SG是从美国纽约州立大学布法罗分校和约翰R. Oishei基金会的资金支持。

<html> 1。试剂的制备<ol><li>准备PEMFA缓冲区，使用100mm的管道，2mm的EGTA，1毫米硫酸镁 4 。缓冲区的pH值调整至7.0。 PEMFA缓冲过滤消毒和储存在4 ° C </li><li> PBT的加入为1ml准备TRITON X - 100至100ml 1X PBS（磷酸盐缓冲液）（pH值7.0）。保存在4 ° C。 </li><li>准备1X PBS（pH值7.0）0.01％NaAcide的5％BSA（牛血清白蛋白）。 </li><li>准备的盐溶液，用0.7％NaCl和去离子水0.03％的Triton X - 100。 </li><li>准备结合PEMFA缓冲区4部分第1部分37％的甲醛固定液。 </li></ol> 2。收集胚胎<ol><li>一个笼子里，放置在一个塑料瓶含有琼脂喂养板适配器&gt; 100 苍蝇 ，果蝇的利益应变准备。 </li><li>保留在室温下72小时的苍蝇在黑暗的牢笼，改变板，每8-12小时，使苍蝇得到驯化笼，并且不要把鸡蛋。 </li><li>为实验收集了12个小时的胚胎。从容器中取出培养皿中，并取代它。以取出培养皿和关闭的菜洗的胚胎，使用的盐溶液和一个干净的油漆刷。 </li><li>收集在一个细筛的胚胎。用生理盐水漂洗，除去多余的酵母。 </li><li>使用压舌板轻轻地拿起胚胎和它们放置在一个小玻璃瓶，一顶帽子。关于盐溶液中添加1毫升。 </li></ol> 3。去除卵壳和卵黄膜和固定<ol><li>随着胚胎的收集，准备庚烷50％和50％的固色剂结合的庚烷解决办法。该解决方案将与正庚烷的顶部和底部的固定液分层。 </li><li>在玻璃小瓶，他们在5分钟的盐水洗孵化的胚胎。删除多余的盐水，用巴斯德吸管。 </li><li>用去离子水冲洗的胚胎。 </li><li> Dechorionate胚胎孵化在50％，3分钟漂白。 </li><li>用去离子水彻底冲洗胚胎。 </li><li>孵育30分钟，他们在轻轻摇动庚烷解决办法修复的胚胎。 </li><li>使用巴斯德吸管取代留在管的正庚烷甲醇固定液（底层）。摇大力消除卵黄膜。 devitellinized胚胎将沉到试管底部。 </li><li>收集在一个新的玻璃小瓶，用巴斯德吸管的胚胎。胚胎洗净用甲醇5次，每次5分钟孵化的胚胎。 </li></ol> 4。抗体染色，安装和可视化<ol><li>补充水分在50％PBT解决方案和50％甲醇为5分钟的混合胚胎。然后在100％PBT溶液5分钟补充水分。 </li><li>胚胎在5％BSA孵育0.01％，为1小时NaAcide解决方案，在4 ° C座</li><li>现在，胚胎抗体染色的准备。准备稀释至适当浓度0.01％的NaAcide溶液中使用5％BSA的主要抗体。我们使用一种抗体对半胱氨酸字符串蛋白（CSP），水泡突触蛋白和抗辣根过氧化物酶或辣根过氧化物酶抗体，它承认一个神经元的具体碳水化合物基团在表面的糖蛋白和标签的所有神经元。 </li><li> 4℃过夜孵育的胚胎° C的主要抗体溶液。 </li><li>翌日，洗的胚胎与PBT的10倍，超过一个小时的过程中。 </li><li>准备二次抗体溶液，通过稀释适当浓度。我们使用Alexa的抗体浓度为1:200。 </li><li>孵育两小时的胚胎在4 ° C的二次抗体溶液。 </li><li>洗净的胚胎与PBT的10倍，超过一个小时的过程中。 </li><li>孵育过夜Vectashield安装中等的胚胎。 </li><li>山在玻片上的胚胎，并用指甲油密封盖玻片。使用荧光显微镜可视化胚胎。 </li></ol> 5。代表性的成果<img src="/files/ftp_upload/2150/2150fig1a.jpg" alt="图1" /> 总警司 <img src="/files/ftp_upload/2150/2150fig1b.jpg" alt="图1" /> HRP - FITC <img src="/files/ftp_upload/2150/2150fig1c.jpg" alt="图1" /> 合并 图1：一个显示16-17阶段的胚胎的中枢神经系统，使用抗体对CSP（红色）和HRP（绿色）代表图像。请注意不同的染色的合缝。 <img src="/files/ftp_upload/2150/2150fig2a.jpg" alt="图2" /> 总警司 <img src="/files/ftp_upload/2150/2150fig2b.jpg" alt="图2" /> HRP - FITC “/&gt; 合并 图2：一个阶段16日至17日对CSP（红色）和HRP（绿色）抗体的胚胎PNS的代表图像。需要注意的外周神经元的重复模式。

<ol>
	<li>Bodmer, R., Barbel, S., Sheperd, S., Jack, J. W., Jan, L. Y., Jan, Y. N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Transformation+of+sensory+organs+by+mutations+of+the+cut+locus+of+D.+melanogaster.">Transformation of sensory organs by mutations of the cut locus of D. melanogaster.</a> Cell. 51, 293-307 (1987).</li><li>Bodmer, R., Carretto, R., Jan, Y. N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Neurogenesis+of+the+peripheral+nervous+system+in+Drosophila+embryos:+DNA+replication+patterns+and+cell+lineages.">Neurogenesis of the peripheral nervous system in Drosophila embryos: DNA replication patterns and cell lineages.</a> Neuron. 3, 21-32 (1989).</li><li>Campos-Ortega, J. A., Hartenstein, V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> The Embryonic Development of Drosophila melanogaster. , (1985).</li><li>Carroll, S. B., Scott, M. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Localization+of+the+fushi+tarazu+protein+during+Drosophila+embryogenesis.">Localization of the fushi tarazu protein during Drosophila embryogenesis.</a> Cell. 43, 47-57 (1985).</li><li>Dambly-Chaudi&#232;re, C., Ghysen, A., Jan, Y. N., Jan, L. Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Muscle+connections+between+imaginal+discs+in+Drosophila.">Muscle connections between imaginal discs in Drosophila.</a> Developmental Biology. 113, 288-294 (1986).</li></ol>

果蝇胚胎是一个功能强大的系统，为在神经元发育的调查细胞和分子的变化。在这个协议中，我们已经演示了如何准备整装胚胎进行免疫组化。卡罗尔和斯科特，1985年的协议，我们已经适应了这个协议。该协议还可以适应测试其他神经元抗体，但抗体的优化应该做。此外，胚胎PNS和中枢神经系统的发育缺陷，可以很容易地通过比较不同的突变品系的胚胎可视化。我们已经成功地使用这一?...

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		<th>Material Name</th>
		<th>Type</th>
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		<th>Catalogue Number</th>
		<th>Comment</th>
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<td>37% formaldehyde</td>
<td>&nbsp;</td>
<td>Fisher</td>
<td>BP531-25</td>
<td>&nbsp;</td>
<tr>
<td>CSP</td>
<td>&nbsp;</td>
<td>Developmental Studies Hybridoma Bank</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<tr>
<td>HRP-FITC</td>
<td>&nbsp;</td>
<td>Jackson ImmunoResearch Laboratories</td>
<td>323-095-021</td>
<td>&nbsp;</td>
<tr>
<td>Alexa Fluor 568 Goat Anti-Mouse IgG</td>
<td>&nbsp;</td>
<td>Invitrogen</td>
<td>A-11004</td>
<td>&nbsp;</td>
<tr>
<td>Vectashield Mounting Medium</td>
<td>&nbsp;</td>
<td>Vector Laboratories, Inc.</td>
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visualization of the embryonic nervous system in whole-mount drosophila embryos

果蝇的胚胎是有吸引力的模型系统，为研究神经元发育的细胞和分子基础。在这里，我们描述了果蝇胚胎神经系统使用神经元蛋白的抗体的可视化的过程。由于整个胚胎周围神经和中枢神经系统，在单个细胞水平（Dambly - Chaudière等，1986;。博德默等，1987;。博德默等 ，1989），这些系统的像差以及特点可以很容易地确定使用不同的神经元蛋白的抗体。胚胎发育是在特定的时间收集，以确保在适当的可视化的发展阶段的胚胎。集合后，胚胎的外层，蛋壳膜和周围胚胎的卵黄信封，被删除前固定。胚胎，然后与神经细胞抗体培养和可视化使用荧光标记的抗体。在阶段的胚胎12-17可视化访问胚胎的神经系统。阶段12的中枢神经系统生殖乐队开始阶段15的缩短和明确的格局已经实现的合缝。通过阶段17中枢神经系统的合同和PNS的全面发展（坎波斯，奥尔特加等人1985年）。因此，在PNS和CNS格局的变化可以很容易地观察到在这些发育阶段。

Watch this Scientific Journal Video about Visualization of the Embryonic Nervous System in Whole-mount Drosophila Embryos at JoVE.com

Visualization of the Embryonic Nervous System in Whole-mount Drosophila Embryos

我们描述了程序准备上演<em&gt;果蝇</em&gt;胚胎神经系统的胚胎发育过程中的可视化的胚胎。

在全安装的胚胎神经系统的可视化果蝇胚胎

The Drosophila embryo is an attractive model system for investigating the cellular and molecular basis of neuronal development. Here we describe the ...

visualization-embryonic-nervous-system-whole-mount-drosophila

Research

JoVE Journal

Neuroscience

Visualization of the Embryonic Nervous System in Whole-mount Drosophila Embryos

13.1K 次观看.  SUNY-University at Buffalo. 我们描述了程序准备上演果蝇胚胎神经系统的胚胎发育过程中的可视化的胚胎。

视频: Visualization of the Embryonic Nervous System in Whole-mount Drosophila Embryos

Whole Mount Preparation of the Adult Drosophila Ventral Nerve Cord for Giant Fiber Dye Injection

To analyze the axonal and dendritic morphology of neurons, it is essential to obtain accurate labeling of neuronal structures. Preparing well labeled samples with little to no tissue damage enables us to analyze cell morphology and to compare individual samples to each other, hence allowing the identification of mutant anomalies.
In the demonstrated dissection method the nervous system remains mostly inside the adult fly. Through a dorsal incision, the abdomen and thorax are opened and most of the internal organs are removed. Only the dorsal side of the ventral nerve cord (VNC) and the cervical connective 
(CvC) containing the big axons of the giant fibers (GFs)1 are exposed, while the brain containing the GF cell body and dendrites remains2 in the 
intact head. In this preparation most nerves of the VNC should remain attached to their muscles. 
Following the dissection, the intracellular filling of the giant fiber (GF) with a fluorescent dye is demonstrated. In the CvC the GF axons are located at the dorsal surface and thus can be easily visualized under a microscope with differential interference contrast (DIC) optics. This allows the injection of the GF axons with dye at this site to label the entire GF including the axons and their terminals in the VNC. This method results in reliable and strong staining of the GFs allowing the neurons to be imaged immediately after filling with an epifluorescent microscope. Alternatively, the fluorescent signal can be enhanced using standard immunohistochemistry procedures3 suitable for high resolution confocal microscopy.

Whole Mount Preparation of the Adult Drosophila Ventral Nerve Cord for Giant Fiber Dye Injection

An in vivo dissection of the adult Drosophila ventral nerve cord (VNC) is demonstrated. This particular dissection method causes little damage to the VNC allowing the subsequent labeling of the giant fiber neurons with fluorescent dye for high resolution imaging.

成人整装准备果蝇巨纤维染料注射腹侧神经索

To analyze the axonal and dendritic morphology of neurons, it is essential to obtain accurate labeling of neuronal structures. Preparing well labeled ...

科研

神经科学

Visualization of Proprioceptors in Drosophila Larvae and Pupae

Proprioception is the ability to sense the motion, or position, of body parts by responding to stimuli arising within the body. In fruitflies and other insects proprioception is provided by specialized sensory organs termed chordotonal organs (ChOs) 2. Like many other organs in Drosophila, ChOs develop twice during the life cycle of the fly. First, the larval ChOs develop during embryogenesis. Then, the adult ChOs start to develop in the larval imaginal discs and continue to differentiate during metamorphosis.
The development of larval ChOs during embryogenesis has been studied extensively 10,11,13,15,16. The centerpiece of each ChO is a sensory unit composed of a neuron and a scolopale cell. The sensory unit is stretched between two types of accessory cells that attach to the cuticle via specialized epidermal attachment cells 1,9,14. When a fly larva moves, the relative displacement of the epidermal attachment cells leads to stretching of the sensory unit and consequent opening of specific transient receptor potential vanilloid (TRPV) channels at the outer segment of the dendrite 8,12. The elicited signal is then transferred to the locomotor central pattern generator circuit in the central nervous system.
Multiple ChOs have been described in the adult fly 7. These are located near the joints of the adult fly appendages (legs, wings and halters) and in the thorax and abdomen. In addition, several hundreds of ChOs collectively form the Johnston's organ in the adult antenna that transduce acoustic to mechanical energy 3,5,17,4.
In contrast to the extensive knowledge about the development of ChOs in embryonic stages, very little is known about the morphology of these organs during larval stages. Moreover, with the exception of femoral ChOs 18 and Johnston's organ, our knowledge about the development and structure of ChOs in the adult fly is very fragmentary.
Here we describe a method for staining and visualizing ChOs in third instar larvae and pupae. This method can be applied together with genetic tools to better characterize the morphology and understand the development of the various ChOs in the fly.

Visualization of Proprioceptors in Drosophila Larvae and Pupae

A method to immunostain and visualize chordotonal organs in larvae and pupae of Drosophila melanogaster is described.

可视化本体感受器<em&gt;果蝇</em&gt;幼虫和蛹

Proprioception is the ability to sense the motion, or position, of body parts by responding to stimuli arising within the body. In fruitflies and other ...

Labeling of Single Cells in the Central Nervous System of Drosophila melanogaster

In this article we describe how to individually label neurons in the embryonic CNS of Drosophila melanogaster by juxtacellular injection of the lipophilic fluorescent membrane marker DiI. This method allows the visualization of neuronal cell morphology in great detail. It is possible to label any cell in the CNS: cell bodies of target neurons are visualized under DIC optics or by expression of a fluorescent genetic marker such as GFP. After labeling, the DiI can be transformed into a permanent brown stain by photoconversion to allow visualization of cell morphology with transmitted light and DIC optics. Alternatively, the DiI-labeled cells can be observed directly with confocal microscopy, enabling genetically introduced fluorescent reporter proteins to be colocalised. The technique can be used in any animal, irrespective of genotype, making it possible to analyze mutant phenotypes at single cell resolution.

Labeling of Single Cells in the Central Nervous System of Drosophila melanogaster

We present a technique for labeling single neurons in the central nervous system (CNS) of Drosophila embryos, which allows the analysis of neuronal morphology by either transmitted light or confocal microscopy.

标记的单细胞在中枢神经系统<em&gt;果蝇</em

In this  article we describe how to individually label neurons in the embryonic CNS of Drosophila melanogaster by juxtacellular injection of the ...

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. ...

Direct Intraventricular Delivery of Drugs to the Rodent Central Nervous System

Due to an inability to cross the blood brain barrier, certain drugs need to be directly delivered into the central nervous system (CNS). Our lab focuses specifically on antisense oligonucleotides (ASOs), though the techniques shown in the video here can also be used to deliver a plethora of other drugs to the CNS. Antisense oligonucleotides (ASOs) have the capability to knockdown sequence-specific targets 1 as well as shift isoform ratios of specific genes 2. To achieve widespread gene knockdown or splicing in the CNS of mice, the ASOs can be delivered into the brain using two separate routes of administration, both of which we demonstrate in the video.
The first uses Alzet osmotic pumps, connected to a catheter that is surgically implanted into the lateral ventricle. This allows the ASOs to be continuously infused into the CNS for a designated period of time. The second involves a single bolus injection of a high concentration of ASO into the right lateral ventricle. Both methods use the mouse cerebral ventricular system to deliver the ASO to the entire brain and spinal cord, though depending on the needs of the study, one method may be preferred over the other.

We describe a method to target drugs to the central nervous system by either implanting a catheter or performing a bolus injection into the right lateral ventricle in mice. We focus specifically on the delivery of antisense oligonucleotides. This technique is readily adaptable to other drugs and to rats.

啮齿动物的中枢神经系统药物直接脑室交货

Due to an inability to cross the blood brain barrier, certain drugs need to be directly delivered into the central nervous system (CNS). Our lab focuses ...

Whole Mount Immunolabeling of Olfactory Receptor Neurons in the Drosophila Antenna

Odorant molecules bind to their target receptors in a precise and coordinated manner. Each receptor recognizes a specific signal and relays this information to the brain. As such, determining how olfactory information is transferred to the brain, modifying both perception and behavior, merits investigation. Interestingly, there is emerging evidence that cellular transduction and transcriptional factors are involved in the diversification of olfactory receptor neuron. Here we provide a robust whole mount immunological labeling method to assay in vivo olfactory receptor neuron organization. Using this method, we identified all olfactory receptor neurons with anti-ELAV antibody, a known pan-neural marker and Or49a-mCD8::GFP, an olfactory receptor neuron specifically expressed in Nba neuron using anti-GFP antibody.

Whole Mount Immunolabeling of Olfactory Receptor Neurons in the Drosophila Antenna

Herein we describe the process of whole mount immunostaining of Drosophila antennae, which enables us to better understand the molecular mechanisms involved in the diversification of olfactory receptor neurons (ORN)s.

嗅觉受体神经元在全山免疫标记<em&gt;果蝇</em&gt;天线

Odorant molecules bind to their target receptors in a precise and coordinated manner. Each receptor recognizes a specific signal and relays this ...

In vivo Optogenetic Stimulation of the Rodent Central Nervous System

The ability to probe defined neural circuits in awake, freely-moving animals with cell-type specificity, spatial precision, and high temporal resolution has been a long sought tool for neuroscientists in the systems-level search for the neural circuitry governing complex behavioral states. Optogenetics is a cutting-edge tool that is revolutionizing the field of neuroscience and represents one of the first systematic approaches to enable causal testing regarding the relation between neural signaling events and behavior. By combining optical and genetic approaches, neural signaling can be bi-directionally controlled through expression of light-sensitive ion channels (opsins) in mammalian cells. The current protocol describes delivery of specific wavelengths of light to opsin-expressing cells in deep brain structures of awake, freely-moving rodents for neural circuit modulation. Theoretical principles of light transmission as an experimental consideration are discussed in the context of performing in vivo optogenetic stimulation. The protocol details the design and construction of both simple and complex laser configurations and describes tethering strategies to permit simultaneous stimulation of multiple animals for high-throughput behavioral testing.

In vivo Optogenetic Stimulation of the Rodent Central Nervous System

Optogenetics has become a powerful tool for use in behavioral neuroscience experiments. This protocol offers a step-by-step guide to the design and set-up of laser systems, and provides a full protocol for carrying out multiple and simultaneous in vivo optogenetic stimulations compatible with most rodent behavioral testing paradigms.

在啮齿动物中枢神经系统的体内光遗传学刺激

The ability to probe defined neural circuits in awake, freely-moving animals with cell-type specificity, spatial precision, and high temporal resolution ...

Whole Mount Labeling of Cilia in the Main Olfactory System of Mice

The mouse olfactory system comprises 6-10 million olfactory sensory neurons in the epithelium lining the nasal cavity. Olfactory neurons extend a single dendrite to the surface of the epithelium, ending in a structure called dendritic knob. Cilia emanate from this knob into the mucus covering the epithelial surface. The proteins of the olfactory signal transduction cascade are mainly localized in the ciliary membrane, being in direct contact with volatile substances in the environment. For a detailed understanding of olfactory signal transduction, one important aspect is the exact morphological analysis of signaling protein distribution. Using light microscopical approaches in conventional cryosections, protein localization in olfactory cilia is difficult to determine due to the density of ciliary structures. To overcome this problem, we optimized an approach for whole mount labeling of cilia, leading to improved visualization of their morphology and the distribution of signaling proteins. We demonstrate the power of this approach by comparing whole mount and conventional cryosection labeling of Kirrel2. This axon-guidance adhesion molecule is known to localize in a subset of sensory neurons and their axons in an activity-dependent manner. Whole mount cilia labeling revealed an additional and novel picture of the localization of this protein.

Cilia of olfactory sensory neurons contain proteins of the signal transduction cascade, but a detailed spatial analysis of their distribution is difficult in cryosections. We describe here an optimized approach for whole mount labeling and en face visualization of ciliary proteins.

纤毛对小鼠的主要嗅觉系统整装标签

The mouse olfactory system comprises 6-10 million olfactory sensory neurons in the epithelium lining the nasal cavity. Olfactory neurons extend a single ...

In Situ Ca2+ Imaging of the Enteric Nervous System

Reflex behaviors of the intestine are controlled by the enteric nervous system (ENS). The ENS is an integrative network of neurons and glia in two ganglionated plexuses housed in the gut wall. Enteric neurons and enteric glia are the only cell types within the enteric ganglia. The activity of enteric neurons and glia is responsible for coordinating intestinal functions. This protocol describes methods for observing the activity of neurons and glia within the intact ENS by imaging intracellular calcium (Ca2+) transients with fluorescent indicator dyes. Our technical discussion focuses on methods for Ca2+ imaging in whole-mount preparations of the myenteric plexus from the rodent bowel. Bulk loading of ENS whole-mounts with a high-affinity Ca2+ indicator such as Fluo-4 permits measurements of Ca2+ responses in individual neurons or glial cells. These responses can be evoked repeatedly and reliably, which permits quantitative studies using pharmacological tools. Ca2+ responses in cells of the ENS are recorded using a fluorescence microscope equipped with a cooled charge-coupled device (CCD) camera. Fluorescence measurements obtained using Ca2+ imaging in whole-mount preparations offer a straightforward means of characterizing the mechanisms and potential functional consequences of Ca2+ responses in enteric neurons and glial cells.

In Situ Ca2+ Imaging of the Enteric Nervous System

The enteric nervous system (ENS) is a network of neurons and glia located in the gut wall that controls intestinal reflexes. This protocol describes methods for recording the activity of enteric neurons and glia in live preparations of ENS using Ca2+ imaging.

在肠神经系统的原位 钙成像

Reflex behaviors of the intestine are controlled by the enteric nervous system (ENS). The ENS is an integrative network of neurons and glia in two ...

A Simple One-step Dissection Protocol for Whole-mount Preparation of Adult Drosophila Brains

There is an increasing interest in using Drosophila to model human brain degenerative diseases, map neuronal circuitries in adult brains, and study the molecular and cellular basis of higher brain functions. A whole-mount preparation of adult brains with well-preserved morphology is critical for such whole brain-based studies, but can be technically challenging and time-consuming. This protocol describes an easy-to-learn, one-step dissection approach of an adult fly head in less than 10 s, while keeping the intact brain attached to the rest of the body to facilitate subsequent processing steps. The procedure helps remove most of the eye and tracheal tissues normally associated with the brain that can interfere with the later imaging step, and also places less demand on the quality of the dissecting forceps. Additionally, we describe a simple method that allows convenient flipping of the mounted brain samples on a coverslip, which is important for imaging both sides of the brains with similar signal intensity and quality. As an example of the protocol, we present an analysis of dopaminergic (DA) neurons in adult brains of WT (w1118) flies. The high efficacy of the dissection method makes it particularly useful for large-scale adult brain-based studies in Drosophila.

A Simple One-step Dissection Protocol for Whole-mount Preparation of Adult Drosophila Brains

The adult Drosophila brain is a valuable system for studying neuronal circuitry, higher brain functions, and complex disorders. An efficient method to dissect whole brain tissue from the small fly head will facilitate brain-based studies. Here we describe a simple, one-step dissection protocol of adult brains with well-preserved morphology.

一个简单的一步解剖协议成人全安装准备<em&gt;果蝇</em&gt;脑

There is an increasing interest in using Drosophila to model human brain degenerative diseases, map neuronal circuitries in adult brains, and study the ...