我们要感谢奎因皮亚克大学的凯勒和玛吉实验室的所有成员，提供有用的建议。特别是，我们要感谢Barron L. Lincoln II在凯勒实验室内开发这种损伤测定。我们还要感谢Quinnipiac大学艺术与科学学院授予LC Keller的赠款。

1.试剂和设备的制备<ol><li>制备1X夹层缓冲液（70 mM的氯化钠，5mM的KCl，0.02 mM的氯化钙 ，20mM的MgCl 2的，10mM的碳酸氢钠 ，115毫蔗糖，5mM的海藻糖，5mM的HEPES; pH 7.2）中。 </li><li>准备1克磷酸盐缓冲盐水（PBS）。 </li><li>使用含有0.01％Triton X-100的1x PBS制备1x PBT。 </li><li>准备果汁琼脂平板。 14简要地说，在700毫升混合的H 2 O和高压釜30克琼脂。将0.5g对羟基苯甲酸甲酯溶于10mL乙醇中，并将溶液加入300 mL果汁浓缩物（葡萄或苹果）中。将浓缩物混合到高压灭菌的琼脂溶液中，倒入10 mm x 35 mm培养皿的盖子中。 注意:葡萄琼脂功率预混还可以从各种公司购买（参见材料表 ）。 </li><li>获得大小5镊子，允许一个机械伤害幼虫。 </li><li>使用标准果蝇 CO 2 </s基于麻醉的麻醉装置。 </li></ol> 2.选择果蝇幼虫<ol><li>拿一瓶或一瓶果汁，含有在25°C培养的流浪三龄幼虫。 </li><li>根据外观选择十只个体第二龄或第三龄幼虫。三龄幼虫可以通过前分枝螺旋体的外观鉴定，而二龄幼虫较小，具有更多的杆状前叶。 注意:幼稚园阶段也可以按时间确定。在25°C时，孵化后大约48 h出现第二龄幼虫，大约24 h后出现三龄幼虫。如果有兴趣在NMJ检查神经变性，二龄幼虫需要受伤。 </li></ol> 3.准备果蝇幼虫机械损伤<ol><li>用镊子或画笔仔细挑选个别幼虫，谨慎不要压缩它。 </li><li>将十只幼虫直接放入含有冷1X PBS或解剖缓冲液的玻璃皿中，以去除任何食物残渣并减缓幼虫运动。 </li></ol> 4. 果蝇幼虫的机械损伤和治疗<ol><li>将每个幼虫置于CO 2麻醉装置上。 </li><li>在解剖显微镜下，仔细将每只幼虫卷入其背侧，以便通过角质层观察节段性神经。 </li><li> 位置大小5镊子从嘴钩开始的长度大约从1/2到2/3。一旦定位，夹住包含5个镊子的节段神经的大约1/3的腹角质层。 <ol><li>为了确保幼虫节段性神经受伤，应用足够的力量粉碎节段性神经，但使角膜完好无损。确定正确的力量，粉碎10只幼虫并确保5小时后的死亡率低于50％。 </li></ol></li><li>损伤后，仔细将幼虫转移到含有约0.5g酵母膏的果汁琼脂平板上。放置每只幼虫，使其前端处于酵母糊上，尽管运动性受到破坏，仍可持续喂食。 注意:为了确保幼虫节段性神经受损，转移到琼脂板后可以观察到破坏的幼虫运动。受伤的幼虫在损伤部位后面有效地瘫痪，但仍然可以移动他们的嘴钩和饲料。受伤后动物的高死亡率是常见的，所以最好比实验所需的动物多20-50％。 </li><li>将含有酵母膏和10只受伤幼虫的琼脂平板放入空的100×15mm培养皿的底部。盖上这个petri板，现在用含有湿纸巾的琼脂板上含有受伤的幼虫，确保纸巾不接触幼虫或琼脂板。放这个培养皿放入更大的气密容器中。 </li><li>在伤后特定时期（6,12或24小时）取回幼虫进行解剖。 注意:为了检查损伤对轴突细胞骨架的即时影响，可以在损伤后直接切除幼虫。为了检查轴突运输缺陷，损伤后约6小时可以解剖幼虫。损伤后约12小时可以观察到泛素化蛋白质的积聚，并且可以观察到损伤后24小时，NMJ处的运动神经元变性。 </li></ol> 5. NMJ神经退行性分析<ol><li>解剖果蝇幼虫NMJ如前所述。 15,16简单地说，销幼虫到的Sylgard板，沿背表面切割，圆角与解剖缓冲一滴销以露出体壁肌肉组织。根据抗体，在4％多聚甲醛（PFA）或Bouin&#39;s Solution中修复幼虫身体用于可视化神经元和肌肉。 注意:如果将荧光蛋白标签（如绿色荧光蛋白（GFP））可视化为Bouin固定剂可能太苛刻，则应使用PFA固定。 </li><li>执行免疫荧光，先前已经描述了图11，13，16，同时标记突触前运动神经元和突触后相邻（SSR）肌肉褶皱。使用荧光共轭辣根过氧化物酶（HRP）（1:300） 17可视化突触前膜和轴突，使用抗Bruchpilot（Brp）抗体（nc82; 1:100; Developmental Studies Hybridoma Bank）染色活性区。用抗Dics Large（Dlg）抗体（1:10,000）同时标记突触后肌肉。 18 </li><li>如前所述，进行定量的NMJ神经变性测定。 8 </suP&gt;，9，10，11，12，13，14简言之，同时可视化对于前和突触后隔室单独NMJs。标记有突触后标记但不突触前标记的任何引物指示神经变性的位点（ 图1 ）。每个NMJ的平均引流次数以及每只动物的NMJ平均数量可以被量化。 </li></ol>

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作者没有什么可以披露的。

早先描述的神经元机械损伤可以用于诱导果蝇幼虫节段神经的损伤/应激。 4，5，6，14本实验技术先前已经被用于剖析事件导致神经变性的时间序列，以及检查在运动神经元细胞体的转录变化后的损伤。 5，14此外，该技术已被用在微流控芯片结合，以...

神经元变性发生在正常发育过程中，可由自然衰老过程，损伤，压力或疾病状态引起。 果蝇黑腹果蝇是常见的果蝇，提供了一个简单而强大的模型生物体来研究神经变性，因为促进神经元退化的分子机制有显着的相似之处。这些相似之处突出表明，大约70％的疾病相关人类基因具有果蝇同系物。 1另外，研究人类神经变性疾病的许多测定和技术工具已被开发并在果蝇中使用 。 2，3在果蝇中 ，神经肌肉接头（NMJ）允许细胞和电生理学特性的分析，并已被证明是研究神经肌肉病的重要系统由于可见Ñeuron肌肉连接。 2在本研究中，我们描述了果蝇幼虫体内神经元损伤测定，其允许节段性神经的可重复损伤。这种运动神经元损伤导致细胞事件的时间序列，导致NMJ损伤后24小时的神经变性。导致神经变性的可重复损伤运动神经元的能力具有多种应用，例如鉴定退行性过程所需的特定基因，对神经元损伤的转录反应的解剖以及保护性信号级联的分析。 4，5，6这方法也被结合使用微流体来研究神经元变性和再生在活的动物。 7 我们利用建立的定量测定来检查运动神经元的退化离子在果蝇 NMJ机械损伤后。该测定是基于突触前膜和蛋白质的丢失先于以突触后肌肉膜折叠为特征的亚突触网（SSR）的拆卸。 8，9，10，11，12，13该试验允许在突触前神经元已经失去了连接到相邻的突触后肌肉"突触足迹"的量化。退化过程已被证明是在幼虫发育12中进行的，并且不能通过改变的突触发育或发芽来解释。 8，9，10，11，12中的广告使用机械损伤超过预先存在的突变的优点是它允许解剖导致NMJ神经变性的细胞事件的时间序列。 13

<table><tbody><tr><td>Micro-dissecting scissors</td><td>Fine Science Tools</td><td>15000-08</td><td></td></tr><tr><td>Dumont #3 Forceps</td><td>Fine Science Tools</td><td>11231-30</td><td>Some people prefer size 3, while others prefer size 5</td></tr><tr><td>Dumont #5 Forceps</td><td>Fine Science Tools</td><td>11251-30</td><td>Some people prefer size 3, while others prefer size 5</td></tr><tr><td>CO&lt;sub&gt;2&lt;/sub&gt; Air Tank</td><td>Tech Air</td><td>UN 1013</td><td>Various tank sizes can be purchased</td></tr><tr><td>CO&lt;sub&gt;2&lt;/sub&gt; Anesthetizing Apparatus</td><td>Genesee Scientific</td><td>59-114</td><td></td></tr><tr><td>Stainless-steel pins, size 0.1</td><td>Fine Science Tools</td><td>26002-10</td><td></td></tr><tr><td>SylGard 184 Silicone Elastomer, Base and Curing Agent</td><td>Dow Corning</td><td>3097358-1004</td><td>To pour dissecting plates</td></tr><tr><td>Bouin&#039;s Solution</td><td>Sigma</td><td>HT 10132-1L</td><td>Antibodies should be tested for their efficiency in Bouin&#039;s and PFA</td></tr><tr><td>4% Paraformaldehyde (PFA) in Phosphate-Buffered Saline (PBS)</td><td>Affymetrix</td><td>FLY-8030-20</td><td>Antibodies should be tested for their efficiency in Bouin&#039;s and PFA</td></tr><tr><td>Dissecting Stereo MIcroscope</td><td>AmScope</td><td>SM-1BZ</td><td></td></tr><tr><td>Light Source</td><td>AmScope</td><td>HL150-AY-220V</td><td></td></tr><tr><td>anti- nc82 antibody</td><td>Developmental Studies Hybridoma Bank</td><td>nc82-s</td><td></td></tr><tr><td>anti-discs large antibody</td><td>Developmental Studies Hybridoma Bank</td><td>AF3</td><td></td></tr><tr><td>Alexa Fluor anti-horseradish peroxidase</td><td>Jackson Immunoresearch</td><td>123-545-021; 123-585-021; 123-605-021</td><td>One can you Alexa Fluor&amp;reg; 488, 594 or 647</td></tr><tr><td>Flystuff Grape Juice Agar Premix</td><td>Genesee Scientific</td><td>47-102</td><td></td></tr><tr><td>Microscope slides</td><td>Genesee Scientific</td><td>29-101</td><td></td></tr><tr><td>Glass Coverslips</td><td>Fisher Scientific</td><td>12-545-87</td><td></td></tr><tr><td>Thermo Scientific Nalgene Utility Box</td><td>Fisher Scientific</td><td>03-484C</td><td>Used to create humid chamber for larval recovery</td></tr></tbody></table>

a simple neuronal mechanical injury methodology to study drosophila motor neuron degeneration

神经元的退化发生在正常发育过程中，并且在损伤，压力和疾病的反应中发生。神经元变性的细胞标志在人类和无脊椎动物中是非常相似的，驱动这些过程的分子机制也是如此。 果蝇，果蝇 ，提供了一个强大但简单的遗传模型生物体来研究神经变性疾病的细胞复杂性。事实上，约70％的疾病相关人类基因具有果蝇同系物，并且已经使用苍蝇描述了大量的工具和测定来研究人类神经变性疾病。更具体地说， 果蝇中的神经肌肉接头（NMJ）已经被证明是研究神经肌肉疾病的有效系统，因为能够分析神经元和肌肉之间的结构连接。在这里，我们报告在果蝇的 体内运动神经元损伤测定再次诱导NMJ神经退行24h。使用这种方法，我们描述了导致运动神经元变性的细胞事件的时间序列。损伤方法具有多种应用，并且还被用于鉴定神经变性所需的特定基因并解剖对神经元损伤的转录反应。

使用这里提出的程序，我们已经证明机械神经元损伤允许神经变性事件的时间解剖。 14，第 18的事件的序列先前已表征和与细胞骨架的立即破坏，随后轴突贩卖缺陷，泛素化蛋白的积累和随后的神经变性24小时后损伤开始。在损伤前，WT NMJs显示突触前活动区标记Brp与整个NMJ的突触后标记物Dlg相一致（ 图1A ?...

Watch this Scientific Journal Video about A Simple Neuronal Mechanical Injury Methodology to Study Drosophila Motor Neuron Degeneration at JoVE.com

A Simple Neuronal Mechanical Injury Methodology to Study Drosophila Motor Neuron Degeneration

在这里，我们描述了一种简单且广泛可及的方法来伤害果蝇幼虫的节段性神经，可视化和量化三龄幼虫神经肌肉接头（NMJ）运动神经元的神经变性。

一个简单的神经元机械损伤方法学研究<em&gt;果蝇</em&gt;运动神经元变性

The degeneration of neurons occurs during normal development and in response to injury, stress, and disease. The cellular hallmarks of neuronal ...

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A Simple Neuronal Mechanical Injury Methodology to Study Drosophila Motor Neuron Degeneration

5.5K 次观看.  Quinnipiac University. 在这里，我们描述了一种简单且广泛可及的方法来伤害果蝇幼虫的节段性神经，可视化和量化三龄幼虫神经肌肉接头（NMJ）运动神经元的神经变性。 

视频: A Simple Neuronal Mechanical Injury Methodology to Study Drosophila Motor Neuron Degeneration

Using Microfluidics Chips for Live Imaging and Study of Injury Responses in Drosophila Larvae

Live imaging is an important technique for studying cell biological processes, however this can be challenging in live animals. The translucent cuticle of the Drosophila larva makes it an attractive model organism for live imaging studies. However, an important challenge for live imaging techniques is to noninvasively immobilize and position an animal on the microscope. This protocol presents a simple and easy to use method for immobilizing and imaging Drosophila larvae on a polydimethylsiloxane (PDMS) microfluidic device, which we call the 'larva chip'. The larva chip is comprised of a snug-fitting PDMS microchamber that is attached to a thin glass coverslip, which, upon application of a vacuum via a syringe, immobilizes the animal and brings ventral structures such as the nerve cord, segmental nerves, and body wall muscles, within close proximity to the coverslip. This allows for high-resolution imaging, and importantly, avoids the use of anesthetics and chemicals, which facilitates the study of a broad range of physiological processes. Since larvae recover easily from the immobilization, they can be readily subjected to multiple imaging sessions. This allows for longitudinal studies over time courses ranging from hours to days. This protocol describes step-by-step how to prepare the chip and how to utilize the chip for live imaging of neuronal events in 3rd instar larvae. These events include the rapid transport of organelles in axons, calcium responses to injury, and time-lapse studies of the trafficking of photo-convertible proteins over long distances and time scales. Another application of the chip is to study regenerative and degenerative responses to axonal injury, so the second part of this protocol describes a new and simple procedure for injuring axons within peripheral nerves by a segmental nerve crush.

Using Microfluidics Chips for Live Imaging and Study of Injury Responses in Drosophila Larvae

Drosophila larvae are an attractive model system for live imaging due to their translucent cuticle and powerful genetics. This protocol describes how to utilize a single-layer PDMS device, called the &#39;larva chip&#39; for live imaging of cellular processes within neurons of 3rd instar Drosophila larvae.

使用微流体芯片实时成像和损伤研究的响应<em&gt;果蝇</em&gt;幼虫

Live imaging is an important technique for studying cell biological processes, however this can be challenging in live animals. The translucent cuticle of ...

科研

生物工程

An Injury Paradigm to Investigate Central Nervous System Repair in Drosophila

An experimental method has been developed to investigate the cellular responses to central nervous system (CNS) injury using the fruit-fly Drosophila. Understanding repair and regeneration in animals is a key question in biology. The damaged human CNS does not regenerate, and understanding how to promote the regeneration is one of main goals of medical neuroscience. The powerful genetic toolkit of Drosophila can be used to tackle the problem of CNS regeneration.
A lesion to the CNS ventral nerve cord (VNC, equivalent to the vertebrate spinal cord) is applied manually with a tungsten needle. The VNC can subsequently be filmed in time-lapse using laser scanning confocal microscopy for up to 24 hr to follow the development of the lesion over time. Alternatively, it can be cultured, then fixed and stained using immunofluorescence to visualize neuron and glial cells with confocal microscopy. Using appropriate markers, changes in cell morphology and cell state as a result of injury can be visualized. With ImageJ and purposely developed plug-ins, quantitative and statistical analyses can be carried out to measure changes in wound size over time and the effects of injury in cell proliferation and cell death. These methods allow the analysis of large sample sizes. They can be combined with the powerful genetics of Drosophila to investigate the molecular mechanisms underlying CNS regeneration and repair.

An Injury Paradigm to Investigate Central Nervous System Repair in Drosophila

An injury paradigm using the Drosophila larval ventral nerve cord to investigate central nervous system regeneration and repair is described. Stabbing followed by laser scanning confocal microscopy in time-lapse and fixed specimens, combined with quantitative analysis with purposefully developed software and genetics, are used to investigate the molecular mechanisms of CNS regeneration and repair.

调查中枢神经系统修复中受伤范式<em&gt;果蝇</em

An experimental method has been developed to investigate the cellular responses to central nervous system (CNS) injury using the fruit-fly Drosophila. ...

神经科学

Retrograde Tracing of Drosophila Embryonic Motor Neurons Using Lipophilic Fluorescent Dyes

We describe a technique for retrograde labeling of motor neurons in Drosophila. We use an oil-dissolved lipophilic dye and deliver a small droplet to an embryonic fillet preparation by a microinjector. Each motor neuron whose membrane is contacted by the droplet can then be rapidly labeled. Individual motor neurons are continuously labeled, enabling fine structural details to be clearly visualized. Given that lipophilic dyes come in various colors, the technique also provides a means to get adjacent neurons labeled in multicolor. This tracing technique is therefore useful for studying neuronal morphogenesis and synaptic connectivity in the motor neuron system of Drosophila.

Retrograde Tracing of Drosophila Embryonic Motor Neurons Using Lipophilic Fluorescent Dyes

We describe a method for retrograde tracing of the Drosophila embryonic motor neurons using lipophilic fluorescent dyes.

使用亲脂荧光染料的果蝇胚胎运动神经元的逆行追踪

We describe a technique for retrograde labeling of motor neurons in Drosophila. We use an oil-dissolved lipophilic dye and deliver a small droplet to an ...

发育生物学

Mass Histology to Quantify Neurodegeneration in Drosophila

Progressive neurodegenerative diseases like Alzheimer&#39;s disease (AD) or Parkinson&#39;s disease (PD) are an increasing threat to human health worldwide. Although mammalian models have provided important insights into the underlying mechanisms of pathogenicity, the complexity of mammalian systems together with their high costs are limiting their use. Therefore, the simple but well-established Drosophila model-system&#160;provides an alternative for investigating the molecular pathways that are affected in these diseases. Besides behavioral deficits, neurodegenerative diseases are characterized by histological phenotypes such as neuronal death and axonopathy. To quantify neuronal degeneration and to determine how it is affected by genetic and environmental factors, we use a histological approach that is based on measuring the vacuoles in adult fly brains. To minimize the effects of systematic error and to directly compare sections from control and experimental flies in one preparation, we use the &#39;collar&#39; method for paraffin sections. Neurodegeneration is then assessed by measuring the size and/or number of vacuoles that have developed in the fly brain. This can either be done by focusing on a specific region of interest or by analyzing the entire brain by obtaining serial sections that span the complete head. Therefore, this method allows one to measure not only severe degeneration but also relatively mild phenotypes that are only detectable in a few sections, as occurs during normal aging.

Mass Histology to Quantify Neurodegeneration in Drosophila

Drosophila is widely used as a model system to study neurodegeneration. This protocol describes a method by which degeneration, as determined by vacuole formation in the brain, can be quantified. It also minimizes effects due to the experimental procedure by processing and sectioning control and experimental flies as one sample.

大众组织学量化在神经退行性变<em&gt;果蝇</em

Progressive neurodegenerative diseases like Alzheimer&#39;s disease (AD) or Parkinson&#39;s disease (PD) are an increasing threat to human health ...

A Drosophila In Vivo Injury Model for Studying Neuroregeneration in the Peripheral and Central Nervous System

The regrowth capacity of damaged neurons governs neuroregeneration and functional recovery after nervous system trauma. Over the past few decades, various intrinsic and extrinsic inhibitory factors involved in the restriction of axon regeneration have been identified. However, simply removing these inhibitory cues is insufficient for successful regeneration, indicating the existence of additional regulatory machinery. Drosophila melanogaster, the fruit fly, shares evolutionarily conserved genes and signaling pathways with vertebrates, including humans. Combining the powerful genetic toolbox of flies with two-photon laser axotomy/dendriotomy, we describe here the Drosophila sensory neuron &#8211; dendritic arborization (da) neuron injury model as a platform for systematically screening for novel regeneration regulators. Briefly, this paradigm includes a) the preparation of larvae, b) lesion induction to dendrite(s) or axon(s) using a two-photon laser, c) live confocal imaging post-injury and d) data analysis. Our model enables highly reproducible injury of single labeled neurons, axons, and dendrites of well-defined neuronal subtypes, in both the peripheral and central nervous system.

A Drosophila In Vivo Injury Model for Studying Neuroregeneration in the Peripheral and Central Nervous System

Here, we present a protocol using the Drosophila sensory neuron - dendritic arborization (da) neuron injury model, which combines in vivo live imaging, two-photon laser axotomy/dendriotomy, and the powerful fly genetic toolbox, as a platform for screening potential promoters and inhibitors of neuroregeneration.

一种研究外周和中枢神经系统 Neuroregeneration 的果蝇体内损伤模型

The regrowth capacity of damaged neurons governs neuroregeneration and functional recovery after nervous system trauma. Over the past few decades, various ...

医学

Morphological and Functional Evaluation of Axons and their Synapses during Axon Death in Drosophila melanogaster

Axon degeneration is a shared feature in neurodegenerative disease and when nervous systems are challenged by mechanical or chemical forces. However, our understanding of the molecular mechanisms underlying axon degeneration remains limited. Injury-induced axon degeneration serves as a simple model to study how severed axons execute their own disassembly (axon death). Over recent years, an evolutionarily conserved axon death signaling cascade has been identified from flies to mammals, which is required for the separated axon to degenerate after injury. Conversely, attenuated axon death signaling results in morphological and functional preservation of severed axons and their synapses. Here, we present three simple and recently developed protocols that allow for the observation of axonal morphology, or axonal and synaptic function of severed axons that have been cut-off from the neuronal cell body, in the fruit fly Drosophila. Morphology can be observed in the wing, where a partial injury results in axon death side-by-side of uninjured control axons within the same nerve bundle. Alternatively, axonal morphology can also be observed in the brain, where the whole nerve bundle undergoes axon death triggered by antennal ablation. Functional preservation of severed axons and their synapses can be assessed by a simple optogenetic approach coupled with a post-synaptic grooming behavior. We present examples using a highwire loss-of-function mutation and by over-expressing dnmnat, both capable of delaying axon death for weeks to months. Importantly, these protocols can be used beyond injury; they facilitate the characterization of neuronal maintenance factors, axonal transport, and axonal mitochondria.

Morphological and Functional Evaluation of Axons and their Synapses during Axon Death in Drosophila melanogaster

Here, we provide protocols to perform three simple injury-induced axon degeneration (axon death) assays in Drosophila melanogaster to evaluate the morphological and functional preservation of severed axons and their synapses.

在果蝇黑色无gaster的Axon死亡期间，对Axon及其突触的形态和功能评估

Axon degeneration is a shared feature in neurodegenerative disease and when nervous systems are challenged by mechanical or chemical forces. However, our ...

Visualizing Synaptic Degeneration in Adult Drosophila in Association with Neurodegeneration

Drosophila serves as a useful model for assessing synaptic structure and function associated with neurodegenerative diseases. While much work has focused on neuromuscular junctions (NMJs) in Drosophila larvae, assessing synaptic integrity in adult Drosophila has received much less attention. Here we provide a straightforward method for dissection of the dorsal longitudinal muscles (DLMs), which are required for flight ability. In addition to flight as a behavioral readout, this dissection allows for the both DLM synapses and muscle tissue to be amenable to structural analysis using fluorescently labeled antibodies for synaptic markers or proteins of interest. This protocol allows for the evaluation of the structural integrity of synapses in adult Drosophila during aging to model the progressive, age-dependent nature of most neurodegenerative diseases.

Visualizing Synaptic Degeneration in Adult Drosophila in Association with Neurodegeneration

The goal of this procedure is to dissect the dorsal longitudinal muscle (DLM) tissue to assess the structural integrity of DLM neuromuscular junctions (NMJs) in neurodegenerative disease models using Drosophila melanogaster.

与神经退化有关的成人 果 蝇的可视化突触变性

Drosophila serves as a useful model for assessing synaptic structure and function associated with neurodegenerative diseases. While much work has focused ...

Assessing the Structural Integrity of the Dorsal Longitudinal Muscle Neuromuscular Junctions in Drosophila

Source: Sidisky, J. M., et al. <a href="https://app.jove.com/v/61363">Visualizing Synaptic Degeneration in Adult Drosophila in Association with Neurodegeneration.</a> J. Vis. Exp. (2020).
This video demonstrates a protocol for assessing the structural integrity of neuromuscular junctions in the dorsal longitudinal muscles of Drosophila. The procedure involves staining neurons and muscles with two distinct antibodies and comparing fluorescence patterns between mutant and wild-type flies. A reduction in neuronal staining in mutant flies indicates neurodegeneration, while the muscle structure remains intact.

评估果蝇背纵肌神经肌肉接头的结构完整性

1. Flash freezing and thorax bisection

	Before beginning the bisections, fill a Dewar flask with liquid nitrogen wearing proper cryo-protective gloves ...

JoVE Encyclopedia of Experiments

Neuropathology

Neuromuscular Disorders

Two-Photon Laser-Induced Neural Injury: A Method to Observe Axon Degeneration and Regeneration in Drosophila Larvae

Source: Li, D., et al. <a href="https://www.jove.com/video/57557/" target="_blank">A Drosophila In Vivo Injury Model for Studying Neuroregeneration in the Peripheral and Central Nervous System</a>. J. Vis. Exp. (2018).
This video describes two-photon laser ablation of axons in a Drosophila melanogaster larva and an example protocol to induce injury and image the injury site.

双光子激光诱导的神经损伤:一种观察果蝇幼虫轴突变性和再生的方法

Source: Li, D., et al. A Drosophila In Vivo Injury Model for Studying Neuroregeneration in the Peripheral and Central Nervous System. J. Vis. Exp. 
...

生物学

黑腹果蝇（果蝇）

幼虫

Mechanically Injuring Drosophila Larvae Segmental Nerves to Induce Neurodegeneration

Source: Danella, E. B., et al. <a href="https://app.jove.com/v/56128">A Simple Neuronal Mechanical Injury Methodology to Study Drosophila Motor Neuron Degeneration.</a> J. Vis. Exp. (2017).
The video demonstrates inducing neurodegeneration in Drosophila larva segmental nerves via mechanical injury. The ventral cuticle of anesthetized larvae is pinched using forceps to cause injury to the segmental nerves. The injured larvae are maintained with food, allowing the injury to cause degeneration of the motor neurons in the nerves, causing disruption of their neuromuscular junctions (NMJs).

机械损伤果蝇幼虫节段神经以诱导神经退行性变

1. Preparation of Reagents and Equipment 

	Prepare 1x Dissection Buffer (70 mM NaCl, 5 mM KCl, 0.02 mM CaCl2, 20 mM MgCl2, 10 mM NaHCO3, 115 mM sucrose, ...

一个简单的神经元机械损伤方法学研究

摘要

探索更多视频

此视频中的章节

使用微流体芯片实时成像和损伤研究的响应

调查中枢神经系统修复中受伤范式

使用亲脂荧光染料的果蝇胚胎运动神经元的逆行追踪

大众组织学量化在神经退行性变

一种研究外周和中枢神经系统 Neuroregeneration 的果蝇体内损伤模型

在果蝇黑色无gaster的Axon死亡期间，对Axon及其突触的形态和功能评估

与神经退化有关的成人果蝇的可视化突触变性

评估果蝇背纵肌神经肌肉接头的结构完整性

双光子激光诱导的神经损伤:一种观察果蝇幼虫轴突变性和再生的方法

机械损伤果蝇幼虫节段神经以诱导神经退行性变

一个简单的神经元机械损伤方法学研究

摘要

探索更多视频

此视频中的章节

使用微流体芯片实时成像和损伤研究的响应

调查中枢神经系统修复中受伤范式

使用亲脂荧光染料的果蝇胚胎运动神经元的逆行追踪

大众组织学量化在神经退行性变

一种研究外周和中枢神经系统 Neuroregeneration 的果蝇体内损伤模型

在果蝇黑色无gaster的Axon死亡期间，对Axon及其突触的形态和功能评估

与神经退化有关的成人 果 蝇的可视化突触变性

评估果蝇背纵肌神经肌肉接头的结构完整性

双光子激光诱导的神经损伤:一种观察果蝇幼虫轴突变性和再生的方法

机械损伤果蝇幼虫节段神经以诱导神经退行性变

与神经退化有关的成人果蝇的可视化突触变性