Name: induction of diffuse axonal brain injury in rats based on rotational acceleration
Uploaded: 2020-05-09T15:00:00
Description: Watch this Scientific Journal Video about Induction of Diffuse Axonal Brain Injury in Rats Based on Rotational Acceleration at JoVE.com

作者感谢内森·克利奥林博士（内盖夫本-古里安大学机械工程系）在生物力学测量方面的协助。此外，我们还感谢乌克兰德尼普罗大学生理学系的奥莱娜·斯潘诺夫斯卡教授、玛丽安娜·库舍里亚瓦教授、马克西姆·克里沃诺索夫教授、达里娜·亚库缅科教授和叶夫根尼亚·贡恰雷克教授对我们的讨论给予的支持和有益贡献。

这些实验是在《赫尔辛基和东京宣言》和欧洲共同体《使用实验动物准则》的建议下进行的。这些实验得到了内盖夫本-古里安大学动物护理委员会的批准。
1. 为实验程序准备大鼠
注：选择成年雄性斯普拉格-达利大鼠体重300-350克。
<ol><li>获得机构动物护理和使用委员会的批准，以进行这些实验。</li>
	<li>将大鼠保持在 22 ± 1 °C 的室温下，12 小时照明和 ...

<ol>
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M., Esiri, M. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Axonal+damage:+a+key+predictor+of+outcome+in+human+CNS+diseases.">Axonal damage: a key predictor of outcome in human CNS diseases.</a> Brain. 126, 515-530 (2003).</li><li>Tang-Schomer, M. D., Johnson, V. E., Baas, P. W., Stewart, W., Smith, D. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Partial+interruption+of+axonal+transport+due+to+microtubule+breakage+accounts+for+the+formation+of+periodic+varicosities+after+traumatic+axonal+injury.">Partial interruption of axonal transport due to microtubule breakage accounts for the formation of periodic varicosities after traumatic axonal injury.</a> Experimental Neurology. 233, 364-372 (2012).</li><li>Johnson, V. E., Stewart, W., Smith, D. 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J., 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=Is+beta-APP+a+marker+of+axonal+damage+in+short-surviving+head+injury.">Is beta-APP a marker of axonal damage in short-surviving head injury.</a> Acta Neuropathologica. 92, 608-613 (1996).</li><li>Wilkinson, A., Bridges, L., Sivaloganathan, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Correlation+of+survival+time+with+size+of+axonal+swellings+in+diffuse+axonal+injury.">Correlation of survival time with size of axonal swellings in diffuse axonal injury.</a> Acta Neuropathologicaogica. 98, 197-202 (1999).</li><li>Thompson, H. J., 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=Lateral+fluid+percussion+brain+injury:+a+15-year+review+and+evaluation.">Lateral fluid percussion brain injury: a 15-year review and evaluation.</a> Journal of Neurotrauma. 22, 42-75 (2005).</li><li>Alder, J., Fujioka, W., Lifshitz, J., Crockett, D. P., Thakker-Varia, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Lateral+fluid+percussion:+model+of+traumatic+brain+injury+in+mice.">Lateral fluid percussion: model of traumatic brain injury in mice.</a> Journal of Visualized Experiments. , e3063 (2011).</li><li>Povlishock, J., Marmarou, A., McIntosh, T., Trojanowski, J., Moroi, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Impact+acceleration+injury+in+the+rat:+evidence+for+focal+axolemmal+change+and+related+neurofilament+sidearm+alteration.">Impact acceleration injury in the rat: evidence for focal axolemmal change and related neurofilament sidearm alteration.</a> Journal of Neuropathology &amp; Experimental Neurology. 56, 347-359 (1997).</li><li>Heath, D. L., Vink, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Impact+acceleration-induced+severe+diffuse+axonal+injury+in+rats:+characterization+of+phosphate+metabolism+and+neurologic+outcome.">Impact acceleration-induced severe diffuse axonal injury in rats: characterization of phosphate metabolism and neurologic outcome.</a> Journal of Neurotrauma. 12, 1027-1034 (1995).</li><li>Lighthall, J. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Controlled+cortical+impact:+a+new+experimental+brain+injury+model.">Controlled cortical impact: a new experimental brain injury model.</a> Journal of Neurotrauma. 5, 1-15 (1988).</li><li>Palmer, A. 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该协议描述了DAI的啮齿动物模型。在DAI中，大脑旋转加速导致剪切效应，触发斧和生化变化，导致在渐进过程中失去六角函数。二次斧子变化是由快速的斧子拉伸损伤产生的，其范围和严重程度44、5、105,10是可变的。在主损伤后几到数天内，生化变化将导致斧功能44、5、105,<...

创伤性脑损伤 （TBI） 是美国死亡和残疾的主要原因。结核病占所有与伤害有关的死亡1，22的约30%。TBI的主要原因因年龄组而异，包括跌倒、运动期间高速碰撞、故意自残、机动车碰撞和袭击11、2、3。2,3
脑扩散性斧头损伤（DAI）是一种特殊类型的TBI，由旋转加速、晃动或爆炸损伤引起的大脑在受伤后瞬间不受限制的头部运动44，5，6，7，8。5,6,7,8DAI涉及广泛的斧子损伤，导致长期神经损伤，与不良的结果，沉重的医疗费用，和33-64%的死亡率1,1，2，4，5，9，10，11。2,4,5,9,10,11尽管最近对DAI的发病机制进行了大量研究，但关于最佳治疗方案11、12、13、14,12,13,14仍没有达成共识。
在过去的几十年里，许多实验模型试图准确地复制DAI11、12、15、1612,15,16的不同方面。11然而，与其他焦伤相比，DAI 具有独特的表现方式，这些模型存在局限性。这些以前的模型不仅在白质区域造成斧伤，而且还会导致焦点脑损伤。临床上，DAI伴有微出血，这可能是白质损伤的主要原因。
只有两种动物模型被证明复制DAI的主要临床特征。Gennarelli和他的同事在1982年制造了第一个侧头旋转装置，使用非冲击头旋转加速诱导昏迷与DAI在非人类灵长类动物模型15。这种灵长类动物模型采用控制的单次旋转来加速和减速，在10-20毫秒内将头部通过60°取代。 这项技术能够模拟意识受损和广泛的斧头损伤，类似于在人类大脑中观察到的严重TBI的影响。然而，灵长类动物模型是非常昂贵的44，11，16。11,16部分基于以前的模型，猪模型旋转加速脑损伤是在1994年（罗斯等人）设计，类似的结果14。
这两种动物模型虽然产生了不同的典型病理学的介绍，但大大增加了DAI发病机制的概念。快速头部旋转被公认为诱导DAI的最好方法，啮齿动物为快速头部旋转研究11、16,16提供了一个更便宜的模型。在这里，我们验证了DAI的简单、可复制和可靠的啮齿动物模型，该模型在不造成颅骨骨折或挫伤的情况下造成广泛的白质损伤。目前的模型将使人们更好地了解DAI的病理生理学和更有效的治疗方法的发展。

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	<tbody>
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			<td height="42" style="height:42px;width:216px;">0.01 M sodium citrate</td>
			<td style="width:121px;">SIGMA - ALDRICH</td>
			<td style="width:161px;"></td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">2.5% normal horse serum</td>
			<td style="width:121px;">SIGMA - ALDRICH</td>
			<td style="width:161px;">H0146</td>
			<td>Liquid</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">4 % buffered formaldehyde solution</td>
			<td style="width:121px;"></td>
			<td style="width:161px;"></td>
			<td></td>
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		<tr height="63">
			<td height="63" style="height:63px;width:216px;">Anti-Amyloid Precursor Protein, C - terminal antibodyproduced in rabbit</td>
			<td style="width:121px;">SIGMA - ALDRICH</td>
			<td style="width:161px;">Lot 056M4867V</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">biotinylated secondary antibody</td>
			<td style="width:121px;">Vector</td>
			<td style="width:161px;">BA-1000-1.5</td>
			<td>10 mM sodium phosphate, pH 7.8, 0.15 M NaCl, 0.08% sodium azide, 3 mg/ml bovine serum albumin</td>
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			<td height="21" style="height:21px;width:216px;">bone-cutting forceps</td>
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			<td height="63" style="height:63px;width:216px;">DAB Peroxidase (HRP) Substrate Kit (with Nickel), 3,3&#8217;-diaminobenzidine</td>
			<td style="width:121px;">vector laboratory</td>
			<td style="width:161px;"></td>
			<td></td>
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		<tr height="21">
			<td height="21" style="height:21px;width:216px;">embedding cassettes</td>
			<td style="width:121px;"></td>
			<td style="width:161px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">ethanol 99.9 %</td>
			<td style="width:121px;">ROMICAL</td>
			<td style="width:161px;"></td>
			<td>Flammable Liquid</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">guillotine</td>
			<td style="width:121px;"></td>
			<td style="width:161px;"></td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Hematoxylin</td>
			<td style="width:121px;">SIGMA - ALDRICH</td>
			<td style="width:161px;">H3136-25G</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Hydrogen peroxide solution</td>
			<td style="width:121px;">Millipore</td>
			<td style="width:161px;">88597-100ML-F</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Isofluran, USP 100%</td>
			<td style="width:121px;">Piramamal Critical Care, Inc</td>
			<td style="width:161px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Olympus BX 40 microscope</td>
			<td style="width:121px;">Olympus</td>
			<td style="width:161px;"></td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">paraffine</td>
			<td style="width:121px;">paraplast plus leica biosystem</td>
			<td style="width:161px;"></td>
			<td>Tissue embedding medium</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;">phosphate-buffered saline (PBS)</td>
			<td style="width:121px;">SIGMA - ALDRICH</td>
			<td style="width:161px;">P5368-10PAK</td>
			<td>Contents of one pouch, when dissolved in one liter of distilled or deionized water, will yield 0.01 M phosphate buffered saline (NaCl 0.138 M; KCl - 0.0027 M); pH 7.4, at 25 &#176;C.</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Streptavidin HRP</td>
			<td style="width:121px;">ABCAM</td>
			<td style="width:161px;">ab64269</td>
			<td>Streptavidin-HRP for use with biotinylated secondary antibodies during IHC / immunohistochemistry.</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">xylene</td>
			<td style="width:121px;"></td>
			<td style="width:161px;"></td>
			<td></td>
		</tr>
	</tbody>
</table>

induction of diffuse axonal brain injury in rats based on rotational acceleration

创伤性脑损伤 （TBI） 是导致死亡和残疾的主要原因。扩散性斧伤 （DAI） 是需要住院的很大一部分 TBI 患者的主要伤害机制。DAI 涉及因晃动、旋转或爆炸伤害而造成的广泛斧头损伤，导致快速的斧头拉伸损伤和与功能恢复的长期影响相关的二次斧头变化。从历史上看，没有焦伤的DAI实验模型一直难以设计。在这里，我们验证了DAI的简单、可重复和可靠的啮齿动物模型，该模型不会造成广泛的白质损伤，而不会造成颅骨骨折或挫伤。

表 1说明了协议时间线。DAI模型的死亡率为0%。Mann-Whitney的一项测试表明，15只DAI大鼠的神经缺陷明显大于干预后48小时15只假鼠（Mdn = 1 vs. 0），U = 22.5，p < 0.001，r = 0.78（见表2）。数据以计数为单位进行测量，并呈现为中位数和 25-75 个百分位数范围。
图1显示了脑组织下部具有代表性的显微图。与对照组（67.46 × 30 vs. 0） 相比，?...

Watch this Scientific Journal Video about Induction of Diffuse Axonal Brain Injury in Rats Based on Rotational Acceleration at JoVE.com

Induction of Diffuse Axonal Brain Injury in Rats Based on Rotational Acceleration

该协议验证了大脑扩散性连体损伤 （DAI） 的可靠、易于执行和可重复的啮齿动物模型，这种模型可引起大范围的白质损伤，而不会造成颅骨骨折或挫伤。

基于旋转加速大鼠漫射性无羟基脑损伤的诱导

Traumatic brain injury (TBI) is a major cause of death and disability. Diffuse axonal injury (DAI) is the predominant mechanism of injury in a large ...

创伤性脑损伤是造成死亡和残疾的主要原因之一。漫射斧脑损伤是TBI后发展的普遍的斧损伤。本研究基于旋转加速机制结果TBI，提出感应漫射轴神经脑损伤的模型。选择体重300至350克的成年雄性斯普拉格-道利大鼠。提供鼠周和水广告。在 6：00 AM 到 12：00 PM 之间执行所有实验。该装置被放置在一个沉重的，稳定的实验室桌子上。该器件由以下部件组成：圆柱体、铁重、由圆柱管组成的旋转机构以及头部固定销。重量附着在一根绳子上，并提升到120厘米的高度。自由下降的重量击中激活旋转机构螺栓。大鼠横向头部旋转装置用于将头部从零快速转向90度。诱导后，弥漫性脑损伤，大鼠被转移到康复室。F 是施加在动物头部的力（以公斤为单位）。资本 M 是力的时刻，K 表示动能。小米是下降重量的质量，g 表示重力加速度，h 表示高度（以厘米为单位）和 D 表示耳针之间的距离（以厘米为单位）。为了评估神经缺陷和等级模型缺陷，我们使用大鼠的神经严重性评分。我们评估移动性、截瘫、光束行走任务、光束行走任务失败。光束平衡任务失败宽1.5厘米。稳定性和光束平衡，光束平衡1.5厘米宽和反射力。受伤后48小时，所有大鼠、损伤组和对照组都进行了深度麻醉，并透粘了0.9%的肝素盐水，随后在0.1 M磷酸盐缓冲液盐水中，500毫升的4%甲醛。灌注后，产生斩首，并立即切除大脑，然后在4%甲醛溶液中固定48小时。然后，当小脑和脑干被解剖时，大脑被堵成五毫米的冠冕部分，从嗅球面到视觉皮层。在石蜡嵌入之后，冠状和下垂部分由塔拉穆斯组成。产生β-淀粉样前体蛋白的免疫化学染色。不同时间的不同大鼠组显示在方案上。实验开始时扩散的斧脑损伤。在48小时，神经严重性评分被检查。在48小时时，所有两组都进行了β-APP的免疫化学染色。两个研究组在弥漫性斧脑损伤后48小时出现神经缺陷。曼-惠特尼的一项测试表明，15只弥漫性脑损伤大鼠的神经缺陷明显较大，而干预后48小时内有15只天真大鼠。代表性的显微器显示神经和神经元免疫活动后，孤立的弥漫性脑损伤在大鼠48小时后受伤。使用β-淀粉样前体蛋白检测出较小的细胞串。在虚假组中，未检测到。在我们的工作中，我们开发了一个简单、可重复、可靠的基于旋转加速机制的弥漫轴伤新模型。这样的模型将有助于更好地了解弥漫性脑损伤的病理生理学，并开发更有效的治疗方法。非常感谢你的关注。

Research

JoVE Journal

Neuroscience

Preparation of Oligomeric &beta;-amyloid1-42 and Induction of Synaptic Plasticity Impairment on Hippocampal Slices

Preparation of Oligomeric ÃƒÅ½Ã‚Â²-amyloid1-42 and Induction of Synaptic Plasticity Impairment on Hippocampal Slices

低聚β-淀粉样蛋白的制备 1-42和突触可塑性损伤诱导海马脑片

Impairment of synaptic connections is likely to underlie the subtle amnesic changes occurring at the early stages of Alzheimer s Disease (AD). ...

科研

神经科学

Chromatin Immunoprecipitation from Dorsal Root Ganglia Tissue following Axonal Injury

染色质免疫沉淀从以下轴索损伤的背根神经节组织

Axons in the central nervous system (CNS) do not regenerate while those in the peripheral nervous system (PNS) do regenerate 
to a limited extent after ...

An Ex Vivo Laser-induced Spinal Cord Injury Model to Assess Mechanisms of Axonal Degeneration in Real-time

An Ex Vivo Laser-induced Spinal Cord Injury Model to Assess Mechanisms of Axonal Degeneration in Real-time

一个<em&gt;离体</em&gt;激光诱导脊髓损伤模型，以评估轴索变性的机制在实时

Injured CNS axons fail to regenerate and often retract away from the injury site. Axons spared from the initial injury may later undergo secondary axonal ...

In Vivo Functional Brain Imaging Approach Based on Bioluminescent Calcium Indicator GFP-aequorin

In Vivo Functional Brain Imaging Approach Based on Bioluminescent Calcium Indicator GFP-aequorin

在基于生物发光钙指标GFP的水母体内脑功能成像方法

Functional in vivo imaging has become a powerful approach to study the function and physiology of brain cells and structures of interest. Recently a new ...

Effects of Blast-induced Neurotrauma on Pressurized Rodent Middle Cerebral Arteries

爆炸性神经损伤对压生啮鼠中脑动脉的影响

Though there have been studies on the histopathological and behavioral effects of blast exposure, fewer have been dedicated to blast&#39;s cerebral ...

Activity-based Training on a Treadmill with Spinal Cord Injured Wistar Rats

脊髓损伤 wistar 大鼠跑步机的作业训练

Spinal cord injury (SCI) results in lasting deficits that include both mobility and a multitude of autonomic-related dysfunctions. Locomotor training (LT) ...

Generation of 3-D Collagen-based Hydrogels to Analyze Axonal Growth and Behavior During Nervous System Development

生成基于3-D胶原蛋白的水凝胶,分析神经系统发育过程中的Axonal生长和行为

This protocol uses natural type I collagen to generate three-dimensional (3-D) hydrogel for monitoring and analyzing the axonal growth. The protocol is ...

Assessment of Long-term Depression Induction in Adult Cerebellar Slices

成人脑切片长期抑郁症诱导评估

Synaptic plasticity provides a mechanism for learning and memory. For cerebellar motor learning, long-term depression (LTD) of synaptic transmissions from ...

Advanced Diffusion Imaging in The Hippocampus of Rats with Mild Traumatic Brain Injury

轻度创伤性脑损伤大鼠海马学的高级扩散成像

Mild traumatic brain injury (mTBI) is the most common type of acquired brain injury. Since patients with traumatic brain injury show a tremendous ...

Murine Model of Controlled Cortical Impact for the Induction of Traumatic Brain Injury

创伤性脑损伤诱导受控皮质冲击的母模型

The Centers for Disease Control and Injury Prevention estimate that almost 2 million people sustain a traumatic brain injury (TBI) every year in the ...