Name: time-lapse live imaging of clonally related neural progenitor cells in the developing zebrafish forebrain
Uploaded: 2011-04-06T13:00:00
Description: Watch this Scientific Journal Video about Time-lapse Live Imaging of Clonally Related Neural Progenitor Cells in the Developing Zebrafish Forebrain at JoVE.com

这项工作是支持theNIH grantNS042626。我们感谢库尔特索恩和加州大学旧金山分校的尼康影像中心协助与成像。

1。斑马鱼胚胎的制备<ol><li>前两天电，成立了野生型使用的分隔独立的男性从女性的鱼交配笼。 </li><li>一天前，电，拉在交配前一天的笼子的分隔。 </li><li>收集胚胎和孵化50个受精胚胎在30毫升胚胎含有0.003％苯基硫脲嘧啶（PTU）的媒介为28.5 ° C。 </li><li>在一天的电，dechorionate手动精细镊子（INOX 5，杜蒙电子，瑞士），当胚胎到达的22hpf发育阶段的胚胎。然后转移胚胎到10ml电振铃中的 3...

<ol>
	<li>Temple, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+development+of+neural+stem+cells.">The development of neural stem cells.</a> Nature. 414, 112-117 (2001).</li><li>G#246;tz, M., Huttner, W. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+cell+biology+of+neurogenesis.">The cell biology of neurogenesis.</a> Nat. Rev. Mol. Cell Biol. 6, 777-788 (2005).</li><li>Cerda, G. A., Thomas, J. E., Allende, M. L., Karlstrom, R. O., Palma, V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Electroporation+of+DNA,+RNA,+and+morpholinos+into+zebrafish+embryos.">Electroporation of DNA, RNA, and morpholinos into zebrafish embryos.</a> Methods. 39, 207-211 (2006).</li><li>Hendricks, M., Jesuthasan, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Electroporation-based+methods+for+in+vivo,+whole+mount+and+primary+culture+analysis+of+zebrafish+brain+development.">Electroporation-based methods for in vivo, whole mount and primary culture analysis of zebrafish brain development.</a> Neural. Dev. 2, 6-6 (2007).</li><li>Tawk, M., Bianco, I. H., Clarke, J. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Focal+electroporation+in+zebrafish+embryos+and+larvae.">Focal electroporation in zebrafish embryos and larvae.</a> Methods Mol. Biol. 546, 145-151 (2009).</li><li>Tawk, M., Tuil, D., Torrente, Y., Vriz, S., Paulin, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=High-efficiency+gene+transfer+into+adult+fish:+a+new+tool+to+study+fin+regeneration.">High-efficiency gene transfer into adult fish: a new tool to study fin regeneration.</a> Genesis. 32, 27-31 (2002).</li></ol>

在这段视频中，我们展示了一个时间推移在一个发展中的斑马鱼的前脑的克隆水平的神经祖细胞的实时成像的方法。我们测试和修改现有的电协议 4-6的基因和荧光标签个别的神经祖细胞。因为只有少数细胞地广人稀一个相对的低电压电标记，克隆相关的子代细胞组成，可以建立一个谱系树。除了EGFP的，神经祖细胞可以subcellularly各种荧光蛋白标记，只需更换与其他荧光标记的EGFP。例如，?...

time-lapse live imaging of clonally related neural progenitor cells in the developing zebrafish forebrain

神经祖细胞的分裂，迁移和分化的精确的模式是正确的大脑发育和功能1,2的关键。为了了解神经祖细胞在体内环境中的复杂行为，时间推移，一个完整的大脑中的神经祖细胞的实时成像是至关重要的。在这段视频中，我们利用斑马鱼胚胎的独特功能，以可视化的前脑的神经祖细胞在体内的发展。我们使用电基因和人口稀少的标签个别的神经祖细胞。简言之，构造编码荧光标记的DNA注射到受精后22小时（HPF），斑马鱼的胚胎和电脉冲，立即交付的前脑脑室。六个小时后，电穿孔的斑马鱼的胚胎被安装在玻璃底培养皿低熔点琼脂糖。荧光标记的神经祖细胞，然后成像下共聚焦显微镜使用水浸渍物镜固定的时间间隔36小时。本方法提供了一种方式来获得脑的神经祖细胞在体内发展的见解，并可以应用到中央神经系统的斑马鱼胚胎的其他部分。

Watch this Scientific Journal Video about Time-lapse Live Imaging of Clonally Related Neural Progenitor Cells in the Developing Zebrafish Forebrain at JoVE.com

Time-lapse Live Imaging of Clonally Related Neural Progenitor Cells in the Developing Zebrafish Forebrain

本视频演示了一种方法，这需要电和共聚焦显微镜相结合的优势，发展中的斑马鱼的前脑执行个别的神经祖细胞的实时成像。<em&gt;在体内</em&gt;脑神经祖细胞克隆的发展的分析，可以实现这种方式。

在发展中国家斑马鱼的前脑的神经祖细胞克隆相关的定时实时成像

Precise patterns of division, migration and differentiation of neural progenitor cells are crucial for proper brain development and function1,2. To ...

Research

JoVE Journal

Neuroscience

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Live-cell Imaging of Sensory Organ Precursor Cells in Intact Drosophila Pupae

在完整的感觉器官前体细胞的活细胞成像果蝇蛹

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科研

神经科学

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There is a substantial body of evidence indicating that new functional neurons are constitutively generated from an endogenous pool of neural stem cells ...

Forebrain Electrophysiological Recording in Larval Zebrafish

斑马鱼幼虫的前脑的电生理记录

Epilepsy affects nearly 3 million people in the United States and up to 50 million people worldwide. Defined as the occurrence of spontaneous unprovoked ...

Motor Nerve Transection and Time-lapse Imaging of Glial Cell Behaviors in Live Zebrafish

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In vivo Postnatal Electroporation and Time-lapse Imaging of Neuroblast Migration in Mouse Acute Brain Slices

In vivo Postnatal Electroporation and Time-lapse Imaging of Neuroblast Migration in Mouse Acute Brain Slices

成神经细胞迁移的小鼠急性脑片在体内电穿孔产后和时间推移成像

The subventricular zone (SVZ) is one of the main neurogenic niches in the postnatal brain. Here, neural progenitors proliferate and give rise to ...

Assessing Cell Cycle Progression of Neural Stem and Progenitor Cells in the Mouse Developing Brain after Genotoxic Stress

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Neurons of the cerebral cortex are generated during brain development from different types of neural stem and progenitor cells (NSPC), which form a ...

Live Imaging of Mitosis in the Developing Mouse Embryonic Cortex

有丝分裂中的不同发育阶段小鼠胚胎皮层实时成像

Although of short duration, mitosis is a complex and dynamic multi-step process fundamental for development of organs including the brain. In the ...

Feeder-free Derivation of Neural Crest Progenitor Cells from Human Pluripotent Stem Cells

从人多能神经嵴前体细胞的无饲养推导干细胞

Human pluripotent stem cells (hPSCs) have great potential for studying human embryonic development, for modeling human diseases in the dish and as a ...

Cell Sorting of Neural Stem and Progenitor Cells from the Adult Mouse Subventricular Zone and Live-imaging of their Cell Cycle Dynamics

神经干细胞和祖细胞来自成年小鼠脑室下区细胞分选和实时成像的细胞周期动力学

Neural stem cells (NSCs) in the subventricular zone of the lateral ventricles (SVZ) sustain olfactory neurogenesis throughout life in the mammalian brain. ...