这项工作是由NIH / NINDS授予4R00的NS 057940，并授予SCBI 200-12从脊髓和脑损伤从印第安纳州卫生署的研究基金的支持。

1。为削弱手术简单的设备<ol><li> （2）我们创建削弱设备由三部分组成（图1）:（1）一个支撑板，允许附件的指导管和一根针，跨颅窗口坐落在手术过程中的不锈钢或塑料制成的;一个指导管保存位置的针头和许可证针轮换;和（3），弯曲成90度〜3毫米的提示，是旋转和插入移动的针。 </li><li>剪下一块1-1.5毫米厚，7〜10 × 30毫米的长方形不锈钢或透明塑料（1），使支撑板。准备一个1.5英寸的22号（BD公司，＃305156）和一个1.5英寸（BD公司，＃305127）25号注射器针头。为了使指导管，切断底胶和22号注射器针头的针尖，其总长度约为31-32毫米（2）。沙的金属端，使其平整，光滑。最后这一指导管的长度应比25号的注射器针头短约5-6毫米。 </li><li>使用氰基丙烯酸酯胶固定到支撑板管的指导，确保针是垂直的金属边缘。 </li><li>指导管插入的25号注射器针头，针90度弯曲的提示（图1）在2.5-3毫米。 </li><li>针到针结束（图1，针停止）的垂直移动范围调整胶合一个小塑料管。最后针移动范围确定有多深，针可以插入到皮质，并应为1.6-1.8毫米P21的大鼠和1.3-1.5毫米为P21的小鼠。这个长度需要针对不同的动物物种和年龄调整。 </li><li>针的上端连接到在同一方向弯曲的提示（图1针指标）的一个小的铜线或小块的磁带。电线或磁带表示在削弱手术针的转动角度（图1）。 </li></ol> 2。动物准备<ol><li>所有手术器械需要进行高压灭菌，用玻璃珠的灭菌，或用70％乙醇消毒灭菌。 </li><li>我们使用只/ SD大鼠在出生后20岁 - 22天（P20 - 22）或小鼠CD1在同一年龄。不同菌株的小鼠或大鼠，可用于具体项目。 </li><li>腹腔注射氯胺酮麻醉鼠标80毫克/公斤和甲苯噻嗪8毫克/公斤。外科手术开始后，动物尾巴捏不响应。在手术过程中，如有必要，一个三分之一的助推器剂量原剂量的麻醉剂鸡尾酒可以得到恢复原来的麻醉状态。 </li><li>切电气头发修剪头发与头皮的动物。麻醉过程中动物保护从干燥的眼睛，应用到少量的眼药膏。使用10％的碘伏溶液，70％的乙醇消毒头皮。 </li><li>一个立体定位仪上的装载动物的头部保持在一个稳定的固定位置。在整个手术中，我们不断在加热垫，以防止体温过低的动物。 </li><li>请使用手术刀的头皮上的中线前后的切口，从波长延伸到两眼之间。使用止血，一边拉扯皮肤，充分暴露左侧颅骨。 </li><li>请在左侧头骨上的一个长方形切割，并用手术刀刮去骨膜。这一步将减少出血和方便的颅骨上钻。 </li></ol> 3。制作削弱<ol><li>添加少量的无菌生理盐水，暴露颅骨面积，然后用几个Q - 提示清洁血液和干燥区域。 </li><li>在手术显微镜下，开始钻探一个长方形的凹​​槽，在左侧头骨中心（〜5 × 7毫米，大鼠，4 × 5在小鼠毫米）（约高于左侧感觉运动皮层）。应用生理盐水的头骨上的下降，有利于钻井和散热。钻骨的深度约2 / 3是后，除去多余的盐水，清洁棉条钻探面积。慢慢地，仔细地钻更深，直到骨的中心部分是一个镊子温柔的抚摸后可移动。 </li><li>小心地取出中央的骨片，骨的边缘上插入了一个钳的锐利尖端，并慢慢抬起钳揭露左半球。 </li><li>在手术显微镜下，按住削弱设备，东方针在矢状方向和支撑板略有尾端垂直中线，倾斜设备，以维持针尖和目标皮层区域的可视化。在颅窗口中，针尖对准一个地区1-2毫米，外侧矢状缝，避免大血管的直接渗透。 </li><li>通过硬脑膜下PIA针插入在水平方向上，以备用血管。坐削弱设备的底部到颅窗口都边缘，使针定位正常的皮质表面（图1）。颅窗口的两个边缘上休息的设备将显着减少或消除在以下过程中颤抖的手。电梯下方的软针，然后慢慢降低它来创建一个transcortical削减直到针头不能深入。旋转〜135 °远离中线创建了半圈白质/深层第六削弱。然后再次提高针下方PIA。倾斜的设备落后，退出针。 </li><li> ，一个可以重复上述步骤（步骤2.5），但没有转动针头，以便创建一个额外的的在窗口外侧边缘transcortical削减。这将创建一个更完整的皮质隔离。 </li><li>广场上的一块塑料薄膜保护颅窗口（6 × 6毫米），缝合头皮。从麻醉中完全恢复之前，放置在一个加热垫的动物。 </li></ol> 4。代表性的成果: 日冕皮质切片可以确认削弱手术的成功。在手术2周后饮片，transcortical并削弱削减明显的显微镜下的低功耗目标（图2）。的部分孤立的皮层通常会变得稍薄，癫痫活动，可以在大多数领域的潜力录制（图3）皮质片检测。 相比之下，白质或深，皮质层的大洞形成戏剧性的损毁皮层，或一个错误的削减，高于或低于脑白质变薄，使大脑无法进一步的实验。 <img src="/files/ftp_upload/2840/2840fig1.jpg" alt="图1"/> 图1。削弱设备的结构及应用 。一个指导管（2）粘到一个支撑板（1）不锈钢或透明塑料制成的。插入一个注射器针头（3）通过引导管和弯曲〜3毫米的小费。一针停止制成的塑料管粘到年底，使针垂直移动范围是有限的〜1.2毫米〜1.5毫米P21的小鼠和大鼠分别使用的针头上。小铜线段下方的手柄固定针弯针的方向指标。请注意，设备倾斜，并在接触既颅窗口的边缘，使切割可以使软脑膜表面平行。 <img src="/files/ftp_upload/2840/2840fig2.jpg" alt="图2"/> 图2。削弱片的一个形象代表 。 A.准备两个星期后，在P48大鼠削弱病变的一个切片的荧光图像。标记荧光染料DII切割伤口。右侧的白色箭头指示transcortical的切割，底部的箭头表示，虽然第四层之间的边界和白质通过削弱。 <img src="/files/ftp_upload/2840/2840fig3.jpg" alt="图3"/> 图3。场潜在的记录从一个削弱的削弱脑片的片领域的潜力录音表现出癫痫的活动，表明受伤皮层组织的过度兴奋。

<ol>
	<li>Hoffman, S. N., Salin, P. A., Prince, D. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Chronic+neocortical+epileptogenesis+in+vitro.">Chronic neocortical epileptogenesis in vitro.</a> J Neurophysiol. 71, 1762-1773 (1994).</li><li>Topolnik, L., Steriade, M., Timofeev, I. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Hyperexcitability+of+intact+neurons+underlies+acute+development+of+trauma-related+electrographic+seizures+in+cats+in+vivo.">Hyperexcitability of intact neurons underlies acute development of trauma-related electrographic seizures in cats in vivo.</a> Eur J Neurosci. 18, 486-496 (2003).</li><li>Graber, K., Prince, D. 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> Models of Seizures and Epilepsy. , 477-493 (2005).</li><li>Nita, D. A., Cisse, Y., Timofeev, I., Steriade, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Increased+propensity+to+seizures+after+chronic+cortical+deafferentation+in+vivo.">Increased propensity to seizures after chronic cortical deafferentation in vivo.</a> J Neurophysiol. 95, 902-913 (2006).</li><li>Sharpless, S. K., Halpern, L. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+electrical+excitability+of+chronically+isolated+cortex+studied+by+means+of+permanently+implanted+electrodes.">The electrical excitability of chronically isolated cortex studied by means of permanently implanted electrodes.</a> Electroencephalogr Clin Neurophysiol. 14, 244-255 (1962).</li><li>Echlin, F. A., Battista, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Epileptiform+Seizures+from+Chronic+Isolated+Cortex.">Epileptiform Seizures from Chronic Isolated Cortex.</a> Arch Neurol. 9, 154-170 (1963).</li><li>Prince, D. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Epileptogenic+neurons+and+circuits.">Epileptogenic neurons and circuits.</a> Adv Neurol. 79, 665-684 (1999).</li><li>Marin-Padilla, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Developmental+neuropathology+and+impact+of+perinatal+brain+damage.+II:+white+matter+lesions+of+the+neocortex.">Developmental neuropathology and impact of perinatal brain damage. II: white matter lesions of the neocortex.</a> J Neuropathol Exp Neurol. 56, 219-235 (1997).</li><li>Jin, X., Prince, D. A., Huguenard, J. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Enhanced+excitatory+synaptic+connectivity+in+layer+v+pyramidal+neurons+of+chronically+injured+epileptogenic+neocortex+in+rats.">Enhanced excitatory synaptic connectivity in layer v pyramidal neurons of chronically injured epileptogenic neocortex in rats.</a> J Neurosci. 26, 4891-4900 (2006).</li><li>Li, H., Prince, D. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Synaptic+activity+in+chronically+injured,+epileptogenic+sensory-motor+neocortex.">Synaptic activity in chronically injured, epileptogenic sensory-motor neocortex.</a> J Neurophysiol. 88, 2-12 (2002).</li><li>Salin, P., Tseng, G. F., Hoffman, S., Parada, I., Prince, D. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Axonal+sprouting+in+layer+V+pyramidal+neurons+of+chronically+injured+cerebral+cortex.">Axonal sprouting in layer V pyramidal neurons of chronically injured cerebral cortex.</a> J Neurosci. 15, 8234-8245 (1995).</li><li>Avramescu, S., Nita, D. A., Timofeev, I. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Neocortical+post-traumatic+epileptogenesis+is+associated+with+loss+of+GABAergic+neurons.">Neocortical post-traumatic epileptogenesis is associated with loss of GABAergic neurons.</a> J Neurotrauma. 26, 799-812 (2009).</li><li>Avramescu, S., Timofeev, I. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Synaptic+strength+modulation+after+cortical+trauma:+a+role+in+epileptogenesis.">Synaptic strength modulation after cortical trauma: a role in epileptogenesis.</a> J Neurosci. 28, 6760-6772 (2008).</li></ol>

没有利益冲突的声明。

削弱模型是一个高效的系统研究外伤性癫痫。一个典型的手术只需要20-30分钟才能完成，而诱发或自发痫活动，可以在大多数动物的切片手术后两周 1-2记录。更重要的是，这个模型模拟方面的变化，如出血，炎症，水肿，干切断和神经元死亡7的脑外伤。不仅被观察到癫痫活动在鼠类和其他动物，但也已在人类遭受可比皮质病变8癫痫发作记录。已经取得了重大进展，澄清在近年来的底层机制。自发和诱发发作的癫痫样放电被记录在大脑切片的病灶&gt; 2周后，发现这些活动都是起源于大鼠脑皮层V层1。电路重组，GABA能interneurons和失控的损失，在神经细胞膜的兴奋性增加，增加兴奋性突触耦合的证据还表明，尤其是皮质V层2,9-13。 在这里，我们引入了一个削弱病变的新手段。当一个执行放手削弱手术，手发抖经常会导致难以稳定和精确的皮层病变。虽然做法和经验，可提高手术的质量，在病变的深度和质量存在显著变异。我们这里介绍的设备是有利于在三个方面。首先，颅窗口的边缘上，该设备在很大程度上消除了手晃动手术过程中的问题，从而使其能够通过细腻的脑组织一个顺利晋级。二，针插入的深度和旋转程度，可仔细控制，这使得有可能削减在白质层第六下方更精确和一致。第三，皮质半球的表面是弯曲的内侧比外侧（图1），这可能会导致缺少有针对性的白质高，如果没有相应的调整针的角度。休息颅窗口的设备，针横向倾斜，并自动调整切割角度，使针的旋转总是平行的皮质表面和精确的病变（图2）。使用此设备的一个潜在的问题是，它可能会干扰针，在显微镜下直接可视化。向尾方向稍微倾斜设备时穿透脑膜和皮层中，就可以解决这个问题。只要针是降低到白质，设备需要进行调整，以成为垂直于皮层表面，和休息的头骨。此时，看着针指标是足够的监察针旋转。总之，这几项优势和建设相对缓和，削弱模式将变得更方便和有用的外伤性癫痫的研究。

<table border="1"><tbody><tr><td> 材料名称 </td><td> 类型 </td><td> 公司 </td><td> 目录编号 </td></tr><tr><td> Foredom微电机套件</td><td>设备</td><td> Foredom </td><td> K.1070 </td></tr><tr><td> 1.5英寸的22号注射器针头</td><td>材料</td><td> BD公司</td><td> 305156 </td></tr><tr><td> 1.5英寸的25号注射器针头</td><td>材料</td><td> BD公司</td><td> 305127 </td></tr><tr><td>氰基丙烯酸酯胶</td><td>材料</td><td>特德佩拉</td><td> 14450 </td></tr></tbody></table>

preparing undercut model of posttraumatic epileptogenesis in rodents

部分隔离皮层（"削弱"）是外伤性癫痫动物模型。手术过程涉及切割通过感觉运动皮层和下面的白质（削弱），使大脑皮层的特定区域，主要是从邻近的皮层和1-3皮层区域隔离。手术后两个星期或以上的延迟后，痫样放电可以被记录在啮齿类动物的大脑切片1;和电气或行为癫痫发作，可在体内从其他物种， 如猫和猴4-6观察。这口井建立的动物模型是有效的生成和模仿脑外伤的几个重要特点。然而，它在技术上是具有挑战性的尝试使精确放手的小鼠患脑皮质病变。基于最初在大卫博士王子的实验室设在斯坦福大学1的程序，在这里我们提出一个改进的技术来执行这种模式在小鼠和大鼠的准备手术。我们演示了如何做一个简单的手术设备，并用它来获得更好的控制切割的深度和角度，以产生更精确和一致的结果。该设备是容易做，程序是快速学习。这种动物模型的生成提供了一个有效率的制度，为外伤性癫痫的机制的研究。

Watch this Scientific Journal Video about Preparing Undercut Model of Posttraumatic Epileptogenesis in Rodents at JoVE.com

Preparing Undercut Model of Posttraumatic Epileptogenesis in Rodents

部分隔离皮层（"削弱"）是外伤性癫痫的有效的动物模型。在这里，我们将演示如何使一个新的手术设备，并用它来进行更精确和一致的病变，产生这种模式。

准备在鼠害外伤性癫痫的底切模型

Partially isolated cortex ("undercut") is an animal model of posttraumatic epileptogenesis. The surgical procedure involves cutting through the ...

preparing-undercut-model-of-posttraumatic-epileptogenesis-in-rodents

Nanyang Technological University

Research

JoVE Journal

Neuroscience

9.3K 次观看.  Indiana University School of Medicine. 部分隔离皮层（"削弱"）是外伤性癫痫的有效的动物模型。在这里，我们将演示如何使一个新的手术设备，并用它来进行更精确和一致的病变，产生这种模式。

视频: Preparing Undercut Model of Posttraumatic Epileptogenesis in Rodents

Habituation and Prepulse Inhibition of Acoustic Startle in Rodents

The acoustic startle response is a protective response, elicited by a sudden and intense acoustic stimulus. Facial and skeletal muscles are activated within a few milliseconds, leading to a whole body flinch in rodents1. Although startle responses are reflexive responses that can be reliably elicited, they are not stereotypic. They can be modulated by emotions such as fear (fear potentiated startle) and joy (joy attenuated startle), by non-associative learning processes such as habituation and sensitization, and by other sensory stimuli through sensory gating processes (prepulse inhibition), turning startle responses into an excellent tool for assessing emotions, learning, and sensory gating, for review see 2, 3. The primary pathway mediating startle responses is very short and well described, qualifying startle also as an excellent model for studying the underlying mechanisms for behavioural plasticity on a cellular/molecular level3.
We here describe a method for assessing short-term habituation, long-term habituation and prepulse inhibition of acoustic startle responses in rodents. Habituation describes the decrease of the startle response magnitude upon repeated presentation of the same stimulus. Habituation within a testing session is called short-term habituation (STH) and is reversible upon a period of several minutes without stimulation. Habituation between testing sessions is called long-term habituation (LTH)4. Habituation is stimulus specific5. Prepulse inhibition is the attenuation of a startle response by a preceding non-startling sensory stimulus6. The interval between prepulse and startle stimulus can vary from 6 to up to 2000 ms. The prepulse can be any modality, however, acoustic prepulses are the most commonly used.
Habituation is a form of non-associative learning. It can also be viewed as a form of sensory filtering, since it reduces the organisms' response to a non-threatening stimulus. Prepulse inhibition (PPI) was originally developed in human neuropsychiatric research as an operational measure for sensory gating7. PPI deficits may represent the interface of "psychosis and cognition" as they seem to predict cognitive impairment8-10. Both habituation and PPI are disrupted in patients suffering from schizophrenia11, and PPI disruptions have shown to be, at least in some cases, amenable to treatment with mostly atypical antipsychotics12, 13. However, other mental and neurodegenerative diseases are also accompanied by disruption in habituation and/or PPI, such as autism spectrum disorders (slower habituation), obsessive compulsive disorder, Tourette's syndrome, Huntington's disease, Parkinson's disease, and Alzheimer's Disease (PPI)11, 14, 15 Dopamine induced PPI deficits are a commonly used animal model for the screening of antipsychotic drugs16, but PPI deficits can also be induced by many other psychomimetic drugs, environmental modifications and surgical procedures.

Habituation and prepulse inhibition of startle are operational measures of sensory gating. Sensory gating is disrupted in schizophrenia, and some other mental disorders and neurodegenerative diseases. We here describe a standard protocol to assess short-term and long-term habituation as well as prepulse inhibition of acoustic startle responses in rats and mice.

在鼠害声不惊人死不休的习惯和前脉冲抑制

The acoustic startle response is a protective response, elicited by a sudden and intense acoustic stimulus. Facial and skeletal muscles are activated ...

科研

神经科学

Large-scale Recording of Neurons by Movable Silicon Probes in Behaving Rodents

A major challenge in neuroscience is linking behavior to the collective activity of neural assemblies. Understanding of input-output relationships of neurons and circuits requires methods with the spatial selectivity and temporal resolution appropriate for mechanistic analysis of neural ensembles in the behaving animal, i.e. recording of representatively large samples of isolated single neurons. Ensemble monitoring of neuronal activity has progressed remarkably in the past decade in both small and large-brained animals, including human subjects1-11. Multiple-site recording with silicon-based devices are particularly effective because of their scalability, small volume and geometric design.
Here, we describe methods for recording multiple single neurons and local field potential in behaving rodents, using commercially available micro-machined silicon probes with custom-made accessory components. There are two basic options for interfacing silicon probes to preamplifiers: printed circuit boards and flexible cables. Probe supplying companies (<a href="http://www.neuronexustech.com/" >http://www.neuronexustech.com/</a>; <a href="http://www.sbmicrosystems.com/">http://www.sbmicrosystems.com/</a>; <a href="http://www.acreo.se/">http://www.acreo.se/</a>) usually provide the bonding service and deliver probes bonded to printed circuit boards or flexible cables. Here, we describe the implantation of a 4-shank, 32-site probe attached to flexible polyimide cable, and mounted on a movable microdrive. Each step of the probe preparation, microdrive construction and surgery is illustrated so that the end user can easily replicate the process.

We describe methods for large-scale recording of multiple single units and local field potential in behaving rodents with silicon probes. Drive fabrication, probe attachment to the drive and probe implantation processes are illustrated in sufficient details for easy replication.

移动硅探针的神经行为鼠害的大型记录

A major challenge in neuroscience is linking behavior to the collective activity of neural assemblies. Understanding of input-output relationships of ...

A Low Cost Setup for Behavioral Audiometry in Rodents

In auditory animal research it is crucial to have precise information about basic hearing parameters of the animal subjects that are involved in the experiments. Such parameters may be physiological response characteristics of the auditory pathway, e.g. via brainstem audiometry (BERA). But these methods allow only indirect and uncertain extrapolations about the auditory percept that corresponds to these physiological parameters. To assess the perceptual level of hearing, behavioral methods have to be used. A potential problem with the use of behavioral methods for the description of perception in animal models is the fact that most of these methods involve some kind of learning paradigm before the subjects can be behaviorally tested, e.g. animals may have to learn to press a lever in response to a sound. As these learning paradigms change perception itself 1,2 they consequently will influence any result about perception obtained with these methods and therefore have to be interpreted with caution. Exceptions are paradigms that make use of reflex responses, because here no learning paradigms have to be carried out prior to perceptual testing. One such reflex response is the acoustic startle response (ASR) that can highly reproducibly be elicited with unexpected loud sounds in na&iuml;ve animals. This ASR in turn can be influenced by preceding sounds depending on the perceptibility of this preceding stimulus: Sounds well above hearing threshold will completely inhibit the amplitude of the ASR; sounds close to threshold will only slightly inhibit the ASR. This phenomenon is called pre-pulse inhibition (PPI) 3,4, and the amount of PPI on the ASR gradually depends on the perceptibility of the pre-pulse. PPI of the ASR is therefore well suited to determine behavioral audiograms in na&iuml;ve, non-trained animals, to determine hearing impairments or even to detect possible subjective tinnitus percepts in these animals. In this paper we demonstrate the use of this method in a rodent model (cf. also ref. 5), the Mongolian gerbil (Meriones unguiculatus), which is a well know model species for startle response research within the normal human hearing range (e.g. 6).

A fast and inexpensive method for the behavioral determination of hearing parameters like hearing thresholds, hearing impairments or phantom perceptions (subjective tinnitus) is described. It uses pre-pulse inhibition of the acoustic startle response and can be easily implemented in a personal computer using a programmable AD/DA-converter and a piezo sensor.

在啮齿动物的行为测听的低成本设置

In auditory animal research it is crucial to have precise information about basic hearing parameters of the animal subjects that are involved in the ...

Recording and Analysis of Circadian Rhythms in Running-wheel Activity in Rodents

When rodents have free access to a running wheel in their home cage, voluntary use of this wheel will depend on the time of day1-5. Nocturnal rodents, including rats, hamsters, and mice, are active during the night and relatively inactive during the day. Many other behavioral and physiological measures also exhibit daily rhythms, but in rodents, running-wheel activity serves as a particularly reliable and convenient measure of the output of the master circadian clock, the suprachiasmatic nucleus (SCN) of the hypothalamus. In general, through a process called entrainment, the daily pattern of running-wheel activity will naturally align with the environmental light-dark cycle (LD cycle; e.g. 12 hr-light:12 hr-dark). However circadian rhythms are endogenously generated patterns in behavior that exhibit a ~24 hr period, and persist in constant darkness. Thus, in the absence of an LD cycle, the recording and analysis of running-wheel activity can be used to determine the subjective time-of-day. Because these rhythms are directed by the circadian clock the subjective time-of-day is referred to as the circadian time (CT). In contrast, when an LD cycle is present, the time-of-day that is determined by the environmental LD cycle is called the zeitgeber time (ZT).
Although circadian rhythms in running-wheel activity are typically linked to the SCN clock6-8, circadian oscillators in many other regions of the brain and body9-14 could also be involved in the regulation of daily activity rhythms. For instance, daily rhythms in food-anticipatory activity do not require the SCN15,16 and instead, are correlated with changes in the activity of extra-SCN oscillators17-20. Thus, running-wheel activity recordings can provide important behavioral information not only about the output of the master SCN clock, but also on the activity of extra-SCN oscillators. Below we describe the equipment and methods used to record, analyze and display circadian locomotor activity rhythms in laboratory rodents.

Circadian rhythms in voluntary wheel-running activity in mammals are tightly coupled to the molecular oscillations of a master clock in the brain. As such, these daily rhythms in behavior can be used to study the influence of genetic, pharmacological, and environmental factors on the functioning of this circadian clock.

运行轮活动在啮齿类动物的昼夜节律的记录和分析

When rodents have free access to a running wheel in their home cage, voluntary use of this wheel will depend on the time of day1-5. Nocturnal rodents, ...

Deep Brain Stimulation with Simultaneous fMRI in Rodents

In order to visualize the global and downstream neuronal responses to deep brain stimulation (DBS) at various targets, we have developed a protocol for using blood oxygen level dependent (BOLD) functional magnetic resonance imaging (fMRI) to image rodents with simultaneous DBS. DBS fMRI presents a number of technical challenges, including accuracy of electrode implantation, MR artifacts created by the electrode, choice of anesthesia and paralytic to minimize any neuronal effects while simultaneously eliminating animal motion, and maintenance of physiological parameters, deviation from which can confound the BOLD signal.&#160;Our laboratory has developed a set of procedures that are capable of overcoming most of these possible issues. For electrical stimulation, a homemade tungsten bipolar microelectrode is used, inserted stereotactically at the stimulation site in the anesthetized subject. In preparation for imaging, rodents are fixed on a plastic headpiece and transferred to the magnet bore. For sedation and paralysis during scanning, a cocktail of dexmedetomidine and pancuronium is continuously infused, along with a minimal dose of isoflurane; this preparation minimizes the BOLD ceiling effect of volatile anesthetics. In this example experiment, stimulation of the subthalamic nucleus (STN) produces BOLD responses which are observed primarily in ipsilateral cortical regions, centered in motor cortex. Simultaneous DBS and fMRI allows the unambiguous modulation of neural circuits dependent on stimulation location and stimulation parameters, and permits observation of neuronal modulations free of regional bias. This technique may be used to explore the downstream effects of modulating neural circuitry at nearly any brain region, with implications for both experimental and clinical DBS.

This protocol describes a standard method for simultaneous functional magnetic resonance imaging and deep brain stimulation in the rodent. The combined use of these experimental tools allows for the exploration of global downstream activity in response to electrical stimulation at virtually any brain target.

深部脑刺激与同步功能磁共振成像在鼠害

In order to visualize the global and downstream neuronal responses to deep brain stimulation (DBS) at various targets, we have developed a protocol for ...

Acrylic Resin Molding Based Head Fixation Technique in Rodents

Head fixation is a technique of immobilizing animal's head by attaching a head-post on the skull for rigid clamping. Traditional head fixation requires surgical attachment of metallic frames on the skull. The attached frames are then clamped to a stationary platform resulting in immobilization of the head. However, metallic frames for head fixation have been technically difficult to design and implement in general laboratory environment. In this study, we provide a novel head fixation method. Using a custom-made head fixation bar, head mounter is constructed during implantation surgery. After the application of acrylic resin for affixing implants such as electrodes and cannula on the skull, additional resins applied on top of that to build a mold matching to the port of the fixation bar. The molded head mounter serves as a guide rails, investigators conveniently fixate the animal&#8217;s head by inserting the head mounter into the port of the fixation bar. This method could be easily applicable if implantation surgery using dental acrylics is necessary and might be useful for laboratories that cannot easily fabricate CNC machined metal head-posts.

Traditional head fixation techniques have used metallic frames attached on the skull as an interface for head fixation apparatus. This protocol demonstrates a frameless, acrylic resin molding based head fixation technique for behavioral tasks in rodents.

在鼠害丙烯酸树脂模塑基头固定技术

Head fixation is a technique of immobilizing animal's head by attaching a head-post on the skull for rigid clamping. Traditional head fixation requires ...

Transcranial Electrical Brain Stimulation in Alert Rodents

Transcranial electrical brain stimulation can modulate cortical excitability and plasticity in humans and rodents. The most common form of stimulation in humans is transcranial direct current stimulation (tDCS). Less frequently, transcranial alternating current stimulation (tACS) or transcranial random noise stimulation (tRNS), a specific form of tACS using an electrical current applied randomly within a pre-defined frequency range, is used. The increase of noninvasive electrical brain stimulation research in humans, both for experimental and clinical purposes, has yielded an increased need for basic, mechanistic, safety studies in animals. This article describes a model for transcranial electrical brain stimulation (tES) through the intact skull targeting the motor system in alert rodents. The protocol provides step-by-step instructions for the surgical set-up of a permanent epicranial electrode socket combined with an implanted counter electrode on the chest. By placing a stimulation electrode into the epicranial socket, different electrical stimulation types, comparable to tDCS, tACS, and tRNS in humans, can be delivered. Moreover, the practical steps for tES in alert rodents are introduced. The applied current density, stimulation duration, and stimulation type may be chosen depending on the experimental needs. The caveats, advantages, and disadvantages of this set-up are discussed, as well as safety and tolerability aspects.

This protocol describes a surgical set-up for a permanent epicranial electrode socket and an implanted chest electrode in rodents. By placing a second electrode into the socket, different types of transcranial electrical brain stimulation can be delivered to the motor system in alert animals through the intact skull.

警惕啮齿动物的经颅电脑刺激

Transcranial electrical brain stimulation can modulate cortical excitability and plasticity in humans and rodents. The most common form of stimulation in ...

Cannula Implantation into the Cisterna Magna of Rodents

Cisterna magna cannulation (CMc) is a straightforward procedure that enables direct access to the cerebrospinal fluid (CSF) without operative damage to the skull or the brain parenchyma. In anesthetized rodents, the exposure of the dura mater by blunt dissection of the neck muscles allows the insertion of a cannula into the cisterna magna (CM). The cannula, composed either by a fine beveled needle or borosilicate capillary, is attached via a polyethylene (PE) tube to a syringe. Using a syringe pump, molecules can then be injected at controlled rates directly into the CM, which is continuous with the subarachnoid space. From the subarachnoid space, we can trace CSF fluxes by convective flow into the perivascular space around penetrating arterioles, where solute exchange with the interstitial fluid (ISF) occurs. CMc can be performed for acute injections immediately following the surgery, or for chronic implantation, with later injection in anesthetized or awake, freely moving rodents. Quantitation of tracer distribution in the brain parenchyma can be performed by epifluorescence, 2-photon microscopy, and magnetic resonance imaging (MRI), depending on the physico-chemical properties of the injected molecules. Thus, CMc in conjunction with various imaging techniques offers a powerful tool for assessment of the glymphatic system and CSF dynamics and function. Furthermore, CMc can be utilized as a conduit for fast, brain-wide delivery of signaling molecules and metabolic substrates that could not otherwise cross the blood brain barrier (BBB).

Here we describe a protocol to perform cisterna magna cannulation (CMc), a minimally invasive way to deliver tracers, substrates and signaling molecules into the cerebrospinal fluid (CSF). Combined with different imaging modalities, CMc enables glymphatic system and CSF dynamics assessment, as well as brain-wide delivery of various compounds.

将套管植入啮齿动物的麦格纳池中

Cisterna magna cannulation (CMc) is a straightforward procedure that enables direct access to the cerebrospinal fluid (CSF) without operative damage to ...

Intracranial Pharmacotherapy and Pain Assays in Rodents

Pain is a salient sensory experience with affective and cognitive dimensions. However, central mechanisms for pain remain poorly understood, hindering the development of effective therapeutics. Intracranial pharmacology presents an important tool for understanding the molecular and cellular mechanisms of pain in the brain, as well as for novel treatments. Here we present a protocol that integrates intracranial pharmacology with pain behavior testing. Specifically, we show how to infuse analgesic drugs into a select brain region, which may be responsible for pain modulation. Furthermore, to determine the effect of the candidate drug in the central nerve system, pain assays are performed after intracranial treatment. Our results demonstrate that intracranial administration of analgesic drugs in a targeted region can provide relief of pain in rodents. Thus, our protocol successfully demonstrates that intracranial pharmacology, combined with pain behavior testing, can be a powerful tool for the study of pain mechanisms in the brain.

Here we present a protocol to perform intracranial pharmacological experiments followed by pain behavior assays in rodents. This protocol allows researchers to deliver molecular and cellular targets in the brain, for pharmacologic agents in the treatment of pain.

颅内药物治疗和啮齿动物疼痛检测

Pain is a salient sensory experience with affective and cognitive dimensions. However, central mechanisms for pain remain poorly understood, hindering the ...

The Power of Interstimulus Interval for the Assessment of Temporal Processing in Rodents

Temporal processing deficits have been implicated as a potential elemental dimension of higher-level cognitive processes, commonly observed in neurocognitive disorders. Despite the popularization of prepulse inhibition (PPI) in recent years, many current protocols promote using a percent of control measure, thereby precluding the assessment of temporal processing. The present study used cross-modal PPI and gap prepulse inhibition (gap-PPI) to demonstrate the benefits of employing a range of interstimulus intervals (ISIs) to delineate effects of sensory modality, psychostimulant exposure, and age. Assessment of sensory modality, psychostimulant exposure, and age reveals the utility of an approach varying the interstimulus interval (ISI) to establish the shape of the ISI function, including increases (sharper curve inflections) or decreases (flattening of the response amplitude curve) in startle amplitude. Additionally, shifts in peak response inhibition, suggestive of a differential sensitivity to the manipulation of ISI, are often revealed. Thus, the systematic manipulation of ISI affords a critical opportunity to evaluate temporal processing, which may reveal the underlying neural mechanisms involved in neurocognitive disorders.

Temporal processing, a preattentive process, may underlie deficits in higher-level cognitive processes, including attention, commonly observed in neurocognitive disorders. Using prepulse inhibition as an exemplar paradigm, we present a protocol for manipulating interstimulus interval (ISI) to establish the shape of the ISI function to provide an assessment of temporal processing.