Funding for this work was provided by Shriners Hospitals and CIRM fellowships to AMC and AM. We would also like to acknowledge Plemel et al.7 for first designing and publishing the calibrated forceps model.

所有动物的程序及动物护理方法应该由该机构的机构动物护理和使用委员会（IACUC）批准。 1.手术准备<ol><li>收集所有必要的外科手术工具和试剂:镊子，剪刀瓦纳，roungeurs，牵引器，手术刀，剪刀，缝合，皮肤缝合器，棉签，无菌生理盐水，手术海绵和异氟醚。准备和高压灭菌的外科手术工具的完整包在手术前。如果执行手术在一个以上的小鼠，制备和高压灭菌的每只动物的工具之一包，或高压灭菌手术器械的一个分组和消毒用工具消毒器在手术之间。通常，无菌工具之一包可以用在最多5只小鼠，如果玻璃珠工具terilizer用于清洁在两者之间手术（5只动物后的工具必须重新清洗并在使用前灭菌）的工具。请与您当地的IACUC管理员instituti检查具体的指导方针。 </li><li>消毒手术区域用70％异丙醇擦拭。建立手术单，以确保在手术过程中的无菌场被维持。 </li><li>权衡手术前各小鼠。施用0.05毫克/公斤体重丁丙诺啡的剂量，皮下。 </li><li>通过在异氟醚机的感应腔室的剂量为4％施用异氟烷麻醉动物。 </li><li>一旦动物被麻醉，敷药膏对眼睛，以防止脱水，设置在加热垫的动物，在37℃，并确保该鼠标的头部适当地坐落在麻醉锥。 （注:使用热源，不会造成热灼伤; 也就是说 ，循环水毯，热水袋，或等效。）此时辖2％异氟醚的剂量动物。 </li><li>剃鼠标的背表面上，围绕着预定切口位置约1厘米。 </li><li>消毒切口s伊特通过用70％异丙醇擦拭洗涤，然后用碘溶液（10％聚维酮碘，1％有效碘）。重复3次。 </li></ol> 2.背椎板切除术<ol><li>在作出一个切口，确保动物正常通过检查用脚趾或尾巴捏法见长麻醉。 </li><li>使沿背棘与手术刀和刀片切口，然后检查反射了。拱起背部，帮助妥善可视化地标，如椎骨之间的界限。 </li><li>通过皮肤切开。插入牵开器保持皮肤和筋膜从脊髓背面。清除在脊髓任一侧的组织以暴露覆盖脊柱的肌肉。 注:外科医生应熟悉解剖标志。例如，肩胛骨的劣角对应用T7。鼠标的脊柱的自然曲线的顶部是T12和可以用作一个参考点。 </li><li>与对罗珀照明，确定椎骨之间的空间。发现T10的后端与切断肌肉和筋膜垂直于椎间盘空间。切刚够露出T10的棘突和后部薄层。 </li><li>使用一对细尖杜蒙＃5镊子，去除一些从椎板和棘突的组织的，以暴露小纱条脊髓。必要时，用组织钳脊柱稳定。 </li><li>通过插入一对小瓦瑙剪刀沿着椎骨的背外侧侧面和刚椎板下方的一侧执行椎板切除术。 <ol><li>使小，小心剪刀通过椎板的侧面切割。确保没有压力施加到脊髓。 </li><li>重复另一侧。 </li><li>轻轻推压，停止作为必要的止血用棉条或外科用纱布，同时注意不要施加压力的脊髓。 注:准备发泡胶在事件出血需要被控制浸泡在无菌盐水中。 </li><li>一旦切口已经作出，抬起该椎骨的背侧并轻轻清除任何组织的附件。使用适当的手段来控制，如果必要的出血。 </li></ol></li></ol> 3.脊髓压迫<ol><li>使用roungeurs或椎板钳，确保了脊髓的侧面是自由椎体骨，使得校准钳子用于压缩损伤可对脊髓的两侧位于。钳子的臂必须能够被放置在脊髓和相邻侧的钳子的前端硬膜外空间内必须能够到达椎管的楼层。 <ol><li>确保脊髓认为可视性良好。 </li></ol></li><li>位于校准杜蒙＃5镊子大约在脊髓的暴露段的中间。回想一下，部队的武器PS必须放置在硬膜外腔的相邻侧面和尖端必须接触椎管的底板，以产生重现的伤害。 </li><li>仔细压迫脊髓直到隔板连接。固定到位，持续15秒。 </li><li>轻轻释放压缩力和从脊髓取出校准的钳子。无菌盐水应该用来恢复稳态之前伤口闭合。 </li></ol> 4.创伤闭合<ol><li>仔细缝合肌肉层在脊髓，同时注意不要破坏或施加压力的脊髓。 </li><li>使用任一缝合线或U形钉来闭合皮肤在伤口上。 </li><li>如果使用气体麻醉，开始逐渐/关闭麻醉剂。 </li><li>辖0.1毫升每10g体重乳酸林格氏来帮助在手术时，动物是昏昏欲睡，从损伤恢复的手术后帐户脱水。该解决方案应该是WÄRMEd可在注射前的体温。 </li><li>鼠标放置在一个床上用品，免费笼。笼应搁在一个加热垫（如在1.1.5中描述）的方式，使一半的笼区域是在垫的顶部，而另一半被搁在一个RT计数器，为了得到鼠标的气候方案，一旦它走动。请注意，以确保"回收笼"坐落在一个安静的环境。密切监测动物，直到它恢复意识，此时鼠标可以被转移到一个普通笼子里的床上用品。 </li></ol> 5.术后护理<ol><li>手术后，按照需要来管理疼痛症状给药0.05毫克/千克体重丁丙诺啡皮下注射，每12小时一个剂量的第3天手术后，再。 </li><li>辖乳酸林格氏（每10g体重皮下注射0.1ml）中的第3的剂量 - 手术后第5天。给这个剂量，如果/当一个进制开始显示出这个初始时间段以外脱水的迹象。 </li><li>手动表达使用克雷德机动每日两次的动物的膀胱。轻轻触诊动物的腹部找到膀胱，然后应用温和的下行压力，直到膀胱是空的。 <ol><li>如果膀胱不空或尿液是血性或阴天，辖50毫克/公斤按体重拜有利的，以动物通过interperitoneal注射10天。 </li></ol></li><li>监测动物对自噬，脱水和过度减肥（体重大于20％）的迹象。如果动物遇到任何这些症状，请与兽医立即用问候治疗方案咨询，或安乐死的动物以人道的方式下IACUC准则。 <ol><li>鉴于流动性可能在手术后的权利的限制，采取必要的步骤，以确保动物获得食物和水。预包装湿的食物，还有一s的水凝胶，可以提供给动物在这些实例。 </li></ol></li></ol> 6.评估组织损伤的压缩损伤引起的<ol><li>如在步骤1.4和1.5中所述麻醉动物。检查麻醉深度脚趾捏和监控角膜眨眼反射。当动物是不敏感的刺激，继续执行步骤6.2。 </li><li>执行一个心内灌注10。 <ol><li>暴露胸腔和针头插入左心室。冲洗掉现有的流体用20 - 30毫升冰冷的磷酸盐缓冲盐水（PBS），随后加入15 - 40毫升冰冷的4％多聚甲醛（PFA）的。 </li></ol></li><li>取出脊髓。 <ol><li>切开皮肤背侧脊髓和清除周围的脊柱的长度的任何多余的组织。 </li><li>消费脊柱和切去任何剩余的组织。椎板切除术和我的实际水平njury可以通过计数肋的确认。 </li><li>使用万纳剪刀和镊子来置换脊柱的小部分在一个尾至头端的方向。继续砍，直到线暴露足以让安全移除。切割的位置和方法的可视化可以通过使用一个立体镜的来促进。 </li></ol></li><li>放置脊髓组织在4％PFA。允许组织向定位后在该溶液中24小时，在4℃下。 </li><li> Cryoprotect组织孵育在30％蔗糖24小时，在4℃。 </li><li>在OCT嵌入组织。简单地说，采取组织从30％蔗糖孵化和删除任何多余的解决方案。将组织放入盛有OCT一个cryomold孵育1小时，在4℃。 <ol><li>除去从4℃的模具中，确认组织的方向取向，将模具中的2-甲基丁烷浅盘（预冷却1小时在干冰），并允许OCT来完全固化。如果使用即时或STO继续干冰再在-80℃。 </li></ol></li><li>切割组织成使用低温恒温器20微米的矢状切面。直接安装到组织的幻灯片。储存在-20℃直到使用。 </li><li>执行苏木和曙红（H＆E）染色。 <ol><li>简要地说，再水合所述组织（5分钟，2次），染色用苏木精（2.5分），并用水（1分钟，2次）。 </li><li>孵育组织中的50％，然后70％的乙醇（每次3分钟），染色与曙红（45秒），并且温育在90％以上（5秒），95％（5秒）脱水，100％乙醇（2分钟）和异丙醇（2分钟）。 </li><li>清除二甲苯（5分钟，3次）。 注:H＆E染色将随具体的组织厚度和条件。因此，标准化是在进行试验的组织标本前需要。 </li></ol></li><li>覆盖组织与薄带Permount（约100微米）和盖玻片。按下盖玻片上的各方，确保流体的正确分布。让幻灯片干O / N。 </li><li>可见ualize使用使用附带的软件数码显微镜拍摄的图像组织切片。 </li></ol>

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The authors have nothing to disclose.

的脊髓损伤模型的选择是在设计实验，以确定对脊髓损伤的人类病例的治疗的疗效很重要。这些实验需要的动物模型，该模型是高度可再现的，以限制可变性可能导致不确定的数据。他们也应该是临床相关正确地评价人的条件，他们的造型。为此目的，在一个横切选择的压缩或挫伤性损伤是更临床相关的3。然而，撞击和体重下降的装置为挫伤受伤需要使用昂贵和复杂的机械。与此相反，脊髓损伤的校准钳子模型利用改性镊子，很容易从普通实验室材料来组装，并在手术仅需要一个标准的背侧椎板切除后额外的步骤，以暴露脊髓。然而，使用这种方法的一个缺点是，压缩力总是施加横向而非背侧，如是最常见于脊髓损伤9，和压缩损伤的方法，使用影响组织的更大的喙-尾程度比挫伤模型1,2-产生人类临床病例。该模型已被证明了该技术的发起者，并且我们，以生成可再现的SCI 7,11，并且非常适合于小鼠的大小。此外，这种损伤模型允许动物手术后及治疗方法使用多种行为测试，如巴索鼠标规模为运动和冯弗雷头发测试，以验证动物的人群共享相同的损伤程度进行评估和神经功能缺损7,11-13。这些相同的技术也可以用来评估的过程中调查研究给予动物治疗的功效，满足对用于评估疗法的SCI 2,7-动物模型的一般标准。 制备校准钳子的方法S为损伤模型简单，能够实现与各种不同的方法。我们已经使用了隔板11的方法，由Plemel 7所公布，并且还修改使用小螺钉，它不仅提供了一个更简单的方法，用于产生压缩装置钳子，也允许为多功能性在调整最终压缩宽度受益比较研究。在创造钳子选项的范围几乎是无限的，只要该间隔件（多个）提供始终关闭所述钳子以相同距离的稳定装置，并能承受高压灭菌和消毒。此视频中所描述的手术方法是在用户之间高度重现，但是有必要执行椎板切除和缝合动物后的程序已被执行，以使​​脊髓不会受到任何附加的压缩力，可能会增加当护理采取受伤的严重程度和混淆未来的实验。 </p&gt; 用适当的训练和练习，压迫损伤的校准钳子模型非常适合于在模仿在人体中观察到的2,3,7-临床病例的小鼠进行脊髓损伤。由于易于制造镊子，产生不同程度损伤严重程度的小鼠能够容易地完成。这将是大有裨益观察不同程度的转基因小鼠严重的遗传效应对SCI以及评价干细胞移植的疗效小鼠。大多数的文献中的研究已经在大鼠上进行，由于它们的大小，这通常使得手术更容易执行。然而由Plemel 等 7本视频出版，由我们描述的方法应该使SCI要对小鼠以极大的方便和可重复性进行。

脊髓损伤的动物模型可用于评估旨在减轻损害创伤到脊髓的后果的治疗范例的功效有价值的工具。出实验的必要性，这些模型必须提供运动和感觉的行为重复的缺陷，随意调节产生不同程度的伤害，并证明损害严重程度与神经功能缺损的观察到的程度。有SCI三种主要类型有伤害的鲜明特色:切断，挫伤和压缩1。简要地说，一个横断损伤是裂伤至脊髓，挫伤损伤起因于施加到背侧脊髓的简要，焦力，并且当有害的力被施加到脊髓发生压迫损伤，并且也可以是被称为挤压伤2。 完全横断伤是在人类3临床上较少见，而挫伤了ð压缩损伤是比较常见的。压缩损伤产生类似于被人的SCI发现引起，例如，肿瘤压迫或其他有害的压缩力的结果，并且可以使用的工具的一个简单的阵列来执行。挫伤和压缩损伤是在双方都是一个压缩力两者具有类似的病理特征，如cytoarchitectonic解体相似，并且唤起类似的内源性应答损伤1,4。挫伤损伤模型通常适用于类似SCI的人类病例来自脊柱2,5,6的冲击造成的方式使用特殊的设备，这股力量对脊髓背侧。与此相反，压缩损伤可通过各种施加力背侧或侧面的方法来生成的。的压缩损伤的方法包括校准钳7，动脉瘤夹2，或者直接放置一个重量到脊髓8。的优势动脉瘤夹是它们能提供不同量的力9。添加权重给脊髓直接8的背表面的方法中，需要的重量到位10分钟后，急剧增加了手术的长度，并导致不一致由于重量和运动的放置由于的呼吸动物。由于小尺寸的小鼠，情境动物设计用于大鼠，如撞击用于挫伤受伤使用专门的设备，可能是困难的或导致不一致的受伤7。然而，小鼠是可在一个宽范围的转基因品系，不象较大的动物如大鼠或兔，这对脊髓损伤的研究非常有用的。 使用校准钳压迫脊髓的Plemel方法生成一个可重复使用SCI损伤程度及神经功能缺损7之间的相关度很高。这种手术SCI模型使用一对修饰将在由任一金属的环氧树脂或一些其它障碍物，防止完全关闭一个限定的距离保持分开号5杜蒙钳生成的。这种工程化的间距确保了钳子将始终接近一定的宽度在多次手术和由不同的用户。所述Plemel方法的优点是，以产生校准的钳子的材料可以容易地购得，使用时需要专门的设备组装在实验室。这些镊子可以承受多轮高压灭菌和杀菌的，而且缺乏一个单独的，笨重的设备的简化手术。 在这个视频，我们证明在小鼠脊髓外科手术使用的一对校准镊子以产生一个压缩的损伤。我们也解决与脊髓受伤的实验动物的照顾，以提高他们的生活质量手术后，降低死亡率独特的担忧。

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			<td height="64" style="height:64px;width:101px;">Laminectomy Forceps (Dumont #2)&#160;</td>
			<td style="width:127px;">Fine Science Tools</td>
			<td style="width:123px;">11223-20</td>
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			<td height="43" style="height:43px;width:101px;">Dumont #5 Forceps</td>
			<td style="width:127px;">Fine Science Tools</td>
			<td style="width:123px;">11252-20</td>
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			<td height="22" style="height:22px;width:101px;">Stapler</td>
			<td style="width:127px;">Fine Science Tools</td>
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			<td style="width:123px;">101-2636</td>
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			<td height="43" style="height:43px;width:101px;">Needles (30 G x &#189;)</td>
			<td style="width:127px;">BD Biomedical</td>
			<td style="width:123px;">305106</td>
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calibrated forceps model of spinal cord compression injury

小鼠脊髓压迫损伤是有价值的动物模型脊髓损伤（SCI）和脊髓再生疗法的研究。压迫损伤的校准镊子模型是一个方便，成本低，而且非常重复性的动物模型SCI。我们所用的一对修改的镊子根据由Plemel 等人发表的方法（2008），以横向地压迫脊髓，以0.35毫米的距离。在此视频中，我们将展示一个背椎板切除以暴露脊髓，其次是脊髓与改性钳子的压缩。在视频中，我们也将解决与截瘫实验动物的护理问题。这种损伤模型产生表现出的感觉障碍，以及受损的后肢运动功能的小鼠。此外，受伤的这个方法，在SCI的病理产生一致的畸变，通过免疫组化方法确定。看着这个VID后EO，观众应该能够确定用于各种严重程度的，在小鼠为设计损伤后，以减轻损害对脊髓损伤和/或治疗方法的研究产生的SCI的必要的用品和方法。

我们进行了椎板12小鼠 - 如上所述（25 30克），接着脊髓压缩为0.25毫米（N = 4），0.35毫米（N = 4）和0.55毫米（N = 4）。我们在三个牺牲的动物（N = 6）和7（6）日伤后心内灌注。脊髓从脊柱去除，制备并如上所述进行处理的组织。整个脊髓的图像用的Leica EZ4数字显微镜和附带的软件。脊髓切片图像使用Olympus数码显微镜及配套的软件采取2倍的放大倍率。 我们发现，脊髓受压产生的震中在压缩（ 图1）的部位受伤。伤病的影响，在延髓和尾鳍方向延伸几毫米。损伤的严重性随着所述间隔件之间的距离减小（0.25毫米&gt;0.35毫米&gt;0.55毫米， 图2）。压缩后的三天，有血在伤的震中和损伤后不在场7天周边地区。 0.25毫米和0.35毫米按压产生的空腔，但不是0.55毫米模型。经过7天，背，腹白质主要是缩小尺寸的震中，灰质组织被严重扭曲，和空化呈持续性。采用适当的测试，如巴索鼠标规模为运动功能和冯弗雷头发和乙基氯化物测试感觉功能正如我们在以前的出版物8展示了这些cytoarchitectonical改变被翻译成运动和感觉的改变动物的行为进行评估。 <img alt="图1" src="/files/ftp_upload/52318/52318fig1highres.jpg" width="500" /> 完整脊髓图1代表性图像之前和AF之三伤。 （A）完整的脊髓。（B）后0.35毫米压缩脊髓。箭头表示损伤的边界。星号indentifies损伤震中。 D =背，L =横向。比例尺:0.50毫米<a href="https://jove.unicartagenaproxy.elogim.com/files/ftp_upload/52318/52318fig1highres.jpg" target="_blank">请点击此处查看该图的放大版本。</a> <img alt="图2" src="/files/ftp_upload/52318/52318fig2highres.jpg" width="650" /> 图鼠标脊髓2.代表图像之前和压迫损伤在不同宽度压缩后。 （A）的控制脊髓矢状切面。（B）在一个0.35毫米SCI后第7天压迫损伤（dpi）的震中冠状切面（C，E，G）脊髓3 DPI为0.25宽度矢状切面0.35 0.55毫米（D，F，H）的H＆E矢状切片染色脊髓第7 DPI至0.25，0.35 0.55毫米的宽度。星号indentifies损伤震中。所有部分用H＆E。 D =背，L =横向。比例尺:1.25毫米<a href="https://jove.unicartagenaproxy.elogim.com/files/ftp_upload/52318/52318fig2highres.jpg" target="_blank">请点击此处查看该图的放大版本。</a>

Watch this Scientific Journal Video about Calibrated Forceps Model of Spinal Cord Compression Injury at JoVE.com

Calibrated Forceps Model of Spinal Cord Compression Injury

Spinal cord injury models should be highly reproducible. We demonstrate that the calibrated forceps compression model of spinal cord injury is an easy to use surgical method for generating reproducible injuries to the murine spinal cord.

脊髓压迫损伤的校准钳型

Compression injuries of the murine spinal cord are valuable animal models for the study of spinal cord injury (SCI) and spinal regenerative therapy. The ...

calibrated-forceps-model-of-spinal-cord-compression-injury

Universidad de Cartagena UDEC

Research

JoVE Journal

Medicine

18.4K 次观看.  University of California, Davis. Spinal cord injury models should be highly reproducible. We demonstrate that the calibrated forceps compression model of spinal cord injury is an easy to use surgical method for generating reproducible injuries to the murine spinal cord.

视频: Calibrated Forceps Model of Spinal Cord Compression Injury

Acute and Chronic Tactile Sensory Testing after Spinal Cord Injury in Rats

Spinal cord injury (SCI) impairs sensory systems causing allodynia1-8. To identify cellular and molecular causes of allodynia, sensitive and valid sensory testing in rat SCI models is needed. However, until recently, no single testing approach had been validated for SCI so that standardized methods have not been implemented across labs. Additionally, available testing methods could not be implemented acutely or when severe motor impairments existed, preventing studies of the development of SCI-induced allodynia3. Here we present two validated sensory testing methods using von Frey Hair (VFH) monofilaments which quantify changes in tactile sensory thresholds after SCI4-5. One test is the well-established Up-Down test which demonstrates high sensitivity and specificity across different SCI severities when tested chronically5. The other test is a newly-developed dorsal VFH test that can be applied acutely after SCI when allodynia develops, prior to motor recovery4-5. Each VFH monofilament applies a calibrated force when touched to the skin of the hind paw until it bends. In the up-down method, alternating VFHs of higher or lower forces are used on the plantar L5 dermatome to delineate flexor withdrawal thresholds. Successively higher forces are applied until withdrawal occurs then lower force VFHs are used until withdrawal ceases. The tactile threshold reflects the force required to elicit withdrawal in 50% of the stimuli. For the new test, each VFH is applied to the dorsal L5 dermatome of the paw while the rat is supported by the examiner. The VFH stimulation occurs in ascending order of force until at least 2 of 3 applications at a given force produces paw withdrawal. Tactile sensory threshold is the lowest force to elicit withdrawal 66% of the time. Acclimation, testing and scoring procedures are described. Aberrant trials that require a retest and typical trials are defined. Animal use was approved by Ohio State University Animal Care and Use Committee.

We describe two tactile sensory testing methods for acute or chronic periods of spinal cord injury in rats. These validated procedures can detect the development and maintenance of allodynia-like sensations.

在大鼠脊髓损伤后的急性和慢性触觉测试

Spinal cord injury (SCI) impairs sensory systems causing allodynia1-8. To identify cellular and molecular causes of allodynia, sensitive and valid sensory ...

科研

医学

A Contusive Model of Unilateral Cervical Spinal Cord Injury Using the Infinite Horizon Impactor

While the majority of human spinal cord injuries occur in the cervical spinal cord, the vast majority of laboratory research employs animal models of spinal cord injury (SCI) in which the thoracic spinal cord is injured. Additionally, because most human cord injuries occur as the result of blunt, non-penetrating trauma (e.g. motor vehicle accident, sporting injury) where the spinal cord is violently struck by displaced bone or soft tissues, the majority of SCI researchers are of the opinion that the most clinically relevant injury models are those in which the spinal cord is rapidly contused.1 Therefore, an important step in the preclinical evaluation of novel treatments on their way to human translation is an assessment of their efficacy in a model of contusion SCI within the cervical spinal cord. Here, we describe the technical aspects and resultant anatomical and behavioral outcomes of an unilateral contusive model of cervical SCI that employs the Infinite Horizon spinal cord injury impactor.
Sprague Dawley rats underwent a left-sided unilateral laminectomy at C5. To optimize the reproducibility of the biomechanical, functional, and histological outcomes of the injury model, we contused the spinal cords using an impact force of 150 kdyn, an impact trajectory of 22.5&deg; (animals rotated at 22.5&deg;), and an impact location off of midline of 1.4 mm. Functional recovery was assessed using the cylinder rearing test, horizontal ladder test, grooming test and modified Montoya's staircase test for up to 6 weeks, after which the spinal cords were evaluated histologically for white and grey matter sparing.
The injury model presented here imparts consistent and reproducible biomechanical forces to the spinal cord, an important feature of any experimental SCI model. This results in discrete histological damage to the lateral half of the spinal cord which is largely contained to the ipsilateral side of injury. The injury is well tolerated by the animals, but does result in functional deficits of the forelimb that are significant and sustained in the weeks following injury. The cervical unilateral injury model presented here may be a resource to researchers who wish to evaluate potentially promising therapies prior to human translation.

A reliable and repeatable way to produce a cervical unilateral spinal cord injury using the Infinite Horizon impactor is described. The method takes advantage of a custom designed frame and clamp to stabilize the spine. The standardized procedure and biomechanical injury parameters result in sufficient and sustained injuries.

单侧颈脊髓损伤的挫伤模型，利用无限的地平线冲击

While the majority of human spinal cord injuries occur in the cervical spinal cord, the vast majority of laboratory research employs animal models of ...

A Contusion Model of Severe Spinal Cord Injury in Rats

The translational potential of novel treatments should be investigated in severe spinal cord injury (SCI) contusion models. A detailed methodology is described to obtain a consistent model of severe SCI. Use of a stereotactic frame and computer controlled impactor allows for creation of reproducible injury. Hypothermia and urinary tract infection pose significant challenges in the post-operative period. Careful monitoring of animals with daily weight recording and bladder expression allows for early detection of post-operative complications. The functional results of this contusion model are equivalent to transection models. The contusion model can be utilized to evaluate the efficacy of both neuroprotective and neuroregenerative approaches.

A contusion model of severe spinal cord injury is described. Detailed pre-operative, operative and post-operative steps are described to obtain a consistent model.

挫伤模型脊髓严重损伤的保护作用

The translational potential of novel treatments should be investigated in severe spinal cord injury (SCI) contusion models. A detailed methodology is ...

Evaluation of Respiratory Muscle Activation Using Respiratory Motor Control Assessment (RMCA) in Individuals with Chronic Spinal Cord Injury

During breathing, activation of respiratory muscles is coordinated by integrated input from the brain, brainstem, and spinal cord. When this coordination is disrupted by spinal cord injury (SCI), control of respiratory muscles innervated below the injury level is compromised1,2 leading to respiratory muscle dysfunction and pulmonary complications. These conditions are among the leading causes of death in patients with SCI3. Standard pulmonary function tests that assess respiratory motor function include spirometrical and maximum airway pressure outcomes: Forced Vital Capacity (FVC), Forced Expiratory Volume in one second (FEV1), Maximal Inspiratory Pressure (PImax) and Maximal Expiratory Pressure (PEmax)4,5. These values provide indirect measurements of respiratory muscle performance6. In clinical practice and research, a surface electromyography (sEMG) recorded from respiratory muscles can be used to assess respiratory motor function and help to diagnose neuromuscular pathology. However, variability in the sEMG amplitude inhibits efforts to develop objective and direct measures of respiratory motor function6. Based on a multi-muscle sEMG approach to characterize motor control of limb muscles7, known as the voluntary response index (VRI)8, we developed an analytical tool to characterize respiratory motor control directly from sEMG data recorded from multiple respiratory muscles during the voluntary respiratory tasks. We have termed this the Respiratory Motor Control Assessment (RMCA)9. This vector analysis method quantifies the amount and distribution of activity across muscles and presents it in the form of an index that relates the degree to which sEMG output within a test-subject resembles that from a group of healthy (non-injured) controls. The resulting index value has been shown to have high face validity, sensitivity and specificity9-11. We showed previously9 that the RMCA outcomes significantly correlate with levels of SCI and pulmonary function measures. We are presenting here the method to quantitatively compare post-spinal cord injury respiratory multi-muscle activation patterns to those of healthy individuals.

Evaluation of Respiratory Muscle Activation Using Respiratory Motor Control Assessment RMCA in Individuals with Chronic Spinal Cord Injury

The purpose of this publication is to present our original work on a multi-muscle surface electromyographic approach to quantitatively characterize respiratory muscle activation patterns in individuals with chronic spinal cord injury using vector-based analysis.

评价呼吸肌慢性脊髓损伤个人使用呼吸电机控制评估（RMCA）

During breathing, activation of respiratory muscles is coordinated by integrated input from the brain, brainstem, and spinal cord. When this coordination ...

A Murine Model of Cervical Spinal Cord Injury to Study Post-lesional Respiratory Neuroplasticity

A cervical spinal cord injury induces permanent paralysis, and often leads to respiratory distress. To date, no efficient therapeutics have been developed to improve/ameliorate the respiratory failure following high cervical spinal cord injury (SCI). Here we propose a murine pre-clinical model of high SCI at the cervical 2 (C2) metameric level to study diverse post-lesional respiratory neuroplasticity. The technique consists of a surgical partial injury at the C2 level, which will induce a hemiparalysis of the diaphragm due to a deafferentation of the phrenic motoneurons from the respiratory centers located in the brainstem. The contralateral side of the injury remains intact and allows the animal recovery. Unlike other SCIs which affect the locomotor function (at the thoracic and lumbar level), the respiratory function does not require animal motivation and the quantification of the deficit/recovery can be easily performed (diaphragm and phrenic nerve recordings, whole body ventilation). This pre-clinical C2 SCI model is a powerful, useful, and reliable pre-clinical model to study various respiratory and non-respiratory neuroplasticity events at different levels (molecular to physiology) and to test diverse putative therapeutic strategies which might improve the respiration in SCI patients.

Respiratory failure is the leading cause of death following a cervical spinal cord injury. Having a reproducible, quantifiable, and reliable pre-clinical animal model of respiratory failure induced by a partial cervical injury will help to understand the subsequent respiratory and non-respiratory neuroplasticity and allow testing putative repair strategies.

颈脊髓损伤的小鼠模型研究后皮损呼吸神经可塑性

A cervical spinal cord injury induces permanent paralysis, and often leads to respiratory distress. To date, no efficient therapeutics have been developed ...

Controlled Cortical Impact Model for Traumatic Brain Injury

Every year over a million Americans suffer a traumatic brain injury (TBI). Combined with the incidence of TBIs worldwide, the physical, emotional, social, and economical effects are staggering. Therefore, further research into the effects of TBI and effective treatments is necessary. The controlled cortical impact (CCI) model induces traumatic brain injuries ranging from mild to severe. This method uses a rigid impactor to deliver mechanical energy to an intact dura exposed following a craniectomy. Impact is made under precise parameters at a set velocity to achieve a pre-determined deformation depth. Although other TBI models, such as weight drop and fluid percussion, exist, CCI is more accurate, easier to control, and most importantly, produces traumatic brain injuries similar to those seen in humans. However, no TBI model is currently able to reproduce pathological changes identical to those seen in human patients. The CCI model allows investigation into the short-term and long-term effects of TBI, such as neuronal death, memory deficits, and cerebral edema, as well as potential therapeutic treatments for TBI.

Traumatic brain injuries (TBIs) remain a serious health problem. Using the controlled cortical impact surgery model, research on the effects of TBI and possible treatment methods may be performed.

控制皮层影响模型创伤性脑损伤

Every year over a million Americans suffer a traumatic brain injury (TBI).  Combined with the incidence of TBIs worldwide, the physical, emotional, ...

A Radio-telemetric System to Monitor Cardiovascular Function in Rats with Spinal Cord Transection and Embryonic Neural Stem Cell Grafts

High thoracic or cervical spinal cord injury (SCI) can lead to cardiovascular dysfunction. To monitor cardiovascular parameters, we implanted a catheter connected to a radio transmitter into the femoral artery of rats that underwent a T4 spinal cord transection with or without grafting of embryonic brainstem-derived neural stem cells expressing green fluorescent protein. Compared to other methods such as cannula insertion or tail-cuff, telemetry is advantageous to continuously monitor blood pressure and heart rate in freely moving animals. It is also capable of long term multiple data acquisitions. In spinal cord injured rats, basal cardiovascular data under unrestrained condition and autonomic dysreflexia in response to colorectal distension were successfully recorded. In addition, cardiovascular parameters before and after SCI can be compared in the same rat if a transmitter is implanted before a spinal cord transection. One limitation of the described telemetry procedure is that implantation in the femoral artery may influence the blood supply to the ipsilateral hindlimb.

We present a protocol for using a radio-telemetric system to record cardiovascular parameters in T4 spinal cord transected rats eight weeks after embryonic brainstem neural stem cell grafting into the lesion site. Telemetry is an advanced technique to accurately evaluate cardiovascular function in conscious freely moving spinal cord injured rats.

无线电遥测系统，以监测大鼠脊髓横断和胚胎神经干细胞移植在心血管功能

High thoracic or cervical spinal cord injury (SCI) can lead to cardiovascular dysfunction. To monitor cardiovascular parameters, we implanted a catheter ...

Neural Stem Cell Transplantation in Experimental Contusive Model of Spinal Cord Injury

Spinal cord injury is a devastating clinical condition, characterized by a complex of neurological dysfunctions. Animal models of spinal cord injury can be used both to investigate the biological responses to injury and to test potential therapies. Contusion or compression injury delivered to the surgically exposed spinal cord are the most widely used models of the pathology. In this report the experimental contusion is performed by using the Infinite Horizon (IH) Impactor device, which allows the creation of a reproducible injury animal model through definition of specific injury parameters. Stem cell transplantation is commonly considered a potentially useful strategy for curing this debilitating condition. Numerous studies have evaluated the effects of transplanting a variety of stem cells. Here we demonstrate an adapted method for spinal cord injury followed by tail vein injection of cells in CD1 mice. In short, we provide procedures for: i) cell labeling with a vital tracer, ii) pre-operative care of mice, iii) execution of a contusive spinal cord injury, and iv) intravenous administration of post mortem neural precursors. This contusion model can be utilized to evaluate the efficacy and safety of stem cell transplantation in a regenerative medicine approach.

Spinal cord injury is a traumatic condition that causes severe morbidity and high mortality. In this work we describe in detail a contusion model of spinal cord injury in mice followed by a transplantation of neural stem cells.

在脊髓损伤的实验挫伤型号神经干细胞移植

Spinal cord injury is a devastating clinical condition, characterized by a complex of neurological dysfunctions. Animal models of spinal cord injury can ...

Contrast Enhanced Ultrasound Imaging for Assessment of Spinal Cord Blood Flow in Experimental Spinal Cord Injury

Reduced spinal cord blood flow (SCBF) (i.e., ischemia) plays a key role in traumatic spinal cord injury (SCI) pathophysiology and is accordingly an important target for neuroprotective therapies. Although several techniques have been described to assess SCBF, they all have significant limitations. To overcome the latter, we propose the use of real-time contrast enhanced ultrasound imaging (CEU). Here we describe the application of this technique in a rat contusion model of SCI. A jugular catheter is first implanted for the repeated injection of contrast agent, a sodium chloride solution of sulphur hexafluoride encapsulated microbubbles. The spine is then stabilized with a custom-made 3D-frame and the spinal cord dura mater is exposed by a laminectomy at ThIX-ThXII. The ultrasound probe is then positioned at the posterior aspect of the dura mater (coated with ultrasound gel). To assess baseline SCBF, a single intravenous injection (400 &#181;l) of contrast agent is applied to record its passage through the intact spinal cord microvasculature. A weight-drop device is subsequently used to generate a reproducible experimental contusion model of SCI. Contrast agent is re-injected 15 min following the injury to assess post-SCI SCBF changes. CEU allows for real time and in-vivo assessment of SCBF changes following SCI. In the uninjured animal, ultrasound imaging showed uneven blood flow along the intact spinal cord. Furthermore, 15 min post-SCI, there was critical ischemia at the level of the epicenter while SCBF remained preserved in the more remote intact areas. In the regions adjacent to the epicenter (both rostral and caudal), SCBF was significantly reduced. This corresponds to the previously described &#8220;ischemic penumbra zone&#8221;. This tool is of major interest for assessing the effects of therapies aimed at limiting ischemia and the resulting tissue necrosis subsequent to SCI.

Contrast Enhanced Ultrasound imaging is a reliable in-vivo tool for quantifying spinal cord blood flow in an experimental rat spinal cord injury model. This paper contains a comprehensive protocol for application of this technique in association with a contusion model of thoracic spinal cord injury.

超声造影成像脊髓血流量的评估实验脊髓损伤

Reduced spinal cord blood flow (SCBF) (i.e., ischemia) plays a key role in traumatic spinal cord injury (SCI) pathophysiology and is accordingly an ...

Photothrombosis-induced Focal Ischemia as a Model of Spinal Cord Injury in Mice

Spinal cord injury (SCI) is a devastating clinical condition causing permanent changes in sensorimotor and autonomic functions of the spinal cord (SC) below the site of injury. The secondary ischemia that develops following the initial mechanical insult is a serious complication of the SCI and severely impairs the function and viability of surviving neuronal and non-neuronal cells in the SC. In addition, ischemia is also responsible for the growth of lesion during chronic phase of injury and interferes with the cellular repair and healing processes. Thus there is a need to develop a spinal cord ischemia model for studying the mechanisms of ischemia-induced pathology. Focal ischemia induced by photothrombosis (PT) is a minimally invasive and very well established procedure used to investigate the pathology of ischemia-induced cell death in the brain. Here, we describe the use of PT to induce an ischemic lesion in the spinal cord of mice. Following retro-orbital sinus injection of Rose Bengal, the posterior spinal vein and other capillaries on the dorsal surface of SC were irradiated with a green light resulting in the formation of a thrombus and thus ischemia in the affected region. Results from histology and immunochemistry studies show that PT-induced ischemia caused spinal cord infarction, loss of neurons and reactive gliosis. Using this technique a highly reproducible and relatively easy model of SCI in mice can be achieved that would serve the purpose of scientific investigations into the mechanisms of ischemia induced cell death as well as the efficacy of neuroprotective drugs. This model will also allow exploration of the pathological changes that occur following SCI in live mice like axonal degeneration and regeneration, neuronal and astrocytic Ca2+ signaling using two-photon microscopy.

Photothrombosis is a minimally invasive and highly reproducible procedure to induce focal ischemia in the spinal cord and serves as a model of spinal cord injury in mice.

Photothrombosis引起的局部脑缺血脊髓损伤的小鼠模型

Spinal cord injury (SCI) is a devastating clinical condition causing permanent changes in sensorimotor and autonomic functions of the spinal cord (SC) ...