Functional Development of Premotor Neurons That Mediate the Vestibulo-Ocular Reflex

摘要 大脑的前庭神经元将周围的平衡感觉转化为反射命令，稳定姿势，步态和凝视。在这里，我们提出了一系列的实验，以测试流行的模型，如何发展中央前庭神经元来正确地传递感官信息，以特定的运动神经元，使凝视稳定。我们的建议有三个目标，每个目标都针对一个关于中枢神经元发育的特定假设： 该领域的当前模型提出，负责垂直/扭转前庭眼反射的中央前庭神经元通过响应已连接到其目标肌肉的眼外运动神经元分泌的线索而采取两种命运之一。我们的初步数据表明，相反，前庭神经元的命运可能会独立于这些信号。我们建议利用斑马鱼制备的光学可及性，快速外部开发和分子可追踪性。为了直接量化，在体内，中央前庭神经元的时空发展，我们将使用一个出生日期的技术，我们以前优化运动神经元。我们将在正常情况下以及眼神经和滑车神经的光学损伤后进行。 其次，目前的前庭发育模型提出了半规管在决定中枢前庭神经元的感觉选择性方面的重要作用。然而，我们的工作在幼体斑马鱼支持的想法，运河输入是一个正常的垂直/扭转VOR的，这表明感官选择性的发展是独立的运河投入。为了定义前庭神经元调谐，我们将使用我们开发的新型电生理制剂直接测量发育中前庭神经元的感觉选择性。我们将提供前庭刺激（翻译）完整的斑马鱼，同时记录从中央前庭神经元细胞内。同样，我们将测量兴奋性和抑制性突触输入对前庭神经元的反应，以确定上游前庭信号如何塑造其目标的反应。 最后，大多数脊椎动物前庭神经元接受来自两个终末器官的输入，耳石和半规管。延迟出现的功能半规管输入的幼斑马鱼提供了一个独特的机会，以确定是否需要感官活动，以建立适当的连接。我们将首先测量神经管传入神经元的电生理特性，以确定斑马鱼的电活动的紧急模式。接下来，我们将确定在发育过程中，耳道和耳石传入神经何时正确会聚。我们将通过首先在单通道传入中表达光敏通道来实现。然后，我们将记录中央前庭神经元，定义它们的感觉调谐。然后，我们将定义特定通道传入神经之间的连通性概率， 调谐到特定的旋转轴，与类似的和正交的中央前庭神经元。 总之，这些实验的影响将是确定何时以及如何解剖专业化和感觉选择性出现在前庭神经元负责凝视稳定。这些数据是评估和治疗异常发育以及改善急性中枢扰动（例如中风后）的先决条件。

Abstract The brain's vestibular neurons transform peripheral balance sensations into reflexive commands, stabilizing posture, gait, and gaze. Here we propose a series of experiments to test prevailing models of how developing central vestibular neurons come to properly relay sensory information to particular motoneurons, enabling gaze stabilization. Our proposal has three aims, each addressing a specific hypothesis about central neuron development: The current model in the field proposes that central vestibular neurons responsible for the vertical/torsional vestibuloocular reflex adopt one of two fates by responding to cues secreted from extraocular motoneurons that have wired to their target muscles. Our preliminary data suggests instead that vestibular neuron fate may instead proceed independently of such signals. We propose to leverage the optical accessibility, rapid external development, and molecular tractability of the zebrafish preparation. To directly quantify, in vivo, the spatiotemporal development of central vestibular neurons, we will use a birthdating technique we have previously optimized for motoneurons. We will do so both under normal conditions, as well as following optical lesions of oculomotor and trochlear nerves. Next, the current model of vestibular development proposes a vital role for the semicircular canals in determining the sensory selectivity of central vestibular neurons. However, our work in the larval zebrafish supports the idea that canal input is dispensable for a normal vertical/torsional VOR, suggesting the development of sensory selectivity is independent of canal in put. To define vestibular neuron tuning, we will directly measure the sensory selectivity of developing vestibular neurons using a novel electrophysiological preparation we have developed. We will provide vestibular stimuli (translation) to intact zebrafish while recording intracellularly from central vestibular neurons. Similarly, we will measure the response of the excitatory and inhibitory synaptic inputs to vestibular neurons to determine how the upstream vestibular signals shape the response of their target. Finally, most vertebrate vestibular neurons receive input from two end organs, the otoliths and the semicircular canals. The delayed emergence of functional semicircular canal input in the larval zebrafish provides a unique opportunity to determine whether or not sensory activity is required to establish proper connectivity. We will first measure the electrophysiological properties of canal afferent neurons as zebrafish develop to define emergent patterns of electrical activity. Next, we will determine when during development the canal and otolith afferents properly converge. We will do so by first expressing a light-sensitive channel in single canal afferents. We will then record from central vestibular neurons, defining their sensory tuning. We will then define the probability of connectivity between specific canal afferents, each tuned to a particular axis of rotation, with similarly and orthogonally tuned central vestibular neurons. Together, the impact of these experiments will be to define when and how anatomical specialization and sensory selectivity emerge in the vestibular neurons responsible for gaze stabilization. Such data are a prerequisite to evaluate and treat abnormal development, and to ameliorate acute central perturbations, such as follow stroke.