Longitudinal axon guidance by long range cues in the embryonic brain

描述（由申请人提供）：本提案的总体目标是确定在胚胎脊椎动物脑中建立第一个纵向轴突通路的分子机制。纵向轴突形成CNS中的主要通信高速公路，在脑区域和脊髓之间传递所有信号。在早期的大脑发育过程中，先驱轴突通过选择特定的路径和精确的长距离生长，建立了一个最初的简单纵向束阵列。我们的初步结果已经确定了Slit/Robo信号作为纵向指导的主要决定因素，特别是Slit 1和Slit 2分泌的信号蛋白及其Robo 1和Robo 2受体。我们发现，所有的先驱纵向束严重破坏小鼠胚胎携带Slit或Robo突变，由于广泛的背腹徘徊和其他错误。该提案的总体目标是定义Slit/Robo信令如何控制纵向制导。实验系统是在早期小鼠和鸡胚胎中的先驱轴突的简单有组织的阵列，在完整的胚胎和培养的轴突中使用一系列功能测定。目标1。确定分泌的Slit线索在引导纵向轴突中的功能。我们对Slit突变胚胎的初步分析确定了Slit 1和2提供了关键线索。我们假设，裂缝功能作为直接的远程指导信号，以定向和定位轴突，或作为强制性的允许信号，轴突响应其他指导线索。为了区分这些机制，我们将研究Slit突变小鼠的轴突投射，在外植体培养中用Slit挑战轴突，并在体内错误表达Slit。目标二。测试Robo家族的Slit受体在纵向引导中的功能。我们的证据表明，Robo 1和Robo 2的轴突突变体不能沿着沿着精确的通路导航。这些结果表明Robo 1和2介导Slit信号传导。我们建议，机器人设置的位置不同的纵向道通过调制的异构体的特定组合或不同的表达水平。为了测试这些Robo机制，我们将研究Robo突变小鼠的轴突，并在特定的轴突群体中错误表达Robo。 这项计划的主要目标是确定神经线路如何在胚胎大脑发育过程中形成。对发育过程中神经元生长机制的新见解将为大脑中的出生缺陷以及如何在创伤或疾病后刺激和引导神经系统的功能再生提供重要见解。

DESCRIPTION (provided by applicant): The overall goal of this proposal is to define the molecular mechanisms that establish the first longitudinal axon pathways in the embryonic vertebrate brain. Longitudinal axons form the main communication highways in the CNS, transmitting all signals between brain regions and spinal cord. During early brain development, pioneer axons establish an initial simple array of longitudinal tracts by choosing specific pathways and accurately growing long distances. Our preliminary results have identified Slit/Robo signaling as a major determinant of longitudinal guidance, specifically the Slit1 and Slit2 secreted signaling proteins and their Robo1 and Robo2 receptors. We show that all pioneer longitudinal tracts are severely disrupted in mouse embryos carrying Slit or Robo mutations, due to extensive dorsal-ventral wandering and other errors. The overall goal of the proposal is to define how Slit/Robo signaling controls longitudinal guidance. The experimental system is the simple organized array of pioneer axons in early mouse and chick embryos, using a range of functional assays in intact embryos and with cultured axons. Aim 1. Determine the function of secreted Slit cues in guiding longitudinal axons. Our initial analysis of Slit mutant embryos identifies Slit1 and 2 as providing critical cues. We hypothesize that Slits function either as direct long-range instructive signals to orient and position axons, or as obligatory permissive signals for axons to respond to other guidance cues. To distinguish between these mechanisms, we will study axon projections in Slit mutant mice, challenge axons with Slits in explant cultures, and mis-express Slits in vivo. Aim 2. Test the function of the Robo family of Slit receptors in longitudinal guidance. Our evidence shows that axons mutant for Robo1 and 2 are unable to navigate along precise pathways. These results indicate that Robo1 and 2 mediate Slit signaling. We propose that Robos set the position of different longitudinal tracts via modulation of either specific combinations of isoforms or by different expression levels. To test these Robo mechanisms, we will study axons in Robo mutant mice, and mis-express Robos in specific axon populations. The main goal of this proposal is to define how neural wiring can form during embryonic brain development. New insights into neuron growth mechanisms during development will provide significant insights into birth defects in the brain, as well as how functional regeneration of the nervous system could be stimulated and guided following trauma or disease.