The Role of Cajal-Retzius Neurons in Postnatal Cortical Circuit Assembly

描述(由申请人提供)：为了更好地了解精神分裂症、双相情感障碍、精神发育迟滞或自闭症患者的智力障碍和情绪障碍的机制，有必要首先研究正常大脑发育过程中皮质回路是如何组装的。Cajal-Retzius(CR)神经元长期以来一直与皮质发育有关，并被认为在其中几种疾病中发挥了作用。最有文献记载的CR神经元的功能是协调神经元迁移和皮质分层。这是通过它们分泌卷轴蛋白来实现的，尽管皮质中其他产生卷轴蛋白的细胞也可能起到作用。有趣的是，CR神经元在出生后的新皮质中显示出自发的相关活动，并已知与发育中的锥体神经元的顶端树突棘进行化学和电突触。因此，可想而知，CR神经元可能通过锥体神经元树突参与突触的形成和皮层神经元自发振荡的出现。我们希望通过活体双光子成像对未成熟的皮质神经回路的结构和功能进行验证。具体地说，我们将成像不同品系的转基因小鼠中表达绿色荧光蛋白(GFP)的CR和锥体神经元，以及加载了荧光钙指示剂的皮质神经元，以检测神经元的放电。我们已经确定了一种转基因小鼠(Ebf2-GFP)，在其中GFP在几乎所有的CR神经元中都有表达。此外，我们还获得了一株在相同细胞中表达Cre重组酶的小鼠(Ebf2：GFPiCre)。使用最先进的双光子显微镜，我们打算选择性地杀死CR神经元，或者使用光遗传学和CRE-Lox技术来遗传地沉默它们的活动，然后检查这种有针对性的CR神经元扰动对新皮质发育的影响。我们将研究CR消融/沉默是否影响锥体神经元顶端树突和棘突的成熟和/或锥体神经元网络的自发活动。这些研究解决了关于大脑皮层发育的基本问题。这些实验旨在填补关于CR神经元的知识空白，并确定这些细胞在皮质回路组装中的新角色。这些数据应该会解决因先前研究的方法学缺陷而引起的关于其功能的长期争议。我们的工作还将产生新的想法，即皮质回路中的细微缺陷可能会导致几种神经精神障碍，从而有助于找到这些毁灭性疾病的更好治疗方法。 与公共健康相关：拟议的研究将调查大脑回路在发育过程中是如何在对情绪、认知和创造力以及学习和记忆重要的区域组装的。这些实验旨在产生新的想法，即大脑连接中的细微变化如何导致破坏性的神经精神疾病，如精神分裂症、自闭症、精神发育迟滞或双相情感障碍。

DESCRIPTION (provided by applicant): To better understand the mechanisms of intellectual impairment and emotional disturbances in individuals with schizophrenia, bipolar disorder, mental retardation or autism, it is essential to first study how cortical circuits are assembled during normal brain development. Cajal-Retzius (CR) neurons have long been implicated in cortical development and are thought to play a role in several of these disorders. The best documented function of CR neurons is in orchestrating neuronal migration and cortical lamination. This is achieved through their secretion of reelin, though other reelin-producing cells in the cortex could also play a role. Interestingly, CR neurons show spontaneous correlated activity in the postnatal neocortex and are known to make both chemical and electrical synapses with apical dendritic spines of developing pyramidal neurons. Therefore, it is conceivable that CR neurons could play a role in synaptogenesis by pyramidal neuron dendrites and in the emergence of spontaneous oscillations in cortical neurons. We want to test this hypothesis using in vivo 2-photon imaging of the structure and function of immature cortical circuits. Specifically, we will image CR and pyramidal neurons expressing the green fluorescent protein (GFP) in different lines of transgenic mice, as well as cortical neurons loaded with fluorescent calcium indicators to detect neuronal firing. We have identified a line of transgenic mice (Ebf2-GFP) in which GFP is expressed in virtually all CR neurons. In addition, we obtained a line of mice (Ebf2:GFPiCre) that expresses the Cre recombinase in the same cells. Using state-of- the-art 2-photon microscopy, we intend to selectively kill CR neurons, or genetically silence their activity using optogenetics and Cre-Lox technologies, and then examine the repercussions of such targeted perturbations of CR neurons on neocortical development. We will investigate whether CR ablation/silencing affects the maturation of apical dendrites and spines of pyramidal neurons and/or the spontaneous activity of networks of pyramidal neurons. These studies address fundamental questions about cortical development. The experiments are designed to fill knowledge gaps about CR neurons and identify novel roles for these cells in cortical circuit assembly. These data should resolve longstanding controversies about their function that arose from methodological shortcomings of prior studies. Our work will also generate new ideas about how subtle defects in cortical circuits might contribute to several neuropsychiatric disorders and thereby help to find improved treatments for these devastating diseases. PUBLIC HEALTH RELEVANCE: The proposed studies will investigate how brain circuits are assembled during development in areas important for emotion, cognition and creativity, as well as for learning and memory. The experiments are designed to generate new ideas about how subtle alterations in brain wiring could result in devastating neuropsychiatric disorders such as schizophrenia, autism, mental retardation or bipolar disorder.