Roles of Ryk and EphB Signaling in Branch Morphogenesis and Topographic Mapping

描述（由申请人提供）：神经系统从一组未分化的细胞发育成精确互连的神经元的精细网络。这个网络的创建和组织是由一系列分子引导线索指导和定义的，引导轴突从一个目标到另一个目标，直到它们到达指定的目的地。了解这些指导线索在将轴突引导到其目的地的过程中如何相互作用，使我们更接近于全面了解精确的神经回路是如何形成的，这反过来可能会导致更好的分子药物靶点和改进的治疗策略，以鼓励在脑损伤后重组神经移植物的脑区域的准确指导。 在地形图绘制中，轴突根据某些性质（如声音频率或空间位置）被发送到CNS内一致有序的位置;这些地图通常在整个大脑回路中逐点重复。例如，视网膜神经节细胞（RGC）将轴突从视网膜发送到视顶盖，基于其原始视网膜位置在顶盖表面上形成有序连接。虽然肝配蛋白最初被认为是唯一的分子发挥作用，在产生这个地图，最近的排斥Wnt梯度，反对有吸引力的肝配蛋白B梯度的一个映射轴上，被确定。阐明这些Wnt和ephrinB梯度，更值得注意的是它们各自的受体在RGCs中的梯度如何相互作用，将揭示相反的分子引导线索如何在生长锥内和通过对分支形态发生的影响来指导和定义地形图。该提案将通过电穿孔受体构建体的实时成像和视网膜外植体培养中的免疫组织化学来检查RGC生长锥中Ryk和EphB受体的相互作用，长期视网膜外植体和分离的视网膜培养以观察分支形态的变化，和使用增益-和损失-在鸡胚视网膜中用电穿孔的受体构建体和RNAi构建体进行功能研究，以观察体内映射的变化。 公共卫生相关性：需要精确的地形图来正确分类传入的感觉信号，并形成一致的有组织的电路，沿着这些电路在CNS内发送。因此，通过更好地了解对映射至关重要的受体如何相互作用以创建这种秩序，我们可以深入了解大脑中有多少回路是精确生成的。然后，这些信息还可以用于生产药物，以改善中风或其他损伤后大脑的准确重新布线，以及鼓励神经移植患者准确的新回路形成。

DESCRIPTION (provided by applicant): The nervous system develops from a set of undifferentiated cells into an elaborate network of precisely interconnected neurons. The creation and organization of this network is directed and defined by a series of molecular guidance cues, leading axons from target to target until they have reached their specified destinations. Understanding how these guidance cues interact in the process of directing axons to their destinations brings us closer to a full understanding of how precise neural circuits are formed, which may, in turn, lead to better molecular drug targets and improved therapeutic strategies for encouraging the accurate guidance of reorganizing brain areas of neural grafts after brain injury. In topographic mapping, axons are sent to consistently ordered positions within the CNS based on certain qualities, such as frequency of sound or spatial position; these maps are often repeated site-by-site throughout brain circuits. Retinal ganglion cells (RGCs), for example, send axons from the retina to the optic tectum, forming orderly connections on the tectal surface based on their original retinal locations. While ephrins were initially thought to be the only molecules to play a role in generating this map, recently a repulsive Wnt gradient, countering the attractive ephrinB gradient on one mapping axis, was identified. Elucidating how these Wnt and ephrinB gradients, and more notably the gradients of their respective receptors in the RGCs, interact will shed light on how opposing molecular guidance cues direct and define the topographic map, both within the growth cone and through effects on branch morphogenesis. This proposal will examine the interaction of Ryk and EphB receptors in RGC growth cones through live imaging of electroporated receptor constructs and immunohistochemistry in retinal explant culture, with long-term retinal explant and dissociated retinal culture to observe changes in branch morphology, and with use gain- and loss-of-function studies with electroporated receptor constructs and RNAi constructs in embryonic chick retina to observe changes to mapping in vivo. PUBLIC HEALTH RELEVANCE: Accurate topographic mapping is required to correct sort incoming sensory signals and form the consistently organized circuit along which they are sent within the CNS. Thus, by understanding better how receptors crucial to mapping interact to create this order, we may gain insight into how many circuits in the brain are precisely generated. This information could then also be used to generate drugs to improve the accurate rewiring of the brain after stroke or other injury, as well as to encourage accurate new circuit formation in patients with neural grafts.