Name: manual drainage of the zebrafish embryonic brain ventricles
Uploaded: 2012-12-16T12:00:00
Description: Watch this Scientific Journal Video about Manual Drainage of the Zebrafish Embryonic Brain Ventricles at JoVE.com

这项工作是由美国国家精神卫生研究所和美国国家科学基金会的支持。特别感谢仁Gutzman博士，博士阿曼达迪金森和其他许多有益的讨论和建设性的批评的的西伯实验室成员，并奥利维尔Paugois专家鱼的饲养。

1。准备显微注射针和细胞电<ol><li>根据制造商的指示，填写Eppendorf公司CellTram油微量注射器装置与矿物油。 </li><li>准备显微注射针拉毛细管萨特仪器针拔出器。 </li><li>微操作机器人连接到Eppendorf公司CellTram上安装针。 </li><li>小心打破针尖。为针尖大小均匀，用千分尺测量，或比较的参考针。 </li><li>填写针的长度与油，使用CellTram，推动油价下跌的长度，小心，以避免任何气泡。 </li></ol><p class="...

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	<li>Chodobski, A., Szmydynger-Chodobska, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Choroid+plexus:+target+for+polypeptides+and+site+of+their+synthesis.">Choroid plexus: target for polypeptides and site of their synthesis.</a> Microsc. Res. Tech. 52, 65-82 (2001).</li><li>Redzic, Z. B., Preston, J. E., Duncan, J. A., Chodobski, A., Szmydynger-Chodobska, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+choroid+plexus-cerebrospinal+fluid+system:+from+development+to+aging.">The choroid plexus-cerebrospinal fluid system: from development to aging.</a> Curr. Top. Dev. Biol. 71, 1-52 (2005).</li><li>Brown, P. D., Davies, S. L., Speake, T., Millar, I. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Molecular+mechanisms+of+cerebrospinal+fluid+production.">Molecular mechanisms of cerebrospinal fluid production.</a> Neuroscience. 129, 957-970 (2004).</li><li>Praetorius, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Water+and+solute+secretion+by+the+choroid+plexus.">Water and solute secretion by the choroid plexus.</a> Pflugers Arch. 454, 1-18 (2007).</li><li>Speake, T., Whitwell, C., Kajita, H., Majid, A., Brown, P. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mechanisms+of+CSF+secretion+by+the+choroid+plexus.">Mechanisms of CSF secretion by the choroid plexus.</a> Microsc. Res. Tech. 52, 49-59 (2001).</li><li>Garcia-Lecea, M., Kondrychyn, I., Fong, S. H., Ye, Z. R., Korzh, V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=In+vivo+analysis+of+choroid+plexus+morphogenesis+in+zebrafish.">In vivo analysis of choroid plexus morphogenesis in zebrafish.</a> PLoS One. 3, e3090 (2008).</li><li>Welss, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Secretory+activity+of+the+inner+layer+of+the+embryonic+mid-brain+of+the+chick,+as+revealed+by+tissue+culture.">Secretory activity of the inner layer of the embryonic mid-brain of the chick, as revealed by tissue culture.</a> The Anatomical Record. 58, 299-302 (1934).</li><li>Saunders, N. R., Habgood, M. D., Dziegielewska, K. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Barrier+mechanisms+in+the+brain,+II.+Immature+brain.">Barrier mechanisms in the brain, II. Immature brain.</a> Clin. Exp. Pharmacol. Physiol. 26, 85-91 (1999).</li><li>Zheng, W., Chodobski, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> The blood-cerebrospinal fluid barrier. , (2005).</li><li>Salehi, Z., Mashayekhi, F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+role+of+cerebrospinal+fluid+on+neural+cell+survival+in+the+developing+chick+cerebral+cortex:+an+in+vivo+study.">The role of cerebrospinal fluid on neural cell survival in the developing chick cerebral cortex: an in vivo study.</a> Eur. J. Neurol. 13, 760-764 (2006).</li><li>Parada, C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Embryonic+cerebrospinal+fluid+collaborates+with+the+isthmic+organizer+to+regulate+mesencephalic+gene+expression.">Embryonic cerebrospinal fluid collaborates with the isthmic organizer to regulate mesencephalic gene expression.</a> J. Neurosci. Res. 82, 333-345 (2005).</li><li>Mashayekhi, F., Salehi, Z. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+importance+of+cerebrospinal+fluid+on+neural+cell+proliferation+in+developing+chick+cerebral+cortex.">The importance of cerebrospinal fluid on neural cell proliferation in developing chick cerebral cortex.</a> Eur. J. Neurol. 13, 266-272 (2006).</li><li>Martin, C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=FGF2+plays+a+key+role+in+embryonic+cerebrospinal+fluid+trophic+properties+over+chick+embryo+neuroepithelial+stem+cells.">FGF2 plays a key role in embryonic cerebrospinal fluid trophic properties over chick embryo neuroepithelial stem cells.</a> Dev. Biol. 297, 402-416 (2006).</li><li>Martin, C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Early+embryonic+brain+development+in+rats+requires+the+trophic+influence+of+cerebrospinal+fluid.">Early embryonic brain development in rats requires the trophic influence of cerebrospinal fluid.</a> Int. J. Dev. Neurosci. 27, 733-740 (2009).</li><li>Gato, A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Embryonic+cerebrospinal+fluid+regulates+neuroepithelial+survival,+proliferation,+and+neurogenesis+in+chick+embryos.">Embryonic cerebrospinal fluid regulates neuroepithelial survival, proliferation, and neurogenesis in chick embryos.</a> Anat. Rec. A Discov. Mol. Cell Evol. Biol. 284, 475-484 (2005).</li><li>Desmond, M. E., Levitan, M. L., Haas, A. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Internal+luminal+pressure+during+early+chick+embryonic+brain+growth:+descriptive+and+empirical+observations.">Internal luminal pressure during early chick embryonic brain growth: descriptive and empirical observations.</a> Anat. Rec. A Discov. Mol. Cell Evol. Biol. 285, 737-747 (2005).</li><li>Alonso, M. I., Martin, C., Carnicero, E., Bueno, D., Gato, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cerebrospinal+fluid+control+of+neurogenesis+induced+by+retinoic+acid+during+early+brain+development.">Cerebrospinal fluid control of neurogenesis induced by retinoic acid during early brain development.</a> Dev. Dyn. 240, 1650-1659 (2011).</li><li>Miyan, J. A., Zendah, M., Mashayekhi, F., Owen-Lynch, P. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cerebrospinal+fluid+supports+viability+and+proliferation+of+cortical+cells+in+vitro,+mirroring+in+vivo+development.">Cerebrospinal fluid supports viability and proliferation of cortical cells in vitro, mirroring in vivo development.</a> Cerebrospinal Fluid Res. 3, 2 (2006).</li><li>Mashayekhi, F., Bannister, C. M., Miyan, J. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Failure+in+cell+proliferation+in+the+germinal+epithelium+of+the+HTx+rats.">Failure in cell proliferation in the germinal epithelium of the HTx rats.</a> Eur. J. Pediatr. Surg. 11, S57-S59 (2001).</li><li>Lehtinen, M. K., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+cerebrospinal+fluid+provides+a+proliferative+niche+for+neural+progenitor+cells.">The cerebrospinal fluid provides a proliferative niche for neural progenitor cells.</a> Neuron. 69, 893-905 (2011).</li><li>Zappaterra, M. D., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+comparative+proteomic+analysis+of+human+and+rat+embryonic+cerebrospinal+fluid.">A comparative proteomic analysis of human and rat embryonic cerebrospinal fluid.</a> J. Proteome. Res. 6, 3537-3548 (2007).</li><li>Parvas, M., Parada, C., Bueno, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+blood-CSF+barrier+function+controls+embryonic+CSF+protein+composition+and+homeostasis+during+early+CNS+development.">A blood-CSF barrier function controls embryonic CSF protein composition and homeostasis during early CNS development.</a> Dev. Biol. 321, 51-63 (2008).</li><li>Parada, C., Gato, A., Bueno, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mammalian+embryonic+cerebrospinal+fluid+proteome+has+greater+apolipoprotein+and+enzyme+pattern+complexity+than+the+avian+proteome.">Mammalian embryonic cerebrospinal fluid proteome has greater apolipoprotein and enzyme pattern complexity than the avian proteome.</a> J. Proteome Res. 4, 2420-2428 (2005).</li><li>Gato, A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Analysis+of+cerebro-spinal+fluid+protein+composition+in+early+developmental+stages+in+chick+embryos.">Analysis of cerebro-spinal fluid protein composition in early developmental stages in chick embryos.</a> J. Exp. Zool. A Comp. Exp. Biol. 301, 280-289 (2004).</li><li>Westerfield, M., Sprague, J., Doerry, E., Douglas, S., Grp, Z. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+Zebrafish+Information+Network+(ZFIN):+a+resource+for+genetic,+genomic+and+developmental+research.">The Zebrafish Information Network (ZFIN): a resource for genetic, genomic and developmental research.</a> Nucleic Acids Res. 29, 87-90 (2001).</li><li>Gutzman, J. H., Sive, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Zebrafish+Brain+Ventricle+Injection.">Zebrafish Brain Ventricle Injection.</a> J. Vis. Exp. (26), e1218 (2009).</li><li>Parada, C., Gato, A., Bueno, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=All-trans+retinol+and+retinol-binding+protein+from+embryonic+cerebrospinal+fluid+exhibit+dynamic+behaviour+during+early+central+nervous+system+development.">All-trans retinol and retinol-binding protein from embryonic cerebrospinal fluid exhibit dynamic behaviour during early central nervous system development.</a> Neuroreport. 19, 945-950 (2008).</li><li>Parada, C., Escola-Gil, J. C., Bueno, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Low-density+lipoproteins+from+embryonic+cerebrospinal+fluid+are+required+for+neural+differentiation.">Low-density lipoproteins from embryonic cerebrospinal fluid are required for neural differentiation.</a> J. Neurosci. Res. 86, 2674-2684 (2008).</li><li>Kramer-Zucker, A. G., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cilia-driven+fluid+flow+in+the+zebrafish+pronephros,+brain+and+Kupffer's+vesicle+is+required+for+normal+organogenesis.">Cilia-driven fluid flow in the zebrafish pronephros, brain and Kupffer's vesicle is required for normal organogenesis.</a> Development. 132, 1907-1921 (2005).</li><li>Lowery, L. A., Sive, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Initial+formation+of+zebrafish+brain+ventricles+occurs+independently+of+circulation+and+requires+the+nagie+oko+and+snakehead/atp1a1a.1+gene+products.">Initial formation of zebrafish brain ventricles occurs independently of circulation and requires the nagie oko and snakehead/atp1a1a.1 gene products.</a> Development. 132, 2057-2067 (2005).</li><li>Lowery, L. A., De Rienzo, G., Gutzman, J. H., Sive, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Characterization+and+classification+of+zebrafish+brain+morphology+mutants.">Characterization and classification of zebrafish brain morphology mutants.</a> Anat. Rec. (Hoboken). 292, 94-106 (2009).</li></ol>

使用这种技术的手动排水eCSF的从斑马鱼脑室用于确定eCSF在开发过程中的要求将是有益的。此外，这种技术将允许在胚胎发育过程中描述的eCSF蛋白更新。在此期间，不同的蛋白质的鉴定将能够进一步调查到CSF和大脑发育过程中的潜在作用的功能。在脊椎动物中，在eCSF（FGF2，IGF2，视黄酸和载脂蛋白）的一些因素有证明作用在神经上皮细胞存活，增殖和神经细胞13,17,20,23,27,28。然而，似乎有...

<table border="1"><tbody><tr><td> 试剂名称 </td><td> 公司 </td><td> 目录编号 </td></tr><tr><td> Eppendorf公司CellTram油</td><td> Eppendorf公司</td><td> 516 000.025 </td></tr><tr><td>矿物油</td><td>西格玛</td><td> M8410 </td></tr><tr><td>三卡因粉</td><td>西格玛</td><td> A5040 </td></tr><tr><td>毛细管</td><td> FHC公司</td><td> 30-30-1 </td></tr></tbody></table>

manual drainage of the zebrafish embryonic brain ventricles

脑脊液（CSF）是一种富含蛋白质的脑室内所含的流体。这是目前早期脊椎动物的胚胎发育过程中，坚持在整个生命。成人CSF认为，以减轻大脑，排除废物，并进行分泌的分子1,2。在成人和年龄较大的胚胎中，大多数的CSF是由脉络丛，一系列的高度血管化的分泌位于脑室3-5相邻的区域。在斑马鱼，脉络丛完全形成受精后144小时（HPF）6。在此之前，在包括鼠标，一个显着量的胚胎CSF（eCSF）的斑马鱼和其他脊椎动物胚胎是本。这些数据和研究表明，在鸡的神经上皮分泌在开发的早期，可能是之前的主要来源eCSF发展脉络丛7。 eCSF包含了大约3倍以上的蛋白质吨韩成人CSF，这表明它可能具有重要作用，在发展8,9。研究表明，在小鸡和老鼠分泌因子eCSF，流体压力，或这些的组合，是非常重要的神经发生，基因表达，细胞增殖和细胞生存的神经上皮10-20中。人，大鼠，小鼠和小鸡eCSF的蛋白质组学分析，已经确定了许多蛋白质​​可能是必要的CSF功能。这些包括胞外基质成分，载脂蛋白，渗透压调节蛋白和蛋白参与细胞死亡和增殖21-24。然而，复杂的功能在很大程度上是未知的eCSF。 我们已经开发出一种方法，用于除去从斑马鱼脑室eCSF，从而允许识别eCSF组件，并在开发过程中进行分析的eCSF规定的。虽然可以采集更多eCSF，从其他脊椎动物系统W第i个较大的胚胎，eCSF可以收集到的斑马鱼发育的早期阶段，与遗传或环境条件下，导致脑室体积异常或形态。拆卸和回收的eCSF允许质谱分析，调查eCSF功能，并重新选择因素破入脑室分析它们的功能。因此，斑马鱼早期胚胎的可访问性可以在开发过程中的eCSF功能进行详细的分析。

一个排水脑室的一个例子示出在图1B-C。作为他们缺乏eCSF的（ 图1B和C），脑室倒塌。可以看出在背侧的图像（ 图1B-C，以及图2A-D）的后脑神经上皮确实保留其特征的形态，而且似乎是开放，尽管缺乏eCSF可能是由于强大的铰接点。然而，横向次（ 图2A&#39;-D&#39;）表明已后脑脑室沥干，的存在下，一薄的柔性顶板上皮折叠腹侧一致。在?...

Watch this Scientific Journal Video about Manual Drainage of the Zebrafish Embryonic Brain Ventricles at JoVE.com

Manual Drainage of the Zebrafish Embryonic Brain Ventricles

我们提出一个方法，收集脑脊液（CSF），并建立系统，缺乏的胚胎斑马鱼大脑脑室系统内脑脊液。 CSF的组成及其胚胎大脑发育过程中的要求，这使得进一步的检查。

手动排水的斑马鱼胚胎脑室

Cerebrospinal fluid  (CSF) is a protein rich fluid contained within the brain ventricles. It is  present during early vertebrate embryonic development and ...

Research

JoVE Journal

Neuroscience

An Assay for Permeability of the Zebrafish Embryonic Neuroepithelium

的斑马鱼胚胎神经上皮通透性的测定

The brain ventricular system is conserved among vertebrates and is composed of a series of interconnected cavities called brain ventricles, which form ...

科研

神经科学

Micromanipulation of Gene Expression in the Adult Zebrafish Brain Using Cerebroventricular Microinjection of Morpholino Oligonucleotides

显微吗啉代寡核苷酸脑室微量注射成年斑马鱼大脑中的基因表达

Manipulation of gene expression in tissues is  required to perform functional studies. In this paper, we demonstrate the  cerebroventricular ...

Patch Clamp Recordings from Embryonic Zebrafish Mauthner Cells

从胚胎斑马鱼的毛特纳细胞膜片钳记录

Mauthner cells (M-cells) are large  reticulospinal neurons located in the hindbrain of teleost fish. They are key  neurons involved in a characteristic ...

A Comprehensive Protocol for Manual Segmentation of the Medial Temporal Lobe Structures

一个全面协议的内侧颞叶结构的手动分割

The present paper describes a comprehensive protocol for manual tracing of the set of brain regions comprising the medial temporal lobe (MTL): amygdala, ...

Isolation and Culture of Endothelial Cells from the Embryonic Forebrain

从胚胎前脑内皮细胞的分离和培养

Embryonic brain endothelial cells can serve as an important tool in the study of angiogenesis and neurovascular development and interactions. The two ...

Electrophysiological Recording in the Brain of Intact Adult Zebrafish

在完整的成年斑马鱼大脑电生理记录

Previously, electrophysiological studies in adult zebrafish have been limited to slice preparations or to eye cup preparations and electrorentinogram ...

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 ...

Conditional Genetic Transsynaptic Tracing in the Embryonic Mouse Brain

有条件的遗传跨突触标记在胚胎小鼠脑

Anatomical path tracing is of pivotal importance to decipher the relationship between brain and behavior. Unraveling the formation of neural circuits ...