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

描述(申请人提供)：神经系统从一组未分化的细胞发育成一个由精确相互连接的神经元组成的复杂网络。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，引导轴突从一个靶点到另一个靶点，直到它们到达指定的目的地。了解这些引导信号在将轴突引导到目的地的过程中如何相互作用，可以让我们更充分地理解精确的神经电路是如何形成的，这反过来可能导致更好的分子药物靶点和改进的治疗策略，以鼓励对脑损伤后神经移植物的大脑区域进行准确的指导。在地形图中，轴突根据某些性质，如声音频率或空间位置，被发送到中枢神经系统内一致有序的位置；这些地图通常在整个大脑回路中逐个地点重复。例如，视网膜神经节细胞(RGC)将轴突从视网膜发送到视顶盖，根据它们最初的视网膜位置在顶盖表面形成有序的连接。虽然eparin最初被认为是唯一在生成这一图谱中发挥作用的分子，但最近发现了一种排斥性的Wnt梯度，它抵消了一个映射轴上吸引人的ewitinB梯度。阐明这些Wnt和ewitinB梯度是如何相互作用的，更值得注意的是，它们在RGC中各自受体的梯度，将有助于揭示相反的分子指导信号如何指导和定义地形图，无论是在生长锥体内，还是通过对分支形态发生的影响。这项建议将通过视网膜外植体培养中Ryk和EphB受体结构的实时成像和免疫组织化学来研究RGC生长锥中Ryk和EphB受体的相互作用，与长期的视网膜外植体和分离的视网膜培养观察分支形态的变化，并通过使用电穿孔受体结构和RNAi结构在胚胎鸡视网膜中的使用获得和功能丧失的研究来观察体内映射的变化。 与公共卫生相关：需要准确的地形图来正确分类传入的感觉信号，并形成它们在中枢神经系统内发送的一致组织的电路。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来创建这种顺序，我们可能会深入了解大脑中有多少电路是精确生成的。然后，这些信息还可以用来产生药物，以改善中风或其他损伤后大脑的准确重新连接，以及鼓励神经移植患者准确地形成新的回路。

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.