Name: multi-electrode array recordings of human epileptic postoperative cortical tissue
Uploaded: 2014-10-26T14:00:00.000+00:00
Duration: 13 min 14 s
Description: Watch this Scientific Journal Video about Multi-electrode Array Recordings of Human Epileptic Postoperative Cortical Tissue at JoVE.com

这项工作是由ANR（计划相思神经科学），FRC（联合会倒拉RECHERCHE河畔乐Cerveau）支持补助，巴黎市（诞生计划）的，INSERM和LaPitié妇女救济院医院（临床转化研究合同），以NR，从Neuropôle德RECHERCHE Francilien（NERF）以ED，从UNIVERSITE皮埃尔与玛丽·居里UPMC（方案融合），并从研究所杜Cerveau等è拉Moelle epiniere（巴黎），生长激素

注:这里所描述的协议遵循法国的"法国精品行业Consultatif国家科特迪瓦Ethique"的指导方针，并宣布由INSERM一个征集活动。 1.手术切除癫痫人体组织<ol><li>患者<ol><li>取得患者难治性癫痫患脑组织。手术前获得书面知情同意书。 </li><li>对于所有操作的癫痫患者进行了广泛的手术前的后处理，包括神经系统检查，神经心理学评估，表面脑电图和神经影像（MRI和有时18 FDG-PET扫描的; 图1A），接着切除的组织的外科手术后的组织学检查（ 图1B） 。注:手术中指出，当不同的探索本地化类似的发作起始区，当其去除的好处超过了手术风险。 </li></ol></li><li>手术<ol><li>麦ntain抗癫痫药物预操作性。与吸入七氟醚（6％递送率和100％的吸入的O 2分），静脉内舒芬太尼（0.3微克/千克）和antracurium（0.5毫克/千克），接着用静脉内舒芬太尼维修感应:在全身麻醉下进行手术（0.5微克/千克/小时）和吸入七氟醚（1，MAC）。 </li><li>使用神经导航系统，以允许在受损皮层（ 图1C）的精确定位。后的皮肤切口，执行中的骨随后用高洒钻头开颅钻孔内。轻轻地提升骨瓣，然后打开用剪刀（ 图1D）硬脑膜。 </li><li>使用术中超声，以便在手术显微镜下识别的异常皮层，和显微器械和超声波抽吸。 </li><li>凝结和根据脑沟限制切断的PIO-蛛网膜平面。使用软膜下剥离至Release来自各地的受损区域的软皮质。 </li><li>切正常皮质下白质完成一件"整块"切除术，不遗余力，尽可能的血管。避免创伤性操作的皮质。 </li><li>切成1厘米厚的片出了切除的组织的沿一个方向正交的软膜表面，包括沟底部及过渡皮质。 </li></ol></li></ol> 2.人工脑脊液（学联）的切片准备，孵化和录音<ol><li>学联的切片准备<ol><li>制备1升的蔗糖为基础的ACSF 7,8，含有（以mM计）:250蔗糖，3的KCl，25的NaHCO 3，10 D-葡萄糖，1的CaCl 2，10的MgCl 2;溶解这些盐在去离子水和充氧与卡波金的溶液至少10分钟。 </li><li>放〜700毫升蔗糖的ACSF中，在-80℃下进行50-60分钟（或-150℃下进行15-20分钟）。不断的休息根据氧合冰的解决方案。注:ACSF中为片制剂可贮存于4℃;在这种情况下，加在实验的CaCl 2和MgCl 2的那一天，将其冷却之前。 </li></ol></li><li>学联的切片孵化和录音<ol><li>准备至少3L的记录的ACSF 7,8的，它含有（以mM计）:124氯化钠，氯化钾3，26的NaHCO 3，10 D-葡萄糖，1.6的CaCl 2，1.3 MgCl 2的;溶解这些盐在去离子水和充氧与卡波金的溶液。在添加氯化镁 ，保留一些学联在0 镁离子学联进行录音。注:孵化/记录的ACSF可贮存于4℃;在这种情况下，添加的CaCl 2和MgCl 2的实验的日子。 </li></ol></li></ol> 3.设备切片孵化和录音<ol><li>接口室孵化片<ol><li>采用脑片室维持生活片体外&lt;/ em&gt;的接口模式。注:该腔室是由一低部，其中包含了加热和传感元件和一个陶瓷鼓泡器，并填充有蒸馏水，和上部，其中，所述片上休息晶状体组织中的平坦区域的片材中的腔室的中心（ 图2A - B）中 。 </li><li>使用两个单独的细的PTFE管中，通过该preoxygenated ACSF进入腔室。注:将管螺旋中被加热的水并到达腔室的上部;然后将溶液排出通过毛细作用带镜头薄纸，该输送溶液进入出口孔装置。 </li><li>连接接口室的聚四氟乙烯管，以蠕动泵，以允许与preoxygenated介质的切片的连续灌注。 </li><li>由通过陶瓷扩散器与卡波金（95％O 2和5％CO 2）的充氧维持高氧张力。注:此温热和湿润的混合气体达到片在通过适当的孔腔的上部。 </li><li>通过将腔室与温度控制器，通过一个特殊的双端连接线，具有连接器，用于所述腔室的加热元件和一个外部温度传感器的一端保持在所述腔室的上部的温度。在附近放置切片传感器在整个温育（ 图2C）检查温度。 </li></ol></li><li>多电极阵列记录<ol><li>使用平面多电极阵列120的氮化钛电极（直径30微米）绝缘用氮化硅和内部参比电极，布置在一个12×12的矩阵与200μm的中心到中心的间距。注意:用不同的电极直径和间距的其它配置是可能的。特别是，在MEA芯片可以加宽来样较大的人体组织的切片。 </li><li>获得的电信号在10kHz使用MEA2100-120系统（前和音响滤波器放大器，通过专用的软件（MC_Rack）集成的数据采集和模拟 - 数字转换器，16位的数据的分辨率，输入电压范围和带宽，通过软件可调）。注:MEA2100探头被赋予一个金属板，通过磁持有人可加热灌注元素固定的，在整个录制过程持续灌注切片。 </li><li>持组织向下在MEA由小自制铂锚，这确保了片和电极之间有良好的电耦合。 </li></ol></li></ol> 4.编写界面商会和工具解剖和切片<ol><li>接口室<ol><li>装满蒸馏水的界面室的下段。确保水的水平完全沉浸在加热元件和为2至4mm的腔室的下部和上部部分之间的连接处下方。在加热开关前，每天都要检查这个层面上，为了避免加热元件的损害。 </li><li>该腔室连接到一个卡波金源，赋予了二次流量调节器对气体压力进行微调。 </li><li>切镜头纸的片材，以适合的堰板腔的平坦区域，并且其定位接近输入解管，让来自输入线路的任何气泡逃逸到达片之前。 </li><li>切两片滞留棉线的，只要片透镜的组织，并将它们沿腔的平坦区域的主面的位置。注意:这有助于略微提高腔室中的灌注溶液的水平。 </li><li>设置蠕动泵的流速为1毫升/分钟，并启动泵。 </li><li>启动水的氧合作用在腔室的底部部分。 </li><li>将温度控制器的温度在37℃，并打开它。盖上一片封口膜的界面室的上部，并等待至少30分钟的Temperat .. [温度URE增加和稳定。 </li></ol></li><li>工具解剖和切片<ol><li>准备解剖套件包括两个细镊子，铲子，一个小勺子和一个刀片;两个玻璃培养皿中，两个电刷以及氰基丙烯酸酯胶。 </li><li>设置的vibratome的参数:厚度400微米;频率，70 - 90赫兹;振幅，0.9 - 1毫米，速度:0.1毫米/秒。 </li></ol></li></ol> 5.人力皮质切片制备<ol><li>取下-80℃冷冻蔗糖为基础的学联，混匀，以获得一种搪塑的。与卡波含氧它，把〜250毫升玻璃瓶，并保持它在杜瓦瓶中装满冰块，而要去医院的手术室，以获得组织。在此期间，保持静止，在-20℃下。 注:在手术中，神经外科医生解剖皮层组织的一小块。立即放入冰冷的蔗糖学联样品转移到实验室。 </li><li>保持所需要的从医院转移到实验室到最多30分钟的时间。在实验室中，以蔗糖为基础的含氧学联出​​-20℃的冰箱中，混合它有行贿，填补了培养皿和的vibratome缓冲盘，并同时启动与卡波充氧。小心取出一块人体组织从瓶中，用勺子，把它的皮氏培养皿（ 图2D）。 </li><li>使用镊子，小心地取出血栓，血管及脑膜，避免切割过程中的阻力。除去脑膜，包括在软质，从组织块的侧面开始，注意不要损伤大脑皮质表面。用刀片，切割组织，以获得平坦的表面，这将在对标本的vibratome板胶合的一侧。注:脑膜，包括在软质，除去来自组织块的侧面开始，注意不要损伤大脑皮质表面。 </li><li>如果TI块ssue大于MEA的腔室，在样品板（ 图2E）胶合之前切下一块的相应尺寸。 </li><li>胶组织，以获得横向皮质切片，放入缓冲盘切400微米厚的薄片，在低速（0.1毫米/秒）（ 图2F）。 </li><li>镜头切与组织切片维度件;用刮铲和刷子，传送片上，这些透镜的组织和开始记录（ 图2G - I）的前孵育它们在接口室中于37℃下至少1小时。连续地存储在相同的条件下的切片，直到使用。注:对镜头纸放置片是为了便于片底侧的两个灌注和从接口室切片的记录前除去的重要步骤。 <ol><li>在传送片的镜头纸，管理起来非常仔细，避免与触摸皮质层区域刷。 </li><li>制备切片一次一个，并立即他们每个传送到接口室，以确保适当的氧气供应和温度尽可能快地切片切除组织的块。 </li></ol></li></ol> 6.准备MEA设置为录音<ol><li>堵塞灌注系统以蠕动泵与MEA系统到计算机。含氧录音学联。将放大器内部的空MEA芯片。 </li><li>设置温度控制器，用于加热套管元件达到37℃，在MEA室并开始灌注在5-6毫升/分钟。注意:通常是必要的，设置温度3-4℃，比所述一个需要的话，根据房间温度和流率。 <ol><li>检查在MEA室内的温度用细温度计，将其放置在尽可能接近该中心，而不触及电极面积。 </li></ol></li><li>开始录制仿真软件e和检查所有的MEA的电极连接好，并且有因灌注无噪音。 </li><li>开始MEA_Monitor检查摄像机表的功能。 </li></ol> 7.转移切片从界面室以MEA芯片记录<ol><li>倒一些温热的玻璃培养皿记录的ACSF;用钳子，采取从接口室切片，通过镜头的组织抓住它，并把它放在培养皿。仔细分离从组织切片。如果在接口室中的溶液的水平期间的温育时间是经过精心调节，限幅将分离从镜头纸浸没它在溶液中;否则，用一个小刷子，方便拆卸。 </li><li>填充的MEA芯片与一些ACSF和用抹刀，切片转移到它。用塑料巴斯德吸管，轻轻地取下的解决方案，使切片粘附在电极区域，并与铂ANCH保持它在的地方或。加入1 ml录音学联，将MEA芯片进入放大器，并在5-6毫升/分钟开始灌注（ 图2J - L）。 </li><li>检查在MEA记录区域中的切片位置使用MEA_Monitor，如果需要调整它，以便从所述皮质层来记录并拍照。开始录制。 </li></ol>

<ol>
	<li>Huberfeld, G., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Glutamatergic+pre-ictal+discharges+emerge+at+the+transition+to+seizure+in+human+epilepsy.">Glutamatergic pre-ictal discharges emerge at the transition to seizure in human epilepsy.</a> Nature Neuroscience. 14 (5), 627-634 (2011).</li><li>Kwan, P., Brodie, M. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Early+identification+of+refractory+epilepsy.">Early identification of refractory epilepsy.</a> New England Journal of Medicine. 342 (5), 314-319 (2000).</li><li>Van Pelt, J., Corner, M. A., Wolters, P. S., Rutten, W. L. C., Ramakers, G. J. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Longterm+stability+and+developmental+changes+in+spontaneous+network+burst+firing+patterns+in+dissociated+rat+cerebral+cortex+cell+cultures+on+multielectrode+arrays.">Longterm stability and developmental changes in spontaneous network burst firing patterns in dissociated rat cerebral cortex cell cultures on multielectrode arrays.</a> Neuroscience Letters. 361 (1-3), 86-89 (2004).</li><li>Sun, J. -. J., Luhmann, H. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Spatio-temporal+dynamics+of+oscillatory+network+activity+in+the+neonatal+mouse+cerebral+cortex:+Network+oscillations+in+neonatal+cerebral+cortex.">Spatio-temporal dynamics of oscillatory network activity in the neonatal mouse cerebral cortex: Network oscillations in neonatal cerebral cortex.</a> European Journal of Neuroscience. 26 (7), 1995-2004 (2007).</li><li>Dossi, E., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Functional+Regeneration+of+the+ex-vivo+Reconstructed+Mesocorticolimbic+Dopaminergic+System.">Functional Regeneration of the ex-vivo Reconstructed Mesocorticolimbic Dopaminergic System.</a> Cerebral cortex. 23 (12), 2905-2922 (1991).</li><li>Darbon, P., Scicluna, L., Tscherter, A., Streit, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mechanisms+controlling+bursting+activity+induced+by+disinhibition+in+spinal+cord+networks.">Mechanisms controlling bursting activity induced by disinhibition in spinal cord networks.</a> The European journal of neuroscience. 15 (4), 671-683 (2002).</li><li>Cohen, I., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=On+the+Origin+of+Interictal+Activity+in+Human+Temporal+Lobe+Epilepsy+in+Vitro.">On the Origin of Interictal Activity in Human Temporal Lobe Epilepsy in Vitro.</a> Science. 298 (5597), 1418-1421 (1126).</li><li>Huberfeld, G., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Perturbed+chloride+homeostasis+and+GABAergic+signaling+in+human+temporal+lobe+epilepsy.">Perturbed chloride homeostasis and GABAergic signaling in human temporal lobe epilepsy.</a> The Journal of neuroscience. 27 (37), 9866-9873 (2007).</li><li>Maier, N., Morris, G., Johenning, F. W., Schmitz, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=An+Approach+for+Reliably+Investigating+Hippocampal+Sharp+Wave-Ripples+In+Vitro.">An Approach for Reliably Investigating Hippocampal Sharp Wave-Ripples In Vitro.</a> PLoS ONE. 4 (9), (2009).</li><li>H&#225;jos, N., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Maintaining+network+activity+in+submerged+hippocampal+slices:+importance+of+oxygen+supply.">Maintaining network activity in submerged hippocampal slices: importance of oxygen supply.</a> European Journal of Neuroscience. 29 (2), 319-327 (2009).</li><li>Wittner, L., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+epileptic+human+hippocampal+cornu+ammonis+2+region+generates+spontaneous+interictal-like+activity+in+vitro.">The epileptic human hippocampal cornu ammonis 2 region generates spontaneous interictal-like activity in vitro.</a> Brain. 132 (11), 3032-3046 (2009).</li><li>Dzhala, V., Valeeva, G., Glykys, J., Khazipov, R., Staley, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Traumatic+alterations+in+GABA+signaling+disrupt+hippocampal+network+activity+in+the+developing+brain.">Traumatic alterations in GABA signaling disrupt hippocampal network activity in the developing brain.</a> The Journal of neuroscience. 32 (12), 4017-4031 (2012).</li><li>Menendezdela de la Prida, L., Huberfeld, G., Cohen, I., Miles, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Threshold+Behavior+in+the+Initiation+of+Hippocampal+Population+Bursts.">Threshold Behavior in the Initiation of Hippocampal Population Bursts.</a> Neuron. 49 (1), 131-142 (2006).</li><li>Avoli, M., Drapeau, C., Louvel, J., Pumain, R., Olivier, A., Villemure, J. -. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Epileptiform+activity+induced+by+low+extracellular+magnesium+in+the+human+cortex+maintained+in+vitro.">Epileptiform activity induced by low extracellular magnesium in the human cortex maintained in vitro.</a> Annals of neurology. 30 (4), 589-596 (1991).</li><li>Heinemann, U., Dreier, J., Leschinger, A., Stabel, J., Draguhn, A., Zhang, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effects+of+anticonvulsant+drugs+on+hippocampal+neurons.">Effects of anticonvulsant drugs on hippocampal neurons.</a> Hippocampus. 4 (3), 291-296 (1994).</li><li>Li Zhang, C., Dreier, J. P., Heinemann, U. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Paroxysmal+epileptiform+discharges+in+temporal+lobe+slices+after+prolonged+exposure+to+low+magnesium+are+resistant+to+clinically+used+anticonvulsants.">Paroxysmal epileptiform discharges in temporal lobe slices after prolonged exposure to low magnesium are resistant to clinically used anticonvulsants.</a> Epilepsy research. 20 (2), 105-111 (1995).</li><li>Buzs&#225;ki, G., Anastassiou, C. A., Koch, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+origin+of+extracellular+fields+and+currents+-+EEG,+ECoG,+LFP+and+spikes.">The origin of extracellular fields and currents - EEG, ECoG, LFP and spikes.</a> Nature Reviews Neuroscience. 13 (6), 407-420 (2012).</li></ol>

提供了用于制作视频和开放存取出版的赞助多声道系统。

Pharmacoresistant癫痫病是一种罕见的疾病，它可以在体外培养人体组织进行研究。这使得研究人类癫痫的皮层，这显示只有部分转载在动物模型中特定的缺陷。这里介绍的方法可以准备和记录人术后组织离体保存与细胞活力和网络，使他们自发地产生癫痫活动。护比记录在体内类似的活动是非常重要的研究病理活动发生的机制。此外，这种方法允许探索人体组织和​​避免疾病的非完美的动物模型。然而，研究人体组织需要神经外科医生和实验室实验之间的同步。组织运输需要特定小心不要被创伤。另外，检体的数量和组织是有限的。最后，获得适当的控制的组织是迈ñ关注。有了这个准备，术后组织自发产生发作样放电正常学联7,8。发作样的事件，也可以在改性，proconvul​​sive ACSF中引起使癫痫发作起始和转换的机制，从发作状态癫痫发作可以调查。 人体组织可保持存活长达10小时的接口条件。片被认为是可行的，当观察到多单位活动或场电位自发并且当这样的活动是通过细胞外钾的增加和/或镁的减少增加的兴奋诱发。虽然变异性活动发生，可能反映了病理和皮质区的差异，我们只能在健康的切片呈现自发性发作探讨了组织癫痫的特点及诱发发作期放电。为了保存组织活力和活性，一个界面室用于存储吨他切片在36-37℃下恢复录制与MEA系统之前。事实上，几个小组已经清楚地表明相对于标准烧杯存储和温度对网络活动的保护的重要性接口基于存储系统的优点，如自发尖波涟漪或胆碱诱导的振荡9,10。片接口存储先前已用于记录从人类海马和subicular片1,11癫痫活动。与当前的MEA技术，从切片在界面条件的恢复期后，网络活动，在37℃下记录，在浸没的条件下，在高流动速率（5-6毫升/分钟）存在下，与MEA系统。 MEA芯片的直径减小（1.8厘米），其限定一个小容积室（&lt;1.5毫升），用增加的流量一起，提高了氧气供应的片，它已被证明是一个重要的因素为自发的和帕的rmacologically引起的网络活动，9,10。另外，循环的ACSF的减少量的方便的药理试验。 在切片过程是但一个创伤的组织12。两个神经元体系结构和氯化物稳态出现在组织表面（50微米）被扰动。记录MEA芯片，这样的组织大多是表面上无熔深大，可能引起创伤领域的活动的起源。但是，我们的数据表明，细胞外场电位检测到的局部被记录在最MEA的电极和以前的工作表明，它们被集成在从记录站点13中的 100至200微米的距离，表明记录在我们的制剂发作不太可能是受创伤的地区生产。此外，与钨电极使深层渗透进行的研究中，记录在人体组织中的癫痫样活动是相似对那些癫痫患者1,7,8观察。 离体组织的记录的另一个限制是不同脑区之间的连接的干扰，从而限制了动态neuromodulations。这也许可以解释为什么在这些组织中不发作状事件自发记录，但是需要通过离子操纵或药理学刺激增强的兴奋性来触发。因此，在该协议中，癫痫样事件是通过结合于细胞外K + 3至6毫米的变化，减少了外部的Mg 2+为1.3毫米的Mg 2+ -free ACSF中，为了提高组织的兴奋性诱导并去除镁离子依赖的 NMDA受体块。事实上，先前已经表明，癫痫样活动诱导使用的Mg 2+学联类似于记录在体内 14的脑电图癫痫人类新皮层和海马。此外，它已被证明在颞叶切片获得的癫痫样放电产生耐药性临床使用的抗惊厥药物长时间暴露于镁2+的ACSF 15,16之后，从而提供了一种模型， 在体外研究pharmacoresistant癫痫样事件。 多边环境协定允许两个记录，场电位和多单元的活动，包括在神经元动作电位体外的。因此，多边环境协定的较脑电图，探讨其在体内的神经元合奏的同步活动产生的场电位，但不给接入单个神经元的行为17一个强大的电工具。虽然最近的微电极能够记录在体内多单位的活动，包括在神经元动作电位，它们是侵入性的，所以它们的使用大多限于研究目的在颅内录音。特别是，记录的MEA代表所选择的技术研究癫痫的事件的时空格局，控制癫痫发作和传播的经典著作和新的抗癫痫药物的作用机制。值得注意的是，解开的细胞类型和癫痫放电的信令基础，穗排序技术和药理学试验应与MEA的技术进行组合。虽然多边环境协定可以给访问各个尖峰，他们不提供有关突触和生物物理特性的信息。在将来，其它技术应该被耦合到MEA的记录，以便更好样品细胞行为，网络活动和突触信号。例如，荧光成像解开神经元或神经胶质细胞的行为，以及离子动力学特性，可以用的MEA的录音进行组合。进一步事后组织学分析还可以揭示类型的细胞，蛋白质或受体的特异性改变，使得癫痫活动的位置可以用的具体的重排相关联神经结构。所述的MEA系统还可以嵌入在一个膜片钳设置以与人口活动相关联的单个细胞或电导。在未来，光遗传学工具也可以使用，条件是人类的切片可以培养在长期，作为执行用于器官切片，所以特定细胞类型的转染或感染可以被执行。

癫痫是一种慢性疾病，其中癫痫发作，这是间歇性的图案放电持续几秒钟到几十秒钟的临床表现相关的脑电图（EEG）记录，中断的发作状态，特点是神经元同步放电持续数十毫秒的存在并呼吁发作事件1。它会影响约1％的世界人口，虽然癫痫发作被控制在广大的患者，癫痫患者大约有三分之一不显示抗癫痫药2充分反应。在这种情况下，所谓的pharmacoresistant癫痫，其机制仍有待清楚地识别时，手术切除确定为癫痫发作区大脑的特定部位的仍然是唯一的替代治疗的患者给予积极的成果。因此，从手术切除的标本得到opportunity研究发作间放电和癫痫发作的产生和传播的上可行的人类中枢神经组织离体的机制，以及pharmacoresistance。 多电极阵列（MEAs）中，包括在空间上分布的微电极的布置，允许电生理学活性的同时刺激和记录来自多个站点的组织的，从而提供了一个极好的方法来研究自发和诱发活性的时空图案。该技术中，首先施加到监测神经元细胞培养物的活性3的发育变化，然后适于急性和器官型脑和脊髓切片4 - 6，目前被认为有价值的电生理工具。 目前的协议描述了如何从手术切除的组织准备的人脑片和多边环境协定可靠的前六记录VO发作，并从这些片癫痫样活动。这种技术因此提供了一种方法，以解决潜在的癫痫活动引发，繁殖和抗癫痫药物同时在蜂窝网络和水平的影响的基本机制。所有用于获取，编制，维护和记录人类切片的过程进行了详细的描述。

<table><tbody><tr><td>Brain Slice Chamber-2: Interface</td><td>AutoMate Scientific, Inc.</td><td>S-BSC2</td><td>It requires separate temperature controller</td></tr><tr><td>2-channel Temperature Controller</td><td>Multichannel Systems, Germany</td><td>TC02</td><td>It allows to plug in and use 2 interface chambers.&amp;nbsp;</td></tr><tr><td>Special cable for connection of TC01 to S-BSC2 with external reference PT100</td><td>Multichannel Systems, Germany</td><td>CA3</td><td>The reference PT100 is placed near the slices&amp;nbsp;</td></tr><tr><td>6pin plug-connector with PT100</td><td>Multichannel Systems, Germany</td><td>TS-PT100</td><td>External reference for CA3 cable</td></tr><tr><td>MEA workstation for recording data from 120-electrode MEAs</td><td>Multichannel Systems, Germany</td><td>MEA2100-120</td><td>It includes MEA2100-120 headstage and MEA2100 interface board. www.multichannelsystems.com</td></tr><tr><td>Microelectrode array for MEA2100-120</td><td>Multichannel Systems, Germany</td><td>120MEA200/30</td><td>Electrode spacing: 200 &amp;micro;m; electrode diameter: 30 &amp;micro;m; glass ring: 6 mm high. Different configurations (spacing, diameter, ring) possible.&amp;nbsp;</td></tr><tr><td>Video Microscope Table</td><td>Multichannel Systems, Germany</td><td>MEA-VMT-1</td><td>Table with a video microscope underneath to image the electrode field of the MEAs in an amplifier placed on top of the table and transfer the image to a computer.</td></tr><tr><td>Perfusion cannula</td><td>Multichannel Systems, Germany</td><td>PH01</td><td>Heatable perfusion cannula with temperature sensor; temperature can be programmed with TC02 controller</td></tr><tr><td>MC_Rack</td><td>Multichannel Systems, Germany</td><td></td><td>Software for data acquisition and recordings</td></tr><tr><td>Magnetic Perfusion Holder</td><td>Multichannel Systems, Germany</td><td>MPH</td><td>Magnetic perfusion holder for PH01 element to fix the perfusion cannula and connect the perfusion system to the amplifier&#039;s ground</td></tr><tr><td>Neuroexplorer</td><td>Nex Technologies</td><td></td><td>Software for data analysis; info@neuroexplorer.com</td></tr><tr><td>Peristaltic pump (drive unit)</td><td>Gilson</td><td>F155001</td><td>0,01 to 48 rp</td></tr><tr><td>Peristaltic pump (pump head)</td><td>Gilson</td><td>F117800</td><td>R2 two channel</td></tr><tr><td>Ultrasonic aspirator</td><td>Integra Life sciences, USA</td><td>Cusa Excel +</td><td>It allows blunt subpial dissection of the cortex</td></tr><tr><td>Neuronavigation</td><td>Isis Solutions, France</td><td>Surgiscope</td><td>It allows real time identification of the brain structure on the preoperative MRI</td></tr><tr><td>Vibratome HM 650 V</td><td>Microm</td><td></td><td>Block slicing into 400 &amp;mu;m thick slices</td></tr></tbody></table>

multi-electrode array recordings of human epileptic postoperative cortical tissue

癫痫，影响约1％的人口，包括一组特征为周期性发生癫痫发作的神经性疾病，这扰乱正常脑功能的影响。尽管与现有的抗癫痫药物靶向神经功能的治疗，对癫痫患者有三分之一是pharmacoresistant。在这种情况下，手术切除的脑区产生惊厥仍然是唯一的替代治疗。研究人类癫痫组织中作出了贡献，在过去10年时间理解新的致痫机制。的确，这些组织产生自发性发作癫痫放电以及其中可以记录与经典电生理技术，药理学诱导的发作的事件。值得注意的是，多电极阵列（多边环境协定），这是微加工设备中嵌入的空间排列的微电极阵列，提供了独特的机会，同时刺激并记录字段POTEntials，以及从组织的不同区域的多个神经元的动作电位。因此，多边环境协定的录音提供了一个很好的方法来研究自发性发作的时空格局和诱发癫痫样的事件和潜在的癫痫发作和传播的机制。在这里，我们描述了如何从手术切除的组织准备的人脑片和多边环境协定发作和发作般的事件在体外进行录制。

在灌注与正常记录的ACSF的组织（如前面所述）-7,8-人皮质切片的活动的记录开始。在这种状态下，当组织在手术过程中被完好地保存并稍后从医院和切片制备组织转移过程中，人们可以观察到自发活动，在小发作样的活动，由小簇的MEA的电极检测出的形式。发作样放电包括在一个场电位，通常是双相的，包括一个初始急剧负偏转，达到几十微伏，随后较长相对波持续几十毫秒。快的多单元活动通常嵌入在字段电位主要是在初始部分。发作间样活性的代表性实例示于图3Aii，其中记录了一个MEA的电极迹线被示出。 记录从癫痫发作体外人体皮质切片，必须灌注它们与一个"癫痫"的ACSF，其中的Mg 2+不存在和K +升高到6毫米，以提高组织的兴奋性1（K + 3至6毫米的增加独自不足以诱发癫痫发作;数据未示出）。一旦有0 镁离子6毫米的K +学联灌注开始，脑片的活性逐渐增加，首检出现在15〜20分钟（15.4±1.7分钟，N = 10，5例; 图3艾 ）。我们诱发发作样活性，如图3中所示，在患者进行测试的75％，并在所有的记录片的66.7％。 癫痫活动从MEA芯片的相邻电极被记录，并且场的潜在事件的发生动力学的比较研究允许发生和传播他们的网站。一位代表扣押的嗡嗡声一片记录一个大脑皮层与MEA系统示于图3B（记录由单个的MEA电极发作的代表性跟踪被显示在更高的时间分辨率， 如图3 AIII）。注意，癫痫发作是从MEA芯片的几乎所有的电极记录的，从而表明它能够传播到一个大的皮层区域。此外，在寻找从每个电极记录的单一的痕迹，所以能够观察到的发作横跨组织的逐渐传播。事实上，它开始先在皮质区覆盖电极阵列的右半部分，并且它逐渐扩散到覆盖左半侧区域（。即 ，比较从电极L6和B6的迹线; 图3B）。 <img alt="图1" fo:content-width="3in" src="/files/ftp_upload/51870/51870fig1highres.jpg" width="300" /> 图1:皮质发育不良的。ð其手术去除泰勒焦型皮质发育不良（白色箭头），嵌在左额叶脑沟可视化的术前MRI（A），后来被组织学检查（B）MAP激酶的标签确认;比例尺:200微米）示出与下白质的不完全迁移皮质dyslamination，异常的神经元和在轨道上的神经元。在手术中，在发育不良是根据与一个神经导航系统（C）的 MRI数据本地化。可视化手术（D）中含不典型增生的脑回的。 <img alt="图2" fo:content-width="4in" src="/files/ftp_upload/51870/51870fig2highres.jpg" width="400" /> 图2:设备和过程对人类大脑皮质切片制备的界面室用来维持人体皮质切片的离体 （A）的;切片休息晶状体组织中的平坦区域的片材在所述腔室（B）中 ，其中的温度传感器（C，左），通过一个双连接到温度控制器（C，向右，向上）的上半部分端线（C，右，下），置于保持37℃，整个切片的培养时间。一旦得到，将组织的切除块的培养皿中充满冰冷蔗糖基ACSF（D）和去除血凝块，血管和脑膜，以方便切割。如果组织块大于MEA室，隔离片的适当的尺寸，用刀片（E）和把它粘在所述的vibratome样品板（F）中 。切400微米厚的薄片，用抹刀（G）的帮助下转移他们的镜头纸（H）的作品，并培养他们在接口室（I）。之前的记录，圣雷莫已经透镜组织浸没在培养皿中充满人工脑脊液的切片，并用刮铲（J）传送到ACSF中填充的MEA芯片。除去溶液，让切片附着在MEA（K），并保持在适当的位置使用铂锚。放置在放大器（L）的 MEA中，开始灌注和记录。 <img alt="图3" fo:content-width="3in" src="/files/ftp_upload/51870/51870fig3highres.jpg" width="300" /> 图3:在人脑片痫性发作的MEA记录（A）记录由MEA的电极人类大脑皮层切片活动的代表痕迹显示在面板上我。在正常的ACSF的存在下​​，能够观察到间期状自发活性（小灰色矩形，放大面板ⅱ）;然后，当与0的Mg 2+ 6毫米的K +的ACSF灌流开始后，第一seizu重新出现了〜15分钟的延迟之后，并随后在2-3分钟间隔的其他事件。第三小组展示检获的大灰色矩形面板中的我和缩放比例。蓝色迹线示出了活动的变焦，用蓝线和箭头所示。面板IV）显示过滤，在250赫兹面板三所示的数据）高通，揭示多单位活动时放大（蓝色线）。（二）扣押在0存在的Mg 2+ 6毫米ķ代表录音+学联所有MEA的电极。每平方表示12×12的MEA阵列的一个电极和显示了一个80秒的时间窗口;虚线表示皮质表面的相对向的MEA阵列的位置。

Watch this Scientific Journal Video about Multi-electrode Array Recordings of Human Epileptic Postoperative Cortical Tissue at JoVE.com

Multi-electrode Array Recordings of Human Epileptic Postoperative Cortical Tissue

在这里，我们将介绍如何执行人类癫痫的皮层组织的多电极阵列记录。发作间期和发作性事件的癫痫组织切除，切片准备，多电极阵列记录都证明了细节。

人类癫痫术后皮层组织的多电极阵列录音

Epilepsy, affecting about 1% of the population, comprises a group of neurological disorders characterized by the periodic occurrence of seizures, which ...

multi-electrode-array-recordings-human-epileptic-postoperative

Universidad San Sebastian

Research

JoVE Journal

Medicine

20.5K 次观看.  CNRS UMR 7241, INSERM U1050, Collège de France. 在这里，我们将介绍如何执行人类癫痫的皮层组织的多电极阵列记录。发作间期和发作性事件的癫痫组织切除，切片准备，多电极阵列记录都证明了细节。 

视频: Multi-electrode Array Recordings of Human Epileptic Postoperative Cortical Tissue

Recording Human Electrocorticographic (ECoG) Signals for Neuroscientific Research and Real-time Functional Cortical Mapping

Neuroimaging studies of human cognitive, sensory, and motor processes are usually based on noninvasive techniques such as electroencephalography (EEG), magnetoencephalography or functional magnetic-resonance imaging. These techniques have either inherently low temporal or low spatial resolution, and suffer from low signal-to-noise ratio and/or poor high-frequency sensitivity. Thus, they are suboptimal for exploring the short-lived spatio-temporal dynamics of many of the underlying brain processes. In contrast, the invasive technique of electrocorticography (ECoG) provides brain signals that have an exceptionally high signal-to-noise ratio, less susceptibility to artifacts than EEG, and a high spatial and temporal resolution (i.e., &lt;1 cm/&lt;1 millisecond, respectively). ECoG involves measurement of electrical brain signals using electrodes that are implanted subdurally on the surface of the brain. Recent studies have shown that ECoG amplitudes in certain frequency bands carry substantial information about task-related activity, such as motor execution and planning1, auditory processing2 and visual-spatial attention3. Most of this information is captured in the high gamma range (around 70-110 Hz). Thus, gamma activity has been proposed as a robust and general indicator of local cortical function1-5. ECoG can also reveal functional connectivity and resolve finer task-related spatial-temporal dynamics, thereby advancing our understanding of large-scale cortical processes. It has especially proven useful for advancing brain-computer interfacing (BCI) technology for decoding a user's intentions to enhance or improve communication6 and control7. Nevertheless, human ECoG data are often hard to obtain because of the risks and limitations of the invasive procedures involved, and the need to record within the constraints of clinical settings. Still, clinical monitoring to localize epileptic foci offers a unique and valuable opportunity to collect human ECoG data. We describe our methods for collecting recording ECoG, and demonstrate how to use these signals for important real-time applications such as clinical mapping and brain-computer interfacing. Our example uses the BCI2000 software platform8,9 and the SIGFRIED10 method, an application for real-time mapping of brain functions. This procedure yields information that clinicians can subsequently use to guide the complex and laborious process of functional mapping by electrical stimulation.
Prerequisites and Planning:
Patients with drug-resistant partial epilepsy may be candidates for resective surgery of an epileptic focus to minimize the frequency of seizures. Prior to resection, the patients undergo monitoring using subdural electrodes for two purposes: first, to localize the epileptic focus, and second, to identify nearby critical brain areas (i.e., eloquent cortex) where resection could result in long-term functional deficits. To implant electrodes, a craniotomy is performed to open the skull. Then, electrode grids and/or strips are placed on the cortex, usually beneath the dura. A typical grid has a set of 8 x 8 platinum-iridium electrodes of 4 mm diameter (2.3 mm exposed surface) embedded in silicon with an inter-electrode distance of 1cm. A strip typically contains 4 or 6 such electrodes in a single line. The locations for these grids/strips are planned by a team of neurologists and neurosurgeons, and are based on previous EEG monitoring, on a structural MRI of the patient's brain, and on relevant factors of the patient's history. Continuous recording over a period of 5-12 days serves to localize epileptic foci, and electrical stimulation via the implanted electrodes allows clinicians to map eloquent cortex. At the end of the monitoring period, explantation of the electrodes and therapeutic resection are performed together in one procedure.
In addition to its primary clinical purpose, invasive monitoring also provides a unique opportunity to acquire human ECoG data for neuroscientific research. The decision to include a prospective patient in the research is based on the planned location of their electrodes, on the patient's performance scores on neuropsychological assessments, and on their informed consent, which is predicated on their understanding that participation in research is optional and is not related to their treatment. As with all research involving human subjects, the research protocol must be approved by the hospital's institutional review board. The decision to perform individual experimental tasks is made day-by-day, and is contingent on the patient's endurance and willingness to participate. Some or all of the experiments may be prevented by problems with the clinical state of the patient, such as post-operative facial swelling, temporary aphasia, frequent seizures, post-ictal fatigue and confusion, and more general pain or discomfort.
At the Epilepsy Monitoring Unit at Albany Medical Center in Albany, New York, clinical monitoring is implemented around the clock using a 192-channel Nihon-Kohden Neurofax monitoring system. Research recordings are made in collaboration with the Wadsworth Center of the New York State Department of Health in Albany. Signals from the ECoG electrodes are fed simultaneously to the research and the clinical systems via splitter connectors. To ensure that the clinical and research systems do not interfere with each other, the two systems typically use separate grounds. In fact, an epidural strip of electrodes is sometimes implanted to provide a ground for the clinical system. Whether research or clinical recording system, the grounding electrode is chosen to be distant from the predicted epileptic focus and from cortical areas of interest for the research. Our research system consists of eight synchronized 16-channel g.USBamp amplifier/digitizer units (g.tec, Graz, Austria). These were chosen because they are safety-rated and FDA-approved for invasive recordings, they have a very low noise-floor in the high-frequency range in which the signals of interest are found, and they come with an SDK that allows them to be integrated with custom-written research software. In order to capture the high-gamma signal accurately, we acquire signals at 1200Hz sampling rate-considerably higher than that of the typical EEG experiment or that of many clinical monitoring systems. A built-in low-pass filter automatically prevents aliasing of signals higher than the digitizer can capture. The patient's eye gaze is tracked using a monitor with a built-in Tobii T-60 eye-tracking system (Tobii Tech., Stockholm, Sweden). Additional accessories such as joystick, bluetooth Wiimote (Nintendo Co.), data-glove (5th Dimension Technologies), keyboard, microphone, headphones, or video camera are connected depending on the requirements of the particular experiment.
Data collection, stimulus presentation, synchronization with the different input/output accessories, and real-time analysis and visualization are accomplished using our BCI2000 software8,9. BCI2000 is a freely available general-purpose software system for real-time biosignal data acquisition, processing and feedback. It includes an array of pre-built modules that can be flexibly configured for many different purposes, and that can be extended by researchers' own code in C++, MATLAB or Python. BCI2000 consists of four modules that communicate with each other via a network-capable protocol: a Source module that handles the acquisition of brain signals from one of 19 different hardware systems from different manufacturers; a Signal Processing module that extracts relevant ECoG features and translates them into output signals; an Application module that delivers stimuli and feedback to the subject; and the Operator module that provides a graphical interface to the investigator.
A number of different experiments may be conducted with any given patient. The priority of experiments will be determined by the location of the particular patient's electrodes. However, we usually begin our experimentation using the SIGFRIED (SIGnal modeling For Realtime Identification and Event Detection) mapping method, which detects and displays significant task-related activity in real time. The resulting functional map allows us to further tailor subsequent experimental protocols and may also prove as a useful starting point for traditional mapping by electrocortical stimulation (ECS).
Although ECS mapping remains the gold standard for predicting the clinical outcome of resection, the process of ECS mapping is time consuming and also has other problems, such as after-discharges or seizures. Thus, a passive functional mapping technique may prove valuable in providing an initial estimate of the locus of eloquent cortex, which may then be confirmed and refined by ECS. The results from our passive SIGFRIED mapping technique have been shown to exhibit substantial concurrence with the results derived using ECS mapping10.
The protocol described in this paper establishes a general methodology for gathering human ECoG data, before proceeding to illustrate how experiments can be initiated using the BCI2000 software platform. Finally, as a specific example, we describe how to perform passive functional mapping using the BCI2000-based SIGFRIED system.

Recording Human Electrocorticographic ECoG Signals for Neuroscientific Research and Real-time Functional Cortical Mapping

We present a method for collecting electrocorticographic signals for research purposes from humans who are undergoing invasive epilepsy monitoring. We show how to use the BCI2000 software platform for data collection, signal processing and stimulus presentation. Specifically, we demonstrate SIGFRIED, a BCI2000-based tool for real-time functional brain mapping.

神经科学的研究和实时功能皮层映射记录人力Electrocorticographic（ECOG）信号

Neuroimaging studies of human cognitive, sensory, and motor processes are usually based on noninvasive techniques such as electroencephalography (EEG), ...

科研

神经科学

Simultaneous Eye Tracking and Single-Neuron Recordings in Human Epilepsy Patients

Intracranial recordings from patients with intractable epilepsy provide a unique opportunity to study the activity of individual human neurons during active behavior. An important tool for quantifying behavior is eye tracking, which is an indispensable tool for studying visual attention. However, eye tracking is challenging to use concurrently with invasive electrophysiology and this approach has consequently been little used. Here, we present a proven experimental protocol to conduct single-neuron recordings with simultaneous eye tracking in humans. We describe how the systems are connected and the optimal settings to record neurons and eye movements. To illustrate the utility of this method, we summarize results that were made possible by this setup. This data shows how using eye tracking in a memory-guided visual search task allowed us to describe a new class of neurons called target neurons, whose response was reflective of top-down attention to the current search target. Lastly, we discuss the significance and solutions to potential problems of this setup. Together, our protocol and results suggest that single-neuron recordings with simultaneous eye tracking in humans are an effective method to study human brain function. It provides a key missing link between animal neurophysiology and human cognitive neuroscience.

We describe a method to conduct single-neuron recordings with simultaneous eye tracking in humans. We demonstrate the utility of this method and illustrate how we used this approach to obtain neurons in the human medial temporal lobe that encode targets of a visual search.

人类癫痫患者同步眼动追踪和单神经记录

Intracranial recordings from patients with intractable epilepsy provide a unique opportunity to study the activity of individual human neurons during ...

行为学

Investigating the Function of Deep Cortical and Subcortical Structures Using Stereotactic Electroencephalography: Lessons from the Anterior Cingulate Cortex

Stereotactic Electroencephalography (SEEG) is a technique used to localize seizure foci in patients with medically intractable epilepsy. This procedure involves the chronic placement of multiple depth electrodes into regions of the brain typically inaccessible via subdural grid electrode placement. SEEG thus provides a unique opportunity to investigate brain function. In this paper we demonstrate how SEEG can be used to investigate the role of the dorsal anterior cingulate cortex (dACC) in cognitive control. We include a description of the SEEG procedure, demonstrating the surgical placement of the electrodes. We describe the components and process required to record local field potential (LFP) data from consenting subjects while they are engaged in a behavioral task. In the example provided, subjects play a cognitive interference task, and we demonstrate how signals are recorded and analyzed from electrodes in the dorsal anterior cingulate cortex, an area intimately involved in decision-making. We conclude with further suggestions of ways in which this method can be used for investigating human cognitive processes.

Stereotactic Electroencephalography (SEEG) is an operative technique used in epilepsy surgery to help localize seizure foci. It also affords a unique opportunity to investigate brain function. Here we describe how SEEG can be used to investigate cognitive processes in human subjects.

调查深皮层的功​​能和皮层下结构采用立体定向脑电图:由前扣带皮层的经验教训

Stereotactic Electroencephalography (SEEG) is a technique used to localize seizure foci in patients with medically intractable epilepsy. This procedure ...

Fabrication of Ti3C2 MXene Microelectrode Arrays for In Vivo Neural Recording

Implantable microelectrode technologies have been widely used to elucidate neural dynamics at the microscale to gain a deeper understanding of the neural underpinnings of brain disease and injury. As electrodes are miniaturized to the scale of individual cells, a corresponding rise in the interface impedance limits the quality of recorded signals. Additionally, conventional electrode materials are stiff, resulting in a significant mechanical mismatch between the electrode and the surrounding brain tissue, which elicits an inflammatory response that eventually leads to a degradation of the device performance. To address these challenges, we have developed a process to fabricate flexible microelectrodes based on Ti3C2 MXene, a recently discovered nanomaterial that possesses remarkably high volumetric capacitance, electrical conductivity, surface functionality, and processability in aqueous dispersions. Flexible arrays of Ti3C2 MXene microelectrodes have remarkably low impedance due to the high conductivity and high specific surface area of the Ti3C2 MXene films, and they have proven to be exquisitely sensitive for recording neuronal activity. In this protocol, we describe a novel method for micropatterning Ti3C2 MXene into microelectrode arrays on flexible polymeric substrates and outline their use for in vivo micro-electrocorticography recording. This method can easily be extended to create MXene electrode arrays of arbitrary size or geometry for a range of other applications in bioelectronics and it can also be adapted for use with other conductive inks besides Ti3C2 MXene. This protocol enables simple and scalable fabrication of microelectrodes from solution-based conductive inks, and specifically allows harnessing the unique properties of hydrophilic Ti3C2 MXene to overcome many of the barriers that have long hindered the widespread adoption of carbon-based nanomaterials for high-fidelity neural microelectrodes.

Fabrication of Ti3C2 MXene Microelectrode Arrays for In Vivo Neural Recording

We describe here a method for fabricating Ti3C2 MXene microelectrode arrays and utilizing them for in vivo neural recording.

用于Vivo神经记录的 Ti3C2 MXene 微电极阵列的制造

Implantable microelectrode technologies have been widely used to elucidate neural dynamics at the microscale to gain a deeper understanding of the neural ...

生物工程

Simultaneous Recordings of Cortical Local Field Potentials, Electrocardiogram, Electromyogram, and Breathing Rhythm from a Freely Moving Rat

Monitoring the physiological dynamics of the brain and peripheral tissues is necessary for addressing a number of questions about how the brain controls body functions and internal organ rhythms when animals are exposed to emotional challenges and changes in their living environments. In general experiments, signals from different organs, such as the brain and the heart, are recorded by independent recording systems that require multiple recording devices and different procedures for processing the data files. This study describes a new method that can simultaneously monitor electrical biosignals, including tens of local field potentials in multiple brain regions, electrocardiograms that represent the cardiac rhythm, electromyograms that represent awake/sleep-related muscle contraction, and breathing signals, in a freely moving rat. The recording configuration of this method is based on a conventional micro-drive array for cortical local field potential recordings in which tens of electrodes are accommodated, and the signals obtained from these electrodes are integrated into a single electrical board mounted on the animal's head. Here, this recording system was improved so that signals from the peripheral organs are also transferred to an electrical interface board. In a single surgery, electrodes are first separately implanted into the appropriate body parts and the target brain areas. The open ends of all of these electrodes are then soldered to individual channels of the electrical board above the animal's head so that all of the signals can be integrated into the single electrical board. Connecting this board to a recording device allows for the collection of all of the signals into a single device, which reduces experimental costs and simplifies data processing, because all data can be handled in the same data file. This technique will aid the understanding of the neurophysiological correlates of the associations between central and peripheral organs.

This study introduces a method for the simultaneous recording of local field potentials in the brain, electrocardiograms, electromyograms, and breathing signals of a freely moving rat. This technique, which reduces experimental costs and simplifies data analysis, will contribute to the understanding of the interactions between the brain and peripheral organs.

自由运动大鼠皮质局部电位、心电图、肌电图和呼吸节律的同步记录

Monitoring the physiological dynamics of the brain and peripheral tissues is necessary for addressing a number of questions about how the brain controls ...

Recording and Modulation of Epileptiform Activity in Rodent Brain Slices Coupled to Microelectrode Arrays

Temporal lobe epilepsy (TLE) is the most common partial complex epileptic syndrome and the least responsive to medications. Deep brain stimulation (DBS) is a promising approach when pharmacological treatment fails or neurosurgery is not recommended. Acute brain slices coupled to microelectrode arrays (MEAs) represent a valuable tool to study neuronal network interactions and their modulation by electrical stimulation. As compared to conventional extracellular recording techniques, they provide the added advantages of a greater number of observation points and a known inter-electrode distance, which allow studying the propagation path and speed of electrophysiological signals. However, tissue oxygenation may be greatly impaired during MEA recording, requiring a high perfusion rate, which comes at the cost of decreased signal-to-noise ratio and higher oscillations in the experimental temperature. Electrical stimulation further stresses the brain tissue, making it difficult to pursue prolonged recording/stimulation epochs. Moreover, electrical modulation of brain slice activity needs to target specific structures/pathways within the brain slice, requiring that electrode mapping be easily and quickly performed live during the experiment. Here, we illustrate how to perform the recording and electrical modulation of 4-aminopyridine (4AP)-induced epileptiform activity in rodent brain slices using planar MEAs. We show that the brain tissue obtained from mice outperforms rat brain tissue and is thus better suited for MEA experiments. This protocol guarantees the generation and maintenance of a stable epileptiform pattern that faithfully reproduces the electrophysiological features observed with conventional field potential recording, persists for several hours, and outlasts sustained electrical stimulation for prolonged epochs. Tissue viability throughout the experiment is achieved thanks to the use of a small-volume custom recording chamber allowing for laminar flow and quick solution exchange even at low (1 mL/min) perfusion rates. Quick MEA mapping for real-time monitoring and selection of stimulating electrodes is performed by a custom graphic user interface (GUI).

We illustrate how to perform recording and electrical modulation of 4-aminopyridine-induced epileptiform activity in rodent brain slices using microelectrode arrays. A custom recording chamber maintains tissue viability throughout prolonged experimental sessions. Live electrode mapping and selection of stimulating pairs are performed by a custom graphical user interface.

鼠脑切片偶联微电极阵列癫痫活性的记录与调控

Temporal lobe epilepsy (TLE) is the most common partial complex epileptic syndrome and the least responsive to medications. Deep brain stimulation (DBS) ...

Surgical Training for the Implantation of Neocortical Microelectrode Arrays Using a Formaldehyde-fixed Human Cadaver Model

This protocol describes a procedure to assist surgeons in training for the implantation of microelectrode arrays into the neocortex of the human brain. Recent technological progress has enabled the fabrication of microelectrode arrays that allow recording the activity of multiple individual neurons in the neocortex of the human brain. These arrays have the potential to bring unique insight onto the neuronal correlates of cerebral function in health and disease. Furthermore, the identification and decoding of volitional neuronal activity opens the possibility to establish brain-computer interfaces, and thus might help restore lost neurological functions. The implantation of neocortical microelectrode arrays is an invasive procedure requiring a supra-centimetric craniotomy and the exposure of the cortical surface; thus, the procedure must be performed by an adequately trained neurosurgeon. In order to provide an opportunity for surgical training, we designed a procedure based on a human cadaver model. The use of a formaldehyde-fixed human cadaver bypasses the practical, ethical and financial difficulties of surgical practice on animals (especially non-human primates) while preserving the macroscopic structure of the head, skull, meninges and cerebral surface and allowing realistic, operating room-like positioning and instrumentation. Furthermore, the use of a human cadaver is closer to clinical daily practice than any non-human model. The major drawbacks of the cadaveric simulation are the absence of cerebral pulsation and of blood and cerebrospinal fluid circulation. We suggest that a formaldehyde-fixed human cadaver model is an adequate, practical and cost-effective approach to ensure proper surgical training before implanting microelectrode arrays in the living human neocortex.

We designed a procedure in which a formaldehyde-fixed human cadaver is used to assist neurosurgeons in training for the implantation of microelectrode arrays into the neocortex of the human brain.

应用甲醛固定人尸体模型植入皮层微电极阵列的外科训练

This protocol describes a procedure to assist surgeons in training for the implantation of microelectrode arrays into the neocortex of the human brain. ...

医学

Chronic Implantation of Whole-cortical Electrocorticographic Array in the Common Marmoset

Electrocorticography (ECoG) allows the monitoring of electrical field potentials from the cerebral cortex with high spatiotemporal resolution. Recent development of thin, flexible ECoG electrodes has enabled conduction of stable recordings of large-scale cortical activity. We have developed a whole-cortical ECoG array for the common marmoset. The array continuously covers almost the entire lateral surface of cortical hemisphere, from the occipital pole to the temporal and frontal poles, and it captures whole-cortical neural activity in one shot. This protocol describes a chronic implantation procedure of the array in the epidural space of the marmoset brain. Marmosets have two advantages regarding ECoG recordings, one being the homologous organization of anatomical structures in humans and macaques, including frontal, parietal, and temporal complexes. The other advantage is that the marmoset brain is lissencephalic and contains a large number of complexes, which are more difficult to access in macaques with ECoG, that are exposed to the brain surface.These features allow direct access to most cortical areas beneath the surface of the brain. This system provides an opportunity to investigate global cortical information processing with high resolutions at a sub-millisecond order in time and millimeter order in space.

We have developed a whole-cortical electrocorticographic array for the common marmoset that continuously covers almost the entire lateral surface of cortex, from the occipital pole to the temporal and frontal poles. This protocol describes a chronic implantation procedure of the array in the epidural space of the marmoset brain.

常见马莫塞的全皮质电皮质阵列的慢性植入

Electrocorticography (ECoG) allows the monitoring of electrical field potentials from the cerebral cortex with high spatiotemporal resolution. Recent ...

Subdural Soft Electrocorticography (ECoG) Array Implantation and Long-Term Cortical Recording in Minipigs

Neurological impairments and diseases can be diagnosed or treated using electrocorticography (ECoG) arrays. In drug-resistant epilepsy, these help delineate the epileptic region to resect. In long-term applications such as brain-computer interfaces, these epicortical electrodes are used to record the movement intention of the brain, to control the robotic limbs of paralyzed patients. However, current stiff electrode grids do not answer the need for high-resolution brain recordings and long-term biointegration. Recently, conformable electrode arrays have been proposed to achieve long-term implant stability with high performance. However, preclinical studies for these new implant technologies are needed to validate their long-term functionality and safety profile for their translation to human patients. In this context, porcine models are routinely employed in developing medical devices due to their large organ sizes and easy animal handling. However, only a few brain applications are described in the literature, mostly due to surgery limitations and integration of the implant system on a living animal.
Here, we report the method for long-term implantation (6 months) and evaluation of soft ECoG arrays in the minipig model. The study first presents the implant system, consisting of a soft microfabricated electrode array integrated with a magnetic resonance imaging (MRI)-compatible polymeric transdermal port that houses instrumentation connectors for electrophysiology recordings. Then, the study describes the surgical procedure, from subdural implantation to animal recovery. We focus on the auditory cortex as an example target area where evoked potentials are induced by acoustic stimulation. We finally describe a data acquisition sequence that includes MRI of the whole brain, implant electrochemical characterization, intraoperative and freely moving electrophysiology, and immunohistochemistry staining of the extracted brains.
This model can be used to investigate the safety and function of novel design of cortical prostheses; mandatory preclinical study to envision translation to human patients.

Subdural Soft Electrocorticography ECoG Array Implantation and Long-Term Cortical Recording in Minipigs

Here, we present a method for the long-term performance and safety assessment of soft subdural electrode arrays in a minipig model, describing surgical method and tools, postoperative magnetic resonance imaging, electrophysiology of the auditory cortex, electrochemical properties of the implant, and postmortem immunochemistry.

硬膜下软皮质电图 （ECoG） 阵列植入和小型猪的长期皮质记录

Neurological impairments and diseases can be diagnosed or treated using electrocorticography (ECoG) arrays. In drug-resistant epilepsy, these help ...

<meta charset="utf8"></head><body>记录大脑组件中的网络活动：多电极阵列方法

A Multi-Electrode Array Platform for Modeling Epilepsy Using Human Pluripotent Stem Cell-Derived Brain Assembloids

Human brain organoids are three-dimensional (3D) structures derived from human pluripotent stem cells (hPSCs) that recapitulate&#160;aspects of fetal brain development. The fusion of dorsal with ventral regionally specified brain organoids in vitro generates assembloids, which have functionally integrated microcircuits with excitatory and inhibitory neurons. Due to their structural complexity and diverse population of neurons, assembloids have become a useful in vitro tool for studying aberrant network activity. Multi-electrode array (MEA) recordings serve as a method for capturing electrical field potentials, spikes, and longitudinal network dynamics from a population of neurons without compromising cell membrane integrity. However, adhering assembloids onto the electrodes for long-term recordings can be challenging due to their large size and limited contact surface area with the electrodes. Here, we demonstrate a method to plate assembloids onto MEA plates for recording electrophysiological activity over a 2-month span. Although the current protocol utilizes human cortical organoids, it can be broadly adapted to organoids differentiated to model other brain regions. This protocol establishes a robust, longitudinal, electrophysiological assay for studying the development of a neuronal network, and this platform has the potential to be used in drug screening for therapeutic development in epilepsy.

<meta charset="utf8"></head><body>作者聚焦：通过基于多电极阵列的人脑组件长期电生理监测推进遗传性癫痫研究

This protocol aims to stably plate dorsal-ventral fused assembloids on multi-electrode arrays for modeling epilepsy in vitro.

使用人类多能干细胞来源的脑组件对癫痫进行建模的多电极阵列平台

Human brain organoids are three-dimensional (3D) structures derived from human pluripotent stem cells (hPSCs) that recapitulate&#160;aspects of fetal ...

人类癫痫术后皮层组织的多电极阵列录音

摘要

探索更多视频

此视频中的章节

神经科学的研究和实时功能皮层映射记录人力Electrocorticographic（ECOG）信号

人类癫痫患者同步眼动追踪和单神经记录

调查深皮层的功能和皮层下结构采用立体定向脑电图:由前扣带皮层的经验教训

用于Vivo神经记录的 Ti₃C₂ MXene 微电极阵列的制造

自由运动大鼠皮质局部电位、心电图、肌电图和呼吸节律的同步记录

鼠脑切片偶联微电极阵列癫痫活性的记录与调控

应用甲醛固定人尸体模型植入皮层微电极阵列的外科训练

常见马莫塞的全皮质电皮质阵列的慢性植入

硬膜下软皮质电图（ECoG）阵列植入和小型猪的长期皮质记录

使用人类多能干细胞来源的脑组件对癫痫进行建模的多电极阵列平台

人类癫痫术后皮层组织的多电极阵列录音

摘要

探索更多视频

此视频中的章节

神经科学的研究和实时功能皮层映射记录人力Electrocorticographic（ECOG）信号

人类癫痫患者同步眼动追踪和单神经记录

调查深皮层的功​​能和皮层下结构采用立体定向脑电图:由前扣带皮层的经验教训

用于Vivo神经记录的 Ti3C2 MXene 微电极阵列的制造

自由运动大鼠皮质局部电位、心电图、肌电图和呼吸节律的同步记录

鼠脑切片偶联微电极阵列癫痫活性的记录与调控

应用甲醛固定人尸体模型植入皮层微电极阵列的外科训练

常见马莫塞的全皮质电皮质阵列的慢性植入

硬膜下软皮质电图 （ECoG） 阵列植入和小型猪的长期皮质记录

使用人类多能干细胞来源的脑组件对癫痫进行建模的多电极阵列平台

调查深皮层的功能和皮层下结构采用立体定向脑电图:由前扣带皮层的经验教训

用于Vivo神经记录的 Ti₃C₂ MXene 微电极阵列的制造

硬膜下软皮质电图（ECoG）阵列植入和小型猪的长期皮质记录