CD133 神经干细胞对大脑皮层不对称发育的研究

The brain is responsible for processing multiple modalities of sensory information, controlling motor output, and mediating higher-order cognitive functions. Brain asymmetry is a conserved feature of vertebrate nervous system and is thought to be an advantageous mechanism for efficiently processing information.It displays anatomical and molecular left-right (L-R) asymmetries that correlate with their functional specialization in particular cognitive processes. Hemispheric specialization for a given cognitive function reflects differences in the neural circuits of specific neuron clusters. Although tremendous progresses have been achieved to decipher the complex neural networks using single gene lineage tracing or transcriptome analysis, the heterogeneity and the complexity of L-R asymmetrically distributed, functional neural networks are still largely unknown. Recently, by using an inducible CD133 lineage reporter mouse line, we found that E8.5-labeled, CD133+ neural stem cell (CD133+ NSC) lineages distributed asymmetrically in L-R neocortex of the brain, suggesting potential involvement of CD133+ NSCs in L-R asymmetrical brain development. In this proposal, we aim to (1) determine molecular signatures of CD133+ NSC-derived, L-R asymmetrically distributed cells in developing mouse cortex; (2) Validate sequencing result using immunohistochemistry and in situ hybridization analyses on critical genes involved in L-R asymmetry within E8.5 labeled CD133 lineage (ZsGreen positive) during embryonic development and to determine whether such asymmetry is sustained in the adult brain; (3) investigate functions of those asymmetrically neuron clusters in cortical and subcortical regions of the left hemisphere using reversible neuronal silencing approach. Accomplishment of this study will reveal the cellular and molecular signatures and spatiotemporal distributions of E8.5-labeled, CD133+ NSC lineages that contribute to L-R asymmetrical cortical development. This work will shed some light on using lineage specific stem cells combined with single cell transcriptome analysis to build a comprehensive road map to the understanding of L-R neural connections in the brain. Recognition of this fundamental pattern of brain lateralization in vertebrate species has allowed important questions to be asked about functional lateralization in nonhuman species and the findings applied to understanding lateralization in the human brain. This stud will also provide invaluable information to help researchers to understand molecular etiology of human neurological disorders that show disrupted normal brain asymmetry. For example, reduced and reversed anatomical brain asymmetry has been reported in individuals with schizophrenia, autism or dyslexia, suggesting a potential indirect relationship between the causes of these disorders and the asymmetrical development of the human cerebral cortex.

大脑偏侧性在哺乳动物中广泛存在，但长期以来由于大脑细胞类型复杂多样并缺乏有效的神经细胞异质性研究工具，何种类型的细胞参与大脑偏侧性发育及分子机制未知。哺乳动物的神经发生起源于神经上皮细胞，CD133神经干细胞直接或间接产生了大脑的多种神经细胞类型。CD133+NSC不对称特异表达在E8.5顶面神经上皮细胞,我们在前期研究中发现E8.5CD133+NSC谱系细胞在神经发育各阶段大脑中偏侧性分布。为阐明CD133+NSC谱系细胞对大脑偏侧性发育的作用，我们将利用液滴单细胞RNA测序技术解析E8.5CD133+NSC谱系神经元的类型和标志基因，其次利用原位杂交验证标志基因在胚胎和成体神经发生阶段的表达分布情况，并利用透明脑结合光片显微镜技术观测表达标志基因的神经元类群在大脑中的三维分布，最后利用果蝇抑咽侧体神经肽受体系统抑制非对称分布神经元的活性，通过小鼠行为学实验验证功能。

摘要在当前的研究中，我们展示了小鼠前脑神经发生关键阶段 (E10-18) 期间神经谱系扩展/转变的关键时间窗。然后我们证明左右不对称基因表达在大脑发育中相当普遍。通过单细胞转录组学分析和 ISH，我们证明了产前和成人大脑右侧 CeA 中 Ebf1+ 细胞的不对称偏侧化。此外，我们确定了一组在 CeA 中特异性表达的基因。作为原理证明，本研究将展示非对称侧化 CeA Ebf1+ 细胞在调节情绪行为中的功能。这项研究还将为研究其他大脑区域的偏侧化铺平道路，例如前额叶皮层和视觉皮层。继续当前的研究以了解模型动物的功能偏侧化可能有助于了解与患有神经发育障碍 (例如 ASD) 的患者的不对称大脑活动相关的一些分子调节。