Name: zebrafish brain ventricle injection
Uploaded: 2009-04-06T00:00:00
Description: Watch this Scientific Journal Video about Zebrafish Brain Ventricle Injection at JoVE.com

作者要感谢劳拉安妮洛厄谁开发这项技术在实验室中。 HS是由NIHMH70926支持。詹妮弗Gutzman支持CSBi /默克博士后。

协议第1部分：准备显微注射<ol><li>准备使用萨特仪器针拉马拉毛细管针注射。 </li><li>用荧光染料（如得克萨斯州红葡聚糖）填写针。 </li><li>芒针在显微和显微注射器具。 </li><li>切成适当大小的针的角度，创建一个斜角提示。 </li><li>测量液滴大小，并检查它每注入1 - 2nl之间的石油。 </li></ol>第2部分：准备胚胎<ol><li>注入30分钟，比利益阶段的早期胚胎。例如...

<ol>
	<li>Gutzman, J. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Formation+of+the+zebrafish+midbrain-hindbrain+boundary+constriction+requires+laminin-dependent+basal+constriction.">Formation of the zebrafish midbrain-hindbrain boundary constriction requires laminin-dependent basal constriction.</a> Mech. Dev. 125, 974-983 (2008).</li><li>Lowery, L. A. <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><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>Kimmel, C. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Stages+of+embryonic+development+of+the+zebrafish.">Stages of embryonic development of the zebrafish.</a> Dev. Dyn. 203, 253-310 (1995).</li><li>Westerfield, M. <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 Zebrafish Book: A guide for the laboratory use of zebrafish. , (1995).</li></ol>

在这段视频中，我们演示了如何注入荧光染料（得克萨斯州红葡聚糖），斑马鱼脑室。这种方法是用来在脑室周围神经上皮空间可视化，并确定脑室空间的形状以及周围脑组织的形状是非常有用。它使我们能够更好地了解脑室形成，随着时间的推移活胚胎脑的形态发生过程。这项技术是一个极好的工具，为研究脑室形成缺陷和脑的形态发生突变体的初步鉴定和研究。

<!DOCTYPE html PUBLIC "-//W3C//DTD XHTML 1.0 Transitional//EN" "http://www.w3.org/TR/xhtml1/DTD/xhtml1-transitional.dtd">
<html xmlns="http://www.w3.org/1999/xhtml" xml:lang="en" lang="en">	
		
		<div>
		<table border="1">
		<thead>
		<tr>
		<th>Material Name</th>
		<th>Type</th>
		<th>Company</th>
		<th>Catalogue Number</th>
		<th>Comment</th>
		</tr>
		</thead>
		<tbody>
		
<tr>
<td>Forceps</td>
<td>Tool</td>
<td>Fine Science Tools</td>
<td>11252-20</td>
<td>&nbsp;</td>
<tr>
<td>Capillary Tubes (needles)</td>
<td>Tools</td>
<td>Frederick Haer and Co.</td>
<td>30-30-1</td>
<td>&nbsp;</td>
<tr>
<td>Agarose-Coated dishes</td>
<td>Tools</td>
<td>Agarose (Seakem GTG; Lonza) Dishes (Corning) </td>
<td>Agarose (50027) Dishes (430166) </td>
<td>Make agarose approximately 2-3 mm deep in the dish depending on the stage of embryo you are injecting</td>
</tr>
<tr>
<td>Dissecting Microscope</td>
<td>Microscope</td>
<td>Zeiss Stereomicroscope Stemi SV 6 </td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<tr>
<td>Microscope Camera with Fluorescence capabilities and with image capture software</td>
<td>Tools</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<tr>
<td>1-200 &mu; pipette tips</td>
<td>Tools</td>
<td>Tip One, US Scientific</td>
<td>1111-0806</td>
<td>These are the perfect size to poke the holes in the agarose and have the embryos fit perfectly.</td>
</tr>
<tr>
<td>Photoshop</td>
<td>Image Analysis</td>
<td>Adobe</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<tr>
<td>Embryo Loop</td>
<td>Tools</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>Make these yourself with a glass pipette and hair or fishing line and seal with beeswax.</td>
</tr>
<tr>
<td>Microinjector</td>
<td>Tools</td>
<td>Medical Systems Research Products Harvard Apparatus</td>
<td>PL1-100</td>
<td>&nbsp;</td>
<tr>
<td>Needle Puller</td>
<td>Tools</td>
<td>Sutter Instruments</td>
<td>&nbsp;</td>
<td>Settings (Heat 785, Pull 70, Velocity 40, Time 70).</td>
</tr>
<tr>
<td>Micromanipulator</td>
<td>Tools</td>
<td>Narishige</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<tr>
<td>Fluorescent dye Texas-Red Dextran 70,000MW</td>
<td>Reagent</td>
<td>Molecular Probes</td>
<td>D1830</td>
<td>Other Molecular weights are available if desired.</td>
</tr>
<tr>
<td>Tricaine (3-aminobenzoic acid ethyl ester)</td>
<td>Reagent</td>
<td>Sigma</td>
<td>A-5040</td>
<td>&nbsp;</td>
<tr>
<td>Embryo Media</td>
<td>Reagent</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>According to Westerfield (5)</td>
</tr>		</tbody>
		</table>
		</div>

zebrafish brain ventricle injection

正确的大脑心室形成胚胎大脑发育过程中是必需的正常的脑功能。脑室高度保守的大脑内充满脑脊液腔。在斑马鱼，神经管形成后，神经上皮经历了一系列的收缩和褶皱填充液体造成脑室形成。为了了解心室形成过程中，上皮形状的变化，在同一时间发生，我们需要一个方法来可视化脑组织相比，心室空间。然而，透明的斑马鱼胚胎的性质，使得它很难区分脑室空间组织。因此，我们制定了脑室注射技术脑室空间充满了一种荧光染料和明和荧光显微镜成像。明和荧光图像，然后处理，并在Photoshop中叠加。这种技术允许可视化与荧光染料心室的空间，比较明形象的神经上皮的形状。在斑马鱼的脑脑室注射可以采用18小时受精后通过早期的幼体阶段。我们已经广泛使用这种技术在脑室形成和形态发生的研究以及在脑的形态发生突变体（1-3）特征。

Watch this Scientific Journal Video about Zebrafish Brain Ventricle Injection at JoVE.com

Zebrafish Brain Ventricle Injection

神经管形成后，神经上皮收缩和皱褶而胚胎脑脊液（eCSF）形成的胚胎脑室管填充。我们开发这个脑室注射技术，以更好地可视化的充满液体的空间，相反，在一个活的胚胎神经上皮形状。

斑马鱼脑室注射液

Proper brain ventricle formation during embryonic brain development is required for normal brain function.  Brain ventricles are the highly conserved ...

Research

JoVE Journal

Biology

Live Imaging of the Zebrafish Embryonic Brain by Confocal Microscopy

现场共聚焦显微镜对斑马鱼的胚胎脑成像

In this video, we demonstrate the method our lab has developed to analyze the cell shape changes and rearrangements required to bend and fold the ...

科研

生物学

Retro-orbital Injection in Adult Zebrafish

在成年斑马鱼的眼窝射出

Drug treatment of whole animals is an essential tool in any model system for pharmacological and chemical genetic studies. Intravenous (IV) injection is ...

Transplantation of GFP-expressing Blastomeres for Live Imaging of Retinal and Brain Development in Chimeric Zebrafish Embryos

移植绿色荧光蛋白表达对视网膜和大脑发育的实时成像嵌合斑马鱼胚胎的卵裂球

Cells change extensively in their locations and property during embryogenesis. These changes are regulated by the interactions between the cells and their ...

Intraperitoneal Injection into Adult Zebrafish

腹腔注射到成年斑马鱼

A convenient method for chemically treating zebrafish is to introduce the reagent into the tank water, where it will be taken up by the fish. However, ...

Direct Delivery of MIF Morpholinos Into the Zebrafish Otocyst by Injection and Electroporation Affects Inner Ear Development

通过注射和电穿孔的斑马鱼Otocyst的MIF Morpholinos直接运抵影响内耳发展

In recent years, electroporation has become a popular technique for in vivo transfection of DNA, RNA, and morpholinos into various tissues, including the ...

In vivo Electroporation of Morpholinos into the Adult Zebrafish Retina

In vivo Electroporation of Morpholinos into the Adult Zebrafish Retina

在体内电穿孔Morpholinos到成年斑马鱼视网膜

Many devastating inherited eye diseases result in progressive and irreversible blindness because humans cannot regenerate dying or diseased retinal ...

In vivo Electroporation of Morpholinos into the Regenerating Adult Zebrafish Tail Fin

In vivo Electroporation of Morpholinos into the Regenerating Adult Zebrafish Tail Fin

再生的成年斑马鱼尾鳍在吗啉代入体内电穿孔

Certain species of urodeles and teleost fish can regenerate their tissues. Zebrafish have become a widely used model to study the spontaneous regeneration ...

Gavaging Adult Zebrafish

Gavaging成年斑马鱼

The zebrafish has become an important in vivo model in biomedical research. Effective methods must be developed and utilized to deliver compounds or ...

Reverse Genetic Morpholino Approach Using Cardiac Ventricular Injection to Transfect Multiple Difficult-to-target Tissues in the Zebrafish Larva

反向遗传吗啉使用方法心脏心室射出转染多难靶组织中的斑马鱼幼虫

The zebrafish is an important model to understand the cell and molecular biology of organ and appendage regeneration. However, molecular strategies to ...

Nephrotoxin Microinjection in Zebrafish to Model Acute Kidney Injury

肾毒素显微注射斑马鱼到模型急性肾损伤

The kidneys are susceptible to harm from exposure to chemicals they filter from the bloodstream. This can lead to organ injury associated with a rapid ...