我们想承认，这种技术最初所描述的斯巴达等 ，2012年，并很容易地被使用在我们的实验室。

*所有材料连同各自的制造商和/或供应商下面列出的协议。 1。大会植入<ol><li>准备光纤的热固化性环氧树脂的混合物，通过添加固化剂，100毫克1克树脂。 </li><li>测量并削减约35毫米的125微米的光纤，100微米的核心，它的楔尖硬质合金划线得分。文士垂直放置在一个单一的，单向运动的光纤和得分。切割纤维将完全损坏的光纤纤芯。 </li><li>插入一个LC陶瓷插芯钻入副为127微米，凸侧朝下。 </li><li>插入到套圈的光纤。光纤应顺利滑动，和边缘伸出的凸端的套箍（ 图1a）。 </li><li>应用一滴的热固化性的光纤环氧树脂的平坦的端部和热用热枪直到变成黑色环氧树脂。环氧填充，因为它被加热并在固化前的套圈。环氧固化在1分钟之内的恒热应用。 </li><li>清理任何环氧树脂沿着套圈的两侧，它会妨碍与联接器的接口。 </li><li>波兰语氧化铝抛光垫的抛光表上（ 图1b）上的套圈用LC光纤抛光的光盘（FOPD）的凸端。圆形旋转图案和抛光四个等级按以下顺序：5，3，1 0.3微米砂砾。 </li><li>切割到适当的长度，使得它的目标感兴趣的区域的平坦的端的光纤。长度可以采用脑立体定位图谱确定。 </li><li>测试植入物，通过将其连接到所述激光通过下面描述的耦合器线。抛光的植入物的端部插入到耦合器的套筒，并应使直接接触相对的套箍。的植入物应能保持10毫瓦的光输出，测量的尖端处的第Ë植入物纤维。一个坏的植入物将光纤的前端附近有一个弱的焦点。 </li><li>存储成品植入物（ 图1c）的泡沫，直到使用。 </li></ol> 2。大会的光纤连接器电源线<ol><li>准备上述光纤的热固化性环氧树脂的混合物。 </li><li>测量和切成适当的长度为220μm的光纤200微米的核心，它的楔尖硬质合金划线得分。纤维的长度应允许自由移动鼠标周围的外壳，但不允许鼠标咀嚼通过光纤。 </li><li>插入到分叉管的长度略长于光纤的光纤长度。管应当具有比光纤的内径稍大。 </li><li>地带〜25毫米的光纤的一端，将其插入到多模FC MM的金属端套圈组件与230微米孔，直到它停止。光纤应坚持通过ferru乐端（ 图2a）。 </li><li>氰基丙烯酸酯（超级胶水）的金属端固定连接。覆盖与连接器引导和打磨套圈端与FC FOPD。进行圆形旋转模式和抛光四个等级顺序如下：5，3，1，0.3微米粒度（ 图2b）。 </li><li>剥去，并插入成LC陶瓷插芯（230微米内径孔）与凸状端部的前端的光纤的另一端。环氧树脂平端和热滴一滴，直至痊愈。 </li><li>波兰语的凸端的套箍上使用FC FOPD氧化铝抛光表如上所述。 </li><li>滑动一个LC的套箍套筒超过凸状的端部的套圈，直到套筒的中点。 </li><li>地点热缩管在分叉管，套管和热安全和保护的连接（ 图2c）。 </li><li>测试的耦合器，通过将其连接到所述激光源和测量的光的OUTPUT斯达康通过耦合器，用分光光度计。之间的激光输出与测得的耦合器输出的光损失应不超过30％。 </li></ol> 3。植入手术 *这是一个提示程序。仪器是无菌的，但不需要手套是由于不断操作仪器和设备之间的。 <ol><li>麻醉的小鼠腹膜内注射氯胺酮/赛拉嗪的混合物100和10毫克/公斤，分别使用30号针头。 </li><li>电推剪，刮头皮。擦拭头皮其次由Betasept擦拭（4％的氯己定溶液），2分钟，重复一次，用70％异丙醇。 </li><li>将鼠标在sterotaxic钻机及固定头，确保头骨水平。眼药膏眼睛，防止干燥和术后疼痛。根据“谅解备忘录”的生理状态与氧气的稀释挥发的异氟醚（1-3％，维持麻醉本身，应持续监测响应的尾部捏）。 </li><li>通过中线的头皮做一个切口，暴露颅骨眼轨道的lambda。推开结缔组织是必要的。 </li><li>塞拉芬夹来保存肌肤，保持访问头盖骨（ 图3a）。 </li><li>蚀刻与牙科采撷头盖骨的整个表面的方格图案。清洗碎片用无菌生理盐水。彻底干燥。 </li><li>过氧化氢（3％）的暴露颅骨，用棉签〜2-3秒微孔。多次洗净并晾干。此外，锚可以拧入头盖骨，作为斯巴达等中描述的。 （2012年）。 </li><li>同样，蚀刻的整个头骨与牙齿挑方格图案和用生理盐水洗去杂物。彻底干燥。 </li><li>使用的旋转工具，使一个小的毛刺孔开颅（&lt;1毫​​米直径），用无菌钻头（灭菌）以上该地区的利益，脑立体定位图谱校准前囟门和lambda确定。要小心，不要破坏硬膜任何组织或损坏。洗去杂物，彻底晾干。 </li><li>光纤套箍（植入物）插入到探头保持器，并连接到立体定位臂。 </li><li>代替直接上方的感兴趣的区域，使用立体定位臂（ 图3b）中的植入物定位。如果插入的脑组织中的光学纤维，纤维应在〜2毫米/分钟的速率缓慢​​前进。本的套圈应休息的剩余头盖骨。 </li><li>准备牙科水泥的混合物。该混合物应具有足够低的粘度，很容易地适用于头盖骨。混合物将可用于2-4分钟。 </li><li>用无菌牙签，应用薄，甚至跨头盖骨和到植入物的下部层的牙科用粘固剂。牙科水泥的基础层应覆盖尽可能多的表面区域的颅骨可能。与皮肤接触的鼠标，不要让牙齿水泥。这将导致增加困难缝合以及刺激鼠标。如果牙科用粘固剂不进入与皮肤接触，允许它的部分干燥，使得整个层的水泥可以设定前剥离。 </li><li>允许将其完全晾干。 </li><li>甚至应用层的牙科水泥和种植体周围形成一个小土丘头盖骨，让每个层完全干燥（ 图3c）。离开〜3-5毫米的凸端的套圈干净的水泥，以便顺利，通畅的连接。 </li><li>在土堆的牙科水泥和植入物周围使用无菌，一次性使用的丝绸编织缝合线（6-0）用C22针，缝合头皮。删除后7天。可选：使用兽医缝合后的其他具有约束力的债券。要确保使用最小的兽医债券的。过量可导致严重的皮肤损害，因搔抓。 </li><li>立即出现Ŷ手术后，鼠标应皮下注射与Ketprophen（5毫克/公斤），以减少术后疼痛。这应24小时后重复。外用止痛剂（布比卡因）和抗生素（抗生素软膏）应用缝合的皮肤的植入物和周围的基。 </li><li>关在笼子里加热毯将鼠标从麻醉中恢复。将鼠标在无菌术，术后恢复笼。恢复笼子里不应该包含任何床上用品，以保持温度，避免窒息。鼠标可以返回到原来的笼子或新的笼一次清醒。 </li></ol>正确的组装的光纤植入物和耦合器的查询结果在光源和在感兴趣的区域中的光纤的端部之间的最小光子损耗。精心打磨的光纤传输光在一个统一的，同心圆（ 图2d）。经过仔细的注入和缝合，植入物不会造成明显的刺激鼠标和没有任何显着降解的光纤或透射的光量为长期的研究（ 图3d，&gt; 1个月，未发表的观测）可以留在原地。注入或缝合不当可引起刺激，并可能导致在鼠标刮伤通过其头皮，露出的牙科用粘固剂，或破损，从牙科用粘固剂的套圈由于持续的操纵。 图4可以看出，在组装的系统的示意性框图。 <img alt="图1" src="/files/ftp_upload/50004/50004fig1.jpg" /> 图1。大会的植入式光纤。 （a）在光纤插入套箍，略微伸出的箭头所表示的凸端。 （b）在凸端的套箍使用FOPD上逐渐变细的等级的抛光表抛光。 （三）成品植入纤维的选择IC。 <a href="https://www.jove.com/files/ftp_upload/50004/50004fig1large.jpg" target="_blank">点击这里查看大图</a> 。 <img alt="图2" src="/files/ftp_upload/50004/50004fig2.jpg" /> 图2。大会使用的光纤耦合器，光纤旋转接头系绳给植入物。 （一）光纤通过环箍组装体的粘附。 （b）在套箍的装配侧插入FOPD和抛光使用逐渐变细的等级的研磨纸。 （c）在套箍套筒装配在套箍和热缩管固定。 （d）在完成的光纤耦合器应该产生的同心的光以最小的光子损耗。 <img alt="图3" src="/files/ftp_upload/50004/50004fig3.jpg" /> 图3。外科手术植入的光纤。 （a）在整个表面的cranium的暴露和结缔组织被清零。 （b）在与立体定位臂的位置被保持在光纤植入。 （三）牙科水泥固定光纤植入到头骨。 （D）植入后1个月，皮肤愈合植入物周围，没有任何迹象的刺激。 <img alt="图4" src="/files/ftp_upload/50004/50004fig4.jpg" /> 图4的功能的系统的示意图。

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光遗传学是一个功能强大的新技术，使特定的神经元亚型前所未有的控制。这可以被利用来调节神经回路的解剖和时间精度，同时也避免了细胞类型不分青红皂白，通过电极的电刺激侵袭作用。植入的光纤允许一致的，慢性刺激的神经回路在多个会话中保持清醒，用最少的组织损伤的小鼠行为。该系统最初是由斯巴达等 。和修改，以适应我们的目的，进了一步超越了植入套管和修复光纤在...

<table border="1"><tbody><tr><td> 试剂或设备的名称 </td><td> 公司 </td><td> 目录＃ </td><td> 评论 </td></tr><tr><td> LC插芯袖的</td><td>精密光纤产品（PFP） </td><td> SM-CS125S </td><td> 1.25 mm内径</td></tr><tr><td> FC MM预装配连接器</td><td>亲民党</td><td> MM-CON2004-2300 </td><td> 230微米套圈</td></tr><tr><td>米勒FOPD-LC光盘</td><td>亲民党</td><td> M1-80754 </td><td>对于LC套圈</td></tr><tr><td>分叉管</td><td>亲民党</td><td> FF9-250 </td><td> 900微米外径，内径250微米</td></tr><tr><td> MM LC棒套圈1.25毫米</td><td>亲民党</td><td>的MM-FER2007C-1270 </td><td> 127微米ID口径</td></tr><tr><td> MM LC棒套圈1.25毫米</td><td>亲民党</td><td> MM-FER2007C-2300 </td><td> 230微米ID缸径</td></tr><tr><td>可热固化的环氧树脂，固化剂和树脂</td><td>亲民党</td><td> ET-353ND-16OZ </td><td></td></tr><tr><td> FC / PC和SC / PC连接器研磨盘</td><td> Thorlabs公司</td><td> D50-FC </td><td>对于FC套圈</td></tr><tr><td>数字光功率和能量计</td><td> Thorlabs公司</td><td> PM100D </td><td>分光光度计</td></tr><tr><td>软磨片</td><td> Thorlabs公司</td><td> NRS913 </td><td> 9“×13”50硬度计</td></tr><tr><td>三氧化二铝研磨（抛光）表0.3，1，3，5微米糁</td><td> Thorlabs公司</td><td> LFG03P，LFG1P，LFG3P，LFG5P </td><td></td></tr><tr><td>标准硬包层多模光纤</td><td> Thorlabs公司</td><td> BFL37-200 </td><td>低OH，200微米的核心，0.37 NA </td></tr><tr><td>光纤剥线工具</td><td> Thorlabs公司</td><td> T10S13 </td><td>包钢/公司于：200微米/ 300微米</td></tr><tr><td>硅/二氧化硅光纤</td><td> Polymicro技术</td><td> FVP100110125 </td><td>高哦，紫外线增强，0.22 NA </td></tr><tr><td> 1×光纤旋转接头</td><td>多立克镜头</td><td> FRJ_FC-FC </td><td></td></tr><tr><td>单光纤跳线</td><td>多立克镜头</td><td> MFP_200/230/900-0.37_2m_FC-FC </td><td></td></tr><tr><td>热缩管，1/8英寸</td><td>盟军电子</td><td> 689-0267 </td><td></td></tr><tr><td>热风枪</td><td>盟军电子</td><td> 972-6966 </td><td> 250 W，750〜800°F </td></tr><tr><td>棉签涂药</td><td>清教徒的医疗产品公司</td><td> 806-WC </td><td></td></tr><tr><td> VetBond组织粘合剂</td><td>菲舍尔科学</td><td> 19-027136 </td><td></td></tr><tr><td>闪存义齿基丙烯酸</td><td>耶茨Motloid </td><td> ColdPourPowder +液体</td><td></td></tr><tr><td>新华社微型虹膜剪</td><td> INTEGRA Miltex </td><td> 18-1392 </td><td> 3-1/2“（8.9厘米），直，15毫米刀片</td></tr><tr><td>约翰霍普金斯大学牛头犬钳</td><td> INTEGRA Miltex </td><td> 7-290 </td><td> 1-1/2“（3.8厘米），弯曲</td></tr><tr><td> MEGA-扭矩电动实验室汽车</td><td>向量</td><td> EL-S </td><td></td></tr><tr><td>豹锉球＃1 </td><td>克拉克森实验室</td><td> 77.1006 </td><td></td></tr><tr><td>紫蓝色激光系统</td><td> CrystaLaser </td><td> CK473-050-O </td><td>波长：473纳米</td></tr><tr><td>激光电源</td><td> CrystaLaser </td><td> CL-2005 </td><td></td></tr><tr><td>的杜蒙＃2椎板钳</td><td> F国家统计局科学工具</td><td> 11223-20 </td><td></td></tr><tr><td>探测器</td><td>精细科学工具</td><td> 10140-02 </td><td></td></tr><tr><td> 10“直止血钳</td><td> EXCELTA </td><td> 35-PH </td><td></td></tr><tr><td>虎钳加权基地</td><td> ALTEX电子</td><td> PAN381 </td><td></td></tr></tbody></table>

fiber-optic implantation for chronic optogenetic stimulation of brain tissue

阐明神经元的连接模式一直是临床和基础神经科学的一个挑战。电已经分析模式的突触连接的金标准，但配对的电生理记录可以既繁琐和实验限制。光遗传学的发展引入一个优雅的方法来刺激神经 ​​元和电路中，无论是在体外和在体内 2,3。通过利用驱动视蛋白表达的离散的神经元群体的细胞类型特异性的启动子活性，其中一个可以精确地刺激基因定义的神经元的亚型，在不同的电路4-6。很好的描述的方法，以刺激神经元，包括电刺激和/或药理操纵，往往型细胞不加区别，侵入性的，并且可能会损坏周围组织。这些限制可能会改变正常的突触功能和/或电路的行为。此外，由于操纵的性质，目前的方法通常是急性的和终端。光遗传学得到的能力，以刺激神经元的相对无害的方式，而在转基因的目标神经元。 体内光遗传学研究，涉及的大多数目前使用的是通过植入插管6,7引导的光纤，但是，这种方法的局限性包括受损的脑组织与重复插入的光纤，和潜在的断裂的纤维内部的套管。由于光遗传学领域的蓬勃发展，更可靠的方法，以方便长期研究，以最小的附带组织损伤的慢性刺激是必要的。在这里，我们提供我们修改后的协议作为视频文章补充的方法，有效和优雅的描述斯巴达等 。的麻醉小鼠的头盖骨上的光纤植入物和其永久固定的制作，以及组装的纤维植入物连接到光纤耦合器的光源。的植入物，与光纤连接的固态激光器，允许一个有效的方法，对长期photostimulate功能的神经元的电路与使用小型，可拆卸的，系绳的组织破坏少9。永久固定的光纤植入提供稳定，长期在体内 optogenetic清醒，行为10用最少的组织损伤小鼠的神经回路的研究。

Watch this Scientific Journal Video about Fiber-optic Implantation for Chronic Optogenetic Stimulation of Brain Tissue at JoVE.com

Fiber-optic Implantation for Chronic Optogenetic Stimulation of Brain Tissue

现在的光遗传学的发展提供了手段精确地刺激基因定义的神经元和电路，既<em在体外</em&gt;和<em在体内</em&gt;。在这里，我们描述的组件和一个光纤为慢性光刺激脑组织植入。

光纤植入慢性Optogenetic刺激脑组织

Elucidating patterns of neuronal connectivity has been a challenge for both clinical and basic neuroscience. Electrophysiology has been the gold standard ...

fiber-optic-implantation-for-chronic-optogenetic-stimulation-brain

Research

JoVE Journal

Neuroscience

44.4K 次观看.  Baylor College of Medicine (BCM). 现在的光遗传学的发展提供了手段精确地刺激基因定义的神经元和电路，既

视频: Fiber-optic Implantation for Chronic Optogenetic Stimulation of Brain Tissue

Combining Transcranial Magnetic Stimulation and fMRI to Examine the Default Mode Network

The default mode network is a group of brain regions that are active when an individual is not focused on the outside world and the brain is at "wakeful rest."1,2,3 It is thought the default mode network corresponds to self-referential or "internal mentation".2,3
It has been hypothesized that, in humans, activity within the default mode network is correlated with certain pathologies (for instance, hyper-activation has been linked to schizophrenia 4,5,6 and autism spectrum disorders 7 whilst hypo-activation of the network has been linked to Alzheimer's and other neurodegenerative diseases 8). As such, noninvasive modulation of this network may represent a potential therapeutic intervention for a number of neurological and psychiatric pathologies linked to abnormal network activation. One possible tool to effect this modulation is Transcranial Magnetic Stimulation: a non-invasive neurostimulatory and neuromodulatory technique that can transiently or lastingly modulate cortical excitability (either increasing or decreasing it) via the application of localized magnetic field pulses.9
In order to explore the default mode network's propensity towards and tolerance of modulation, we will be combining TMS (to the left inferior parietal lobe) with functional magnetic resonance imaging (fMRI). Through this article, we will examine the protocol and considerations necessary to successfully combine these two neuroscientific tools.

In this article, we examine the methodology and considerations relevant to the combination of TMS and fMRI to examine the effects of brain stimulation on the default network.

结合经颅磁刺激和功能磁共振成像检查默认模式网络

The default mode network is a group of brain regions that are active when an individual is not focused on the outside world and the brain is at "wakeful ...

科研

神经科学

TMS: Using the Theta-Burst Protocol to Explore Mechanism of Plasticity in Individuals with Fragile X Syndrome and Autism

Fragile X Syndrome (FXS), also known as Martin-Bell Syndrome, is a genetic abnormality found on the X chromosome.1,2 Individuals suffering from FXS display abnormalities in the expression of FMR1 - a protein required for typical, healthy neural development.3 Recent data has suggested that the loss of this protein can cause the cortex to be hyperexcitable thereby affecting overall patterns of neural plasticity.4,5
In addition, Fragile X shows a strong comorbidity with autism: in fact, 30% of children with FXS are diagnosed with autism, and 2 - 5% of autistic children suffer from FXS.6
Transcranial Magnetic Stimulation (a non-invasive neurostimulatory and neuromodulatory technique that can transiently or lastingly modulate cortical excitability via the application of localized magnetic field pulses 7,8) represents a unique method of exploring plasticity and the manifestations of FXS within affected individuals. More specifically, Theta-Burst Stimulation (TBS), a specific stimulatory protocol shown to modulate cortical plasticity for a duration up to 30 minutes after stimulation cessation in healthy populations, has already proven an efficacious tool in the exploration of abnormal plasticity.9,10 
Recent studies have shown the effects of TBS last considerably longer in individuals on the autistic spectrum - up to 90 minutes.11 This extended effect-duration suggests an underlying abnormality in the brain's natural plasticity state in autistic individuals - similar to the hyperexcitability induced by Fragile X Syndrome.
In this experiment, utilizing single-pulse motor-evoked potentials (MEPs) as our benchmark, we will explore the effects of both intermittent and continuous TBS on cortical plasticity in individuals suffering from FXS and individuals on the Autistic Spectrum.

In this article, we examine the effects of Theta-Burst TMS stimulation on cortical plasticity in individuals suffering from Fragile X syndrome and individuals on the autistic spectrum.

TMS：使用的Theta突发协议，以探索个人与脆性X综合征与孤独症的可塑性Mechasnism

Fragile X Syndrome (FXS), also known as Martin-Bell Syndrome, is a genetic abnormality found on the X chromosome.1,2 Individuals suffering from FXS ...

State-Dependency Effects on TMS:  A Look at Motive Phosphene Behavior

Transcranial magnetic stimulation (TMS) is a non-invasive neurostimulatory and neuromodulatory technique that can transiently or lastingly modulate cortical excitability (either increasing or decreasing it) via the application of localized magnetic field pulses.1,2 Within the field of TMS, the term state dependency refers to the initial, baseline condition of the particular neural region targeted for stimulation. As can be inferred, the effects of TMS can (and do) vary according to this primary susceptibility and responsiveness of the targeted cortical area.3,4,5

In this experiment, we will examine this concept of state dependency through the elicitation and subjective experience of motive phosphenes. Phosphenes are visually perceived flashes of small lights triggered by electromagnetic pulses to the visual cortex. These small lights can assume varied characteristics depending upon which type of visual cortex is being stimulated. In this particular study, we will be targeting motive phosphenes as elicited through the stimulation of V1/V2 and the V5/MT+ complex visual regions.6

In this article, we examine the effects of visually relevant state dependency on TMS induced motive phosphenic presentations.

依赖国家对TMS的：一看动机幻视行为

Transcranial magnetic stimulation (TMS) is a non-invasive neurostimulatory and neuromodulatory technique that can transiently or lastingly modulate ...

The NeuroStar TMS Device: Conducting the FDA Approved Protocol for Treatment of Depression

The Neuronetics NeuroStar Transcranial Magnetic Stimulation (TMS) System is a class II medical device that produces brief duration, pulsed magnetic fields. These rapidly alternating fields induce electrical currents within localized, targeted regions of the cortex which are associated with various physiological and functional brain changes.1,2,3
In 2007, O'Reardon et al., utilizing the NeuroStar device, published the results of an industry-sponsored, multisite, randomized, sham-stimulation controlled clinical trial in which 301 patients with major depression, who had previously failed to respond to at least one adequate antidepressant treatment trial, underwent either active or sham TMS over the left dorsolateral prefrontal cortex (DLPFC). The patients, who were medication-free at the time of the study, received TMS five times per week over 4-6 weeks.4
The results demonstrated that a sub-population of patients (those who were relatively less resistant to medication, having failed not more than two good pharmacologic trials) showed a statistically significant improvement on the Montgomery-Asberg Depression Scale (MADRS), the Hamilton Depression Rating Scale (HAMD), and various other outcome measures. In October 2008, supported by these and other similar results5,6,7, Neuronetics obtained the first and only Food and Drug Administration (FDA) approval for the clinical treatment of a specific form of medication-refractory depression using a TMS Therapy device (FDA approval K061053).
In this paper, we will explore the specified FDA approved NeuroStar depression treatment protocol (to be administered only under prescription and by a licensed medical profession in either an in- or outpatient setting).

In this article, we examine the methodology and considerations relevant to the FDA approved depression treatment protocol using the Neuronetics NeuroStar TMS device.

NeuroStar TMS的设备进行美国FDA批准用于治疗抑郁症的议定书“

The Neuronetics NeuroStar Transcranial Magnetic Stimulation (TMS) System is a class II medical device that produces brief duration, pulsed magnetic ...

Selective Viral Transduction of Adult-born Olfactory Neurons for Chronic in vivo Optogenetic Stimulation

Local interneurons are continuously regenerated in the olfactory bulb of adult rodents1-3. In this process, called adult neurogenesis, neural stem cells in the walls of the lateral ventricle give rise to neuroblasts that migrate for several millimeters along the rostral migratory stream (RMS) to reach and incorporate into the olfactory bulb. To study the different steps and the impact of adult-born neuron integration into preexisting olfactory circuits, it is necessary to selectively label and manipulate the activity of this specific population of neurons. The recent development of optogenetic technologies offers the opportunity to use light to precisely activate this specific cohort of neurons without affecting surrounding neurons4,5. Here, we present a series of procedures to virally express Channelrhodopsin2(ChR2)-YFP in a temporally restricted cohort of neuroblasts in the RMS before they reach the olfactory bulb and become adult-born neurons. In addition, we show how to implant and calibrate a miniature LED for chronic in vivo stimulation of ChR2-expressing neurons.

Selective Viral Transduction of Adult-born Olfactory Neurons for Chronic in vivo Optogenetic Stimulation

Adult-born neurons of the olfactory bulb can be optogenetically controlled using Channelrhodopsin2-expressing lentiviral injection in the rostral migratory stream and chronic photostimulation with an implanted miniature LED.

慢性成人出生的嗅觉神经元的选择性病毒转导在体内 Optogenetic刺激

Local interneurons are continuously regenerated in the olfactory bulb of adult rodents1-3. In this process, called adult neurogenesis, neural stem cells ...

Optogenetic Stimulation of Escape Behavior in Drosophila melanogaster

A growing number of genetically encoded tools are becoming available that allow non-invasive manipulation of the neural activity of specific neurons in Drosophila melanogaster1. Chief among these are optogenetic tools, which enable the activation or silencing of specific neurons in the intact and freely moving animal using bright light. Channelrhodopsin (ChR2) is a light-activated cation channel that, when activated by blue light, causes depolarization of neurons that express it. ChR2 has been effective for identifying neurons critical for specific behaviors, such as CO2 avoidance, proboscis extension and giant-fiber mediated startle response2-4. However, as the intense light sources used to stimulate ChR2 also stimulate photoreceptors, these optogenetic techniques have not previously been used in the visual system. Here, we combine an optogenetic approach with a mutation that impairs phototransduction to demonstrate that activation of a cluster of loom-sensitive neurons in the fly's optic lobe, Foma-1 neurons, can drive an escape behavior used to avoid collision. We used a null allele of a critical component of the phototransduction cascade, phospholipase C-&beta;, encoded by the norpA gene, to render the flies blind and also use the Gal4-UAS transcriptional activator system to drive expression of ChR2 in the Foma-1 neurons. Individual flies are placed on a small platform surrounded by blue LEDs. When the LEDs are illuminated, the flies quickly take-off into flight, in a manner similar to visually driven loom-escape behavior. We believe that this technique can be easily adapted to examine other behaviors in freely moving flies.

Optogenetic Stimulation of Escape Behavior in Drosophila melanogaster

Genetically encoded optogenetic tools enable noninvasive manipulation of specific neurons in the Drosophila brain. Such tools can identify neurons whose activation is sufficient to elicit or suppress particular behaviors. Here we present a method for activating Channelrhodopsin2 that is expressed in targeted neurons in freely walking flies.

Optogenetic刺激的逃避行为<em&gt;果蝇</em

A  growing number of genetically encoded tools are becoming available that allow non-invasive  manipulation of the neural activity of specific neurons in ...

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 ...

Optogenetic Stimulation of the Auditory Nerve

Direct electrical stimulation of spiral ganglion neurons (SGNs) by cochlear implants (CIs) enables open speech comprehension in the majority of implanted deaf subjects1-6. Nonetheless, sound coding with current CIs has poor frequency and intensity resolution due to broad current spread from each electrode contact activating a large number of SGNs along the tonotopic axis of the cochlea7-9. Optical stimulation is proposed as an alternative to electrical stimulation that promises spatially more confined activation of SGNs and, hence, higher frequency resolution of coding. In recent years, direct infrared illumination of&#160;the cochlea has been used to evoke responses in the auditory nerve10. Nevertheless it requires higher energies than electrical stimulation10,11 and uncertainty remains as to the underlying mechanism12. Here we describe a method based on optogenetics to stimulate SGNs with low intensity blue light, using transgenic mice with neuronal expression of channelrhodopsin 2 (ChR2)13 or virus-mediated expression of the ChR2-variant CatCh14. We used micro-light emitting diodes (&#181;LEDs) and fiber-coupled lasers to stimulate ChR2-expressing SGNs through a small artificial opening (cochleostomy) or the round window. We assayed the responses by scalp recordings of light-evoked potentials (optogenetic auditory brainstem response: oABR) or by microelectrode recordings from the auditory pathway and compared them with acoustic and electrical stimulation.

Cochlear implants (CIs) enable hearing by direct electrical stimulation of the auditory nerve. However, poor frequency and intensity resolution limits the quality of hearing with CIs. Here we describe optogenetic stimulation of the auditory nerve in mice as an alternative strategy for auditory research and developing future CIs.

听觉神经的光遗传学刺激

Direct electrical stimulation of spiral ganglion neurons (SGNs) by cochlear implants (CIs) enables open speech comprehension in the majority of implanted ...

Microelectrode Guided Implantation of Electrodes into the Subthalamic Nucleus of Rats for Long-term Deep Brain Stimulation

Deep brain stimulation (DBS) is a widely used and effective therapy for several neurologic disorders, such as idiopathic Parkinson&#8217;s disease, dystonia or tremor. DBS is based on the delivery of electrical stimuli to specific deep anatomic structures of the central nervous system. However, the mechanisms underlying the effect of DBS remain enigmatic. This has led to an interest in investigating the impact of DBS in animal models, especially in rats. As DBS is a long-term therapy, research should be focused on molecular-genetic changes of neural circuits that occur several weeks after DBS. Long-term DBS in rats is challenging because the rats move around in their cage, which causes problems in keeping in place the wire leading from the head of the animal to the stimulator. Furthermore, target structures for stimulation in the rat brain are small and therefore electrodes cannot easily be placed at the required position. Thus, a set-up for long-lasting stimulation of rats using platinum/iridium electrodes with an impedance of about 1 M&#937; was developed for this study. An electrode with these specifications allows for not only adequate stimulation but also recording of deep brain structures to identify the target area for DBS. In our set-up, an electrode with a plug for the wire was embedded in dental cement with four anchoring screws secured onto the skull. The wire from the plug to the stimulator was protected by a stainless-steel spring. A swivel was connected to the circuit to prevent the wire from becoming tangled. Overall, this stimulation set-up offers a high degree of free mobility for the rat and enables the head plug, as well as the wire connection between the plug and the stimulator, to retain long-lasting strength.

A method for implanting electrodes into the subthalamic nucleus (STN) of rats is described. Better localization of the STN was achieved by using a microrecording system. Furthermore, a stimulation set-up is presented that is characterized by long-lasting connections between the head of the animal and the stimulator.

微电极引导电极植入到大鼠丘脑底核的长期深部脑刺激

Deep brain stimulation (DBS) is a widely used and effective therapy for several neurologic disorders, such as idiopathic Parkinson&#8217;s disease, ...

Ex Vivo Optogenetic Dissection of Fear Circuits in Brain Slices

Optogenetic approaches are now widely used to study the function of neural populations and circuits by combining targeted expression of light-activated proteins and subsequent manipulation of neural activity by light. Channelrhodopsins (ChRs) are light-gated cation-channels and when fused to a fluorescent protein their expression allows for visualization and concurrent activation of specific cell types and their axonal projections in defined areas of the brain. Via stereotactic injection of viral vectors, ChR fusion proteins can be constitutively or conditionally expressed in specific cells of a defined brain region, and their axonal projections can subsequently be studied anatomically and functionally via ex vivo optogenetic activation in brain slices. This is of particular importance when aiming to understand synaptic properties of connections that could not be addressed with conventional electrical stimulation approaches, or in identifying novel afferent and efferent connectivity that was previously poorly understood. Here, a few examples illustrate how this technique can be applied to investigate these questions to elucidating fear-related circuits in the amygdala. The amygdala is a key region for acquisition and expression of fear, and storage of fear and emotional memories. Many lines of evidence suggest that the medial prefrontal cortex (mPFC) participates in different aspects of fear acquisition and extinction, but its precise connectivity with the amygdala is just starting to be understood. First, it is shown how ex vivo optogenetic activation can be used to study aspects of synaptic communication between mPFC afferents and target cells in the basolateral amygdala (BLA). Furthermore, it is illustrated how this ex vivo optogenetic approach can be applied to assess novel connectivity patterns using a group of GABAergic neurons in the amygdala, the paracapsular intercalated cell cluster (mpITC), as an example.

Ex Vivo Optogenetic Dissection of Fear Circuits in Brain Slices

Optogenetic approaches are widely used to manipulate neural activity and assess the consequences for brain function. Here, a technique is outlined that&#160;upon in vivo expression of the optical activator Channelrhodopsin, allows for ex vivo analysis of synaptic properties of specific long range and local neural connections in fear-related circuits.

在脑片恐惧电路体外光遗传学解剖

Optogenetic approaches are now widely used to study the function of neural populations and circuits by combining targeted expression of light-activated ...