Development of the Layer 5 Pyramidal Neuron Subgroup Expressing Er81

在发育中的大脑皮层中，锥体神经元在生发区生成并向发育中的皮层板迁移，最早生成的占据最深层，最晚生成的占据更浅层。在生发区，锥体神经元的类型是由基因表达的差异调控控制的，其中一些已经被确定。我们还不知道特定基因的表达如何导致轴突生长、目标选择或生理特性的特定模式。Er81在皮质发育的早期阶段在分裂细胞的心室区表达，随后在皮质板有丝分裂后的L5神经元中表达。在Pax6突变体和Ngn2突变体中，Er81的表达被破坏，这意味着Er81受这两个基因的调控。这表明Er81在神经元分化和皮层层压/迁移中的早期作用，类似于它在嗅球发育中的作用。在后期，大多数表达L5 Er81的神经元也表达Ctip2，但不表达Otx1。Ctip2控制皮质脊髓轴突生长的形成，而Otx1控制丘/桥突的生长。虽然一些研究表明ER81是Ctip2的下游，但我们的初步芯片和qPCR数据显示ER81控制Ctip2，这被ER81 KO中Ctip2的上调所证实。Ctip2的缺失导致CST轴突的缺失，而表达减少导致皮质锥体神经元的错误规范，这表明Ctip2和Er81控制了轴突规范和L5投射侧支退出的过程。在皮层外，Er81在L5神经元的靶核（如上丘和下橄榄）中特异性表达，为研究皮层连通性规范提供了令人兴奋的途径。Er81在脊髓回路的形成中也有类似的作用，其中Er81由特定的运动和感觉神经元表达，突变体中这些相互连接的形成存在缺陷。Er81和Ctip2均受Fezf2调控。在Fezf2突变体中，存在L5 Ctip2表达细胞的错误规范，这些细胞表达Tbr1和Satb2，迁移到L6，并通过前连合/胼胝体发送轴突。Satb2在L5胼胝体投射神经元中表达，抑制Ctip2的表达。在Er81 KO小鼠中，我们发现表达Ctip2的胼胝体突起神经元数量增加，这表明Er81通过抑制Satb2来控制Ctip2。可见Er81在皮层连通性的建立中起核心作用。在过去的几年里，我们对皮质神经元规范的分子控制的理解有所提高，我们已经产生了一些非常令人兴奋的初步数据，支持拟议的实验。虽然这种复杂的基因网络调节着特定锥体细胞的发育，但我们对这些网络的变化如何允许特定区域的连接（这是大脑连接的基本过程）知之甚少。由于脑发育的临床相关研究发现了与导致脑功能改变的发育障碍相关的新基因，因此了解正常转录网络的细节至关重要。虽然现在就认为了解特定基因如何影响大脑皮层的连接可能会使未来的治疗成为可能还为时过早，但如果我们要在大脑修复中使用基于细胞的治疗方法，这将是必不可少的。目前的研究几乎完全集中在从先天和实验诱导的干细胞中产生可识别的神经细胞类别，尚未了解这些细胞如何在其连通性方面受到控制。

In the developing cerebral cortex, pyramidal neurons are generated in the germinal zones and migrate into the developing cortical plate, with the earliest generated occupying the deepest layers, and the last generated the more superficial layers. Within the germinal zones the type of pyramidal neuron generated is controlled by differential regulation of gene expression, some of which have been identified. We do not yet know how expression of particular genes leads to particular patterns of axon outgrowth, target selection, or physiological properties. Er81 is expressed at early stages of cortical development in the ventricular zone in dividing cells and then later in post-mitotic L5 neurons in the cortical plate. In both Pax6 mutants and Ngn2 mutants the expression of Er81 is disrupted, implying Er81 is regulated by both of these genes. This suggests an early role for Er81 in neuronal differentiation and cortical lamination/migration, similar to its role in olfactory bulb development. At later stages, most L5 Er81 expressing neurons also express Ctip2 but not Otx1. Ctip2 controls the formation of corticospinal axon outgrowth, whilst Otx1 controls outgrowth of collicular/pontine projections. Whilst some studies have shown ER81 to be down-stream of Ctip2, our preliminary microarray and qPCR data show that Er81 controls Ctip2, which is confirmed by the up-regulation of Ctip2 seen in the Er81 KO. Loss of Ctip2 results in a loss of CST axons, whilst reduced expression results in mis-specification of cortical pyramidal neurons, suggesting that Ctip2 and Er81 control the processes of axonal specification and collateral withdrawal of L5 projections. Outside the cortex, Er81 is expressed specifically in target nuclei of L5 neurons, such as the superior colliculus and inferior olive, offering an exciting avenue for investigation of specification of cortical connectivity. A similar role has been shown for Er81 in formation of circuitry in the spinal cord where ER81 is expressed by specific motor and sensory neurons and there are defects in the formation of these interconnections in mutants. Er81 and Ctip2 are both regulated by Fezf2. In Fezf2 mutants there is a mis-specification of the L5 Ctip2 expressing cells, which instead express Tbr1 and Satb2, migrate to L6 and send axons through the anterior commissure/corpus callosum. Satb2 is expressed in L5 callosal projection neurons and represses the expression of Ctip2. In the Er81 KO mouse we showed an increase in the number of Ctip2-expressing neurons with callosal projections suggesting that Er81 controls Ctip2 by suppressing Satb2. Er81 can be seen to play a central role in the establishment of cortical connectivity. In the last few years our understanding of the molecular control of cortical neuronal specification has improved and we have produced some very exciting preliminary data underpinning the proposed experiments. Whilst this complex network of genes regulates the development of particular pyramidal cells, we have very little idea about how changes in these networks allow for specific regional connectivity which is a fundamental process of brain wiring. As clinically related studies of brain development identify new gene associations with developmental disorders that result in altered brain function, it is essential to understand the details of the normal transcriptional networks. Whist it is perhaps premature to suggest that knowledge of how specific genes influence cortical wiring could enable future therapies, this will be essential if we are ever to use cell based therapies in brain repair. Current research has focused almost exclusively on the generation of identifiable neuronal cell classes from both innate and experimentally induced stem cells, no understanding of how such cells might be controlled in terms of their connectivity has yet been made.