We thank the thousands of brain donors, patients, families, and neuroscientists around the world who, during the last two centuries and through their generous gifts and intellectual efforts, helped to discover how the human brain works, to understand devastating brain diseases, and to develop treatments thereof. We particularly thank Mrs. Cecilia V. Feltis for editing and reviewing this manuscript.

涉及死后人体组织程序已被机构审查委员会审查，并在45 CFR（联邦法规）豁免。 注:本协议描述了完成在人类神经病理学研究死后的大脑评估对称bihemispheric脑部切割程序。的装置，仪器，材料和必要执行人类大脑切割建筑材料的详细描述将被排除。对于脑解剖材料和用品在单研究者决定被选择并且是基于允许或在各研究机构批准尸检工具。集此过程所需的工具和材料最小的材料/设备表中描述。具体切割程序和疑似传染性脑疾病，如人类克雅氏病的预防措施，是这个手稿的目标之外，并从其他来源获得7。 1.对称双大脑半球切割注意:确保大脑已经接收了必要的组织固定（使用，例如，中性缓冲的10％福尔马林），为期两到三周的，这取决于periagonal，代谢（ 即，pH值），和组织保存（ 即，温度）的条件。然而，对于成像病理学相关研究，固定的周期较长（&gt;5.4周）已建议8。 <ol><li>放置在面向研究者平坦表面脑，与朝向相反的方向相对于所述研究者的正面极。 </li><li>将这样的方式大脑以允许所有皮质脑回和整个大脑（ 图1a）的沟的完整和清晰的可视化。 </li><li>先来说一下脑膜异常，宏观半球不对称性（焦点，叶，或萎缩的广义半球现象可能的指标），宏观tissutal病变（ 即吐沫RS或症），先天性畸形，脉管异常，以及任何其他可能的异常或脑表面的不寻常的演示。 注:有关如何评估一个人的大脑的详细说明，请参阅市售的神经病理学教科书和尸检手册9,10。 </li></ol> 2.协议序列<ol><li>面对正面极点从调查路程，半球（端脑）所面临的研究者肤浅的方面。多少就拿多少数码照片根据需要在每一个特定的情况下记录可能的宏观异常现象，并考虑到可能的临床 - 神经解剖和切割后的注意事项。有一个研究助理拿数码照片垂直于大脑捕捉整个皮质表面（ 图1a - C）。 </li><li>马克前和使用的墨水或彩色针中央后皮质脑回切割前脑（ 图1b </s仲&gt;）。 注:此过程有利于电机和切割后体感皮层主要的更直接的认识。 </li><li>旋转180度的脑，同时保持其朝向相同的方向（即从研究者背向正面极）。仔细检查大脑底部。要特别注意脑血管系统的条件（ 即，基底动脉和椎动脉和Willis环）在其脑干出口/入口水平和颅神经。管理嗅球和大片特别小心，以避免tissutal裂伤，由于他们的极端脆弱性。 <ol><li>多少就拿多少数码照片根据需要在每一个特定的情况下记录可能的宏观异常现象，并考虑到可能的临床 - 神经解剖和切割后的注意事项。有一个研究助理拿数码照片垂直于大脑捕捉到皮层和脑干表面的全部。 </l我&gt; </ol></li><li>朝向脑的碱和使用手术刀，剪在脑桥的上部的水平脑干横向（尽可能接近到大脑的碱）。仔细检查SN（ 即 ，为苍白）11和其他邻近结构12。需注意，可能相比于正常脑13用脑的任何不寻常的外观的音频记录装置中，。 </li><li>再次旋转180度脑并用一把锋利的刀，通过集中切断胼胝体通过内侧纵裂缝，并遵循额枕部方向的两个半球分开。检查每个半球为可能的异常（ 例如，心室扩印，畸形，组织软化，肿瘤等 ）13的每一侧。参见图2a <ol><li>多少就拿多少数码照片根据需要在每一个特定的情况下记录可能的宏观异常和考虑到可能的临床 - 神经解剖和切割后的注意事项。有一个研究助理拿数码照片垂直于大脑捕捉整个皮质表面。注意到脑的任何不寻常的特点的相比于正常脑。 </li></ol></li><li>放置两个半球扁平，躺在其内侧方面，朝向从研究者远额叶， 如图2b所示 。将它们放置在这样一种方式，其中心触摸（也标记为半球不对称的情况下）。 </li><li>用一把锋利的刀，手动通过两个大脑半球切开，开始在额极，并通过半球的整个长度走向两极枕部移动。获得两个系列的脑组织1厘米厚的块（约18板为每个半球）。 </li><li>放置脑板坯在一个单独的平面上的解剖学组织（额枕骨方向）序列。使用白色surfa与拍摄时印有更好的对比度尺子CE。显示两个系列脑板坯在解剖学对称方式（额枕方向），确保它们的冠状表面是直接目视和数字摄影（ 图3a）中可见。使用两侧印刷毫米波网格切割表面来定位大脑结构，尺寸和可能的异常以更精确的方式。 <ol><li>多少就拿多少数码照片根据需要在每一个特定的情况下记录可能的宏观异常现象，并考虑到可能的临床 - 神经解剖和切割后的注意事项。有一个研究助理拿数码照片垂直于大脑捕捉整个皮质表面。记笔记相比于正常脑（可能使用音频记录器设备），大脑中的任何不寻常的方面。 </li></ol></li><li>用锋利的手术刀，解剖手动较小的矩形块脑组织中的每个建立脑区。按照表1所述建议的脑区域收集方案。 <ol><li>放入单独标示histocassettes每个组织块。 注:脑组织中的每块需要被切断，以适应，尽可能，标准histocassette最大体积（30×20×4mm的3）。 </li><li>标签使用每一种情况下的去识别代码和使用特定的神经解剖学标识符histocassettes（不使用随机字母或数字为不同的大脑;相反，始终使用相同的区域的解剖学的名称或相应的数字，如表1所示）。创建去识别码，例如，通过产生随机或半随机数为每一种情况下（ 即 ，BRC 130，其中B停留脑，R停留资源，C停留中心和130是一个渐进的加入或AD160001，其中，AD代表"阿尔茨海默氏症研究"，16是年时进行尸体解剖（2016），和0001逐行加入检体数）。 注意:这一步是为未来的研究非常有帮助的;保持一个传奇，并指定半球（L =左脑，R =右半球）。使用histocassettes两种不同颜色，建立为每个半球特定的颜色。 </li><li>多少就拿多少数码照片根据需要在每一个特定的情况下记录可能macroanomalies，并考虑到可能的临床 - 神经解剖和切割后的注意事项。有一个研究助理拿数码照片垂直于大脑捕捉整个皮质表面。注意到脑的任何不寻常的特点的相比于正常脑。 </li></ol></li><li>拍摄数字图片（多达必要或期望的）整个切脑和相关histocassettes的。 </li><li>冲床（ 例如，通过的Accu冲）小块组织的DNA提取和遗传分析。使用直径5毫米 - 2一记重拳。 注:对于其高的基因组材料的含量，小脑是首选;然而，任何其他区域是好的。 </li><li>再浸入含有相同类型的先前使用，直到组织处理的下一步骤固定液（ 例如，10％的中性缓冲的福尔马林）的脑组织块所有histocassettes。 </li><li>按照人类福尔马林固定，组织处理14的标准程序。 </li></ol> 3.特殊的方法:交替冷冻和固定对称Bihemispheric切割注:在第2描述的对称bihemispheric脑切削协议提供了从未定影，新鲜的大脑切割组织板坯在相同的系统和对称的方式的可能性（当可用时）。 <ol><li>将整个大脑清新倒挂（最好在半球碗状的塑料表面）8 - 10分钟，在-80℃冷冻硬化的脑组织withouŧ挑起生化损伤，以方便手动切割。 </li><li>用一把锋利的刀，切两半球在交替和连续的方式，按照第2节中描述的大脑切割协议，但冻结和修复每隔板（由两个半球和沿额枕部解剖方向）。 <ol><li>此时，不要试图削减各脑区域，如表1所述。切新鲜的特定脑区仅在必要时立即RNA或蛋白质的提取（ 即对基因组或蛋白质组学研究）15。 </li></ol></li><li>切割后，立即冻结，标签和数量每个新鲜组织。占据整个平板系列数码照片;装在一个塑料袋每个切片;收集在一个单独的，一个脑只容器中的板;和容器存放在专用-80℃冰箱中。 注:冰箱应致力于只有人脑组织。只是后来会唱根据需要为每个特定实验勒冷冻大脑区域进行切割。 </li><li>沉浸选择用于在含有固定剂的足够量（3/1体积固定剂/组织块的比例）分离袋固定（10％的中性缓冲的福尔马林或其他固定剂）每隔其它组织板坯。通过连续编号他们跟着一个额枕部序列标签每袋。密封每个袋，拍摄数字图片，并将它们存储在塑料容器中。 </li><li> 2周组织固定后打开含有固定剂袋和如表1中所述切割每个脑区。 </li></ol> 4. Histostain及免疫组化注:本套脑区的切基础上，提出方案（ 表1）足以满足大多数，如果不是全部，当前已建立共识为基础的病理标准AD 16，PD 17，路易体痴呆（DLB）18，额颞叶痴呆（FTD / MND） <suP&gt; 19，逐行Suprabulbar麻痹（PSP）20，多系统萎缩（MSA）21，慢性创伤性脑病（CTE）22， 等等 。 <ol><li>对于每一个脑区和两个半球，请执行下列最低限度histostains的:苏木和曙红（H＆E），甲酚紫（CV;如果形态计量学研究计划，例如），银染色（如"探索性"分析是需要）。 </li><li>对于每一个脑区和两个半球，请执行下列最低限度的免疫组化协议:β淀粉状蛋白（βA），磷酸化头（pTau），磷酸化α突触核蛋白（Pα-SYN）和磷酸化TDP43（pTDP43） ，如上所述14。 注:组织切片，以评估每个脑内此协议的总数为46（如果由两个半球各脑区都可用）。 </li></ol>

作者什么都没有透露。

这种脑切断方法可适于对每个神经病理学实验室的特定需求（例如，通过减少脑区域的数量，以评估每个半球），同时仍保留bihemispheric对称切割的步骤，其主要特征之一。此提出的协议可用于常规程序（研究型神经病理学中心）或只在必要时（具体临床取向研究）。它可以选择性地仅用于特定类型的调查（ 即，免疫组织化学）或分子分析（ 即 ，基因组或蛋白质组分析）。从技术角...

Neuropathologists有科学的特权，智力的荣誉和诊断义务评估人类大脑。几十年来，脑部疾病和重大努力的详细的临床描述，以个体化在人死后的大脑其可能neurohistological相关因素已经开展。从历史上看，这些努力所代表的最有成效的方式由医学科学和神经学尤其是在当今时代先进。由于以前的杰出neuropathologists和他们的奉献精神，决心，奖学金，惊人的能力，以正常和异常的脑组织（通常使用非常残留的工具）之间的区别，我们现在可以研究和目标疾病，如阿尔茨海默Perusini病（不公平只被称为阿尔茨海默氏症病; APD / AD）1，帕金森病（PD）2，克雅氏病（CJD）3，娄格里克病/肌萎缩侧索SclerosiS（ALS）4，和关岛病5，仅举几例。 神经影像学的先进的技术，如高清晰计算机断层扫描（ 即多层螺旋CT扫描，CT血管造影），功能和形态磁共振成像（ 即功能磁共振成像，弥散核磁共振成像，示踪-MRI 等 ），正电子发射断层扫描（PET），基于超声波的成像，等人，都肯定是我们修改关于如何诊断和治疗神经和精神病患者一般做法。尽管如此，虽然神经成像技术能够可视化的人的大脑时活着，他们不提供的机会，在发生的时刻，直接分析细胞，如神经元的高度复杂的细胞和亚细胞结构的;或可视化，标记和量化特定类型的细胞内病变;或精确显示他们的神经解剖学和次区域定位在circuital和分circuital解剖层次。例如，神经成像技术不能在嗅皮层识别或定位在黑质（SN），与帕金森病相关的一个共同的病理学特征，或神经原纤维缠结（NFT）的色素神经元Lewy小体（LB），AD的经典特征和其他脑病变。神经病理学的调查与先进的数码显微镜相结合仍然是不可替代的详细的临床相关性，因此，为明确诊断。 由于人脑的特有anatomo功能特性，特别是它的解剖定位（即，颅骨，天然保护系统，该系统不允许内容的直接检查内部），引入体内神经成像技术非常有帮助临床医生和调查人员发现最初的解答一些这种复杂组织的奥秘。然而，没有临床或neuroimagiNG的方法，可替换的独特的机会，脑组织尸检过程中直接分析。仅有组织收集，保存，和人脑的分类可允许神经元和非神经元细胞，亚细胞成分，细胞内和细胞外的病理病变直接与系统的调查，而大脑内的任何类型的异常进行确认，修改，或重新定义临床诊断和发现新的临床相关性。之一有关的大脑尸检评估表观限制一直是事实，这过程是一个剖方法论。总会有一个持续的神经病理过程（临床表现或不）和机会，如果有的话，在neurohistological级别来定义它之间的延迟。这主要是由于人的大脑没有能力再生本身。它目前还无法获得的脑组织在体内 ，而无需创建PErmanent伤害。因此，是不可能在纵向和神经病理学评估相同的大脑/人。然而，标准化的大脑银行程序和脑捐赠广大市民日益认识到可以极大地持续增加收集和分析案件的数量有助于脑尸检时序问题的解决。以这种方式，可以得到死后的大脑的更充分的数字，以定义病理起源和进展的恒定模式与每个人脑疾病相关的脑损伤的每个特定类型。这将需要从受任何神经精神障碍的患者在所有年龄捐献和尽可能多的大脑尽可能收集，以及来自健康对照受试者。一个可能的方法可以从一般和专门的医疗中心作为一个标准的程序中收集尽可能多的尸检大脑越好。需要对脑捐赠最近已表示那些谁研究老年痴呆症和正常老化6。相同的必要性应由神经字段作为整体来表示。 用于上述和其他原因，正在进行的脑切割程序的更新是必要的。此外，脑切割程序应普遍在不同的神经病理学的研究中心标准化在世界各地，也考虑帐户的可能性聘请当前和未来的生物技术，以更好地调查，并希望能以明确理解的原因和脑部疾病的机制人类。 在这里，主要用于研究目的，我们描述了死后的大脑在人类切对称方法。此过程提出收集比正常完成，并从两个大脑和小脑半球更多脑区。对称双大脑半球切割过程将符合我们当前的人类知识好得多神经解剖学，神经化学，神经生理学和。这种方法还允许的可能性神经病理学分析人类大脑的独特功能，诸如半球专业化和偏侧是与较高的认知和非认知功能一般或专门存在于我们的物种相关联。是否有半球专业化/偏侧和特定类型的脑病变之间的具体发病的关系，或一种特殊的神经精神病发病事件是否是最初，普遍地，或只与特定的半球关联和功能不是目前公知的。通过描述这种大脑对称切割程序，我们的目标是提出人类大脑解剖的更新方法，可以帮助更好地了解一个高度专业化的正常组织和病理条件下，大脑。该方法还考虑到，只存在于人类的那些吗啉官能半球方面。

<table><tbody><tr><td>Copy of signed informed consent allowing autopsy and brain donation for research use.</td><td></td><td></td><td></td></tr><tr><td>Detailed clinical history of the subject which should include a detailed description of any neurologic and psychiatric symptoms and signs.</td><td></td><td></td><td></td></tr><tr><td>Medical or nonmedical video-recordings when available (especially useful in movement disorders field). Next-of-kin&amp;rsquo;s consent required.</td><td></td><td></td><td></td></tr><tr><td>Neuroimaging, neurophysiology, neuropsychiatric and assessment or clinicometric scales.</td><td></td><td></td><td></td></tr><tr><td>Genetic and family history data. Genetic reports review, if neurogenetic diseases were diagnosed.</td><td></td><td></td><td></td></tr><tr><td>Histology Container</td><td>ELECTRON MICROSCOPY SCIENCES</td><td>64233-24</td><td></td></tr><tr><td>Histology Cassettes</td><td>VWR</td><td>18000-142 (orange)</td><td></td></tr><tr><td>Histology Cassettes</td><td>VWR</td><td>18000-132 (navy)</td><td></td></tr><tr><td>Knife Handles and Disposable Blades</td><td>ELECTRON MICROSCOPY SCIENCES</td><td>62560-04</td><td></td></tr><tr><td>Long Blades</td><td>ELECTRON MICROSCOPY SCIENCES</td><td>62561-20</td><td></td></tr><tr><td>Disposable Blade Knife Handles</td><td>ELECTRON MICROSCOPY SCIENCES</td><td>72040-08</td><td></td></tr><tr><td>Scalpel Blades</td><td>ELECTRON MICROSCOPY SCIENCES</td><td>72049-22</td><td></td></tr><tr><td>Accu-Punch 2 mm</td><td>ELECTRON MICROSCOPY SCIENCES</td><td>69038-02&amp;nbsp;</td><td></td></tr><tr><td>Polystyrene Containers &amp;ndash; Sterile</td><td>ELECTRON MICROSCOPY SCIENCES</td><td>64240-12</td><td></td></tr><tr><td>Dissecting Board</td><td>ELECTRON MICROSCOPY SCIENCES</td><td>63307-30</td><td></td></tr><tr><td>Formalin solution, neutral buffered, 10%</td><td>Sigma-Aldrich</td><td>HT501128 SIGMA</td><td></td></tr><tr><td>Hematoxylin Solution, Gill No. 2</td><td>Sigma-Aldrich</td><td>GHS280&amp;nbsp;SIGMA</td><td></td></tr><tr><td>Eosin Y solution, aqueous</td><td>Sigma-Aldrich</td><td>HT1102128&amp;nbsp;SIGMA</td><td></td></tr><tr><td>anti-beta-amyloid</td><td>Covance, Princeton, NJ</td><td>SIG-39220</td><td>1:500</td></tr><tr><td>anti-tau</td><td>Thermo Fisher Scientific</td><td>MN1020</td><td>1:500</td></tr><tr><td>anti-alpha-synuclein</td><td>Abcam</td><td>ab27766</td><td>1:500</td></tr><tr><td>anti-phospho-TDP43</td><td>Cosmo Bio Co.</td><td>TIP-PTD-P02</td><td>1:2000</td></tr><tr><td>Digital Camera</td><td>Any</td><td></td><td></td></tr><tr><td>Head Impulse Sealing machine&amp;nbsp;</td><td>Grainger</td><td>5ZZ35</td><td></td></tr></tbody></table>

symmetric bihemispheric postmortem brain cutting to study healthy and pathological brain conditions in humans

Neuropathologists，有时感到生成在人脑尸检要求的患者描述复杂的神经精神现象，明确的诊断所需要的知识量吓倒。尽管生物医学影像学和的进步已经彻底改变了神经精神领域，他们也产生了误导性的想法，大脑解剖只有一个确定的值。这个假想法在尸检速率的急剧减少，因此，降低的可能性进行更详细的和广泛的神经病理调查，这是必要的，以理解众多正常和病理方面未知的人脑。观察神经精神现象及其可能的相关因素neurohistological相应的本地化/特性之间的相关传统推理方法继续有一个不可否认的价值。在neuropsychi的上下文atric疾病，传统的临床方法仍是最好的可能的方法（通常是唯一可用的），以唯一的神经精神功能链接到其相应的神经病理学基材，因为它在脑组织中的直接身体评估特别依赖。死后大脑的评估是基于大脑切割跨越不同的神经病理学变化中心的程序。脑屑基于存在于每个机构的各种临床和学术突发相对广泛和系统的方式进行。更解剖学包容性和对称双向大脑半球切割的方法至少应该用于研究目的的人类神经病理学连贯调查，深入，与人脑（ 即半球专业化和偏侧特定的特殊性正常和病理条件功能）。这种方法将提供更全面的科尔神经病理良好的特点适用于当前和未来的生物科技和神经成像技术的大脑ction。我们描述了在人类大脑半球病变的差异进行调查并与目前使用的以及未来的生物分子/神经影像学技术的对称双大脑半球切割程序。

协议长度花为单个对称bihemispheric固定脑切削过程中的时间估计为1小时（不包括花费建立解剖台，工具和切割表面的时间;标签; 等 ）。为一个单一的对称双半球交替冻结和固定脑切割过程所需的时间估计需要2小时。它可以至少在4 - 6周获得明确的组织学诊断为单人脑/主题。后从头骨，组织固定的适当时期死后的大脑切除...

Watch this Scientific Journal Video about Symmetric Bihemispheric Postmortem Brain Cutting to Study Healthy and Pathological Brain Conditions in Humans at JoVE.com

Symmetric Bihemispheric Postmortem Brain Cutting to Study Healthy and Pathological Brain Conditions in Humans

有组织的脑切削程序是必要的，以与确定的神经病理学诊断特定神经精神现象相关。脑屑基于各种临床 - 学术突发不同的方式执行。本协议描述的对称bihemispheric脑部切割程序，调查人的大脑半球病变差异，以最大限度地提高当前和未来的生物分子/神经影像学技术。

对称Bihemispheric死后的大脑切割学习健康和病理条件大脑在人

Neuropathologists, at times, feel intimidated by the amount of knowledge needed to generate definitive diagnoses for complex neuropsychiatric phenomena ...

symmetric-bihemispheric-postmortem-brain-cutting-to-study-healthy

Research

JoVE Journal

Medicine

13.9K 次观看.  Biomedical Research Institute of New Jersey (BRInj). 有组织的脑切削程序是必要的，以与确定的神经病理学诊断特定神经精神现象相关。脑屑基于各种临床 - 学术突发不同的方式执行。本协议描述的对称bihemispheric脑部切割程序，调查人的大脑半球病变差异，以最大限度地提高当前和未来的生物分子/神经影像学技术。 

视频: Symmetric Bihemispheric Postmortem Brain Cutting to Study Healthy and Pathological Brain Conditions in Humans

Sex Stratified Neuronal Cultures to Study Ischemic Cell Death Pathways

Sex differences in neuronal susceptibility to ischemic injury and neurodegenerative disease have long been observed, but the signaling mechanisms responsible for those differences remain unclear. Primary disassociated embryonic neuronal culture provides a simplified experimental model with which to investigate the neuronal cell signaling involved in cell death as a result of ischemia or disease; however, most neuronal cultures used in research today are mixed sex. Researchers can and do test the effects of sex steroid treatment in mixed sex neuronal cultures in models of neuronal injury and disease, but accumulating evidence suggests that the female brain responds to androgens, estrogens, and progesterone differently than the male brain. Furthermore, neonate male and female rodents respond differently to ischemic injury, with males experiencing greater injury following cerebral ischemia than females. Thus, mixed sex neuronal cultures might obscure and confound the experimental results; important information might be missed. For this reason, the Herson Lab at the University of Colorado School of Medicine routinely prepares sex-stratified primary disassociated embryonic neuronal cultures from both hippocampus and cortex. Embryos are sexed before harvesting of brain tissue and male and female tissue are disassociated separately, plated separately, and maintained separately. Using this method, the Herson Lab has demonstrated a male-specific role for the ion channel TRPM2 in ischemic cell death. In this manuscript, we share and discuss our protocol for sexing embryonic mice and preparing sex-stratified hippocampal primary disassociated neuron cultures. This method can be adapted to prepare sex-stratified cortical cultures and the method for embryo sexing can be used in conjunction with other protocols for any study in which sex is thought to be an important determinant of outcome.

Primary disassociated embryonic hippocampal neuronal cultures are useful for investigating the signaling mechanisms involved in neuron death. Sexing the embryos before the isolation and dissociation of the hippocampus allows the preparation of separate male and female cultures, which enables the researcher to identify and investigate sex-specific cell signaling.

Sex differences in neuronal susceptibility to ischemic injury and neurodegenerative disease have long been observed, but the signaling mechanisms ...

科研

神经科学

3D Modeling of the Lateral Ventricles and Histological Characterization of Periventricular Tissue in Humans and Mouse

The ventricular system carries and circulates cerebral spinal fluid (CSF) and facilitates clearance of solutes and toxins from the brain. The functional units of the ventricles are ciliated epithelial cells termed ependymal cells, which line the ventricles and through ciliary action are capable of generating laminar flow of CSF at the ventricle surface. This monolayer of ependymal cells also provides barrier and filtration functions that promote exchange between brain interstitial fluids (ISF) and circulating CSF. Biochemical changes in the brain are thereby reflected in the composition of the CSF and destruction of the ependyma can disrupt the delicate balance of CSF and ISF exchange. In humans there is a strong correlation between lateral ventricle expansion and aging. Age-associated ventriculomegaly can occur even in the absence of dementia or obstruction of CSF flow. The exact cause and progression of ventriculomegaly is often unknown; however, enlarged ventricles can show regional and, often, extensive loss of ependymal cell coverage with ventricle surface astrogliosis and associated periventricular edema replacing the functional ependymal cell monolayer. Using MRI scans together with postmortem human brain tissue, we describe how to prepare, image and compile 3D renderings of lateral ventricle volumes, calculate lateral ventricle volumes, and characterize periventricular tissue through immunohistochemical analysis of en face lateral ventricle wall tissue preparations. Corresponding analyses of mouse brain tissue are also presented supporting the use of mouse models as a means to evaluate changes to the lateral ventricles and periventricular tissue found in human aging and disease. Together, these protocols allow investigations into the cause and effect of ventriculomegaly and highlight techniques to study ventricular system health and its important barrier and filtration functions within the brain.

Using MRI scans (human), 3D imaging software, and immunohistological analysis, we document changes to the brain&#8217;s lateral ventricles. Longitudinal 3D mapping of lateral ventricle volume changes and characterization of periventricular cellular changes that occur in the human brain due to aging or disease are then modeled in mice.

脑室周围组织的侧脑室及组织学特性对人类和鼠标3D建模

The ventricular system carries and circulates cerebral spinal fluid (CSF) and facilitates clearance of solutes and toxins from the brain. The functional ...

High-resolution Structural Magnetic Resonance Imaging of the Human Subcortex In Vivo and Postmortem

The focus of this study was to test the resolution limits of structural MRI of a postmortem brain compared to living human brains. The resolution of structural MRI in vivo is ultimately limited by physiological noise, including pulsation, respiration and head movement. Although imaging hardware continues to improve, it is still difficult to resolve structures on the millimeter scale. For example, the primary visual sensory pathways synapse at the lateral geniculate nucleus (LGN), a visual relay and control nucleus in the thalamus that normally is organized into six interleaved monocular layers. Neuroimaging studies have not been able to reliably distinguish these layers due their small size that are less than 1 mm thick.
The resolving limit of structural MRI, in a postmortem brain was tested using multiple images averaged over a long duration (~24 h). The purpose was to test whether it was possible to resolve the individual layers of the LGN in the absence of physiological noise. A proton density (PD)1 weighted pulse sequence was used with varying resolution and other parameters to determine the minimum number of images necessary to be registered and averaged to reliably distinguish the LGN and other subcortical regions. The results were also compared to images acquired in living human brains. In vivo subjects were scanned in order to determine the additional effects of physiological noise on the minimum number of PD scans needed to differentiate subcortical structures, useful in clinical applications.

High-resolution Structural Magnetic Resonance Imaging of the Human Subcortex In Vivo and Postmortem

Here we present a protocol to determine the minimum number images that needed to be registered and averaged to resolve subcortical structures and test whether the individual layers of the LGN could be resolved in the absence of physiological noise.

高分辨率结构磁共振人类皮质下的成像<I&gt;在体内</I&gt;与死亡

The focus of this study was to test the resolution limits of structural MRI of a postmortem brain compared to living human brains. The resolution of ...

Utilizing 3D Printing Technology to Merge MRI with Histology: A Protocol for Brain Sectioning

Magnetic resonance imaging (MRI) allows for the delineation between normal and abnormal tissue on a macroscopic scale, sampling an entire tissue volume three-dimensionally. While MRI is an extremely sensitive tool for detecting tissue abnormalities, association of signal changes with an underlying pathological process is usually not straightforward. In the central nervous system, for example, inflammation, demyelination, axonal damage, gliosis, and neuronal death may all induce similar findings on MRI. As such, interpretation of MRI scans depends on the context, and radiological-histopathological correlation is therefore of the utmost importance. Unfortunately, traditional pathological sectioning of brain tissue is often imprecise and inconsistent, thus complicating the comparison between histology sections and MRI. This article presents novel methodology for accurately sectioning primate brain tissues and thus allowing precise matching between histology and MRI. The detailed protocol described in this article will assist investigators in applying this method, which relies on the creation of 3D printed brain slicers. Slightly modified, it can be easily implemented for brains of other species, including humans.

The overall goal of this protocol is to accurately align magnetic resonance imaging (MRI) image volumes with histology sections via the creation of customized 3D-printed brain holders and slicer boxes.

利用3D打印技术来合并MRI与组织学:一个大脑切片协议

Magnetic resonance imaging (MRI) allows for the delineation between normal and abnormal tissue on a macroscopic scale, sampling an entire tissue volume ...

Stereotactic Atlas-Guided Laser Capture Microdissection of Brain Regions Affected by Traumatic Injury

The ability to isolate specific brain regions of interest can be impeded in tissue disassociation techniques that do not preserve their spatial distribution. Such techniques also potentially skew gene expression analysis because the process itself can alter expression patterns in individual cells. Here we describe a laser capture microdissection (LCM) method to selectively collect specific brain regions affected by traumatic brain injury (TBI) by using a modified Nissl (cresyl violet) staining protocol and the guidance of a rat brain atlas. LCM provides access to brain regions in their native positions and the ability to use anatomical landmarks for identification of each specific region. To this end, LCM has been used previously to examine brain region specific gene expression in TBI. This protocol allows examination of TBI-induced alterations in gene and microRNA expression in distinct brain areas within the same animal. The principles of this protocol can be amended and applied to a wide range of studies examining genomic expression in other disease and/or animal models.

We describe the use of laser capture microdissection to obtain samples of distinct cell populations from different brain regions for gene and microRNA analysis. This technique allows the study of differential effects of traumatic brain injury in specific regions of the rat brain.

立体定向激光捕获创伤性脑区显微解剖

The ability to isolate specific brain regions of interest can be impeded in tissue disassociation techniques that do not preserve their spatial ...

Exploring Deep Space - Uncovering the Anatomy of Periventricular Structures to Reveal the Lateral Ventricles of the Human Brain

Anatomy students are typically provided with two-dimensional (2D) sections and images when studying cerebral ventricular anatomy and students find this challenging. Because the ventricles are negative spaces located deep within the brain, the only way to understand their anatomy is by appreciating their boundaries formed by related structures. Looking at a 2D representation of these spaces, in any of the cardinal planes, will not enable visualisation of all of the structures that form the boundaries of the ventricles. Thus, using 2D sections alone requires students to compute their own mental image of the 3D ventricular spaces. The aim of this study was to develop a reproducible method for dissecting the human brain to create an educational resource to enhance student understanding of the intricate relationships between the ventricles and periventricular structures. To achieve this, we created a video resource that features a step-by-step guide using a fiber dissection method to reveal the lateral and third ventricles together with the closely related limbic system and basal ganglia structures. One of the advantages of this method is that it enables delineation of the white matter tracts that are difficult to distinguish using other dissection techniques. This video is accompanied by a written protocol that provides a systematic description of the process to aid in the reproduction of the brain dissection. This package offers a valuable anatomy teaching resource for educators and students alike. By following these instructions educators can create teaching resources and students can be guided to produce their own brain dissection as a hands-on practical activity. We recommend that this video guide be incorporated into neuroanatomy teaching to enhance student understanding of the morphology and clinical relevance of the ventricles.

This paper demonstrates the effective use of a fiber dissection method to reveal the superficial white matter tracts and periventricular structures of the human brain, in three-dimensional space, to aid student comprehension of ventricular morphology.

探索深空-揭开人脑侧脑室的脑室结构解剖

Anatomy students are typically provided with two-dimensional (2D) sections and images when studying cerebral ventricular anatomy and students find this ...

Visualization of Cortical Modules in Flattened Mammalian Cortices

The cortex of mammalian brains is parcellated into distinct substructures or modules. Cortical modules typically lie parallel to the cortical sheet, and can be delineated by certain histochemical and immunohistochemical methods. In this study, we highlight a method to isolate the cortex from mammalian brains and flatten them to obtain sections parallel to the cortical sheet. We further highlight selected histochemical and immunohistochemical methods to process these flattened tangential sections to visualize cortical modules. In the somatosensory cortex of various mammals, we perform cytochrome oxidase histochemistry to reveal body maps or cortical modules representing different parts of the body of the animal. In the medial entorhinal cortex, an area where grid cells are generated, we utilize immunohistochemical methods to highlight modules of genetically determined neurons which are arranged in a grid-pattern in the cortical sheet across several species. Overall, we provide a framework to isolate and prepare layer-wise flattened cortical sections, and visualize cortical modules using histochemical and immunohistochemical methods in a wide variety of mammalian brains.

This article describes a detailed methodology to obtain flattened tangential sections from mammalian cortices and visualize cortical modules using histochemical and immunohistochemical methods.

扁平哺乳动物皮层皮质模块的可视化研究

The cortex of mammalian brains is parcellated into distinct substructures or modules. Cortical modules typically lie parallel to the cortical sheet, and ...

Comprehensive Autopsy Program for Individuals with Multiple Sclerosis

We describe a rapid tissue donation program for individuals with multiple sclerosis (MS) that requires scientists and technicians to be on-call 24/7, 365 days a year. Participants consent to donate their brain and spinal cord. Most patients were followed by neurologists at the Cleveland Clinic Mellen Center for MS Treatment and Research. Their clinical courses and neurological disabilities are well-characterized. Soon after death, the body is transported to the MS Imaging Center, where the brain is scanned in situ by 3 T magnetic resonance imaging (MRI). The body is then transferred to the autopsy room, where the brain and spinal cord are removed. The brain is divided into two hemispheres. One hemisphere is immediately placed in a slicing box and alternate 1 cm-thick slices are either fixed in 4% paraformaldehyde for two days or rapidly frozen in dry ice and 2-methylbutane. The short-fixed brain slices are stored in a cryopreservation solution and used for histological analyses and immunocytochemical detection of sensitive antigens. Frozen slices are stored at -80 &#176;C and used for molecular, immunocytochemical, and in situ hybridization/RNA scope studies. The other hemisphere is placed in 4% paraformaldehyde for several months, placed in the slicing box, re-scanned in the 3 T magnetic resonance (MR) scanner and sliced into centimeter-thick slices. Postmortem in situ MR images (MRIs) are co-registered with 1 cm-thick brain slices to facilitate MRI-pathology correlations. All brain slices are photographed and brain white-matter lesions are identified. The spinal cord is cut into 2 cm segments. Alternate segments are fixed in 4% paraformaldehyde or rapidly frozen. The rapid procurement of postmortem MS tissues allows pathological and molecular analyses of MS brains and spinal cords and pathological correlations of brain MRI abnormalities. The quality of these rapidly-processed postmortem tissues (usually within 6 h of death) is of great value to MS research and has resulted in many high-impact discoveries.

Multiple sclerosis is an inflammatory demyelinating disease with no cure. Analysis of brain tissue provides important clues to understanding the pathogenesis of the disease. Here we discuss the methodology and downstream analysis of MS brain tissue collected through a unique rapid autopsy program in operation at the Cleveland Clinic.

多发性硬化患者综合解剖计划

We describe a rapid tissue donation program for individuals with multiple sclerosis (MS) that requires scientists and technicians to be on-call 24/7, 365 ...

Abbiategrasso Brain Bank Protocol for Collecting, Processing and Characterizing Aging Brains

In a constantly aging population, the prevalence of neurodegenerative disorders is expected to rise. Understanding disease mechanisms is the key to find preventive and curative measures. The most effective way to achieve this is through direct examination of diseased and healthy brain tissue. The authors present a protocol to obtain, process, characterize and store good quality brain tissue donated by individuals registered in an antemortem brain donation program. The donation program includes a face-to-face empathic approach to people, a collection of complementary clinical, biological, social and lifestyle information and serial multi-dimensional assessments over time to track individual trajectories of normal aging and cognitive decline. Since many neurological diseases are asymmetrical, our brain bank offers a unique protocol for slicing fresh specimens. Brain sections of both hemispheres are alternately frozen (at -80 &#176;C) or fixed in formalin; a fixed slice on one hemisphere corresponds to a frozen one on the other hemisphere. With this approach, a complete histological characterization of all frozen material can be obtained, and omics studies can be performed on histologically well-defined tissues from both hemispheres thus offering a more complete assessment of neurodegenerative disease mechanisms. Correct and definite diagnosis of these diseases can only be achieved by combining the clinical syndrome with the neuropathological evaluation, which often adds important etiological clues necessary to interpret the pathogenesis. This method can be time consuming, expensive and limited as it only covers a limited geographical area. Regardless of its limitations, the high degree of characterization it provides can be rewarding. Our ultimate goal is to establish the first Italian Brain Bank, all the while emphasizing the importance of neuropathologically verified epidemiological studies.

This protocol describes a method to track individual brain-aging trajectories through a brain donation program and proper characterization of brains. Brain donors are involved in a long-term longitudinal study including serial multi-dimensional assessments. The protocol contains a detailed description of brain processing and an accurate diagnostic methodology.

Abbiategrasso 脑库协议，用于收集、处理和描述老化的大脑

In a constantly aging population, the prevalence of neurodegenerative disorders is expected to rise. Understanding disease mechanisms is the key to find ...

Microdissection of Mouse Brain into Functionally and Anatomically Different Regions

The brain is the command center for the mammalian nervous system and an organ with enormous structural complexity. Protected within the skull, the brain consists of an outer covering of grey matter over the hemispheres known as the cerebral cortex. Underneath this layer reside many other specialized structures that are essential for multiple phenomenon important for existence. Acquiring samples of specific gross brain regions requires quick and precise dissection steps.&#160;It is understood that at the microscopic level, many sub-regions exist and likely cross the arbitrary regional boundaries that we impose for the purpose of this dissection.
Mouse models are routinely used to study human brain functions and diseases. Changes in gene expression patterns may be confined to specific brain areas targeting a particular phenotype depending on the diseased state. Thus, it is of great importance to study regulation of transcription with respect to its well-defined structural organization. A complete understanding of the brain requires studying distinct brain regions, defining connections, and identifying key differences in the activities of each of these brain regions. A more comprehensive understanding of each of these distinct regions may pave the way for new and improved treatments in the field of neuroscience. Herein, we discuss a step-by-step methodology for dissecting the mouse brain into sixteen distinct regions. In this procedure, we have focused on male mouse C57Bl/6J (6-8 week old) brain removal and dissection into multiple regions using neuroanatomical landmarks to identify and sample discrete functionally-relevant and behaviorally-relevant&#160;brain regions. This work will help lay a strong foundation in the field of neuroscience, leading to more focused approaches in the deeper understanding of brain function.

We present a hands-on, step-by-step, rapid protocol for mouse brain removal and dissection of discrete regions from fresh brain tissue. Obtaining brain regions for molecular analysis has become routine in many neuroscience labs. These brain regions are immediately frozen to obtain high quality transcriptomic data for system level analysis.

将小鼠大脑显微切割成功能和解剖学上不同的区域

The brain is the command center for the mammalian nervous system and an organ with enormous structural complexity. Protected within the skull, the brain ...