作者要感谢秋里王，语言资源中心，科尔比学院，用于视频处理和埃里克·托马斯，音乐系，科尔比学院技术援助，用于提供背景音乐。该项目由来自国立普通医学科学（P20 GM103423-12）国家研究资源中心，INBRE（P20RR016463-12），资金支持，健康和科学部资助，科尔比学院（STA）的国民研究院。 JL和LWM由来自夏季学者基金，科尔比学院资助。

1， 果蝇惊吓引起的步态分析<ol><li>药物治疗<ol><li>稳重的固定化所需的数字1-3（约8-12）日龄雄蝇使用二氧化碳 ，并将其输送到含有药物的补充食品瓶。注:另麻醉剂如乙醚或冰可用于镇静苍蝇，使数量和处理。 </li><li>允许蝇从镇静在水平位置上恢复20分钟（或直到恢复）与小瓶（防止蝇卡住上的食物），然后竖直放置的管形瓶中在12小时黑暗，12小时光照培养箱中在25℃下对于实验的其余部分。 </li></ol></li><li>实验设置<ol><li>通过标记周围的圆瓶用永久性记号笔分这双瓶设置成6.33厘米三个相等的部分。 </li><li>后药物暴露3天后，转移蝇无麻醉入底小瓶中，并很快将样品瓶在开口上。用胶带将两小瓶加上透明胶带。 </li><li>让苍蝇来适应新的环境15分钟。 </li><li>在白色背景上把试管放入，并成立了数码相机从双瓶装置，在视图中的计时器适当的距离。确保整个装置是在一个单一的图像帧可见的，并且所有的苍蝇都在焦点。保持试验之间是一致的帧，马克相机和小瓶的位置。 </li></ol></li><li>流动性分析<ol><li>清晰地显示试验次数，药物治疗，并在摄像头视图计时器。 </li><li>牢固地挖掘双小瓶装置对对接的3倍，并确保所有的苍蝇落入到小瓶的底部。同时启动定时器。 </li><li>每隔5秒，1分钟，取该装置的图片。注:另外，视频可以被捕获并停在适当的时间间隔进行测量。 </li><li>允许蝇恢复原状，持续1分钟。 </li><li&gt;重复2次以上每次试验之间1分钟的恢复时间。注意:每个设备需要5分钟才能完成数据采集。保持试验间攻类似的力量。多个（至少3个）装置能够同时处理容易。 </li></ol></li><li>数据分析<ol><li>查看照片和在每个部分随时间记录蝇的数目。计算苍蝇在每个部分随时间的百分比。注释:重复此用相同的苍蝇整个过程在感兴趣2或3个时间点，例如，3天，5和7。如果太多苍蝇死整个实验过程中，可以按比例增加原试验编号，以补偿对于死亡率。使用适当的统计分析的数据进行比较。 </li></ol></li></ol> 2， 果蝇自发活动分析<ol><li>食品制作<ol><li>重组3克瞬间果蝇培养基用15ml去离子水和所需的鱼藤酮（或感兴趣的另一种药物）的剂量。 </li><li>一旦食物混合，成为坚定的（约5分钟），谨慎地装载食物是约为1厘米，高为制造商提供透明管（3毫米x 65毫米）。通过小心地放置在管垂直于食品和扭转，直到它们可以与管内的食品被移除添加灌注食品的管中的药物。注意:它有利于把一个手指放在该管的开口，以形成真空。食品中不应含有任何气泡或有凹凸不平的表面的苍蝇能成为卡住。 </li></ol></li><li>实验装置<ol><li>放置在管的末端最接近食品的塑料盖。推在管尽可能少的塑料帽，因为它可以在小瓶内建立一个气泡，如果推到有力。 </li><li>稳重1天岁的雄蝇以CO 2，并小心将1男飞入各管画笔。重复这取决于所需的试验次数。 </li><li>堵塞该管最远离食品用小棉球，这可以从较大的​​商店手卷的端购买棉球。 </li><li>允许苍蝇与管子来恢复在水平位置15分钟，确保所有的苍蝇都活着和活性。插入管放入DAM，并确保所有的管子都在同一位置相对于水坝。注意:可以将它们与监测的区域在所述小瓶的中间，或对所有的小瓶推到一边，以使管的端部正在被监视。注:在这个变化的方法请参见讨论。 </li></ol></li><li>数据收集<ol><li>将大坝在12小时黑暗，12小时光照培养箱设定为25℃。大坝连接到数据采集系统。打开坝软件，并根据喜好选择彬长至10分钟。开始收集数据，并允许该程序收集的数据为7天。注意:滨长度可以根据需要进行调整。 </li><li>数据分析 注:过程的数据，以获得每分钟的计数作为长期自发运动的量度。 <ol><li>打开大坝文件扫描程序和访问监控数据通过点击选择输入数据。 </li><li>选择合适的显示器系列，并选择10分钟的间隔彬长。 </li><li>在输出文件类型选择通道的文件。让所有其他选项为默认值。 </li><li>点击扫描数据保存到指定的文件夹中。 </li><li>在昼夜数据分析软件导入数据以得到每分钟的计数。注意:对于数据分析Clocklab软件常用。其他的选择也可以。 </li></ol></li></ol></li></ol>

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作者什么都没有透露。

在这项研究中，我们描述了两个过程中帕金森病的鱼藤酮诱导的果蝇模型中同时测量长期自发运动和短期惊吓诱发运动。还可以测量在果蝇暴露于已知的建模帕金森氏病如其他药剂，百草枯14，帕金森氏病，例如遗传模型，阿尔法-突触核蛋白突变体15中，并影响运动疾病其他蝇模型这些运动特征。对于这两种方法，替代的方法和修改可以被考虑。?...

运动缺陷是帕金森氏病的主要症状，并在很大程度上引起黑质1的多巴胺能神经元的退化。鱼藤酮是已被广泛研究的模型帕金森运动障碍果蝇 2-6酮类杀虫剂。鱼藤酮通过阻断氧化磷酸化途径，最终导致细胞死亡7导致的氧化损伤。多巴胺能神经元更容易产生鱼藤酮毒性，使得主要马达基于2,7的化学品的影响。通过在果蝇诱发帕金森氏病的症状，我们就可以更好地了解疾病和纠正其症状6,8-11。 果蝇提供了一个很好的模型来研究这种影响，因为它们的基因听话，易于维护，并且具有快速的生命周期。 几项研究已经表明，鱼藤酮引起短期惊跳诱导在果蝇 -当果蝇运动缺陷被保持在鱼藤酮补充食品，他们表现出惊人死不休2-6后以较慢的负geotactic回应。他们未能在小瓶设备，迅速向上攀升对照试验表明了惊吓引起的运动缺陷。 鱼藤酮对长期的效果，自发运动不能很好地说明。 果蝇活动监视器（DAMS）已被成功地用于监测果蝇的昼夜节律运动研究12,13。苍蝇被放置在单独的管，其被加载到DAM。该装置配备有红外线传感器，该计数的次数蝇打破了红外线光束的数目。这些计数可以用作原状的运动和活动12,13的量度。通过将苍蝇在一个坝，鱼藤酮对他们的长期运动的效果可以表征。这项研究描述的方法来MEAS茜短期惊跳诱导运动和长期的自发运动，以便更好地理解的鱼藤酮介导的运动缺陷的影响。对运动缺乏模仿帕金森氏症表征是很重要的，因为它们允许它可以扭转这些运动缺陷等化合物的研究。

<table border="0" cellpadding="0" cellspacing="0" width="628">
	<tbody>
		<tr height="40">
			<td height="40" style="height:40px;width:216px;">Standard narrow vials</td>
			<td style="width:127px;">Genesee Scientific</td>
			<td style="width:119px;">32-120</td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:216px;">Rotenone</td>
			<td style="width:127px;">Sigma</td>
			<td style="width:119px;">R8875</td>
			<td>Store in freezer, make fresh for each experiment</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:216px;">Dimethyl Sulfoxide (DMSO)</td>
			<td style="width:127px;">Sigma</td>
			<td style="width:119px;">D8418</td>
			<td>Solvent for rotenone</td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;width:216px;">Instant Drosophila medium</td>
			<td style="width:127px;">Carolina Biological&#160;</td>
			<td style="width:119px;">Formula 4-24</td>
			<td></td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;width:216px;">Drosophila activity monitor (DAM)</td>
			<td style="width:127px;">Trikinetics</td>
			<td style="width:119px;">DAM2</td>
			<td>trikinetics.com</td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;width:216px;">DAM tubes</td>
			<td style="width:127px;">Trikinetics</td>
			<td style="width:119px;">Tubes 5X65 mm</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;width:216px;"></td>
			<td style="width:127px;"></td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="41">
			<td height="41" style="height:41px;width:216px;">Recipe for Rotenone +food (125 mM dose)</td>
			<td style="width:127px;"></td>
			<td style="width:119px;"></td>
			<td>Make 62.5 mM rotenone stock solution in DMSO by dissolving 25 mg rotenone in 1 ml DMSO&#160;</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;"></td>
			<td style="width:127px;"></td>
			<td style="width:119px;"></td>
			<td>For 125 mM dose, add 10 mM rotenone stock in DMSO to 5 ml water.</td>
		</tr>
	</tbody>
</table>

methods to characterize spontaneous and startle-induced locomotion in a rotenone-induced parkinson's disease model of drosophila

帕金森病是一种神经退行性疾病而导致的多巴胺能神经元的变性，在中枢神经系统中，主要在黑质。这种疾病会导致电机的不足，这表现为强直，震颤和老年痴呆症的人。鱼藤酮是一种杀虫剂，通过抑制线粒体的电子传递链的功能引起的氧化损伤。它也被用来建模帕金森氏病的果蝇 。苍蝇有一个固有的负面geotactic反应，这迫使他们在被吓了一跳向上攀升。它已经确定，鱼藤酮引起早期死亡率和运动缺陷扰乱果蝇的爬它们已被窃听向下之后能力。然而，鱼藤酮对自发运动的效果没有记载。本研究概述了两个敏感，重复性好，高通量检测的表征鱼藤酮诱导的缺陷短期惊跳诱导运动和长期自发运动的果蝇 。这些测定法可便利地适用于表征运动缺陷和治疗剂的效力的其他果蝇模型。

果蝇惊吓引起的步态分析 野生型， 州-S，苍蝇30秒（ 图1）后表现出的双管形瓶装置的仅约88％和苍蝇的顶部和底部部分的5％的鲁棒负geotactic响应分别。暴露于125μM和250μM的鱼藤酮3天蝇表明在果蝇中的顶端部分，并在果蝇中的底部的数目略有增加的数量略有减少。暴露于500μM的鱼藤酮蝇显示显著缺陷在负geotactic反应（P &lt;0.05 ANOVA，邦弗...

Watch this Scientific Journal Video about Methods to Characterize Spontaneous and Startle-induced Locomotion in a Rotenone-induced Parkinson's Disease Model of Drosophila at JoVE.com

Methods to Characterize Spontaneous and Startle-induced Locomotion in a Rotenone-induced Parkinson&#039;s Disease Model of Drosophila

帕金森病是一种神经退行性疾病而导致的多巴胺能神经元在中枢神经系统的变性，造成运动缺陷。鱼藤酮车型帕金森氏症的果蝇 。本文概述了两种检测的特点所造成的鱼藤酮是自发性的，惊吓引起的运动缺陷。

方法来描述自发和惊吓引起的步态中的鱼藤酮诱导的帕金森病模型<em&gt;果蝇</em

Parkinson&#8217;s disease is a neurodegenerative disorder that results from the degeneration of dopaminergic neurons in the central nervous system, ...

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Neuroscience

Methods to Characterize Spontaneous and Startle-induced Locomotion in a Rotenone-induced Parkinson's Disease Model of Drosophila

10.0K 次观看.  Colby College. 帕金森病是一种神经退行性疾病而导致的多巴胺能神经元在中枢神经系统的变性，造成运动缺陷。鱼藤酮车型帕金森氏症的果蝇 。本文概述了两种检测的特点所造成的鱼藤酮是自发性的，惊吓引起的运动缺陷。 

视频: Methods to Characterize Spontaneous and Startle-induced Locomotion in a Rotenone-induced Parkinson's Disease Model of Drosophila

Autologous Blood Injection to Model Spontaneous Intracerebral Hemorrhage in Mice

Investigation of the pathophysiology of injury after intracerebral
hemorrhage (ICH) requires a reproducible animal model. While ICH
accounts for 10-15% of all strokes, there remains no specific
effective therapy. The autologous blood injection model in mice
involves the stereotaxic injection of arterial blood into the basal
ganglia mimicking a spontaneous hypertensive hemorrhage in man. The
response to hemorrhage can then be studied in vivo and the
neurobehavioral deficits quantified, allowing for description of the
ensuing pathology and the testing of potential therapeutic agents. The
procedure described in this protocol uses the double injection
technique to minimize risk of blood reflux up the needle track, no
anticoagulants in the pumping system, and eliminates all dead space
and expandable tubing in the system.

The autologous blood injection model of intracerebral hemorrhage in mice described in this protocol uses the double injection technique to minimize risk of blood reflux up the needle track, no anticoagulants in the pumping system, and eliminates all dead space and expandable tubing in the system.

自体血液的注入模型小鼠自发性脑出血

Investigation of the pathophysiology of injury after intracerebral
hemorrhage (ICH) requires a reproducible animal model.  While ICH
accounts for 10-15% ...

科研

神经科学

Methods to Assay Drosophila Behavior

Drosophila melanogaster, the fruit fly, has been used to study molecular mechanisms of a wide range of human diseases such as cancer, cardiovascular disease and various neurological diseases1. We have optimized simple and robust behavioral assays for determining larval locomotion, adult climbing ability (RING assay), and courtship behaviors of Drosophila. These behavioral assays are widely applicable for studying the role of genetic and environmental factors on fly behavior. Larval crawling ability can be reliably used for determining early stage changes in the crawling abilities of Drosophila larvae and also for examining effect of drugs or human disease genes (in transgenic flies) on their locomotion. The larval crawling assay becomes more applicable if expression or abolition of a gene causes lethality in pupal or adult stages, as these flies do not survive to adulthood where they otherwise could be assessed. This basic assay can also be used in conjunction with bright light or stress to examine additional behavioral responses in Drosophila larvae. Courtship behavior has been widely used to investigate genetic basis of sexual behavior, and can also be used to examine activity and coordination, as well as learning and memory. Drosophila courtship behavior involves the exchange of various sensory stimuli including visual, auditory, and chemosensory signals between males and females that lead to a complex series of well characterized motor behaviors culminating in successful copulation. Traditional adult climbing assays (negative geotaxis) are tedious, labor intensive, and time consuming, with significant variation between different trials2-4. The rapid iterative negative geotaxis (RING) assay5 has many advantages over more widely employed protocols, providing a reproducible, sensitive, and high throughput approach to quantify adult locomotor and negative geotaxis behaviors. In the RING assay, several genotypes or drug treatments can be tested simultaneously using large number of animals, with the high-throughput approach making it more amenable for screening experiments.

Methods to Assay Drosophila Behavior

Drosophila melanogaster is a genetically and behaviorally tractable model system that has been used to understand the molecular and cellular basis of many important biological processes for over a century 1. Drosophila has been well exploited to gain insights into the genetic basis of fly behavior.

对检测方法果蝇行为

Drosophila melanogaster, the fruit fly, has been used to study molecular mechanisms of a wide range of human diseases such as cancer, cardiovascular ...

Local and Global Methods of Assessing Thermal Nociception in Drosophila Larvae

In this article, we demonstrate assays to study thermal nociception in Drosophila larvae. One assay involves spatially-restricted (local) stimulation of thermal nociceptors1,2 while the second involves a wholesale (global) activation of most or all such neurons3. Together, these techniques allow visualization and quantification of the behavioral functions of Drosophila nociceptive sensory neurons.
The Drosophila larva is an established model system to study thermal nociception, a sensory response to potentially harmful temperatures that is evolutionarily conserved across species1,2. The advantages of Drosophila for such studies are the relative simplicity of its nervous system and the sophistication of the genetic techniques that can be used to dissect the molecular basis of the underlying biology4-6 In Drosophila, as in all metazoans, the response to noxious thermal stimuli generally involves a "nocifensive" aversive withdrawal to the presented stimulus7. Such stimuli are detected through free nerve endings or nociceptors and the amplitude of the organismal response depends on the number of nociceptors receiving the noxious stimulus8. In Drosophila, it is the class IV dendritic arborization sensory neurons that detect noxious thermal and mechanical stimuli9 in addition to their recently discovered role as photoreceptors10. These neurons, which have been very well studied at the developmental level, arborize over the barrier epidermal sheet and make contacts with nearly all epidermal cells11,12. The single axon of each class IV neuron projects into the ventral nerve cord of the central nervous system11 where they may connect to second-order neurons that project to the brain.
Under baseline conditions, nociceptive sensory neurons will not fire until a relatively high threshold is reached. The assays described here allow the investigator to quantify baseline behavioral responses or, presumably, the sensitization that ensues following tissue damage. Each assay provokes distinct but related locomotory behavioral responses to noxious thermal stimuli and permits the researcher to visualize and quantify various aspects of thermal nociception in Drosophila larvae. The assays can be applied to larvae of desired genotypes or to larvae raised under different environmental conditions that might impact nociception. Since thermal nociception is conserved across species, the findings gleaned from genetic dissection in Drosophila will likely inform our understanding of thermal nociception in other species, including vertebrates.

Local and Global Methods of Assessing Thermal Nociception in Drosophila Larvae

In this article, we demonstrate assays to study thermal nociception in Drosophila larvae. One assay involves spatially-restricted (local) stimulation of thermal nociceptors1,2 while the second involves a wholesale (global) activation of most or all such neurons3. Together, these techniques allow visualization and quantification of the behavioral functions of Drosophila nociceptive sensory neurons.

局部和全局的方法评估热痛觉<em&gt;果蝇</em&gt;幼虫

In this article, we demonstrate assays to study thermal nociception in Drosophila larvae. One assay involves spatially-restricted (local) stimulation of ...

Multi-unit Recording Methods to Characterize Neural Activity in the Locust (Schistocerca Americana) Olfactory Circuits

Detection and interpretation of olfactory cues are critical for the survival of many organisms. Remarkably, species across phyla have strikingly similar olfactory systems suggesting that the biological approach to chemical sensing has been optimized over evolutionary time1. In the insect olfactory system, odorants are transduced by olfactory receptor neurons (ORN) in the antenna, which convert chemical stimuli into trains of action potentials. Sensory input from the ORNs is then relayed to the antennal lobe (AL; a structure analogous to the vertebrate olfactory bulb). In the AL, neural representations for odors take the form of spatiotemporal firing patterns distributed across ensembles of principal neurons (PNs; also referred to as projection neurons)2,3. The AL output is subsequently processed by Kenyon cells (KCs) in the downstream mushroom body (MB), a structure associated with olfactory memory and learning4,5. Here, we present electrophysiological recording techniques to monitor odor-evoked neural responses in these olfactory circuits.
First, we present a single sensillum recording method to study odor-evoked responses at the level of populations of ORNs6,7. We discuss the use of saline filled sharpened glass pipettes as electrodes to extracellularly monitor ORN responses. Next, we present a method to extracellularly monitor PN responses using a commercial 16-channel electrode3. A similar approach using a custom-made 8-channel twisted wire tetrode is demonstrated for Kenyon cell recordings8. We provide details of our experimental setup and present representative recording traces for each of these techniques.

Multi-unit Recording Methods to Characterize Neural Activity in the Locust Schistocerca Americana Olfactory Circuits

We demonstrate variations of the extracellular multi-unit recording technique to characterize odor-evoked responses in the first three stages of the invertebrate olfactory pathway. These techniques can easily be adapted to examine ensemble activity in other neural systems as well.

多单位记录方法来描述蝗虫的神经活动（<em&gt; Schistocerca美洲</em&gt;）嗅觉电路

Detection and interpretation of olfactory cues are critical for the survival  of many organisms. Remarkably, species across phyla have strikingly similar ...

Determination of the Spontaneous Locomotor Activity in Drosophila melanogaster

Drosophila melanogaster has been used as an excellent model organism to study environmental and genetic manipulations that affect behavior. One such behavior is spontaneous locomotor activity. Here we describe our protocol that utilizes Drosophila population monitors and a tracking system that allows continuous monitoring of the spontaneous locomotor activity of flies for several days at a time. This method is simple, reliable, and objective and can be used to examine the effects of aging, sex, changes in caloric content of food, addition of drugs, or genetic manipulations that mimic human diseases.

Determination of the Spontaneous Locomotor Activity in Drosophila melanogaster

Drosophila melanogaster are useful in studying genetic or environmental manipulations that affect behaviors such as spontaneous locomotor activity.&#160; Here we describe a protocol that utilizes monitors with infrared beams and data analysis software to quantify spontaneous locomotor activity.&#160;

在自发活动的测定<em&gt;果蝇</em

Drosophila melanogaster has been used as an excellent model organism to study environmental and genetic manipulations that affect behavior. One such ...

In Vivo Single-Molecule Tracking at the Drosophila Presynaptic Motor Nerve Terminal

An increasing number of super-resolution microscopy techniques are helping to uncover the mechanisms that govern the nanoscale cellular world. Single-molecule imaging is gaining momentum as it provides exceptional access to the visualization of individual molecules in living cells. Here, we describe a technique that we developed to perform single-particle tracking photo-activated localization microscopy (sptPALM) in Drosophila larvae. Synaptic communication relies on key presynaptic proteins that act by docking, priming, and promoting the fusion of neurotransmitter-containing vesicles with the plasma membrane. A range of protein-protein and protein-lipid interactions tightly regulates these processes and the presynaptic proteins therefore exhibit changes in mobility associated with each of these key events. Investigating how mobility of these proteins correlates with their physiological function in an intact live animal is essential to understanding their precise mechanism of action. Extracting protein mobility with high resolution in vivo requires overcoming limitations such as optical transparency, accessibility, and penetration depth. We describe how photoconvertible fluorescent proteins tagged to the presynaptic protein Syntaxin-1A can be visualized via slight oblique illumination and tracked at the motor nerve terminal or along the motor neuron axon of the third instar Drosophila larva.

In Vivo Single-Molecule Tracking at the Drosophila Presynaptic Motor Nerve Terminal

Here we illustrate how single molecule photo-activated localization microscopy can be carried out on the motor nerve terminal of a live Drosophila melanogaster third instar larva.

在体内果蝇前运动神经末端的单分子跟踪

An increasing number of super-resolution microscopy techniques are helping to uncover the mechanisms that govern the nanoscale cellular world. ...

Quantifying Spontaneous Ca2+ Fluxes and their Downstream Effects in Primary Mouse Midbrain Neurons

Parkinson&#8217;s disease (PD) is a devastating neurodegenerative disorder caused by the degeneration of dopaminergic (DA) neurons. Excessive Ca2+ influx due to the abnormal activation of glutamate receptors results in DA excitotoxicity and has been identified as an important mechanism for DA neuron loss. In this study, we isolate, dissociate, and culture midbrain neurons from the mouse ventral mesencephalon (VM) of ED14 mouse embryos. We then infect the long-term primary mouse midbrain cultures with an adeno-associated virus (AAV) expressing a genetically encoded calcium indicator, GCaMP6f under control of the human neuron-specific synapsin promoter, hSyn. Using live confocal imaging, we show that cultured mouse midbrain neurons display spontaneous Ca2+ fluxes detected by AAV-hSyn-GCaMP6f. Bath application of glutamate to midbrain cultures causes abnormal elevations in intracellular Ca2+ within neurons and this is accompanied by caspase-3 activation in DA neurons, as demonstrated by immunostaining. The techniques to identify glutamate-mediated apoptosis in primary mouse DA neurons have important applications for the high content screening of drugs that preserve DA neuron health.

Quantifying Spontaneous Ca2+ Fluxes and their Downstream Effects in Primary Mouse Midbrain Neurons

Here we present a protocol to measure in vitro Ca2+ fluxes in midbrain neurons and their downstream effects on caspase-3 using primary mouse midbrain cultures. This model can be employed to study pathophysiologic changes related to abnormal Ca2+ activity in midbrain neurons, and to screen novel therapeutics for anti-apoptotic properties.

定量自发 Ca2+ 通量及其在原发性小鼠中脑神经元中的下游效应

Parkinson&#8217;s disease (PD) is a devastating neurodegenerative disorder caused by the degeneration of dopaminergic (DA) neurons. Excessive Ca2+ influx ...

Applying the RatWalker System for Gait Analysis in a Genetic Rat Model of Parkinson's Disease

Parkinson's disease (PD) is a progressive neurodegenerative disorder caused by the loss of dopaminergic (DA) neurons in the substantia nigra pars compacta. Gait abnormalities, including decreased arm swing, slower walking speed, and shorter steps are common in PD patients and appear early in the course of disease. Thus, the quantification of motor patterns in animal models of PD will be important for phenotypic characterization during disease course and upon therapeutic treatment. Most cases of PD are idiopathic; however, the identification of hereditary forms of PD uncovered gene mutations and variants, such as loss-of-function mutations in Pink1 and Parkin, two proteins involved in mitochondrial quality control that could be harnessed to create animal models. While mice are resistant to neurodegeneration upon loss of Pink1 and Parkin (single and combined deletion), in rats, Pink1 but not Parkin deficiency leads to nigral DA neuron loss and motor impairment. Here, we report the utility of FTIR imaging to uncover gait changes in freely walking young (2 months of age) male rats with combined loss of Pink1 and Parkin prior to the development of gross visually apparent motor abnormality as these rats age (observed at 4-6 months), characterized by hindlimb dragging as previously reported in Pink1 knockout (KO) rats.

Here we describe the RatWalker system, built by redesigning the MouseWalker apparatus to accommodate for the increased size and weight of rats. This system uses frustrated total internal reflection (FTIR), high-speed video capture, and open-access analysis software to track and quantify gait parameters.

将RatWalker系统应用于帕金森病遗传大鼠模型中的步态分析

Parkinson's disease (PD) is a progressive neurodegenerative disorder caused by the loss of dopaminergic (DA) neurons in the substantia nigra pars ...

Semi-Quantitative Determination of Dopaminergic Neuron Density in the Substantia Nigra of Rodent Models using Automated Image Analysis

Estimation of the number of dopaminergic neurons in the substantia nigra is a key method in pre-clinical Parkinson's disease research. Currently, unbiased stereological counting is the standard for quantification of these cells, but it remains a laborious and time-consuming process, which may not be feasible for all projects. Here, we describe the use of an image analysis platform, which can accurately estimate the quantity of labeled cells in a pre-defined region of interest. We describe a step-by-step protocol for this method of analysis in rat brain and demonstrate it can identify a significant reduction in tyrosine hydroxylase positive neurons due to expression of mutant &#945;-synuclein in the substantia nigra. We validated this methodology by comparing with results obtained by unbiased stereology. Taken together, this method provides a time-efficient and accurate process for detecting changes in dopaminergic neuron number, and thus is suitable for efficient determination of the effect of interventions on cell survival.

Here we present an automated method for semi-quantitative determination of dopaminergic neuron number in the rat substantia nigra pars compacta.

使用自动图像分析对啮齿动物模型的亚斯坦蒂娅尼格拉多巴明神经元密度进行半定量测定

Estimation of the number of dopaminergic neurons in the substantia nigra is a key method in pre-clinical Parkinson's disease research. Currently, unbiased ...

The 6-hydroxydopamine Rat Model of Parkinson's Disease

Motor symptoms of Parkinson's disease (PD)-bradykinesia, akinesia, and tremor at rest-are consequences of the neurodegeneration of dopaminergic neurons in the substantia nigra pars compacta (SNc) and dopaminergic striatal deficit. Animal models have been widely used to simulate human pathology in the laboratory. Rodents are the most used animal models for PD due to their ease of handling and maintenance. Moreover, the anatomy and molecular, cellular, and pharmacological mechanisms of PD are similar in rodents and humans. The infusion of the neurotoxin, 6-hydroxydopamine (6-OHDA), into a medial forebrain bundle (MFB) of rats reproduces the severe destruction of dopaminergic neurons and simulates PD symptoms. This protocol demonstrates how to perform the unilateral microinjection of 6-OHDA in the MFB in a rat model of PD and shows the motor deficits induced by 6-OHDA and predicted dopaminergic lesions through the stepping test. The 6-OHDA causes significant impairment in the number of steps performed with the contralateral forelimb.

The 6-hydroxydopamine (6-OHDA) model has been used for decades to advance the understanding of Parkinson's Disease. In this protocol, we demonstrate how to perform unilateral nigrostriatal lesions in the rat by injecting 6-OHDA in the medial forebrain bundle, assess motor deficits, and predict lesions using the stepping test.

帕金森病的6-羟基多巴胺大鼠模型

Motor symptoms of Parkinson's disease (PD)-bradykinesia, akinesia, and tremor at rest-are consequences of the neurodegeneration of dopaminergic neurons in ...