MOLECULAR DEVELOPMENT OF CENTRAL RETINAL PATHWAYS

我研究的长期目标是了解一个分子 水平的机制，控制形成精确的神经元 哺乳动物视觉系统中的连接。这里提出的实验 通过研究功能重要的视网膜 视交叉处的神经节细胞轴突通路形成于 胚胎发育对细胞表面分子的理解 参与视网膜轴突引导将使我们能够设计策略， 控制和指导视网膜轴突的生长，因此是至关重要的 受伤后试图重建视觉连接的努力， 疾病此外，由于细胞表面相互作用不仅 重要的视网膜轴突的指导，但也很可能支配 其他视网膜神经元之间的连接模式， 细胞表面分子如何引导这些发育事件是 组织移植修复视神经损伤的策略 连接. 这里提出的研究形成了一套完整的实验，其中 我们使用体外和体内方法来分析 CD 44和L1两种细胞表面分子在胚胎干细胞中表达显著 视网膜轴突和视交叉未来区域的神经元。我们 检验L1和CD 44共同作用，介导相互作用的假设 视网膜轴突和胚胎视交叉神经元之间的联系， 形成“X”形视网膜通路交叉点，称为 视交叉其次，我们还研究了是否CD 44，这是突出 在中线表达，涉及，无论是直接或联合 与其他细胞表面分子，在特定的路由视网膜 轴突进入同侧和对侧视束。这一假设 是基于CD 44调节细胞表面配体-受体 相互作用，并被独特地放置在中线，以影响 决定轴突是否穿过中线进入对侧 视束或避免交叉并同侧突出。 实验上，CD 44和L1作用的分子基础及其作用 使用三种方法研究体内视网膜通路发育。 一种是通过注射直接干扰体内CD 44和L1的功能 将阻断剂注入活的小鼠胚胎中， 对视网膜通路形成的影响。第二，是CD 44的干扰， 胚胎视觉系统L1功能的研究 对生长锥行为的影响。第三、 使用表达截短形式的CD 44和L1的细胞系， 定义介导它们对视网膜轴突的作用的功能域 增长这些研究解决了我们对视觉的理解中的主要空白 通过测试确定的表面分子进行功能性 重要性在体内和解剖机制，以了解分子 中央视觉通路发育的基础。

The long term objective of my research is to understand at a molecular level the mechanisms governing the formation of precise neuronal connections in the mammalian visual system. The experiments proposed here pursue this goal by investigating how the functionally important retinal ganglion cell axon pathways at the optic chiasm are formed during embryonic development. An understanding of the cell surface molecules involved in retinal axon guidance will enable us to devise strategies to control and direct the growth of retinal axons, and is thus fundamental to efforts attempting to re-establish visual connections following injury and disease. Furthermore, since cell surface interactions are not only important for retinal axon guidance, but very likely also govern the patterns of connectivity among other retinal neurons, an understanding of how cell surface molecules direct these developmental events is central to strategies using tissue transplantation to restore visual neuronal connections. The studies proposed here form an integrated set of experiments in which we use both in vitro and in vivo approaches to analyze the function of CD44 and L1, two cell surface molecules prominently expressed on embryonic retinal axons and neurons at the future region of the optic chiasm. We test the hypotheses that L1 and CD44 acting together, mediate interactions between ingrowing retinal axons and embryonic chiasm neurons and govern the formation of the "X" shaped retinal pathway intersection known as the optic chiasm. Secondly, we also examine whether CD44, which is prominently expressed at the midline, is involved, either directly or in conjunction with other cell surface molecules, in the specific routing of retinal axons into the ipsilateral and contralateral optic tracts. This hypothesis is based on the fact that CD44 regulates cell surface ligand-receptor interactions and is uniquely placed at the midline to influence the decision of axons whether to cross the midline into the contralateral optic tract or avoid crossing and project ipsilaterally. Experimentally, the molecular basis of CD44 and L1 action and their role in retinal pathway development in vivo is studied using three approaches. One, is direct perturbation of CD44 and L1 function in vivo by injections of blocking reagents into living mouse embryos and examination of the effects on retinal pathway formation. Second, is interference of CD44 and L1 function in embryonic visual system preparations and analysis of effects on growth cone behavior using time-lapse videomicroscopy. Third, is the use of cell lines expressing truncated forms of CD44 and L1 to define the functional domains mediating their actions on retinal axon growth. These studies address major gaps in our understanding of visual pathway development by testing defined surface molecules for functional importance in vivo and by dissecting the mechanisms to learn the molecular basis for central visual pathways development.