Developmental Influences on the Functional Organization of the Vestibular System

发育对前庭系统功能组织的影响

• 批准号: 8831208
• 负责人: David Schoppik
• 金额: $ 24.9万
• 依托单位: NEW YORK UNIVERSITY SCHOOL OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-12-01 至 2017-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8831208
• 关键词: Ablation; Acute; Anatomy; Animal Model; Architecture; Arousal; Behavior; Behavioral; Biological Models; Cells; Code; Color; Cues; Development; Developmental Biology; Developmental Process; Esthesia; Event; Excision; Eye; Eye Movements; Face; Fishes; Genetic; Goals; Human; Image; Impairment; Individual; Institution; Joints; Label; Laboratories; Labyrinth; Learning; Lesion; Link; Measures; Modeling; Molecular; Monitor; Motor Neurons; Movement; Nervous system structure; Neurons; Output; Pattern; Peptides; Peripheral; Population; Postdoctoral Fellow; Posture; Primates; Property; Recording of previous events; Research; Research Personnel; Rest; Role; Running; Sensory; Series; Shapes; Signal Transduction; Stimulus; Stroke; Sum; Synapses; System; Taste Perception; Testing; Time; Training; Transgenic Organisms; Vertebrates; Weight; Work; Zebrafish; behavior measurement; behavioral impairment; biological systems; design; developmental disease; driving behavior; gain of function; gaze; hypocretin; in vivo; interest; loss of function; member; network models; neural circuit; nonhuman primate; oculomotor; optogenetics; orbit muscle; overexpression; pressure; receptor; relating to nervous system; research study; response; sensor; time use; vestibulo-ocular reflex

DESCRIPTION (provided by applicant): My long-term goal is to run an independent laboratory at a top-tier research institution, where we will use the larval zebrafish vestibular system as a model to understand how populations of neurons coordinate their activity to produce behavior. I postulate that the developmental processes responsible for assembling an aggregate of neurons into a functional circuit will constrain the computations the circuit can perform. Therefore, as we seek to understand how populations of neurons function, we stand to learn much from their history, and the molecular-level events that organize them. The larval zebrafish vestibular system is an ideal place to test this hypothesis. Like other vertebrates, the larval zebrafish uses a highly conserved set of neurons to robustly stabilize gaze and posture in the face of external perturbations. Unlike other vertebrates, though, their vestibular system is comprised of a small number of genetically accessible neurons, each one optically accessible throughout development. I became interested in the larval zebrafish as a model system towards the end of doctoral training. I was studying the primate oculomotor system, and it had become clear that because of the large number of neurons involved, the models of population activity I had generated to explain variability in repeated eye movements were ultimately untestable. I sought a simpler system that would allow control over complete populations of neurons, and found it as a joint post-doc in the Schier and Engert labs at Harvard. I began anew, training as a molecular geneticist of behavior, working to understand the role of a particular peptide, Hypocretin, in modulating arousal. I generated a transgenic line of fish that would let me label the neurons expressing the Hypocretin receptor, in an attempt to identify the downstream targets of the peptide, and possibly the locus of behavioral modulation. The anatomy of the neurons labeled in my transgenic line implicated the vestibular system as a potential substrate, and together with my collaborators, we set out to test this hypothesis. We measured the normal zebrafish vestibuloocular reflex (VOR), traced its anatomical substrates in my line, and showed by ablation and activation that they were necessary and sufficient for behavior. Further, we showed by genetic overexpression of Hypocretin that the VOR is sensitive to this peptide. We are currently concluding a series of electrophysiologcial experiments measuring the effects of Hypocretin on central vestibular neurons. In planning a transition to run a laboratory of my own, I sought to return to the questions of circuit function that I found intractable in non-human primates. However, it became clear that my small taste of developmental biology as a member of the Schier lab had profoundly shaped the way I approached neural circuits: I found that I often wondered not just how aggregates of neurons worked together, but about what forces shaped their fate as members of a functional circuit. It became apparent that the methodological advantages of the zebrafish were uniquely suited to answer such questions, and I sought to frame ones that would allow me to continue to learn and grow as a postdoc, while setting a course for independent work. In this proposal, I begin with a working model of the central vestibular circuit in the larval zebrafish, which proposes that two functional classes of neuron, matching those found in higher vertebrates, are sufficient to explain all of the behaviors and anatomy I had observed to date. The first set of experiments aims to test models of network-level organization and computations in the central vestibular neurons responsible for the VOR. I propose a series of experiments to first anatomically identify the classes of neurons within my model, and then to use ablations and activation of individual neurons to probe their functional roles within the population. In doing so, I will directly test for higher-level network interaction that are, to date, only theoretical propositions. Specifically, I will test whether similarly tuned neurons are redundant (i.e. individually sufficient to produce behavior, but not necessary), whether they act synergistically (i.e. each necessary, but none sufficient by itself), or whether the functional architecture of the larval zebrafish VOR reflects simple linear summation. Beyond making the first direct tests of population-level computations, each of these functional architectures suggests a certain developmental arrangement, which is the subject of my second Aim: To measure and manipulate the central vestibular system during development, to uncover the way it is wired. The central vestibular system faces a deep developmental quandary: it must link peripheral afferents with directionally tuned information to the appropriate pools of motoneurons that produce stabilizing compensatory responses. Because of the difficulty in studying development of the vestibular system in vivo, we have hypotheses of how this di-synaptic coordination might occur, but no model system in which to test them. Along with a collaborator, I have generated a second transgenic line of zebrafish that allows me to color-code each neuron in the peripheral and central vestibular system (Brainbow). I propose to use this line of fish to monitor how the vestibular system comes together. To test hypotheses of trans-synaptic coordination, I will selectively lesion populations of neurons to disrupt information flow monitor the stability of color-coded axonal projections, and measure the behavioral results of my perturbations. Together, these experiments will fill deep gaps in our understanding of how the neurons responsible for the VOR assemble and function. Further, they will fill specific technical and intellectual gaps, preparing me to successfully transition from a postdoctoral trainee to an independent investigator.

描述（由申请人提供）：我的长期目标是在顶级研究机构运行一个独立的实验室，在那里我们将使用幼体斑马鱼前庭系统作为模型，以了解神经元群体如何协调其活动以产生行为。我假设，负责将神经元集合组装成功能电路的发育过程将限制电路可以执行的计算。因此，当我们试图了解神经元群体的功能时，我们可以从它们的历史以及组织它们的分子水平事件中学到很多东西。斑马鱼幼体的前庭系统是检验这一假设的理想场所。像其他脊椎动物一样，斑马鱼幼体使用一组高度保守的神经元来稳定凝视和姿态，以应对外部干扰。然而，与其他脊椎动物不同的是，它们的前庭系统由少量遗传上可接近的神经元组成，每个神经元在整个发育过程中都是光学可接近的。我开始对斑马鱼幼体产生兴趣，并将其作为博士培训结束时的模型系统。当时我正在研究灵长类动物的眼动系统，很明显，由于涉及大量神经元，我为解释重复眼球运动的变异性而建立的群体活动模型最终无法检验。我寻找一个更简单的系统，可以控制整个神经元群体，并在哈佛的希耶和恩格特实验室联合博士后时发现了它。我重新开始，接受行为分子遗传学家的训练，致力于了解一种特殊的肽--下丘脑分泌素在调节唤醒中的作用。我培育了一条转基因鱼，让我标记表达下丘脑泌素受体的神经元，试图确定肽的下游靶点，以及可能的行为调节位点。在我的转基因系中标记的神经元的解剖结构暗示前庭系统是一个潜在的底物，我们与我的合作者一起开始测试这一假设。我们测量了正常的斑马鱼前庭眼反射（VOR），在我的线跟踪其解剖基板，并通过消融和激活表明，他们是必要的和足够的行为。此外，我们通过下丘脑泌素的基因过表达表明VOR对该肽敏感。我们目前正在总结一系列的电生理实验测量Hypocretin对中枢前庭神经元的影响。在计划过渡到经营自己的实验室时，我试图回到我发现在非人类灵长类动物中难以解决的回路功能问题。然而，很明显，作为希耶实验室的一员，我对发育生物学的浅尝辄止，深刻地塑造了我处理神经回路的方式：我发现，我经常想知道的不仅仅是神经元的集合如何协同工作，而是什么力量塑造了它们作为功能回路成员的命运。很明显，斑马鱼的方法论优势非常适合回答这些问题，我试图构建一些可以让我继续学习和成长的博士后，同时为独立工作设定课程。在这篇论文中，我开始以斑马鱼幼鱼中枢前庭回路的工作模型为起点，提出了两种功能性神经元，与高等脊椎动物中发现的神经元相匹配，足以解释我迄今为止观察到的所有行为和解剖学。第一组实验旨在测试负责VOR的中央前庭神经元中的网络级组织和计算模型。我提出了一系列的实验，首先在解剖学上确定我的模型中的神经元类别，然后使用消融和激活单个神经元来探测它们在群体中的功能作用。在这样做的时候，我将直接测试更高层次的网络交互，到目前为止，这只是理论上的命题。具体来说，我将测试是否类似调整 神经元是冗余的（即，单独足以产生行为，但不是必要的），它们是否协同作用（即，每个必要的，但没有足够的本身），或是否斑马鱼幼虫VOR的功能结构反映简单的线性求和。除了对群体水平的计算进行第一次直接测试之外，这些功能架构中的每一个都暗示了某种发育安排，这是我第二个目标的主题：在发育过程中测量和操纵中央前庭系统，揭示它的连接方式。中枢前庭系统面临着一个深层次的发展困境：它必须将具有定向调谐信息的外周传入神经与产生稳定补偿反应的运动神经元相联系。由于在活体中研究前庭系统的发育很困难，我们有关于这种双突触协调如何发生的假设，但没有模型系统来测试它们。沿着，我与一位合作者一起培育出了第二条转基因斑马鱼品系，它使我能够对外周和中枢前庭系统（Brainbow）中的每个神经元进行颜色编码。我建议用这条鱼来监测前庭系统是如何结合在一起的。为了验证跨突触协调的假设，我将选择性地损伤神经元群体以扰乱信息流，监测彩色编码轴突投射的稳定性，并测量我的扰动的行为结果。总之，这些实验将填补我们对负责VOR的神经元如何组装和功能的理解中的深刻空白。此外，他们将填补特定的技术和知识空白，为我从博士后实习生成功过渡到独立调查员做好准备。