Assessing the contact-mediated role of netrin1 in axon guidance

神经回路的建立需要神经元将轴突延伸超过相当大的距离。 距离，利用胚胎环境中的分子线索来定位它们的生长锥。教材 引导因子的实例是netrin 1，其是层粘连蛋白超家族的成员，其首先在脊髓损伤中被表征。 线.经典的研究表明，netrin 1，由底板（FP）细胞产生，通过趋化作用，扩散 在长距离上引导Dcc+连合轴突朝向FP。Netrin 1随后被牵连其中 整个神经系统的指导决策。然而，我们最近的研究表明， netrin 1的关键来源是脊髓脑室区的神经祖细胞（NPC）。而不是作为一个 长程扩散梯度，我们的数据支持一个模型，其中NPC衍生netrin 1局部作用，形成一个 定向路径的轴突，也促进轴突成束。NPC的双极几何结构允许他们 使netrin 1存款在脊髓的基底（软膜）边缘。然后，该netrin 1+底物局部作用， 直接成束的，腹侧轴突通过趋触性延伸，细胞沿着粘附物定向生长 面然而，netrin 1可能介导了一个更复杂的活动，为轴突超越纯粹的粘附：netrin 1是 然后沉积到Dcc+轴突上，这些轴突然后精确地围绕VZ的边界生长，即netrin 1- 表达域。去除一个小区域的netrin 1表达会导致轴突偏离它们的轨迹， 遵循异位netrin 1边界。我们建议将这些活动称为“常春藤”增长 边界，从支撑常春藤（常春藤属：常春藤）生长的墙的类比，它本身没有被穿透 在常春藤旁边Netrin 1可以形成局部生长基质，促进轴突在神经元周围成束状生长。 netrin 1表达域。这种对netrin 1功能的机械基础的重新解释，解释了 为什么使用可溶性netrin 1作为再生因子进展甚微。为了实现这一目标，我们将 评估[1]脊髓NPC建立促进netrin 1+趋触底物的机制 目标1中定向的束状轴突生长，[2] NPC与FP细胞是否具有产生netrins的不同作用 在目的2和[3]中，netrin是否在目的3中的前脑中的轴突引导中具有触针活性。 目的1：确定脊髓NPC建立netrin 1“常春藤”边界的机制 假设：脊髓NPC交通netrin 1软膜表面建立一个haptotactic基板。Netrin 1然后 转移到Dcc+轴突，以促进它们在netrin 1表达区域周围的成束轴突生长。 目的2：确定FP衍生的netrin 1在脊髓中的作用范围 假设：FP-衍生的netrin 1在短距离内作用于穿过中线的束状连合轴突 目的3：确定NPC衍生的netrin 1/3在前脑中的作用 假设：发育中的前脑中的神经祖细胞使用netrin家族建立生长边界 引导间脑和端脑之间的轴突

The establishment of neural circuits requires neurons to extend axons over considerable distances, using molecular cues in the embryonic environment to orient their growth cones. The textbook example of a guidance factor is netrin1, a member of the laminin superfamily first characterized in the spinal cord. Classic studies suggested that netrin1, produced by floor plate (FP) cells, acts by chemotaxis, diffusing over long distances to guide Dcc+ commissural axons towards the FP. Netrin1 was then implicated in guidance decisions throughout the nervous system. However, our studies have recently demonstrated that the key source of netrin1 is neural progenitor cells (NPCs) in the spinal ventricular zone. Rather than acting as a long-range diffusible gradient, our data supports a model in which NPC-derived netrin1 acts locally, forming a directional path for axons that also promotes axonal fasciculation. The bipolar geometry of NPCs permits them to deposit netrin1 at the basal (pial) margins of the spinal cord. This netrin1+ substrate then acts locally to direct fasciculated, ventral axon extension by haptotaxis, the directed growth of cells along an adhesive surface. However, netrin1 may mediate a more complex activity for axons beyond pure adhesion: netrin1 is then deposited onto Dcc+ axons, and these axons then grow precisely around the border of VZ, i.e. a netrin1- expressing domain. Ablating a small region of netrin1 expression causes axons deviate from their trajectory to follow the ectopic netrin1 boundary. We propose to call this collection of activities a “hederal” growth boundary, from the analogy of a wall supporting the growth of ivy (genus: hedera) that is not itself penetrated by the ivy. Netrin1 can form a local growth substrate, and promote the fasciculated growth of axons around a netrin1-expressing domain. This reinterpretation of the mechanistic basis by which netrin1 functions, explains why little progress has been made using soluble netrin1 as a regenerative factor. To further this goal, we will assess [1] the mechanisms by which spinal NPCs establish a netrin1+ haptotactic substrate that promotes directed, fasciculated axonal growth in Aim 1, [2] whether NPC vs FP cells have distinct roles producing netrins in Aim 2 and [3] whether netrins have haptotactic activities in axon guidance in the forebrain in Aim 3. Aim 1: Determine the mechanism by which spinal NPCs establish netrin1 “hederal” boundaries Hypothesis: Spinal NPCs traffic netrin1 to the pial surface to establish a haptotactic substrate. Netrin1 then transfers to Dcc+ axons to promote their fasciculated axon growth around netrin1-expressing regions. Aim 2: Determine the range role of FP-derived netrin1 in the spinal cord Hypothesis: FP-derived netrin1 acts over short range to fasciculate commissural axons crossing the midline Aim 3: Determine the role of NPC-derived netrin1/3 in the forebrain Hypothesis: Neural progenitors in the developing forebrain use the netrin family to establish growth boundaries that guide axons between the diencephalon and telencephalon.