Netrin5 in Mammalian Neurodevelopment

项目摘要/摘要 这项建议的目的是确定一个新的Netrin基因家族成员Netrin5在植物中的功能。 哺乳动物神经系统中运动神经元和感觉神经元的发育。电机的发展 脊髓和脑神经中的感觉神经元得到了很好的研究；然而，主要问题仍然存在， 包括运动神经轴突如何识别腹侧运动出口点，感觉神经轴突如何识别背根入口 区域，以及中枢和外周神经系统之间的边界是如何建立的。的性质 这一边界并不清楚。没有观察到任何物理屏障，如基膜，信号表明 这些重要职能的根本定义并不明确。我们最近确定了Netrin的一名新成员 可能是这样一个信号的基因家族。众所周知，netrins可以指导轴突引导和细胞迁移 发育中的神经系统，在我们对Netrin的研究过程中，我们在小鼠中发现了Netrin5 运动轴突导引中的信号传递。该基因在人类基因组中是保守的，在小鼠中，Netrin5是 在小鼠运动出口点和背根进入区的一群细胞中表达 胚胎脊髓，与三叉神经感觉神经和可能的其他脑神经相关。 这些细胞似乎是边界帽细胞，一种暂时性的神经脊源性细胞群，其功能是 将运动神经元胞体限制在前角，并对背根、雪旺细胞和背角作出贡献 根神经节伤害性神经元群。Netrin5的表达模式表明了这一点 运动和感觉神经元发育中的发育信号和外周轴突的进出 脊髓，可能还有类似的脑神经功能。我们已经为以下项生成了表达式构造 在体外研究中，对缺乏Netrin5表达的基因敲除小鼠进行体内功能评估。在我们的 首先，我们将研究在没有Netrin5的情况下运动神经元和感觉神经元的发育。这些研究 会被边界帽细胞的已知功能所告知，例如将运动神经元胞体限制为 背根节中的腹角或分化为伤害性神经元。我们还将 检查特定Netrin基因尚未参与的已知Netrin依赖过程 下定决心，如滑车神经引导。除了解剖学研究外，我们还将评估运动和 在小鼠的感觉行为中确定丢失Netrin5的功能后果。在我们的第二个 目的：我们将确定NETRIN5蛋白对运动和感觉神经轴突生长和引导的影响。 体外，并确定所需的Netrin受体。这些研究将由表达模式提供信息 体外实验结果将通过培养Netrin受体的神经细胞得到证实。 并通过体内评估相关受体基因敲除菌株的可能表型。 总之，这些实验将确定Netrin5在哺乳动物神经发育中的作用并解决 关于运动神经元和感觉神经元发育的重要而悬而未决的问题。

PROJECT SUMMARY/ABSTRACT The goal of this proposal is to determine the function of a new Netrin gene family member, Netrin5, in the development of motor and sensory neurons in the mammalian nervous system. The development of motor and sensory neurons in the spinal cord and cranial nerves is well studied; however, major questions remain, including how motor axons identify the ventral motor exit point, how sensory axons identify the dorsal root entry zone, and how the boundary between the central and peripheral nervous system is established. The nature of this boundary is unclear. No physical barrier, such as a basal lamina, has been observed, and the signals that underlie these important functions are poorly defined. We recently identified a new member of the Netrin family of genes that may be such a signal. Netrins are well known to direct axon guidance and cell migration in the developing nervous system, and we identified Netrin5 in mice in the course of our studies on Netrin signaling in motor axon guidance. The gene is conserved in the human genome and in mice, Netrin5 is expressed in a population of cells found at the motor exit points and dorsal root entry zones in the mouse embryonic spinal cord, and is associated with the sensory trigeminal nerve and possibly other cranial nerves. These cells appear to be boundary cap cells, a transient neural crest-derived cell population that functions to restrict motor neuron cell bodies to the ventral horn and to contribute to dorsal root Schwann cell and dorsal root ganglia nociceptive neuron populations. The expression pattern of Netrin5 suggests a role for this developmental signal in motor and sensory neuron development and peripheral axon guidance into and out of the spinal cord, and possibly similar functions in cranial nerves. We have generated expression constructs for in vitro studies, and knockout mice lacking Netrin5 expression for an in vivo assessment of its function. In our first Aim, we will examine motor and sensory neuron development in the absence of Netrin5. These studies will be informed by the known function of boundary cap cells, such as restricting motor neuron cell bodies to the ventral horn or differentiating into nociceptive neurons found in the dorsal root ganglia. We will also examine known Netrin-dependent processes for which the specific Netrin gene responsible has not been determined, such as trochlear nerve guidance. In addition to anatomical studies, we will assess motor and sensory behaviors in the mice to determine the functional consequences of the loss of Netrin5. In our second Aim, we will determine the effect of NETRIN5 protein on motor and sensory axon outgrowth and guidance in vitro, and determine the Netrin receptors required. These studies will be informed by the expression patterns of netrin receptors, and in vitro results will be confirmed genetically with neurons cultured from Netrin receptor knockout mice and by an in vivo assessment of possible phenocopy in the relevant receptor knockout strains. Together, these experiments will determine the role of Netrin5 in mammalian neurodevelopment and address important, unanswered questions about motor and sensory neuron development.