1。畜舍和驯化程序<ol><li>男性长埃文斯大鼠，体重300-325克，在动植物公园的到来的时间单独采用标准的塑料笼中含有再生纸颗粒床上用品，用铁丝电网盖容纳瓶装水和食品。 </li><li>老鼠保持反向明暗的时间表（19:00至07:00的灯光），在温度和湿度控制动植物公园。 </li><li>食物和水，可随意在整个实验家笼。 </li><li>老鼠可以使其恢复到手术前一个星期的动植物公园和住房条件。在这段时间内大鼠不受干扰，除了日常的清洁，喂食，体重监测。 </li></ol> 2。静脉导管手术<ol><li>在异氟醚气体麻醉，并使用标准的无菌手术技术，老鼠准备与硅橡胶在右颈静脉导管。到修改后的导管（塑料一）在头骨表面，和导管，导管退出安装的头骨与珠宝商“螺钉和牙科水泥。手术过程中的详细信息，请参阅 7 ，8。 </li><li>在手术结束时，导管冲洗约0.2毫升含有50％的肝素（1000 IU）和50％的葡萄糖（25 g/50毫升）的解决方案。一个塑料拦截器是放置在开放的套管，以保护外部碎片的导管，保持导管通畅。 </li><li>在手术时间，并再次手术后的一天，大鼠的镇痛酮洛芬管理5毫克/公斤，SC。 </li><li>允许大鼠实验手术前至少7天才能恢复。在这段时间里，老鼠留在家里的笼子里不受干扰，除日常清洁和喂养，重量和一般健康状况的日常监测。在复苏期的最后一天，与肝素/葡萄糖溶液（2.2节），以确认导管通畅导管刷新。 </li></ol> 3。行为程序利用MED Associates公司（VT），圣奥尔本斯，提供的设备，所有的行为的程序进行。设备，安置在一个专门的房间，包括一个标准的操作性商会，每一个声音衰减外壳内包含的设置。每个操作性室配有两个可伸缩杆，都升高6.5厘米以上的不锈钢棒地板。一个白色的刺激指示灯位于正上方一杆，输液泵（Razel科学仪器，圣奥尔本斯的刺激轻，激活的同时照明，杆（所谓的“积极的”杠杆）结果的反应， VT）。该泵是位于城外操作性室的架子上，但机箱内的声音衰减。其他杆（所谓的“不活跃”的杠杆）是留在会议厅内扩展到所有的实验程序;响应此杆，而后果是。最后通过加扰器提供恒流，间歇性的，不可避免的，电动电击钢线材的地板上，每室配备。所有的实验条件下（例如，激励演讲，药物注射，电击），以及数据记录和整合，通过的试验室和一台中央计算机操作系统MED Associates的软件（MED - IV）之间的接口完成。 实验在5个阶段进行：1）习惯; 2）可卡因的SA; 3）药物无期; 4）灭绝，和5）测试复职。在所有阶段（为无毒的时期除外），老鼠被转移从房屋室内操作性试验室的日常会话。动物是单独转让，在小塑料容器。食物和水，不具备操作性商会。 <ol><li>习惯<ol><li>二十四小时前的第一可卡因SA届会议（见3.2节），老鼠是允许在一个单一的2 - H会议的操作性商会habituate;在本届会议期间的活动杆缩回。 </li></ol></li><li>可卡因的SA培训<ol><li>每天的训练课之前，老鼠是衡量在住房里。随后他们被转移到操作性商会。 </li><li>一旦被安置在操作性商会，弹簧管总成（延长药物输液泵）是连接到头骨上的套管。 </li><li>老鼠被允许在日常的自我管理可卡因盐酸3 h会议，共8-10天，（连续或两个干预“天”之间的会议5和6）。 </li><li> SA会议之间的早上（9:00-12:00）和下午（13:00-16:00）备用。 SA的会议时间是交替的，因为在随后的实验（即灭绝和测试）阶段的程序都出现吨对所有动物的IME一天（见第3.4和3.5），因此，这种安排可以防止任何一天的时间在随后的毒品寻求行为混杂影响。 </li><li>每次训练前一个5分钟的驯化期，在此期间，动物翻供的积极杠杆的操作性商会。 </li><li>在培训会议开始时，进入会议厅，并扩展活动的杠杆，刺激光（位于上面的积极杠杆）是30照亮第此外，房子光在这个时候被激活，并保持在整个会议期间照亮。总之，这些事件信号的可卡因供应。 </li><li>会议期间，活动杠杆的反应是强化了可卡因的3 - S输液固定比率1加固的时间表（0.23 mg/65微升，IV）。激活输液泵和药物输送的同时，上述活动的杠杆刺激光的20 - S激活。这20 - S光介绍对应一个“超时”期间这额外的积极杠杆的反应被记录，但没有加固。 </li><li>动物被允许最多每3小时会议的50输注。我们经常要求我们的工作标准，在我们的经验，发现，它大大降低，如果不消除，由于过量而死亡的事件。 </li><li>在每天的会议结束时，老鼠的导管冲洗0.2-0.3毫升的肝素葡萄糖溶液（见2.2节）和老鼠返回自己的家乡笼。 </li></ol></li><li>无毒期<ol><li>最终可卡因SA会议后，大鼠不受干扰（除了日常的清洁，喂食，体重监测）最少7天的家庭笼。 </li></ol></li><li>消可卡因训练有素的行为<ol><li>老鼠被转移到09:00-10:00之间的灭绝培训的操作性商会。 </li><li>春天和油管大会是连接到颅骨安装套管。 </li><li> SA的培训条件，每天灭绝的培训是前一个5分钟的驯化期，在此期间，动物翻供的积极杠杆的操作性商会。 </li><li>老鼠然后给出了四个60分钟灭绝会议，30分钟的时间间隔分开。每60分钟的会议是由相同的事件发生在培训班开始（也就是说，房子光照明，延长了积极的杠杆，和30秒以上活跃杠杆的刺激光照明）发起。整个会议的所有条件，目前在南非训练保持（即刺激光响应队伍的照度为20秒和3 - S输液泵的激活），除了积极杠杆的反应不再是钢筋可卡因注射。在30分钟的间隔分开每60分钟灭绝会议，活动杆收回房子的灯熄灭。 </li><li>在最后的60分钟会议结束时，老鼠是返回自己的家乡笼。 </li><li>步骤3.4.1。 3.4.5。连续3天重复。 </li></ol></li><li>电击压力恢复可卡因训练有素的行为<ol><li>大鼠被转移到09:00-10:00之间的操作性商会</li><li>在被安置在操作性室，弹簧和油管总成连接到颅骨安装套管。 </li><li>培训和灭绝的条件，提供了一个5分钟的驯化期，在此期间，大鼠的积极杠杆的操作性商会收回。 </li><li>大鼠，然后给60分钟灭绝的两个会议（见3.4.4）一个30分钟的间隔隔开，在灭绝阶段给予的相同。 </li><li>如果第二个60分钟的会议后的反应以前的活动杆的总人数是小于15，动物都给予复职的考验后，一个额外的30分钟间隔。在这30分钟的时间间隔，电击或电击深水（即，没有电击）管理（见3.5.6）。如果一个标准的15个或更少的积极杠杆的反应是不是取得了第二届会议后，一个额外的灭绝会议（S）的事先测试复职。 </li><li>十几分钟后第二个60分钟灭绝会议（或之后的任何额外的60分钟灭绝会议（S）建立了15个或更少的响应灭绝标准），动物暴露于间歇性的电动电击压力的20分钟（或假电击）在操作性室。电击是在一个活动杆缩回，房子和刺激的灯都熄灭时的时间。电击是根据一个变量的平均间隔40秒（10-70 s范围内）的时间表交付。每个冲击强度为0.9毫安交付，并在持续时间为0.5小号。电击这些参数是基于广泛的试点工作在我们的实验室，建立可靠的恢复，与最低的强度和持续时间电击曝光寻求可卡因。根据这些参数，动物展览自反跳跃的反应，以0.5秒的冲击，并偶尔发出一个简单的发声。虽然整个休克期观察动物，冲击强度足以诱发遇险的迹象更加明显。 </li><li>立即终止后的电击会议，活动的杠杆扩展和老鼠灭绝的条件下（即允许按杆，反应不加强与可卡因，但继续刺激光照明和输液泵的激活导致）。这段时间被定义为复职的考验，持续时间通常在1-3个小时。大多数响应发生在早期测试会话部分（第30分钟，1小时），随后更多的扩展测试期间熄灭。因此，1小时的测试足以捕获的修复效果，较长的测试周期的效果提供更完整的灭绝。 </li><li>当采用重复测量设计，每只大鼠电击和深水电击条件下进行测试。步骤3.5.1。在随后的测试一天重复到3.5.8，管理复职（电击或电击深水）的替代试验。 </li></ol></li></ol> 4。代表性的成果： <img src="/files/ftp_upload/2265/2265fig1.jpg" alt="图1" /> 图1。程式化自我管理培训，灭绝和恢复测试期间的活动和非活动的杠杆活动响应型材。 图1给出了程式化的操作性响应型材，在此处所描述的实验协议的各个阶段中取得的典型。 1。主动杆反应<ol><li> 可卡因SA的培训。图1（左图）显示了一个典型的可卡因SA训练曲线，包括输液和积极杠杆压力机总数。大鼠一般在1-3会话收购可卡因的SA和行为一旦建立，维持稳定的药物摄入率超过以后各届会议（通常低于20％之间的会话方差）。 </li><li>可以看出，在图1（中间面板）灭绝的第1天，当反应是首次非增强，积极杠杆的答复总数最初是高于在灭绝可卡因训练有素的行为。结束训练阶段（对应一个所谓的“灭绝爆裂”）;渐渐地，在四个灭绝环节，响应减小。灭绝的2和3天，可观察到的响应（相对于前一天的最后一次会议期间指出，）有所恢复;然而，反应一般比灭绝的前一天开始减少和灭绝一般当天四次会议的收益更为迅速。 </li><li> 寻求重新可卡因 ，它可以在图1（右面板）看到，20分钟暴露于间歇性电击诱导增加活动的杠杆，相对前灭绝会议结束时指出，加强非响应。电击也导致在深水电击管理的条件下观察到的响应相对增加。因此，恢复电击强调寻求可卡因可表示为：1）非增强反应暴露后，以电击的次数增加，相对反应在灭绝会议之前电击或2）非增强的数量增加深水电击电击相对的反应。 </li></ol> 2。无效杆反应可以看出，在图1（红色线），在整个实验阶段，有非常低的响应水平，与不活动的杠杆相关。然而，在响应此杆的适度增加，有时观察以下接触电击强调。即使在增加在应对发生的案件中，在积极杠杆的增加，响应趋向于按比例更大。

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动物实验在动物保健和加拿大多伦多动物保健委员会大学的加拿大会规定的准则和法规的规定执行。生产此视频赞助MED Associates公司的生产在这篇文章中使用的仪器。

在过去几十年，已发展到3 SA的恢复过程中的变化研究在实验室老鼠和猴子寻求药物复职。定义程序上的差异的差异主要药物SA，物种灭绝和测试会话是否发生在同一天或在不同的日子1。我们在本文中描述的过程就是通常所说的“之间的”会话的变化，因为药物SA出现在日常会话内（即在会议之间的方式），灭绝和测试阶段分开，但灭绝和测试发生在同一天（即在一个内部会议的方式）[例如，7，9] 。另外，在所谓的“会话之间”的程序，实验的各个阶段发生的顺序，如SA，消，和测试会在不同的日子（即会话之间的所有阶段）[例如，8，10 ， 11]。与此相反，在完成“内部会议”的程序，SA培训，灭绝，测试阶段都发生在同一天，例如，12] 。 虽然有电击引起的药物寻求复职程序使用不同的变化复职发表的报告，我们发现这应激（至少在一个可卡因SA的历史动物）是最有效的的灭绝和测试时发生在同一日（即，在“内”会议的方式）。另外，我们不得不寻求使用“会话之间的”完全的程序，成功地获得可靠的电击诱发可卡因复职，但只在动物整个实验8，或灭绝和测试过程中至少设在SA商会的条件13。这间会议的做法，超过在本议定书中所描述的缺点是，它限制，可在一次训练和测试的科目数量，从而降低数据收集的效率。虽然本议定书的条件下可以熄灭，只有一组动物在特定的时间进行测试，一个更大的吞吐量可以达到惊人的SA训练阶段两组大鼠的开始时间一个星期。在这种情况下，第一组的老鼠可以进入无毒的阶段，第二组进入第2周的SA培训，随后，灭绝和测试可以在第一组进行，而第二组是在其药物无相。这种方法可以更有效地利用资源和加快数据采集速度，而导致非常有效和可靠的的电击诱发复职。 更重要的是，在我们的程序，以前增强药物的行为和恢复测试的灭绝发生在与毒品相关的线索的情况。虽然我们从来没有开展药物相关线索的情况下灭绝培训或测试，这是我们的理解，基于个人通信的应激可能实际上是在诱导复职无效的，测试时，在扑灭线索的情况下发生的（至少在动物与可卡因SA的历史情况下）。这个变量系统的检查，和它带来的理论意义，是一个重要的问题，为今后的研究。 程序的最后一个因素值得评论的是，电击强调有关复职的效果似乎是高度敏感的背景下操作。例如，电击强调只有​​在有效地诱导的药物如果是药物SA和灭绝发生在环境管理寻求复职;，电击不恢复在一个新的背景下如果发生接触的应激药物寻求，提示应激和药物的影响复职14，环 ​​境之间的重要互动。 总之，电击恢复程序，如本文中所描述的，已经使用相当大的成功作出澄清的压力和药物复发之间的关系的基本行为和神经生物学机制。事实上，这些程序，修改，以适应药理预处理或针对特定神经元基板的神经化学物质的操作时，可以作为强大的工具，药物强调寻求复职的神经生物学特征。

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<td>Basic Drug Self-Administration Test Package for Rat Package</td>
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<td>Deep extra tall MDF sound attenuating cubicle</td>
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<td>Standard modular test chamber with drug infusion top and waste pan</td>
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<td><a href="http://www.med-associates.com/">Med Associates, Inc</a></td>
<td>ENV-008CT</td>
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<td>Stainless steel grid floor</td>
<td>Basic Drug Self-Administration Test Package for Rat Package Component</td>
<td><a href="http://www.med-associates.com/">Med Associates, Inc</a></td>
<td>ENV-005</td>
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<td>Basic Drug Self-Administration Test Package for Rat Package Component</td>
<td><a href="http://www.med-associates.com/">Med Associates, Inc</a></td>
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<td>Basic Drug Self-Administration Test Package for Rat Package Component</td>
<td><a href="http://www.med-associates.com/">Med Associates, Inc</a></td>
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<td>Basic Drug Self-Administration Test Package for Rat Package Component</td>
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<td>SmartCtrl 8 Input/16 output package</td>
<td>Basic Drug Self-Administration Test Package for Rat Package Component</td>
<td><a href="http://www.med-associates.com/">Med Associates, Inc</a></td>
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<td>Basic Drug Self-Administration Test Package for Rat Package Component</td>
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<td>Basic Drug Self-Administration Test Package for Rat Package Component</td>
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<td>Drug Delivery, Arm assembly </td>
<td>Basic Drug Self-Administration Test Package for Rat Package Component</td>
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<td>PHM-110-SAI</td>
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<td>Square wave aversive stimulator</td>
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<td><a href="http://www.med-associates.com/">Med Associates, Inc</a></td>
<td>ENV-414S</td>
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<td>Patented quick disconnect harness for stainless steel grid floor</td>
<td>Footshock Stress Component</td>
<td><a href="http://www.med-associates.com/">Med Associates, Inc</a></td>
<td>ENV-005-QD</td>
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<td>Shock output cable</td>
<td>Footshock Stress Component</td>
<td><a href="http://www.med-associates.com/">Med Associates, Inc</a></td>
<td>SG-219G-10</td>
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<td>MED-PC IV Software</td>
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a procedure for studying the footshock-induced reinstatement of cocaine seeking in laboratory rats

最阴险的吸毒方面是高复发倾向。动物模型复发，被称为复职程序，已被广泛用于研究神经生物学和现象复发的药物使用。虽然在过去的几十年中出现了程序上的差异，最传统的复职程序是基于对药物的自我管理（SA）模型。在此模型中，动物是训练有素的执行操作性的响应，获得药物。随后，该行为是代扣响应队伍加固熄灭。然后触发由一个独立的事件，如注射培训的药物，重新接触到药物相关线索，或接触到应激1，药物寻求复职。 复职程序最初开发研究急性非培训药物，以寻求恢复的药物在老鼠和猴子1，2的队伍暴露的能力。复职程序，至今已修改，以研究环境的刺激，包括与毒品相关的线索和曝光，以各种形式的压力的作用，在复发的药物，寻求1， 3，4。 在过去的15年，恢复文学的主要重点一直在药物的作用复发的压力。压力的最常用的形式之一，为研究这种关系是轻度的，间歇性的，电动footshocks急性暴露。电击压力的能力，以诱导恢复寻求药物原是沙哈姆和他的同事（1995）表明，在大鼠的静脉注射海洛因SA 5的历史。随后，效果是广义大鼠静脉注射可卡因，甲基安非他明和尼古丁的SA，以及6 SA 3，口服乙醇的历史。 虽然电击引起的药物寻求复职可以实现可靠和有力的，它是一种效果，往往是敏感的某些参数化变量。这些措施包括灭绝和恢复测试会议的安排，电击应力的强度和持续时间，和在场的灭绝和测试复职过程中与毒品相关的线索。在这里，我们提出了一个电击诱发的要求，我们有一致的成功用于研究的压力和可卡因寻求之间的关系可卡因复职的协议。

Watch this Scientific Journal Video about A Procedure for Studying the Footshock-Induced Reinstatement of Cocaine Seeking in Laboratory Rats at JoVE.com

A Procedure for Studying the Footshock-Induced Reinstatement of Cocaine Seeking in Laboratory Rats

动物模型复发，被称为复职程序，已被广泛用于研究复发的药物寻求在应力的作用。在这里，我们报告恢复寻求通过在实验室老鼠轻微，间歇性的电动电击急性暴露可卡因诱导的方法。

在实验室老鼠研究求可卡因电击诱导恢复原状的程序

The most insidious aspect of drug addiction is the high propensity for relapse.  Animal models of relapse, known as reinstatement procedures, have been ...

a-procedure-for-studying-footshock-induced-reinstatement-cocaine

Research

JoVE Journal

Neuroscience

20.7K 次观看.  University of Toronto Scarborough. 动物模型复发，被称为复职程序，已被广泛用于研究复发的药物寻求在应力的作用。在这里，我们报告恢复寻求通过在实验室老鼠轻微，间歇性的电动电击急性暴露可卡因诱导的方法。

视频: A Procedure for Studying the Footshock-Induced Reinstatement of Cocaine Seeking in Laboratory Rats

Operant Sensation Seeking in the Mouse

Operant methods are powerful behavioral tools for the study of motivated behavior. These 'self-administration' methods have been used extensively in drug addiction research due to their high construct validity. Operant studies provide researchers a tool for preclinical investigation of several aspects of the addiction process. For example, mechanisms of acute reinforcement (both drug and non-drug) can be tested using pharmacological or genetic tools to determine the ability of a molecular target to influence self-administration behavior1-6. Additionally, drug or food seeking behaviors can be studied in the absence of the primary reinforcer, and the ability of pharmacological compounds to disrupt this process is a preclinical model for discovery of molecular targets and compounds that may be useful for the treatment of addiction3,7-9. One problem with performing intravenous drug self-administration studies in the mouse is the technical difficulty of maintaining catheter patency. Attrition rates in these experiments are high and can reach 40% or higher10-15. Another general problem with drug self-administration is discerning which pharmacologically-induced effects of the reinforcer produce specific behaviors. For example, measurement of the reinforcing and neurological effects of psychostimulants can be confounded by their psychomotor effects. Operant methods using food reinforcement can avoid these pitfalls, although their utility in studying drug addiction is limited by the fact that some manipulations that alter drug self-administration have a minimal impact on food self-administration. For example, mesolimbic dopamine lesion or knockout of the D1 dopamine receptor reduce cocaine self-administration without having a significant impact on food self-administration 12,16. 
Sensory stimuli have been described for their ability to support operant responding as primary reinforcers (i.e. not conditioned reinforcers)17-22. Auditory and visual stimuli are self-administered by several species18,21,23, although surprisingly little is known about the neural mechanisms underlying this reinforcement. The operant sensation seeking (OSS) model is a robust model for obtaining sensory self-administration in the mouse, allowing the study of neural mechanisms important in sensory reinforcement24. An additional advantage of OSS is the ability to screen mutant mice for differences in operant behavior that may be relevant to addiction. We have reported that dopamine D1 receptor knockout mice, previously shown to be deficient in psychostimulant self-administration, also fail to acquire OSS24. This is a unique finding in that these mice are capable of learning an operant task when food is used as a reinforcer. While operant studies using food reinforcement can be useful in the study of general motivated behavior and the mechanisms underlying food reinforcement, as mentioned above, these studies are limited in their application to studying molecular mechanisms of drug addiction. Thus, there may be similar neural substrates mediating sensory and psychostimulant reinforcement that are distinct from food reinforcement, which would make OSS a particularly attractive model for the study of drug addiction processes. The degree of overlap between other molecular targets of OSS and drug reinforcers is unclear, but is a topic that we are currently pursuing. While some aspects of addiction such as resistance to extinction may be observed with OSS, we have found that escalation 25 is not observed in this model24. Interestingly, escalation of intake and some other aspects of addiction are observed with self-administration of sucrose26. Thus, when non-drug operant procedures are desired to study addiction-related processes, food or sensory reinforcers can be chosen to best fit the particular question being asked. 
In conclusion, both food self-administration and OSS in the mouse have the advantage of not requiring an intravenous catheter, which allows a higher throughput means to study the effects of pharmacological or genetic manipulation of neural targets involved in motivation. While operant testing using food as a reinforcer is particularly useful in the study of the regulation of food intake, OSS is particularly apt for studying reinforcement mechanisms of sensory stimuli and may have broad applicability to novelty seeking and addiction.

In this protocol we describe a method of operant learning using sensory stimuli as a reinforcer in the mouse. It requires no prior training or food restriction, and it allows the study of motivated behavior without the use of a pharmacological or natural reinforcer such as food.

在鼠标的操作性感觉寻求

Operant methods are powerful behavioral tools for the study of motivated behavior.  These 'self-administration' methods have been used extensively in drug ...

科研

神经科学

Muscle Receptor Organs in the Crayfish Abdomen: A Student Laboratory Exercise in Proprioception

The primary purpose of this experiment is to demonstrate primary sensory neurons conveying information of joint movements and positions as proprioceptive information for an animal. An additional objective of this experiment is to learn anatomy of the preparation by staining, dissection and viewing of neurons and sensory structures under a dissecting microscope. This is performed by using basic neurophysiological equipment to record the electrical activity from a joint receptor organ and staining techniques. The muscle receptor organ (MRO) system in the crayfish is analogous to the intrafusal muscle spindle in mammals, which aids in serving as a comparative model that is more readily accessible for electrophysiological recordings. In addition, these are identifiable sensory neurons among preparations. The preparation is viable in a minimal saline for hours which is amenable for student laboratory exercises. The MRO is also susceptible to neuromodulation which encourages intriguing questions in the sites of modulatory action and integration of dynamic signals of movements and static position along with a gain that can be changed in the system.

The primary purpose of this experiment is to understand how primary sensory neurons convey information of joint movements and positions as proprioceptive information for an animal. An additional objective of this report is present the anatomy of the preparation by dissection and viewing of neurons under a dissecting microscope.

小龙虾腹部肌肉受体机关：在本体的学生实验室练习

The primary purpose of this experiment is to demonstrate primary sensory neurons conveying information of joint movements and positions as proprioceptive ...

A General Method for Evaluating Incubation of Sucrose Craving in Rats

For someone on a food-restricted diet, food craving in response to food-paired cues may serve as a key behavioral transition point between abstinence and relapse to food taking 1. Food craving conceptualized in this way is akin to drug craving in response to drug-paired cues. A rich literature has been developed around understanding the behavioral and neurobiological determinants of drug craving; we and others have been focusing recently on translating techniques from basic addiction research to better understand addiction-like behaviors related to food 2-4.
As done in previous studies of drug craving, we examine sucrose craving behavior by utilizing a rat model of relapse. In this model, rats self-administer either drug or food in sessions over several days. In a session, lever responding delivers the reward along with a tone+light stimulus. Craving behavior is then operationally defined as responding in a subsequent session where the reward is not available. Rats will reliably respond for the tone+light stimulus, likely due to its acquired conditioned reinforcing properties 5. This behavior is sometimes referred to as sucrose seeking or cue reactivity. In the present discussion we will use the term "sucrose craving" to subsume both of these constructs.
In the past decade, we have focused on how the length of time following reward self-administration influences reward craving. Interestingly, rats increase responding for the reward-paired cue over the course of several weeks of a period of forced-abstinence. This "incubation of craving" is observed in rats that have self-administered either food or drugs of abuse 4,6. This time-dependent increase in craving we have identified in the animal model may have great potential relevance to human drug and food addiction behaviors.
Here we present a protocol for assessing incubation of sucrose craving in rats. Variants of the procedure will be indicated where craving is assessed as responding for a discrete sucrose-paired cue following extinction of lever pressing within the sucrose self-administration context (Extinction without cues) or as responding for sucrose-paired cues in a general extinction context (Extinction with cues).

Responding for food or drug-paired cues increases over the course of a period of abstinence and this may relate to an increased susceptibility to relapse behaviors. Here we detail a procedure for evaluating this "incubation of craving" in rats that have self-administered sucrose.

评估大鼠蔗糖渴求育成的一个通用方法

For someone on a food-restricted diet, food craving in response to food-paired cues may serve as a key behavioral transition point between abstinence and ...

Rapid Determination of the Thermal Nociceptive Threshold in Diabetic Rats

Painful diabetic neuropathy (PDN) is characterized by hyperalgesia i.e., increased sensitivity to noxious stimulus, and allodynia i.e., hypersensitivity to normally innocuous stimuli1. Hyperalgesia and allodynia have been studied in many different rodent models of diabetes mellitus2. However, as stated by B&ouml;lcskei et al, determination of "pain" in animal models is challenging due to its subjective nature3. Moreover, the traditional methods used to determine behavioral responses to noxious thermal stimuli usually lack reproducibility and pharmacological sensitivity3. For instance, by using the hot-plate method of Ankier4, flinch, withdrawal and/or licking of either hind- and/or fore-paws is quantified as reflex latencies at constant high thermal stimuli (52-55 &deg;C). However, animals that are hyperalgesic to thermal stimulus do not reproducibly show differences in reflex latencies using those supra-threshold temperatures3,5. As the recently described method of B&ouml;lcskei et al.6, the procedures described here allows for the rapid, sensitive and reproducible determination of thermal nociceptive thresholds (TNTs) in mice and rats. The method uses slowly increasing thermal stimulus applied mostly to the skin of mouse/rat plantar surface. The method is particularly sensitive to study anti-nociception during hyperalgesic states such as PDN. The procedures described bellow are based on the ones published in detail by Alm&aacute;si et al 5 and B&ouml;lcskei et al 3. The procedures described here have been approved the Laboratory Animal Care and Use Committee (LACUC), Wright State University.

Here, we describe a rapid reliable and simple procedure to determine the lowest temperature at which rats or mice show nocifensive behavior, i.e. the thermal nociceptive threshold (TNT). This method applies a slowly increasing thermal stimulus allowing precise and reproducible estimation of TNTs with minimum, if any, stress to the animals.

糖尿病大鼠的热痛觉阈值的快速测定

Painful diabetic neuropathy (PDN) is characterized by hyperalgesia i.e., increased sensitivity to noxious stimulus, and allodynia i.e., hypersensitivity ...

Methods for Studying the Mechanisms of Action of Antipsychotic Drugs in Caenorhabditis elegans

Caenorhabditis elegans is a simple genetic organism amenable to large-scale forward and reverse genetic screens and chemical genetic screens. The C. elegans genome includes potential antipsychotic drug (APD) targets conserved in humans, including genes encoding proteins required for neurotransmitter synthesis and for synaptic structure and function. APD exposure produces developmental delay and/or lethality in nematodes in a concentration-dependent manner. These phenotypes are caused, in part, by APD-induced inhibition of pharyngeal pumping1,2. Thus, the developmental phenotype has a neuromuscular basis, making it useful for pharmacogenetic studies of neuroleptics. Here we demonstrate detailed procedures for testing APD effects on nematode development and pharyngeal pumping. For the developmental assay, synchronized embryos are placed on nematode growth medium (NGM) plates containing APDs, and the stages of developing animals are then scored daily. For the pharyngeal pumping rate assay, staged young adult animals are tested on NGM plates containing APDs. The number of pharyngeal pumps per unit time is recorded, and the pumping rate is calculated. These assays can be used for studying many other types of small molecules or even large molecules.

Methods for Studying the Mechanisms of Action of Antipsychotic Drugs in Caenorhabditis elegans

Approaches for testing the effects of antipsychotic drugs (APDs) in Caenorhabditis elegans are demonstrated. Assays are described for testing drug effects on development and viability and on pharyngeal pumping rate. These methods are also applicable for pharmacogenetic experiments with drug classes other than APDs.

方法研究抗精神病药物的作用机制在<em&gt;秀丽隐杆线虫</em

Caenorhabditis elegans is a simple genetic organism amenable to  large-scale forward and reverse genetic screens and chemical genetic screens. The C.  ...

A Procedure for Implanting Organized Arrays of Microwires for Single-unit Recordings in Awake, Behaving Animals

In vivo electrophysiological recordings in the awake, behaving animal provide a powerful method for understanding neural signaling at the single-cell level. The technique allows experimenters to examine temporally and regionally specific firing patterns in order to correlate recorded action potentials with ongoing behavior. Moreover, single-unit recordings can be combined with a plethora of other techniques in order to produce comprehensive explanations of neural function. In this article, we describe the anesthesia and preparation for microwire implantation. Subsequently, we enumerate the necessary equipment and surgical steps to accurately insert a microwire array into a target structure. Lastly, we briefly describe the equipment used to record from each individual electrode in the array. The fixed microwire arrays described are well-suited for chronic implantation and allow for longitudinal recordings of neural data in almost any behavioral preparation. We discuss tracing electrode tracks to triangulate microwire positions as well as ways to combine microwire implantation with immunohistochemical techniques in order to increase the anatomical specificity of recorded results.

Implanting organized arrays of microwires for use in single-unit electrophysiological recordings presents a number of technical challenges.&#160;Methods for performing this technique and the equipment necessary are described.&#160;Also, the beneficial use of organized microwire arrays to record from distinct neural subregions with high spatial selectivity is discussed.

一个程序植入微丝的有组织阵列的单机录音清醒，举止动物

In vivo electrophysiological recordings in the awake, behaving animal provide a powerful method for understanding neural signaling at the single-cell ...

A Simple Way to Measure Alterations in Reward-seeking Behavior Using Drosophila melanogaster

We describe a protocol for measuring ethanol self-administration in fruit flies (Drosophila melanogaster) as a proxy for changes in reward states. We demonstrate a simple way to tap into the fly reward system, modify experiences related to natural reward, and use voluntary ethanol consumption as a measure for changes in reward states. The approach serves as a relevant tool to study the neurons and genes that play a role in experience-mediated changes of internal state. The protocol is composed of two discrete parts: exposing the flies to rewarding and nonrewarding experiences, and assaying voluntary ethanol consumption as a measure of the motivation to obtain a drug reward. The two parts can be used independently to induce the modulation of experience as an initial step for further downstream assays or as an independent two-choice feeding assay, respectively. The protocol does not require a complicated setup and can therefore be applied in any laboratory with basic fly culture tools.

A Simple Way to Measure Alterations in Reward-seeking Behavior Using Drosophila melanogaster

We describe a protocol for inducing rewarding and nonrewarding experiences in fruit flies (Drosophila melanogaster) using voluntary ethanol consumption as a measure for changes in reward states.

一个简单的方法来衡量在改建悬赏寻求行为使用<em&gt;果蝇</em

We describe a protocol for measuring ethanol self-administration in fruit flies (Drosophila melanogaster) as a proxy for changes in reward states. We ...

A Protocol for Measuring Cue Reactivity in a Rat Model of Cocaine Use Disorder

Cocaine use disorder (CUD) follows a trajectory of repetitive self-administration during which previously neutral stimuli gain incentive value. Cue reactivity, the sensitivity to cues previously linked with the drug-taking experience, plays a prominent role in human craving during abstinence. Cue reactivity can be assessed as the attentional orientation toward drug-associated cues that is measurable as appetitive approach behavior in both preclinical and human studies. Herein describes an assessment of cue reactivity in rats trained to self-administer cocaine. Cocaine self-administration is paired with the presentation of discrete cues that act as conditioned reinforcers (i.e., house light, stimulus light, infusion pump sounds). Following a period of abstinence, lever presses in the cocaine self-administration context accompanied by the discrete cues previously paired with cocaine infusion are measured as cue reactivity. This model is useful to explore neurobiological mechanisms underlying cue reactivity processes as well as to assess pharmacotherapies to suppress cue reactivity and therefore, modify relapse vulnerability. Advantages of the model include its translational relevance, and its face and predictive validities.&#160;The primary limitation of the model is that the cue reactivity task can only be performed infrequently and must only be used in short duration (e.g., 1 hour), otherwise rats will begin to extinguish the pairing of the discrete cues with the cocaine stimulus. The model is extendable to any positively reinforcing stimulus paired with discrete cues; though particularly applicable to drugs of abuse, this model may hold future applications in fields such as obesity, where palatable food rewards can act as positively reinforcing stimuli.

Cue reactivity is conceptualized as sensitivity to cues linked with drug-taking experiences that contribute to craving and relapse in abstinent humans. Cue reactivity is modeled in rats by measuring attentional orientation toward drug-associated cues that results in appetitive approach behavior in a cue reactivity test following self-administration and forced abstinence.

可卡因使用障碍大鼠模型中提示反应性测定的一种协议

Cocaine use disorder (CUD) follows a trajectory of repetitive self-administration during which previously neutral stimuli gain incentive value. Cue ...

A Microbiomechanical System for Studying Varicosity Formation and Recovery in Central Neuron Axons

Axonal varicosities are enlarged structures along the shafts of axons with a high degree of heterogeneity. They are present not only in brains with neurodegenerative diseases or injuries, but also in the normal brain. Here, we describe a newly-established micromechanical system to rapidly, reliably, and reversibly induce axonal varicosities, allowing us to understand the mechanisms governing varicosity formation and heterogeneous protein composition. This system represents a novel means to evaluate the effects of compression and shear stress on different subcellular compartments of neurons, different from other in vitro systems that mainly focus on the effect of stretching. Importantly, owing to the unique features of our system, we recently made a novel discovery showing that the application of pressurized fluid can rapidly and reversibly induce axonal varicosities through the activation of a transient receptor potential channel. Our biomechanical system can be utilized conveniently in combination with drug perfusion, live cell imaging, calcium imaging, and patch clamp recording. Therefore, this method can be adopted for studying mechanosensitive ion channels, axonal transport regulation, axonal cytoskeleton dynamics, calcium signaling, and morphological changes related to traumatic brain injury.

This protocol describes a physiologically relevant, pressurized fluid approach for rapid and reversible induction of varicosities in neurons.

中枢神经元轴突静脉曲张形成和恢复的 Microbiomechanical 系统研究

Axonal varicosities are enlarged structures along the shafts of axons with a high degree of heterogeneity. They are present not only in brains with ...

Preparation of Peripheral Nerve Stimulation Electrodes for Chronic Implantation in Rats

Peripheral nerve cuff electrodes have long been used in the neurosciences and related fields for stimulation of, for example, vagus or sciatic nerves. Several recent studies have demonstrated the effectiveness of chronic VNS in enhancing central nervous system plasticity to improve motor rehabilitation, extinction learning, and sensory discrimination. Construction of chronically implantable devices for use in such studies is challenging due to rats&#8217; small size, and typical protocols require extensive training of personnel and time-consuming microfabrication methods. Alternatively, commercially available implantable cuff electrodes can be purchased at a significantly higher cost. In this protocol, we present a simple, low-cost method for construction of small, chronically implantable peripheral nerve cuff electrodes for use in rats. We validate the short and long-term reliability of our cuff electrodes by demonstrating that VNS in ketamine/xylazine anesthetized rats produces decreases in breathing rate consistent with activation of the Hering-Breuer reflex, both at the time of implantation and up to 10 weeks after device implantation. We further demonstrate the suitability of the cuff electrodes for use in chronic stimulation studies by pairing VNS with skilled lever press performance to induce motor cortical map plasticity.

Existing approaches for constructing chronically implantable peripheral nerve cuff electrodes for use in small rodents often require specialized equipment and/or highly trained personnel. In this protocol we demonstrate a simple, low-cost approach for fabricating chronically implantable cuff electrodes, and demonstrate their effectiveness for vagus nerve stimulation (VNS) in rats.

大鼠慢性植入的外周神经刺激电极的制备

Peripheral nerve cuff electrodes have long been used in the neurosciences and related fields for stimulation of, for example, vagus or sciatic nerves. ...