Controlling neural circuits with single-cell resolution in behaving animals

以单细胞分辨率控制行为动物的神经回路

• 批准号: 8559985
• 负责人: Christopher Fang-Yen
• 金额: $ 35万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-08-15 至 2018-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8559985
• 关键词: Algorithms; Animal Model; Animals; Architecture; Axon; Behavior; Behavior monitoring; Behavioral Genetics; Biogenic Amines; Biological Models; Biomedical Engineering; Brain; Caenorhabditis elegans; Calcium; Cells; Code; Computer software; Data; Dendrites; Development; Devices; Dimensions; Disease; Dissection; Dopamine; Drosophila genus; Feedback; Financial compensation; Fluorescence; Food; Future; Genetic; Genetic Models; Goals; Image; Individual; Ion Channel; Ion Pumps; Label; Laboratories; Larva; Lasers; Lead; Light; Lighting; Locomotion; Methods; Modeling; Movement; Nematoda; Nerve; Nerve Degeneration; Nervous system structure; Neurologic; Neurology; Neurons; Neurotransmitters; New York; Opsin; Optics; Organism; Pathway interactions; Pattern; Pennsylvania; Performance; Process; Reagent; Resolution; Role; Serotonin; Shapes; Signal Transduction; Speed; Staging; Structure; Swimming; Synapses; Synaptic Transmission; System; Techniques; Technology; Testing; Time; Universities; Varicosity; Vision; Work; Zebrafish; awake; base; design; digital; dopaminergic neuron; fluorescence imaging; image processing; improved; instrumentation; mutant; nervous system disorder; neural circuit; neuron development; neuronal cell body; neuroregulation; new technology; next generation; optogenetics; public health relevance; relating to nervous system; research study; spatiotemporal; tool

DESCRIPTION (provided by applicant): Neural circuits are the fundamental functional units of the nervous system. A basic understanding of circuit function will provide an important basis for understanding how these circuits malfunction in neurological disorders. The study of neural circuits in small and relatively simple model animals such as C. elegans and Drosophila has many advantages, including genetic manipulability and amenability to optical techniques. Circuit analysis in these organisms has been buoyed by the recent development of 'optogenetic' methods for stimulating and inhibiting neural activity using light-sensitive ion channels and pumps [1]. Progress in optogenetics requires not only development and optimization of new opsin molecules but also new strategies and technologies for perturbing specific opsin-expressing neurons. In this project, we will develop optical and genetic methods for manipulating neural circuits with single- neuron resolution in freely moving C. elegans. This project extends previous work by Dr. Fang-Yen, in which machine vision algorithms and lasers patterned by a digital micromirror device (DMD) were used to achieve spatiotemporal control of neural activity in freely behaving worms [2]. This earlier system was limited to a spatial resolution of about 20-30 microns, which is insufficient to selectively illuminate single neurons in the animal's nerve ring (brain). In this project we will develop a next-generation system capable of resolving single neurons and subcellular features. We will approach this goal in three directions. First, we will develop instrumentation and machine vision algorithms to automatically image and track individual neurons and processes using fluorescence imaging. We will use a dual-magnification optical system to simultaneously track behavior of the entire worm and fluorescence in a smaller region. Second, we will design and implement predictive algorithms to illuminate tracked targets with compensation for the latency due to image processing and data transfer. This system will be designed with real-time feedback such that fine-tuning of its parameters can be done in an automated manner. Third, we will use our system, in combination with other methods, to elucidate the mechanisms of modulation of locomotory behaviors by dopaminergic and serotonergic circuits. By enabling, for the first time, the dynamic perturbation of individual or multiple neurons in a behaving animal, the technology we develop will become an important tool for the analysis of neural circuits, with numerous advantages compared with existing methods. In addition to improving our understanding of the circuit basis of behavior, these studies will help provide a circuit-level context for interpreting genetic mutants, for example in C. elegans models of synaptic transmission, neuronal development, and neurodegeneration. While the focus of this project is on C. elegans, we expect that our methods will be readily extensible to other model organisms. This project will be centered in Dr. Fang-Yen's laboratory but will draw on the expertise of several unpaid consultants at the University of Pennsylvania or nearby. These include Dr. David Raizen (Dept. of Neurology), an expert in C. elegans genetics and behavior, Dr. Brian Chow (Dept. of Bioengineering), an expert on optogenetic reagents, and Dr. Niels Ringstad (New York University), an expert in C. elegans genetics and neurotransmitter signaling.

描述（由申请人提供）：神经回路是神经系统的基本功能单位。 对电路功能的基本理解将为了解这些电路如何在神经系统疾病中发生故障提供重要的基础。 对秀丽隐杆线虫和果蝇等小型和相对简单模型动物中神经回路的研究具有许多优势，包括遗传可操作性和对光学技术的舒适性。 这些生物体中的电路分析是通过使用光敏感的离子通道和泵来刺激和抑制神经活性的最近开发的“光遗传学”方法的促进的[1]。 光遗传学的进展不仅需要开发和优化新的Opsin分子，还需要新的策略和技术来扰动特定的表达OPSIN神经元。 在这个项目中，我们将开发光学和遗传方法，用于在自由移动的秀丽隐杆线虫中使用单神经元分辨率来操纵神经回路。 该项目扩展了Fang-Yen博士的先前工作，其中使用数字微龙器设备（DMD）模仿的机器视觉算法和激光器用于实现自由行为蠕虫神经活动的时空控制[2]。 该较早的系统仅限于大约20-30微米的空间分辨率，这不足以在动物的神经环（大脑）中有选择地照亮单个神经元。 在这个项目中，我们将开发一个能够解决单个神经元和亚细胞特征的下一代系统。 我们将在三个方向上实现这一目标。 首先，我们将开发仪器和机器视觉算法，以使用荧光成像自动对单个神经元和过程进行图像和过程。 我们将使用双磁化光学系统同时跟踪较小区域中整个蠕虫和荧光的行为。 其次，我们将设计和实施预测算法，以照亮跟踪目标，并通过图像处理和数据传输造成的延迟赔偿。 该系统将通过实时反馈设计，以便可以自动化其参数的微调。 第三，我们将使用我们的系统与其他方法结合使用，以阐明通过多巴胺能和血清素能电路调节运动行为的机制。 通过首次使表现动物中个体或多个神经元的动态扰动，我们开发的技术将成为分析神经回路的重要工具，与现有方法相比，具有许多优势。 除了提高我们对行为电路基础的理解外，这些研究还将有助于提供解释的电路级别环境 基因突变体，例如在秀丽隐杆线虫传播模型中，神经元发育和神经变性。 尽管该项目的重点放在秀丽隐杆线虫上，但我们希望我们的方法很容易扩展到其他模型生物体。 该项目将集中在Fang-Yen博士的实验室中，但将借鉴宾夕法尼亚大学或附近的几位未付顾问的专业知识。 其中包括C.秀素遗传学和行为的专家David Raizen博士，Brian Chow博士（生物工程系），光遗传学试剂专家，Niels Ringstad博士（纽约大学），C。ElegransGenetics genetics and Neurotransters和Neurotranster的专家。