REPULSIVE CUES AND THE DEVELOPING VISUAL SYSTEM

视网膜定位图的准确形成对我们的视觉能力至关重要。在这个过程中，视网膜轴突被引导形成大脑中连接的精确地形图，从而允许感知视觉空间。这些指导机制的分子基础尚不清楚。这项建议的长期目标是了解排斥性线索如何引导轴突形成视网膜定位图。最近发现的各种排斥线索的行为，引导轴突形成视网膜定位图。最近发现的各种排斥性线索改变了我们对轴突如何重新引导的理解。这是及时解决如何在体内这些线索的功能。在视网膜顶盖投射的形成过程中，在低等脊椎动物的顶盖中，排斥性引导分子（RGM）和ephrin-A5以高后低前梯度表达。目前的进展已经提出了这两种线索如何在体内作用以形成视网膜顶盖图的具体假设：1）RGM引导初始视网膜顶盖轴突生长。2)Ephrin-A5阻止视网膜轴突离开顶盖，并在初始轴突生长完成后调节侧向分支。拟议的实验将检验这些假设。在活体条件下，观察RGM和ephrin-A5对视网膜顶盖突起的影响。鸡和斑马鱼胚胎将被研究;它们的特点很好，每一个都提供了独特的优势，以实现我们的目标。急性和局部失活的RGM和ephrin-A5在retinotectal地图形成过程中将实现发色团辅助激光灭活（卡利）在最初的轴突生长，后来在侧向分支。用重组逆转录病毒感染将产生慢性错误表达ephrin-A5在鸡顶盖。视网膜顶盖投射的变化将通过高分辨率轴突追踪和实时成像来评估。这两种方法的结合将提供强有力的补充信息，将测试所提出的假设。由于RGM和ephrin-A5可能用于轴突引导和地形秩序的形成，因此所提出的实验具有显著的临床相关性。

Accurate formation of the retinotopic map is critical for our ability to see. During this process, retinal axons are guided to form a precise topographic map of connections in the brain that allow visual space to be perceived. The molecular basis of these guidance mechanisms is not yet known. The long term objective of this proposal is to understand how repulsive cues act to guide axons to form the retinotopic map. The recent discovery of a variety of repulsive cues act to guide axons to form the retinotopic map. The recent discovery of a variety of repulsive cues has modified our understanding of how axons re guided. It is timely to address how these cues function in vivo. Repulsive guidance molecule (RGM) and ephrin-A5 are expressed in high posterior to low anterior gradients in the tectum of lower vertebrates during the formation of the retinotectal projection. Current advances have suggested specific hypotheses of how these two cues act in vivo to shape the retinotectal map: 1) RGM guides initial retinotectal axon outgrowth. 2) Ephrin-A5 prevents retinal axons from exiting the tectum and modulates lateral branching after initial axon outgrowth is completed. The proposed experiments will test these hypotheses. The changes in retinotectal projects caused by perturbing RGM and ephrin-A5 in vivo will be observed. Chick and zebrafish embryos will be studied; they are well characterized and each offers distinct advantages to achieve our goals. Acute and local inactivation of RGM and ephrin-A5 during retinotectal map formation will be achieved by chromophore-assisted laser inactivation (CALI) during initial axon outgrowth and later during lateral branching. Infection with recombinant retrovirus will generate chronic misexpression of ephrin-A5 in the chick tectum. The changes in retinotectal projects will be assessed by high resolution axon tracing and live imaging. The combination of these two approaches will provide strong complementary information that will test the proposed hypotheses. As RGM and ephrin-A5 are likely used for axon guidance and formation of topographic order, the proposed experiments are of significant clinical relevance.