Reeber Stacey L, Sakai Nozomi, Nakada Yuji, Dumas Judy, Dobrenis Kostantin, Johnson Jane E, Kaprielian Zaven
Dominick P. Purpura Department of Neuroscience, Albert Einstein College of Medicine, Bronx, New York 10461, USA.
J Neurosci. 2008 Aug 27;28(35):8698-708. doi: 10.1523/JNEUROSCI.1479-08.2008.
In vertebrate embryos, most spinal commissural axons cross the ventral midline (VM) and project either alongside or significant distances away from the floor plate (FP). The upregulation of repulsive Robo1/2 receptors on postcrossing commissural axons, in mammals, presumably allows these axons to respond to the midline-associated repellents, Slit1-3, facilitating their expulsion from, and prohibiting their reentry into, the FP. Compelling data suggest that Robo3 represses Robo1/2 function on precrossing axons and that Robo1/2 inhibit attractive guidance receptors on postcrossing axons, thereby ensuring that decussated axons are selectively responsive to midline Slits. However, whether Robo1/2 expel decussated commissural axons from the VM and/or prevent their reentry into the FP has not been explicitly established in vivo. Furthermore, some commissural axons do not require Robo1/2 to elaborate appropriate contralateral projections in the mouse spinal cord. Here, we use unilateral in ovo electroporation together with Atoh1 and Neurog1 enhancer elements to visualize, and assess the consequences of manipulating Robo expression on, dl1 and dl2 chick commissural axons. In response to misexpressing a cytoplasmic truncation of Robo1 and/or Robo2, which should block all Robo-ligand interactions, postcrossing commissural axons extend alongside, but do not project away from or reenter the FP. In contrast, misexpression of full-length Robo2 prevents many commissural axons from crossing the VM. Together, these findings support key and selective in vivo roles for Robo receptors in presumably altering the responsiveness of decussated commissural axons and facilitating their expulsion from the VM within the chick spinal cord.
在脊椎动物胚胎中，大多数脊髓连合轴突穿过腹侧中线（VM），并沿着底板（FP）或与底板保持一定距离投射。在哺乳动物中，交叉后连合轴突上排斥性Robo1/2受体的上调，可能使这些轴突对中线相关排斥因子Slit1 - 3产生反应，促进它们从底板排出，并阻止它们重新进入底板。有力的数据表明，Robo3抑制交叉前轴突上的Robo1/2功能，而Robo1/2抑制交叉后轴突上的吸引性导向受体，从而确保交叉的轴突对中线Slits有选择性反应。然而，Robo1/2是否能将交叉的连合轴突从腹侧中线排出和/或防止它们重新进入底板，在体内尚未得到明确证实。此外，一些连合轴突在小鼠脊髓中形成适当的对侧投射并不需要Robo1/2。在这里，我们使用单侧卵内电穿孔技术，结合Atoh1和Neurog1增强子元件，来可视化并评估操纵Robo表达对dl1和dl2鸡连合轴突的影响。作为对错误表达Robo1和/或Robo2的细胞质截短体（这应该会阻断所有Robo - 配体相互作用）的反应，交叉后的连合轴突沿着底板延伸，但不会远离底板投射或重新进入底板。相反，全长Robo2的错误表达会阻止许多连合轴突穿过腹侧中线。总之，这些发现支持了Robo受体在体内具有关键和选择性作用，可能改变交叉连合轴突的反应性，并促进它们从鸡脊髓的腹侧中线排出。
