题：来自单个细胞的信号在空间上不是隔离的。这个问题的解决需要其他技术，包括精细化的基因靶向和数据分析方法。我们计划开发针对中枢神经系统神经元的稀疏 GEVI 靶向方案。稀疏细胞靶向将允许在几乎没有空间重叠的情况下对神经元活动进行成像。我们还计划开发数据分析程序，用于从稀疏标记组织中获得的数据中分离出单细胞反应，并在此基础上在密集目标组织中分离单细胞反应。总之，我们建议开发一种新型的基因定向电压成像工具，该工具在质量上与目前可用的工具不同，并提供数据分析方法，以方便使用来自嵌入功能神经网络的大量神经元的电信号。这里开发的新颖的 GEVI 标记协议以及数据采集和分析算法应该对研究健康和疾病中的大脑功能立即有用且具有影响力。