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（申请人提供）：电（电压）信号是大脑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实验神经科学的最终目标。标准的玻璃和金属电极具有巨大的侵入性，并且它们的使用受到空间分辨率差、覆盖范围有限以及对细胞身份的盲目性的影响。流行的钙敏感指示剂产生的信号被细胞内钙的变化所污染，这些变化与神经元电信号无关，并且仅间接报告具有扭曲的结合和高度扭曲的波形的电信号。通过使用遗传编码电压指示器（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，即使是目前可用的性能最好的GEVI也仅适用于复合突触电位（来自未识别数量的神经元的总和电压信号）的体内监测，但不能同时分辨来自完整神经组织中的许多单个细胞的信号。虽然具有更高灵敏度的更好性能的GEVI有望从正在进行的BRAIN Initiative资助的活动中出现，但仅凭这一点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并不分离。这个问题的解决需要其他技术，包括精确的遗传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方案。稀疏的细胞靶向将允许以很少的空间重叠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单细胞反应，并在此基础上，在密集靶向组织中。总之，我们建议开发一种新的基因导向的电压成像工具，它与目前可用的工具在质量上不同，沿着数据分析方法，以便于使用来自嵌入在功能神经元网络中的大量神经元的电信号。在这里开发的新型GEVI标记协议和数据采集和分析算法应该立即对研究健康和疾病中的大脑功能有用和有效。