Diffuse, spectrally-resolved optical strategies for detecting activity of individual neurons from in vivo mammalian brain with GEVIs

使用 GEVI 检测体内哺乳动物大脑中单个神经元活动的漫反射光谱分辨光学策略

• 批准号: 9395599
• 负责人: Nozomi Nishimura
• 金额: $ 23.55万
• 依托单位: CORNELL UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-09-30 至 2019-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9395599
• 关键词: Action Potentials; Affect; Algorithms; Behavior; Benchmarking; Biological Assay; Bioluminescence; Brain; Calcium; Cells; Code; Collection; Color; DNA; Data; Databases; Dependence; Diffuse; Discrimination; Electrodes; Electrophysiology (science); Face; Fluorescence; Generations; Genetic; Geometry; Head; Image; Individual; Infection; Injectable; Label; Lasers; Lead; Lifting; Light; Lighting; Link; Location; Measures; Methods; Microscope; Microscopy; Mus; Neurons; Optics; Output; Pattern; Performance; Photons; Population; Positioning Attribute; Preparation; Process; Reporter; Reporting; Resolution; Role; Scanning; Signal Transduction; Slice; Somatosensory Cortex; Specificity; Speed; Spike Potential; Stretching; Techniques; Technology; Testing; Time; Translating; Whole-Cell Recordings; adeno-associated viral vector; barrel cortex; base; cell type; data acquisition; density; improved; in vivo; in vivo calcium imaging; indexing; light scattering; miniaturize; neuronal circuitry; new technology; novel; novel strategies; particle; promoter; relating to nervous system; success; temporal measurement; two-photon; voltage

Measuring and understanding the activity of individual neurons is critical for understanding how neuronal circuits function and lead to behavior. Two-photon microscopy of calcium-sensitive indicators has produced insightful data on the role of individual neurons with populations. With the use of head-fixed or miniaturized versions, such optical techniques have lead to links between neural dynamics and behavior. However, these methods have not translated to voltage indicators, so that the understanding of how spikes across populations of cells affects circuits and behavior is lacking. Much of the difficulty is related to the need in two-photon microscopy to scan a small laser focus throughout a volume which results in limited time resolution. Signals from reporters are associated with a neuron because the fluorescence was generated when the laser focus was at the location of that neuron. An alternate strategy based on using multiple colors of indicators to color code neural output is proposed here. This strategy relies entirely on spectral information, so no location information or image formation is required. This enables high-speed data acquisition. This strategy takes advantage of the availability of multiple colors of genetically encoded voltage indicators and associates individual neurons with unique color combinations. A mix of adeno-associated virus vectors, each carrying DNA for a indicator of a particular color, is injected into the brain. Because the infection process is stochastic and neurons are infected by several particles, neurons are labeled by a random combination of colors. In this proposal, the number of colors, delivery methods and analysis of signals is optimized for the identification of individual neurons within a population. For the benchmarking this technology, these novel signals are compared to the performance of multiphoton microscopy and electrophysiology in assays of neural activity.

测量和了解单个神经元的活动对于了解神经元电路如何发挥作用并导致行为至关重要。钙敏感指示器的双光子显微镜已经产生了关于单个神经元与群体的作用的有洞察力的数据。随着头部固定或微型版本的使用，这种光学技术导致了神经动力学和行为之间的联系。然而，这些方法还没有转化为电压指示器，因此缺乏对跨细胞群体的尖峰如何影响电路和行为的了解。许多困难与双光子显微镜的需要有关，需要扫描整个体积的小激光焦点，这导致时间分辨率有限。来自记者的信号与神经元有关，因为荧光是在激光焦点位于神经元的位置时产生的。本文提出了一种基于多颜色指示器对神经输出进行颜色编码的替代策略。这一策略完全依赖于光谱信息，因此不需要位置信息或成像。这就实现了高速数据采集。这一策略利用了多种颜色的遗传编码电压指示器的可用性，并将单个神经元与独特的颜色组合相关联。一组腺相关病毒载体的混合物被注射到大脑中，每个载体携带特定颜色指示剂的DNA。由于感染过程是随机的，神经元受到几个粒子的感染，神经元被随机的颜色组合标记。在这一建议中，颜色的数量、传递方法和信号分析被优化，以识别种群中的单个神经元。为了对这项技术进行基准测试，我们将这些新信号与多光子显微镜和电生理学在神经活动分析中的表现进行了比较。