这个项目是由自然科学和工程研究理事会，加拿大（355881）和加拿大创新基金会的赠款支持。

*最重要的步骤。 1。设备和软件<ol><li>配备两个超轻型的，可伸缩的一侧或食物盛器（ 见表1）定位杆1鼠标操作性商会进行了训练和测试。位于上述各杠杆刺激光，并在笼子的顶部有一个房子光。颗粒机是位于外室。每个分庭内的换气扇声音衰减的房间内。钱伯斯是连接与MedPC四，软件到PC计算机上通过的"智能控制"界面内阁。 </li></ol> 注:当超轻型杠杆不可，建议鼻子戳用于小鼠研究的操作性响应。然而，有两个可供选择，我们建议选择更可靠的鼻子戳反应可以作为一个动机措施超轻杠杆é引发探索嗅探。 <ol start="2"><li>用于运行操作性任务MedPC方案都写在Medstate符号代码，用户可以了解编程手册提供的帮助。编程为用户提供灵活性，修改现有的代码，或产生独特的实验参数。某些代码可以得到免费从医学协会"Medstate符号库" <a href="http://www.mednr.com" target="_blank">http://www.mednr.com</a> 。从医学协会可以购买额外的代码。 </li></ol> 2。鼠标驯化和食品的限制<ol><li>抵达后小鼠组安置至少10天12小时的反向光/暗周期，以适应环境，并提供标准饲料和水的随意访问。在我们的例子中，我们用成年C57BL / 6小鼠从查尔斯河（圣常，质量控制）购买。所有的鼠标操作和行为实验是​​在黑暗的周期。所有亲涉及使用动物的cedures由CRCHUM动物保健委员会批准。 </li><li>小鼠随后被单独安置和食物限制（他们每天的食物摄入量的70％），直到他们达到90-95％，饲养体重，以便收购杠杆记者回应。小鼠在家里笼提供会话终止后，他们每天的食品配额。达到5-10％的体重损失后，每天的食物分配进行调整，以稳定为低体重训练期间的剩余。一旦小鼠已达到收购标准（见下文），他们随意喂养时间表。老鼠恢复失去体重约3-4天。 </li><li>小鼠在试验室进行处理的第一堂训练课之前连续3天。 10-15高脂肪和高糖（HFHS）颗粒进入家庭笼放置，以防止对食物恐新症对操作性performan潜在影响训练开始的前一天CE。我们使用20毫克HFHS，无尘，精密的食物颗粒（14毫克，也可）含48.9％千卡脂肪（生物SERV，Frenchtown，新泽西州）。 </li></ol> 3。操作性训练<ol><li>最初的训练小鼠按一个固定比率（FR）-1加固进度，即单杆按下插座引出的食物颗粒交货的杠杆。只有一杆被指定为"主动"（触发运送食物奖励）和左，右杠杆分配之间的小鼠抵消。重要的是，添加至少5秒超时（TO）FR1号的时间表（FR1/TO-5），在此期间，额外的杠杆按不会导致交货的食物颗粒。这个期限，使小鼠的时间消耗的食物颗粒，但是更长的时间超时如果实验者认为它是必要的小鼠模型，可以使用。每个FR培训会议持续1小时或50颗粒时已交付。 </li><li> *收购标准:&lt;/ STRONG&gt;用于确定收购粮食保持响应操作性的标准包括:（1）赚取的积极响应和奖励的最低数量;（2）（3）的活动和非活动的杠杆之间的歧视措施，并与会议性能稳定。小鼠表现出主动与非杠杆≥3:1的歧视，并取得了连续三届会议≥20％奖励被视为实现收购标准10。我们已经指出，C57BL6小鼠的大部分（75％）需要约7-10天的培训，以达到收购标准。小鼠，没有达到通过这次收购标准，接受进一步的训练额外的5-7天。我们排除测试（〜5％的情况下）小鼠时，有没有额外的第5个训练日的进度。重要的是要牢记操作性收购受损可能是某些药物和遗传干预的结果，因此，应记录。&lt;/ LI&gt; <li>继连续三年获得≥20颗粒会议，日程安排增加活跃的5杆印刷机引发的粮食颗粒交货FR5/TO-5秒。培训的FR5日程上持续了三天。 </li><li>小鼠，然后训练累进比率（PR）加固进度。理查德森和Roberts（1996）11％计算公关测试过程中的响应比时间表可以使用下列公式计算（四舍五入到最接近的整数）: = [5 E（R * 0.2）] - 5 其中 R等于已经获得食物奖励加1号（即下增强剂）。因此，需要获得食物奖励，按照订单的响应:1，2，4，6，9，12，15，20，25，32，40，50，62，77，95等。最终完成的比例是断点。 </li><li>公关会议最多持续1小时。未能在任何10分钟终止在会议期间按杠杆。 perfor钢筋公关时间表曼斯被认为是稳定时，奖励的数量获得了至少连续3天≤10％，在1个小时的会议偏离。 </li></ol> 4。进的比测试和验证<ol><li>为了验证食品保持操作性响应公关任务，可以评估断点响应以下操作，以调节食物奖励。一个相对简单的测试，开展粮食匮乏，这将增加食品鼠标的动机状态，从而增加断点。老鼠必须先进行测试期间随意喂养期间，直到实现稳定，性能基线的公关任务。最后的"基线"一天的食物后，一天从家里笼和老鼠，然后在24小时后开始快速测试的公关任务。 </li><li>我们还调查了抑制食欲的荷尔蒙leptin的外设管理是否会有相反的效果上断点阈值。瘦素（AF Parlow，国家激素和肽计划，NIDDK）溶解在无菌PBS。公关测试进行的PBS在一天的IP注射，然后注射瘦素（5毫克/公斤）的翌日。瘦素注射前一小时公关测试。断点比较采用配对t检验（GraphPad棱镜）。 </li></ol> 5。代表结果测试时，粮食匮乏的能力，以调节断点上一个加固的公关日程响应，我们期望显着增加，如在图1所示的断点。 24小时的食物匮乏的时期，在这个实验中产生的中断点〜3.6倍的增长相比，那些在自由摄食基线从而表明缺粮增加食物的奖赏效应的状态得到。相比之下，外设管理，厌食激素瘦素（5毫克/公斤，IP）前一小时到Testing应减少断点响应如图2所示。 <img alt="图1" src="/files/ftp_upload/3754/3754fig1.jpg" /> 图1。空腹增加食物的奖赏效应。 24 h快速显着增加了一个渐进的比率（PR）的增强小鼠的时间表（N = 4）断点。平均值±标准差; * P &lt;.05。 <img alt="图2" src="/files/ftp_upload/3754/3754fig2.jpg" /> 图2。瘦素的小鼠减少食物奖励。在小鼠（n = 4时），外周血瘦素管理（5毫克/公斤）下降公关断点响应阈值。平均值±标准差; * P &lt;0.05。

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操作性条件反射的任务提供了一种有效的手段，以评估食物的动机性质的变化。本文许可证训练小鼠食物奖励杠杆记者和稳定详述的实验程序响应的加固公关时间表。 测量消耗的食物量不提供相同的读数，因为并不是所有的因素，影响食物的摄入，调节食物的奖赏效应，努力为基础的应对措施。例如，2 -脱氧葡萄糖对大鼠外周血管理消耗利用的葡萄糖增加量免费提供食用的食品，但未能改变破发点，为12食品的公关任务。公关断点响应，也可以反映在食物奖励的质量变化。例如，破发点的正相关，与蔗糖浓度展示方式如下甜味8，增加蔗糖的动机</s达&gt;。在这里，我们表明，24小时剥夺食物大大增加公关小鼠HFHS食品断点。相比之下，我们发现抑制食欲的荷尔蒙leptin的外设管理，显着地降低小鼠为HFHS食品断点。这些研究结果与以前报道的行动粮食匮乏和12-14大鼠瘦素奖励阈值一致。 使用此协议时，重要的是要牢记，粮食收购保持定期培训（每1-2天），并通过实施食品的限制方案，通过增加鼠标的动机状态明显增强操作性响应。此外，确保小鼠表现出相对稳定的一个加固的时间表响应之前移动到更复杂的时间表（如FR1号到FR5），以减少在测试过程中的可变性能的关键。尽管这些措施的应用，这是不寻常，以获得一些变异特别是在不同小鼠之间。出于这个原因，我们强烈建议使用内（重复测量）设计时，可以删除跨学科变异。虽然内受测试调查稳定的遗传改变（即淘汰赛与对照组）的可能，它可以用来检查药物，病变或急性遗传操作（例如，optogenetic）的影响。 肥胖与肥胖有关的疾病是一个日益严重的问题，增加了紧迫感，以确定涉及的分子和神经通路。越来越多的证据突出的重要作用，在食物渴求的大脑奖励电路，增加热量的摄入和体重增加。公关任务是一个有价值的工具，用于测量食物奖赏效应的影响，神经生物学，遗传或药理操纵。

<html><table border="1"><tbody><tr><td> 项目 </td><td> 目录编号 </td><td> 公司 </td><td> 评论 </td></tr><tr><td>小鼠操作性商会</td><td> env-009A的</td><td> <a href="http://www.med-associates.com/" target="_blank">MED公司，公司</a> </td><td> MED联营公司 PO箱319 圣奥尔本斯，佛蒙特州05478， 美国 电话:（802）527-2343 </td></tr><tr><td> 8 inputs/16输入智能 控制连接面板</td><td> SG-716B </td><td> <a href="http://www.med-associates.com/" target="_blank">MED公司，公司</a> </td><td></td></tr><tr><td>超轻型伸缩 鼠标杆</td><td> env-312的</td><td> <a href="http://www.med-associates.com/" target="_blank">MED公司，公司</a> </td><td></td></tr><tr><td>佩莱饮水机</td><td> ENV-203 </td><td>MED-associates.com /"目标="_blank"&gt; MED Associates公司</td><td></td></tr><tr><td>楼光</td><td> env-215的</td><td> <a href="http://www.med-associates.com/" target="_blank">MED公司，公司</a> </td><td></td></tr><tr><td>刺激光</td><td> ENV-221 </td><td> <a href="http://www.med-associates.com/" target="_blank">MED公司，公司</a> </td><td></td></tr><tr><td>无尘精密20 毫克高脂肪及高糖 颗粒</td><td>架F-06649 </td><td>生物-​​Serv的</td><td>生物-​​Serv的 一街8号，1套房 frenchtown，NJ 08825，USA 电话:（800）-996-9908 </td></tr></tbody></table> 表1。具体的设备和材料。

progressive-ratio responding for palatable high-fat and high-sugar food in mice

含有丰富的脂肪和糖的食物，大大有助于过度饮食和肥胖率的不断上升。可口的食物消费，可以产生有益的作用，加强行动的成果协会和加强未来的行为，在取得这些食物。越来越多的证据，能量密度高的食物奖赏效应发挥暴饮暴食和肥胖的发展产生深远的作用，有研究的基因，分子和神经电路，调节食物奖励1,2的兴趣高涨。可以测量的意愿工作，以取得他们如在操作性条件反射的任务3，研究不同的刺激有益的影响。收购和食物奖励措施的操作性模型，在获取食物的自愿行为反应。奖励实力的一个常用的措施是在公关ţ4,5操作性的过程称为累进比率（PR）的加固进度。问，这个问题需要作出操作性反应的越来越多，为每个连续的奖励。的Hodos（1961）的开创性研究表明，作出反应，获得最后的奖励数量，被称为断点，作为奖励强度指数4。尽管操作性的程序，仅在响应速度的变动不能分开奖励强度的变化，表现能力的改变，来自公关时间表的断点是食物的奖赏效应的验证措施。公关任务已被广泛用于评估药物滥用和老鼠的食物（如6-8）有益的影响，但程度较轻，在小鼠9。遗传工程小鼠和饮食诱导的肥胖小鼠模型使用的增加加剧了在老鼠的食物奖励的行为措施的需求。在本文中，我们详细的材料和程序，用来训练老鼠高脂肪回应（杠杆新闻）和高糖食物颗粒上加固公关时间表。我们表明，断点的响应阈值增加剥夺后急性食物，并减少与抑制食欲的荷尔蒙leptin的外设管理，从而验证了使用这种小鼠的食品操作性范式。

Watch this Scientific Journal Video about Progressive-ratio Responding for Palatable High-fat and High-sugar Food in Mice at JoVE.com

Progressive-ratio Responding for Palatable High-fat and High-sugar Food in Mice

本报告详细说明来衡量高脂肪食物的小鼠的奖赏效应的协议，采用累进比例操作性条件反射的任务。

累进比率响应小鼠可口的高脂肪和高糖食品

Foods that are rich in fat and sugar significantly contribute to over-eating and escalating rates of obesity. The consumption of palatable foods can ...

progressive-ratio-responding-for-palatable-high-fat-high-sugar-food

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21.9K 次观看.  University of Montreal. 本报告详细说明来衡量高脂肪食物的小鼠的奖赏效应的协议，采用累进比例操作性条件反射的任务。

视频: Progressive-ratio Responding for Palatable High-fat and High-sugar Food in Mice

A Simple and Inexpensive Method for Determining Cold Sensitivity and Adaptation in Mice

Cold hypersensitivity is a serious clinical problem, affecting a broad subset of patients and causing significant decreases in quality of life. The cold plantar assay allows the objective and inexpensive assessment of cold sensitivity in mice, and can quantify both analgesia and hypersensitivity. Mice are acclimated on a glass plate, and a compressed dry ice pellet is held against the glass surface underneath the hindpaw. The latency to withdrawal from the cooling glass is used as a measure of cold sensitivity.
Cold sensation is also important for survival in regions with seasonal temperature shifts, and in order to maintain sensitivity animals must be able to adjust their thermal response thresholds to match the ambient temperature. The Cold Plantar Assay (CPA) also allows the study of adaptation to changes in ambient temperature by testing the cold sensitivity of mice at temperatures ranging from 30 &#176;C to 5 &#176;C. Mice are acclimated as described above, but the glass plate is cooled to the desired starting temperature using aluminum boxes (or aluminum foil packets) filled with hot water, wet ice, or dry ice. The temperature of the plate is measured at the center using a filament T-type thermocouple probe. Once the plate has reached the desired starting temperature, the animals are tested as described above.
This assay allows testing of mice at temperatures ranging from innocuous to noxious. The CPA yields unambiguous and consistent behavioral responses in uninjured mice and can be used to quantify both hypersensitivity and analgesia. This protocol describes how to use the CPA to measure cold hypersensitivity, analgesia, and adaptation in mice.

The Cold Plantar Assay (CPA) measures cold responsiveness between 30 &#176;C and 5 &#176;C, and can also measure cold adaptation. This protocol describes how to use the CPA to measure cold hypersensitivity, analgesia, and adaptation in mice.

测定冷灵敏度和适应小鼠一种简单廉价的方法

Cold hypersensitivity is a serious clinical problem, affecting a broad subset of patients and causing significant decreases in quality of life.  The cold ...

科研

神经科学

The CApillary FEeder Assay Measures Food Intake in Drosophila melanogaster

For most animals, feeding is an essential behavior for securing survival, and it influences development, locomotion, health and reproduction. Ingestion of the right type and quantity of food therefore has a major influence on quality of life. Research on feeding behavior focuses on the underlying processes that ensure actual feeding and unravels the role of factors regulating internal energy homeostasis and the neuronal bases of decision-making. The model organism Drosophila melanogaster, with its great variety of genetically traceable tools for labeling and manipulating single neurons, allows mapping of neuronal networks and identification of molecular signaling cascades involved in the regulation of food intake. This report demonstrates the CApillary FEeder assay (CAFE) and shows how to measure food intake in a group of flies for time spans ranging from hours to days. This easy-to-use assay consists of glass capillaries filled with liquid food that flies can freely access and feed on. Food consumption in the assay is accurately determined using simple measurement tools. Herein we describe step-by-step the method from setup to successful execution of the CAFE assay, and provide practical examples to analyze the food intake of a group of flies under controlled conditions. The reader is guided through possible limitations of the assay, and advantages and disadvantages of the method compared to other feeding assays in D. melanogaster are evaluated.

The CApillary FEeder Assay Measures Food Intake in Drosophila melanogaster

The CApillary FEeder (CAFE) assay is a simple, budget-friendly, highly reliable method for investigating mechanisms underlying food intake. Used with the highly versatile genetic model organism Drosophila melanogaster, it provides a powerful means of gaining new insights into regulatory mechanisms of food intake.

毛细管馈线分析测量的摄食<em&gt;果蝇</em

For most animals, feeding is an essential behavior for securing survival, and it influences development, locomotion, health and reproduction. Ingestion of ...

An In Ovo Model for Testing Insulin-mimetic Compounds

Elevated blood glucose levels in type 2 diabetes mellitus (T2DM), a complex and multifactorial metabolic disease, are caused by insulin resistance and &#946;-cell failure. Various strategies, including the injection of insulin or the usage of insulin-sensitizing drugs, were pursued to treat T2DM or at least reduce the symptoms. In addition, the application of herbal compounds has attracted increasing attention. Thus, it is necessary to find efficient test systems to identify and characterize insulin-mimetic compounds. Here we developed a modified chick embryo model, which enables testing of synthetic compounds and herbal extracts with insulin-mimetic properties. Using a fluorescence microscopy-based primary screen, which quantifies the translocation of Glucose transporter 4 (Glut4) to the plasma membrane, we were able to identify compounds, mainly herbal extracts, which lead to an increase of intracellular glucose concentrations in adipocytes. However, the efficacy of these substances requires further verification in a living organism. Thus, we used an in-ovo approach to identify their blood glucose-reducing properties. The approval by an ethics committee is not needed since the use of chicken embryos during the first two-thirds of embryonic development is not considered an animal experiment. Here, the application of this model is described in detail.

An In Ovo Model for Testing Insulin-mimetic Compounds

In this study, we describe an in-ovo system as a promising tool to test insulin-mimetic compounds in a living organism. The main advantages include adequate throughput rates and acceptable costs, which enable the identification of phytochemicals with insulin-mimetic properties.

用于测试胰岛素模拟化合物的蛋模型

Elevated blood glucose levels in type 2 diabetes mellitus (T2DM), a complex and multifactorial metabolic disease, are caused by insulin resistance and ...

Data Acquisition and Analysis In Brainstem Evoked Response Audiometry In Mice

Brainstem evoked response audiometry (BERA) is of central relevance in the clinical neurophysiology. As other evoked potential (EP) techniques, such as visually evoked potentials (VEPs) or somatosensory evoked potentials (SEPs), the auditory evoked potentials (AEPs) are triggered by the repetitive presentation of identical stimuli, the electroencephalographic (EEG) response of which is subsequently averaged resulting in distinct positive (p) and negative (n) deflections. In humans, both the amplitude and the latency of individual peaks can be used to characterize alterations in synchronization and conduction velocity in the underlying neuronal circuitries. Importantly, AEPs are also applied in basic and preclinical science to identify and characterize the auditory function in pharmacological and genetic animal models. Even more, animal models in combination with pharmacological testing are utilized to investigate for potential benefits in the treatment of sensorineural hearing loss (e.g., age- or noise-induced hearing deficits). Here we provide a detailed and integrative description of how to record auditory brainstem-evoked responses (ABRs) in mice using click and tone-burst application. A specific focus of this protocol is on pre-experimental animal housing, anesthesia, ABR recording, ABR filtering processes, automated wavelet-based amplitude growth function analysis, and latency detection.

Brainstem evoked response audiometry is an important tool in clinical neurophysiology. Nowadays, brainstem evoked response audiometry is also applied in the basic science and preclinical studies involving both pharmacological and genetic animal models. Here we provide a detailed description of how auditory brainstem responses can be successfully recorded and analyzed in mice.

脑干中的数据采集与分析 在小鼠中引发响应音频测量

Brainstem evoked response audiometry (BERA) is of central relevance in the clinical neurophysiology. As other evoked potential (EP) techniques, such as ...

Quantitative Measurement of Intrathecally Synthesized Proteins in Mice

Cerebrospinal fluid (CSF), a fluid found in the brain and the spinal cord, is of great importance to both basic and clinical science. The analysis of the CSF protein composition delivers crucial information in basic neuroscience research as well as neurological diseases. One caveat is that proteins measured in CSF may derive from both intrathecal synthesis and transudation from serum, and protein analysis of CSF can only determine the sum of these two components. To discriminate between protein transudation from the blood and intrathecally produced proteins in animal models as well as in humans, CSF protein profiling measurements using conventional protein analysis tools must include the calculation of the albumin CSF/serum quotient (Qalbumin), a marker of the integrity of the blood-brain interface (BBI), and the protein index (Qprotein/Qalbumin), an estimate of intrathecal protein synthesis. This protocol illustrates the entire procedure, from CSF and blood collection to quotients and indices calculations, for the quantitative measurement of intrathecal protein synthesis and BBI impairment in mouse models of neurological disorders.

Elevated spinal fluid protein levels can either be the result of diffusion of plasma protein across an altered blood-brain barrier or intrathecal synthesis. An optimized testing protocol is presented in this article that helps to discriminate both cases and provides quantitative measurements of intrathecally synthesized proteins.

小鼠体内合成蛋白的定量测量

Cerebrospinal fluid (CSF), a fluid found in the brain and the spinal cord, is of great importance to both basic and clinical science. The analysis of the ...

Electrocorticographic Recording of Cerebral Cortex Areas Manipulated Using an Adeno-Associated Virus Targeting Cofilin in Mice

The use of electrocorticographic (ECoG) recordings in rodents is relevant to sleep research and to the study of a wide range of neurological conditions. Adeno-associated viruses (AAVs) are increasingly used to improve understanding of brain circuits and their functions. The AAV-mediated manipulation of specific cell populations and/or of precise molecular components has been tremendously useful to identify new sleep regulatory circuits/molecules and key proteins contributing to the adverse effects of sleep loss. For instance, inhibiting activity of the filamentous actin-severing protein&#160;cofilin&#160;using AAV prevents sleep deprivation-induced memory impairment. Here, a protocol is&#160;described that combines the manipulation of cofilin function in a cerebral cortex area with the recording of ECoG activity to examine whether cortical cofilin modulates the wakefulness and sleep ECoG signals. AAV injection is performed during the same surgical procedure as the implantation of ECoG and electromyographic (EMG) electrodes in adult male and female mice. Mice are anesthetized, and their heads are shaved. After skin cleaning and incision, stereotaxic coordinates of the motor cortex are determined, and the skull is pierced at this location. A cannula prefilled with an AAV expressing cofilinS3D, an inactive form of cofilin, is slowly positioned in the cortical tissue. After AAV infusion, gold-covered screws (ECoG electrodes) are screwed through the skull and cemented to the skull with gold wires inserted in the neck muscles (EMG electrodes). The animals are allowed three weeks to recover and to ensure sufficient expression of cofilinS3D. The infected area and cell type are verified using immunohistochemistry, and the ECoG is analyzed using visual identification of vigilance states and spectral analysis. In summary, this combined methodological approach allows the investigation of the precise contribution of molecular components regulating neuronal morphology and connectivity to the regulation of synchronized cerebral cortex activity during wakefulness and sleep.

This article describes a protocol for the manipulation of molecular targets in the cerebral cortex using adeno-associated viruses and for monitoring the effects of this manipulation during wakefulness and sleep using electrocorticographic recordings.

使用腺相关病毒在小鼠中瞄准科菲林操纵的大脑皮层区域的电皮质记录

The use of electrocorticographic (ECoG) recordings in rodents is relevant to sleep research and to the study of a wide range of neurological conditions. ...

A Rapid Food-Preference Assay in Drosophila

To select food with nutritional value while avoiding the consumption of harmful agents, animals need a sophisticated and robust taste system to evaluate their food environment. The fruit fly, Drosophila melanogaster, is a genetically tractable model organism that is widely used to decipher the molecular, cellular, and neural underpinnings of food preference. To analyze fly food preference, a robust feeding method is needed. Described here is a two-choice feeding assay, which is rigorous, cost-saving, and fast. The assay is Petri-dish-based and involves the addition of two different foods supplemented with blue or red dye to the two halves of the dish. Then, ~70 prestarved, 2-4-day-old flies are placed in the dish and&#160;allowed to choose between blue and red foods in the dark for about 90 min. Examination of the abdomen of each fly is followed by the calculation of the preference index. In contrast to multiwell plates, each Petri dish takes only ~20 s to fill and saves time and effort. This feeding assay can be employed to quickly determine whether flies like or dislike a particular food.

A Rapid Food-Preference Assay in Drosophila

We present a protocol for a two-choice feeding assay for flies. This feeding assay is fast and easy to run and is suitable not only for small-scale laboratory research, but also for high-throughput behavioral screens in flies.

果蝇的快速食物偏好分析

To select food with nutritional value while avoiding the consumption of harmful agents, animals need a sophisticated and robust taste system to evaluate ...

Modeling Stroke in Mice: Focal Cortical Lesions by Photothrombosis

Stroke is a leading cause of death and acquired adult disability in developed countries. Despite extensive investigation for novel therapeutic strategies, there remain limited therapeutic options for stroke patients. Therefore, more research is needed for pathophysiological pathways such as post-stroke inflammation, angiogenesis, neuronal plasticity, and regeneration. Given the inability of in vitro models to reproduce the complexity of the brain, experimental stroke models are essential for the analysis and subsequent evaluation of novel drug targets for these mechanisms. In addition, detailed standardized models for all procedures are urgently needed to overcome the so-called replication crisis. As an effort within the ImmunoStroke research consortium, a standardized photothrombotic mouse model using an intraperitoneal injection of Rose Bengal and the illumination of the intact skull with a 561 nm laser is described. This model allows the performance of stroke in mice with allocation to any cortical region of the brain without invasive surgery; thus, enabling the study of stroke in various areas of the brain. In this video, the surgical methods of stroke induction in the photothrombotic model along with&#160;histological analysis are demonstrated.

Described here is the photothrombotic stroke model, where a stroke is produced through the intact skull by inducing permanent microvascular occlusion using laser illumination after administration of a photosensitive dye.

小鼠模型中风:光血栓形成的焦皮质病变

Stroke is a leading cause of death and acquired adult disability in developed countries. Despite extensive investigation for novel therapeutic strategies, ...

Caenorhabditis elegans as a Model System for Discovering Bioactive Compounds Against Polyglutamine-Mediated Neurotoxicity

Age-related misfolding and aggregation of pathogenic proteins are responsible for several neurodegenerative diseases. For example, Huntington&#39;s disease (HD) is principally driven by a CAG nucleotide repeat that encodes an expanded glutamine tract in huntingtin protein. Thus, the inhibition of polyglutamine (polyQ) aggregation and, in particular, aggregation-associated neurotoxicity is a useful strategy for the prevention of HD and other polyQ-associated conditions. This paper introduces generalized experimental protocols to assess the neuroprotective capacity of test compounds against HD using established polyQ transgenic Caenorhabditis elegans models. The AM141 strain is chosen for the polyQ aggregation assay as an age-associated phenotype of discrete fluorescent aggregates can be easily observed in its body wall at the adult stage due to muscle-specific expression of polyQ::YFP fusion proteins. In contrast, the HA759 model with strong expression of polyQ-expanded tracts in ASH neurons is used to examine neuronal death and chemoavoidance behavior. To comprehensively evaluate the neuroprotective capacity of target compounds, the above test results are ultimately presented as a radar chart with profiling of multiple phenotypes in a manner of direct comparison and direct viewing.

Caenorhabditis elegans as a Model System for Discovering Bioactive Compounds Against Polyglutamine-Mediated Neurotoxicity

Here, we present a protocol to assess the neuroprotective activities of test compounds in Caenorhabditis elegans, including polyglutamine aggregation, neuronal death, and chemoavoidance behavior, as well as an exemplary integration of multiple phenotypes.

秀丽隐杆线虫 作为发现生物活性化合物对抗聚谷氨酰胺介导的神经毒性的模型系统

Age-related misfolding and aggregation of pathogenic proteins are responsible for several neurodegenerative diseases. For example, Huntington&#39;s ...

果蝇中的便秘测定

Measuring Constipation in a Drosophila Model of Parkinson's Disease

Non-motor symptoms in Parkinson's disease (PD) are common, difficult to treat, and significantly impair quality of life. One prevalent non-motor symptom is constipation, which can precede the diagnosis of PD by years or even decades. Constipation has been underexplored in animal models of PD and lacks specific therapies. This assay utilizes a Drosophila model of PD in which human alpha-synuclein is expressed under a pan-neuronal driver. Flies expressing alpha-synuclein develop the hallmark features of PD: the loss of dopaminergic neurons, motor impairment, and alpha-synuclein inclusions.
This protocol outlines a method for studying constipation in these flies. Flies are placed on fly food with a blue color additive overnight and then transferred to standard food the following day. They are subsequently moved to new vials with standard fly food every hour for 8 h. Before each transfer, the percentage of blue-colored fecal spots compared to the total fecal spots on the vial wall is calculated. Control flies that lack alpha-synuclein expel all the blue dye hours before flies expressing alpha-synuclein. Additionally, the passage of blue-colored food from the gut can be monitored with simple photography. The simplicity of this assay enables its use in forward genetic or chemical screens to identify modifiers of constipation in Drosophila.

作者聚焦:探索帕金森病的非运动症状 

This protocol presents an assay for modeling constipation in an alpha-synuclein based Drosophila model of Parkinson's disease.

在帕金森病的 果蝇 模型中测量便秘

Non-motor symptoms in Parkinson's disease (PD) are common, difficult to treat, and significantly impair quality of life. One prevalent non-motor symptom ...