Imaging Dynamics and Interactions of Developmental Lineages in the Early Embryo

早期胚胎发育谱系的成像动力学和相互作用

基本信息

批准号：
9031167
负责人：
ALVIN T YEH
金额：
$ 31.73万
依托单位：
TEXAS ENGINEERING EXPERIMENT STATION
依托单位国家：
美国
项目类别：
财政年份：
2015
资助国家：
美国
起止时间：
2015-04-01 至 2019-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9031167
关键词：
4D Imaging Address Age Animal Model Anterior Autistic Disorder Biological Models Brain Brain Stem Caliber Cell Lineage Cell Separation Cell division Cells Cerebellar malformation Cerebellum Congenital Abnormality Defect Detection Development Differentiated Gene Disease Embryo Embryonic Development Ensure Event Fertilization Future Gene Expression Gene Targeting Gene Transfer Techniques Genes Genetic Programming Germ Layers Goals Health Homeobox Genes Hour Human Image In Situ Hybridization Individual Intellectual functioning disability Light Link Methodology Methods Microscopy Midbrain structure Modeling Morphogenesis Neuraxis Optics Pattern Physiologic pulse Population Positioning Attribute Primordium Proliferating Property Proteins Reagent Regulator Genes Reporter Resolution Scanning Signal Transduction Source Specific qualifier value Speed Staging System Technology Tectum Mesencephali Testing Time Transgenic Organisms Vertebrates Zebrafish base cell motility cellular imaging design developmental disease fluorophore gastrulation hindbrain in utero insight live cell imaging malformation man neural plate novel prospective research study spectrograph tool two-photon

项目摘要

DESCRIPTION (provided by applicant): During development, the vertebrate central nervous system is formed by partitioning proliferating primordial cell populations into compartments and gradually refining these populations into distinct functional sub- regions. In the presumptive midbrain and hindbrain, these compartments are lineage restricted by a sharp boundary interface ensuring that cells from these regions do not mix and that they receive region specific signals to properly develop into the tectum and cerebellum, respectively. These region specific signals originate from an organizer, the isthmic organizer, positioned at the midbrain-hindbrain boundary (MHB). Though many of the key regulatory signals of the isthmic or MHB organizer are known, our current understanding of how these regions are initiated, formed, and maintained remains poor. Furthermore, developmental defects of these regions and brain stem have been linked to conditions such as intellectual disabilities, autism, and Chiari malformation. A detailed study of human brain formation in utero to better understand its development and developmental disorders is problematic ethically, but the MHB is one of three evolutionarily conserved organizers of the vertebrate brain. Thus, understanding how the MHB forms in model species such as zebra fish will likely provide valuable insights into human brain development, particularly the tectum and cerebellum. How- ever, the technological capability and methodology to characterize the dynamics and interactions of developmental lineages in the early zebra fish embryo has yet to be demonstrated. This project will develop the technologies and reagents that address fundamental challenges to characterizing the dynamics and interactions of distinct developmental lineages in the zebra fish midbrain and hindbrain and that may be extended to other functional regions and developmental stages and, potentially, other model species. The MHB forms in the zebra fish at an early developmental stage of rapid cell movement through 3D space. At this early developmental stage, cells and their identities are morphologically indistinct and may only be differentiated by gene expression. Thus, a fundamental challenge in characterizing dynamics and interactions of presumptive midbrain and hindbrain cells in MHB formation has been the capability to acquire volumetric images in developing embryos fast enough and with high enough spatial resolution to resolve single cells and to identify the imaged cells by the expression of key regulatory genes. Aim 1 of this project generates transgenic zebra fish with fluorescent reporters that fiducially mark future midbrain and hindbrain cells. Aim 2 of this project develops novel light sheet microscopy based on ultra-short optical pulses. Time-lapse, multimolecular light sheet microscopy will be used to test two main hypotheses: the initially overlapping boundary of mid- brain and hindbrain cells is sharpened through cell sorting; and lineage restriction properties of the MHB are established coincidentally with sharpening of the boundary interface.

描述（由适用提供）：在开发过程中，脊椎动物中枢神经系统是通过将增殖原始细胞种群划分为隔室的，并逐渐将这些种群逐步精制成不同功能子区域的形成。在假定的中脑和后脑脑中，这些隔室受到尖锐的边界界面的限制，可确保这些区域的细胞不混合，并分别接收特定区域的信号以正确地发育到tectum和cerebellum中。这些区域特定的信号源自组织者，地缘组织者，位于中脑 - 脑边界（MHB）。尽管已知静脉或MHB组织者的许多关键监管信号，但我们目前对这些区域如何启动，形成和维护的理解仍然很差。此外，这些区域和脑干的发育缺陷与智力障碍，自闭症和Chiari畸形等疾病有关。对子宫内人脑形成的详细研究，以更好地了解其发育和发育障碍是有问题的，但MHB是脊椎动物大脑的三个进化组成的组织者之一。这就是了解模型物种（例如斑马鱼）中如何形成的MHB可能会为人类脑发育，尤其是tectum和小脑提供宝贵的见解。但是，在早期斑马鱼胚胎中表征发育谱系的动态和相互作用的技术能力和方法尚未得到证明。该项目将开发针对基本挑战的技术和试剂，以表征斑马鱼中脑和后脑在斑马鱼中脑和后脑中不同发育谱系的动态和相互作用，并且可能扩展到其他功能区域和发育阶段以及潜在的其他模型物种。 MHB在经过3D空间的快速细胞运动的早期发育阶段在斑马鱼中形成。在这个早期发育阶段，细胞及其身份在形态上不明显并且只能通过基因表达来区分。这是在表征MHB形成中假定的中脑和后脑细胞的动力学和相互作用方面的基本挑战，这是在足够快地开发胚胎并具有足够高的空间分辨率来解决单个细胞并通过关键调节基因表达鉴定成像细胞的足够高空间分辨率的体积图像的能力。该项目的目标1产生了带有荧光记者的转基因斑马鱼，可信仰未来的中脑和后脑细胞。该项目的目标2基于超短光脉冲开发新型的轻板显微镜。延时的多分子光显微镜将用于检验两个主要假设：中脑和后脑细胞的最初重叠边界通过细胞分选锐化； MHB的谱系限制特性是在边界界面锐化的巧合建立的。