Name: endovascular perforation model for subarachnoid hemorrhage combined with magnetic resonance imaging (mri)
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SL得到了中国国家留学基金委的支持。KT得到了柏林卫生研究所和Sonnenfeld-Stiftung的BIH-MD奖学金的支持。RX由BIH-Charité临床科学家计划提供支持，该计划由Charité-Universitätsmedizin Berlin和柏林卫生研究所资助。我们感谢德国研究基金会（DFG）和柏林夏里特医学院开放获取出版基金的支持。

实验是根据德国柏林的Landesamt fuer Gesundheit und Soziales（LaGeSo）制定的指南和法规进行的（G0063/18）。在这项研究中，使用了C57Bl / 6J雄性（8-12周龄）小鼠，体重为25±0.286克（平均±s.e.m.）。
1. 动物准备
<ol><li>腹腔注射氯胺酮（70mg / kg）和甲苯噻嗪（16mg / kg）诱导麻醉。保持正常的体温，有助于快速诱导深度麻醉。通过疼痛刺激（例如脚趾捏合）测试足够的镇静剂?...

<ol>
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U., 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+ABCs+of+measuring+intracerebral+hemorrhage+volumes.">The ABCs of measuring intracerebral hemorrhage volumes.</a> Stroke. 27 (8), 1304-1305 (1996).</li><li>Leclerc, J. 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=A+comparison+of+pathophysiology+in+humans+and+rodent+models+of+subarachnoid+hemorrhage.">A comparison of pathophysiology in humans and rodent models of subarachnoid hemorrhage.</a> Frontiers in Molecular Neuroscience. 11, 71 (2018).</li><li>Titova, E., Ostrowski, R. P., Zhang, J. 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综上所述，血管内丝穿孔操作诱导的标准化SAH小鼠模型具有轻微的浸润，手术时间短和可接受的死亡率。术后24小时进行MRI，以确保正确的出血部位并排除其他相关的颅内病变。此外，我们对不同的SAH出血等级进行了分类并测量了出血量，从而可以根据出血等级进行进一步的亚组分析。
鼠标的充分定位会影响正确穿孔的成功。小鼠的脖子应略微伸展到手术的另一侧，头部略微...

蛛网膜下腔出血 （SAH） 是由颅内动脉瘤破裂引起的，可构成危及生命的急症，与大量发病率和死亡率相关，约占卒中1，2 的 5%。SAH 患者表现为严重头痛、神经功能障碍和进行性意识障碍3.大约30%的SAH患者在初始出血事件4之后的前30天内死亡。临床上，50% 的患者在早期脑损伤后出现延迟性脑损伤 （DBI）。DBI 的特征是脑缺血延迟和神经功能缺损延迟。目前的研究表明，几种不同因素的协同作用导致神经功能丧失，包括血脑屏障的破坏，小动脉的收缩，微循环功能障碍和血栓形成5，6。
SAH的一个独特方面是发病机制起源于实质外位置，但随后导致实质内有害的级联反应：病理学始于蛛网膜下腔内血液的积聚，引发多种实质内效应，例如神经炎症，神经元和内皮细胞凋亡，皮质扩散去极化和脑水肿形成7，8.
临床研究受到几个因素的限制，这使得动物模型成为一致和准确地模仿疾病病理变化的关键因素。已经提出了不同的SAH模型方案，例如，将自体血液注射到大水箱（ACM）中。此外，改良方法将自体血液分别注射到大水箱和视交叉水箱（APC）中9，10。虽然自体血液注射是模拟蛛网膜下腔出血后血管痉挛和炎症反应病理过程的简单方法，但随后颅内压（ICP）的升高相对较慢，并且没有诱导血脑屏障通透性显着变化11，12。另一种方法，动脉周围血液放置，通常用于大型SAH模型（例如，猴子和狗），涉及在血管周围放置抗凝自体血液或类似的血液制品。可以用显微镜观察动脉的直径变化，作为SAH13后脑血管痉挛的指标。
Barry等人于1979年首次描述了血管内穿孔模型，其中基底动脉在取出颅骨后暴露;然后用钨微电极刺穿动脉，使用微观立体定向技术14。1995年，Bederson和Veelken修改了脑缺血的Zea-Longa模型，建立了血管内穿孔，自15，16以来一直在不断改进。这种方法是基于这样一个事实，即小鼠和人类共享一个相似的颅内血管网络，称为威利斯的圆圈。
对于小鼠模型中SAH的术后评估和分级，已经提出了不同的方法。Sugawara等人开发了一种自2008年以来被广泛使用的分级量表17。该方法根据形态学变化评估SAH的严重程度。然而，对于这种方法，必须在直接视觉下检查小鼠的脑组织形态，因此，必须牺牲小鼠进行评估。此外，已经建立了几种测定体内SAH严重程度 的方法 。方法范围从简单的神经学评分到颅内压监测（ICP），再到各种放射成像技术。此外，MRI 分级已被证明是一种新的非侵入性工具，用于对 SAH 严重程度进行分级，与神经系统评分18，19 相关。
在这里，提出了由血管内穿孔引起的SAH模型的方案，并结合术后MRI。为了建立一个系统来客观化 体内 出血量，我们还开发了一个基于7.0 T高分辨率T2加权MRI的总血容量SAH分级和定量的系统。这种方法可确保正确诱导 SAH 并排除其他病症，如卒中、脑积水或脑出血 （ICH） 和并发症。

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>Eye cream</td>
			<td>Bayer</td>
			<td>815529836</td>
			<td>Bepanthen</td>
		</tr>
		<tr>
			<td>Images analysis software</td>
			<td>ImageJ</td>
			<td></td>
			<td>Bundled with Java 1.8.0_172</td>
		</tr>
		<tr>
			<td>Ligation suture (5-0)</td>
			<td>SMI</td>
			<td></td>
			<td>Silk black USP</td>
		</tr>
		<tr>
			<td>Light source for microscope</td>
			<td>Zeiss</td>
			<td>CL 6000 LED</td>
			<td></td>
		</tr>
		<tr>
			<td>Ketamine</td>
			<td>CP-pharma</td>
			<td>797-037</td>
			<td>100 mg/mL</td>
		</tr>
		<tr>
			<td>MRI</td>
			<td>Bruker</td>
			<td>Pharmascan 70/16</td>
			<td>&#160;7 Tesla</td>
		</tr>
		<tr>
			<td>MRI images acquired software</td>
			<td>Bruker</td>
			<td></td>
			<td>Bruker Paravision 5.1</td>
		</tr>
		<tr>
			<td>Paracetamol (40 mg/mL)</td>
			<td>bene Arzneimittel</td>
			<td>4993736</td>
			<td></td>
		</tr>
		<tr>
			<td>Prolene filament (5-0)</td>
			<td>Erhicon</td>
			<td>EH7255</td>
			<td></td>
		</tr>
		<tr>
			<td>Razor</td>
			<td>Wella</td>
			<td>HS61</td>
			<td></td>
		</tr>
		<tr>
			<td>Surgical instrument (Fine Scissors)</td>
			<td>FST</td>
			<td>14060-09</td>
			<td></td>
		</tr>
		<tr>
			<td>Surgical instrument (forceps#1)</td>
			<td>AESCULAP</td>
			<td>FM001R</td>
			<td></td>
		</tr>
		<tr>
			<td>Surgical instrument (forceps#2)</td>
			<td>AESCULAP</td>
			<td>FD2855R</td>
			<td></td>
		</tr>
		<tr>
			<td>Surgical instrument (forceps#3)</td>
			<td>Hammacher</td>
			<td>HCS 082-12</td>
			<td></td>
		</tr>
		<tr>
			<td>Surgical instrument (Needle holder)</td>
			<td>FST</td>
			<td>91201-13</td>
			<td></td>
		</tr>
		<tr>
			<td>Surgical instrument (Vannas Spring Scissors)</td>
			<td>FST</td>
			<td>15000-08</td>
			<td></td>
		</tr>
		<tr>
			<td>Surgical microscope</td>
			<td>Zeiss</td>
			<td>Stemi 2000 C</td>
			<td></td>
		</tr>
		<tr>
			<td>Ventilation monitoring</td>
			<td>Stony Brook</td>
			<td></td>
			<td>Small Animal Monitoring &#38; Gating System</td>
		</tr>
		<tr>
			<td>Wounding suture(4-0)</td>
			<td>Erhicon</td>
			<td>CB84D</td>
			<td></td>
		</tr>
		<tr>
			<td>Xylavet</td>
			<td>CP-pharma</td>
			<td>797-062</td>
			<td>20 mg/mL</td>
		</tr>
	</tbody>
</table>

endovascular perforation model for subarachnoid hemorrhage combined with magnetic resonance imaging (mri)

模仿蛛网膜下腔出血（SAH）的血管内丝穿孔模型是一种常用的模型 - 然而，该技术可导致高死亡率以及无法控制的SAH体积和其他颅内并发症，如中风或颅内出血。在该协议中，提出了由血管内丝穿孔诱导的标准化SAH小鼠模型，并在手术后24小时结合磁共振成像（MRI）以确保正确的出血部位并排除其他相关的颅内病变。简而言之，用腹腔内氯胺酮/ 甲苯噻嗪（70mg / 16mg / kg体重）注射液麻醉C57BL / 6J小鼠并置于仰卧位。中线颈部切口后，暴露颈总动脉（CCA）和颈动脉分叉，以逆行方式将5-0不可吸收的单丝聚丙烯缝合线插入颈外动脉（ECA）并推进到颈总动脉。然后，将细丝插入颈内动脉 （ICA） 并向前推入脑前动脉 （ACA）。手术恢复后，小鼠在24小时后接受7.0 T MRI。出血量可以通过术后MRI进行量化和分级，从而实现强大的实验性SAH组，并可选择根据血量进行进一步的亚组分析。

死亡率 对于这项研究，共有92只年龄在8-12周之间的雄性C57Bl / 6J小鼠接受SAH手术;在这些中，我们观察到总死亡率为11.9%（n = 12）。死亡率仅发生在手术后的前6-24小时内，表明围手术期死亡率以及SAH出血本身是最可能的促成因素。
SAH出血等级 共有50只小鼠在术后24小时接受MRI以确认SAH并确保检测其他共同发生的病症，包括亚急性缺血性中风和脑积?...

Watch this Scientific Journal Video about Endovascular Perforation Model for Subarachnoid Hemorrhage Combined with Magnetic Resonance Imaging MRI at JoVE.com

Endovascular Perforation Model for Subarachnoid Hemorrhage Combined with Magnetic Resonance Imaging MRI

这里我们提出了一个标准化的SAH小鼠模型，由血管内丝穿孔诱导，术后24小时结合磁共振成像（MRI），以确保正确的出血部位并排除其他相关的颅内病变。

蛛网膜下腔出血结合磁共振成像（MRI）的血管内穿孔模型

The endovascular filament perforation model to mimic subarachnoid hemorrhage (SAH) is a commonly used model - however, the technique can cause a high ...

血管内穿孔模型与术后MRI相结合，可以验证出血并排除其他颅间病变。这种方法的优点之一是可以选择量化出血量以进行进一步的亚组分析。首先剃掉老鼠的脖子毛发。在手术过程中使用眼药膏。用70%乙醇消毒皮肤，然后用Betadine或氯己定，并应用1%利多卡因。将鼠标置于仰卧位置，头部和躯干朝上。通过在四肢上使用胶带来稳定鼠标背侧。拉伸颈部皮肤并抬高它。将颈部皮肤从下巴展开到胸骨的上边缘，并将唾液腺与相邻的结缔组织分开。通过沿着气管一侧分离肌肉来暴露颈总动脉鞘。用牵开器或无菌纱布拭子暴露手术场，分离颈动脉并留下空闲的8-0丝缝线无需结扎，确保不会损伤迷走神经。解剖 ECA 的远端，并在远端位置将血管铰接两次。在两次结扎的细丝段的中点脱离ECA并形成血管残端。在ICA周围留下丝绸缝合线而不关闭它。预先安排一个结扎在ECA树桩周围的细丝。在灯丝插入成功之前不要关闭它。使用缝合线或微夹暂时关闭ICA和CCA。使用微血管剪刀，在ECA中切开一个切口，插入5-0或4-O Prolene丝，并将其推进到CCA中。关闭 ECA 上的连字，同时分离 ICA 和 CCA 上的微夹。轻轻拉回细丝，在颅方向调整ECA残端，同时将细丝通过分叉插入ICA。将细丝尖端向内侧放置，与水平面上的气管中线成约 30 度的角度，并将其推入 ICA 内。在ACA-MCA分岔处，观察到阻力。将灯丝向前移动最多三毫米，并穿孔正确的ACA。快速将细丝拉到 ECA 残端，使血液在蛛网膜下腔内流动。将灯丝放在这个位置10秒钟。取出灯丝。CCA可以事先暂时关闭，以避免过度失血，然后抽取细丝。使用预先安排的缝合线对ECA进行诉讼，如果您之前已经关闭了CCA，请重新打开CCA。肌肉震颤、同侧肌病、气喘吁吁、心律改变和尿失禁的存在可能是手术成功的支持证据。确保没有出血泄漏并对皮肤进行消毒。缝合伤口。将小鼠放在保温箱中，等到动物有足够的意识来进行胸骨卧位。为了缓解疼痛，每毫升对乙酰氨基酚施用1.2毫克。手术24小时后，在小鼠头部谐振器上使用啮齿动物扫描仪进行MRI，将小鼠置于37摄氏度的加热循环水毯上。首先，执行快速参考扫描，获取三个正交切片包。然后使用高分辨率 T2 加权 2D 涡轮旋转回波序列进行成像。将数据转换为DICOM图像格式，并使用Imagej软件进行血栓的SAH分级和体积测定。为了确保正确的出血部位并排除其他颅内病变，术后24小时进行MRI扫描。根据T2加权MRI扫描量化SAH出血等级。在检查的小鼠中，14%的动物被归类为没有SAH或出血的放射学证据的零级出血。大多数动物的出血等级在1到3之间，10%的动物表现出第四级出血，这是沿中风和/或ICH定义的SAH。根据SAH厚度量化每个SAH等级的总出血量，这有助于确定出血区域。基于轴向SAH厚度计算的出血量显示其对应的1至3级亚组存在显着差异。在整个手术过程中，彻底解剖周围的连接组织对于缝合线的正确定位至关重要。确保ECA残端的长度足够，以便您有舒适的工作空间来连接，切开和插入灯丝。切勿在MCA-ACA分岔前感到阻力时将灯丝向前推，以防止血管过早穿孔。

Research

JoVE Journal

Neuroscience

Endovascular Perforation Model for Subarachnoid Hemorrhage Combined with Magnetic Resonance Imaging (MRI)

Multiple-mouse Neuroanatomical Magnetic Resonance Imaging

多鼠神经解剖学磁共振成像

The field of mouse phenotyping with magnetic resonance imaging (MRI) is rapidly growing, motivated by the need for improved tools for characterizing and ...

科研

神经科学

High-resolution Functional Magnetic Resonance Imaging Methods for Human Midbrain

高分辨率人类中脑功能磁共振成像方法

Functional MRI (fMRI) is a widely used tool for non-invasively measuring correlates of human brain activity. However, its use has mostly been focused upon ...

The Use of Magnetic Resonance Spectroscopy as a Tool for the Measurement of Bi-hemispheric Transcranial Electric Stimulation Effects on Primary Motor Cortex Metabolism

磁共振波谱的用途，用于对初级运动皮层新陈代谢双半球经颅电刺激效果的测量工具一

Transcranial direct current stimulation (tDCS) is a neuromodulation technique that has been increasingly used over the past decade in the treatment of ...

High-resolution In Vivo Manual Segmentation Protocol for Human Hippocampal Subfields Using 3T Magnetic Resonance Imaging

High-resolution In Vivo Manual Segmentation Protocol for Human Hippocampal Subfields Using 3T Magnetic Resonance Imaging

高分辨率<em&gt;在体内</em&gt;使用3T磁共振成像手动分割协议为人类海马子字段

The human hippocampus has been broadly studied in the context of memory and normal brain function and its role in different neuropsychiatric disorders has ...

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

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

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 ...

A Magnetic Resonance Imaging Protocol for Stroke Onset Time Estimation in Permanent Cerebral Ischemia

脑缺血中风发作时间估计的磁共振成像协议

MRI provides a sensitive and specific imaging tool to detect acute ischemic stroke by means of a reduced diffusion coefficient of brain water. In a rat ...

A Volumetric Method for Quantification of Cerebral Vasospasm in a Murine Model of Subarachnoid Hemorrhage

小鼠蛛网膜下腔出血模型脑血管痉挛量化的容积法

Subarachnoid hemorrhage (SAH) is a subtype of hemorrhagic stroke. Cerebral vasospasm that occurs in the aftermath of the bleeding is an important factor ...

Double Direct Injection of Blood into the Cisterna Magna as a Model of Subarachnoid Hemorrhage

双直接注射血液到西斯特纳麦格作为苏巴拉奇内德出血的模型

Among strokes, subarachnoid hemorrhage (SAH) consecutive to the rupture of a cerebral arterial aneurysm represents 5-9% but is responsible for about 30% ...

Analysis of Cerebral Vasospasm in a Murine Model of Subarachnoid Hemorrhage with High Frequency Transcranial Duplex Ultrasound

高频颅内双体超声波的苏巴拉赫诺德出血的穆林模型脑血管痉挛分析

Cerebral vasospasm that occurs in the weeks after subarachnoid hemorrhage, a type of hemorrhagic stroke, contributes to delayed cerebral ischemia. A ...

Whole-Brain Single-Cell Imaging and Analysis of Intact Neonatal Mouse Brains Using MRI, Tissue Clearing, and Light-Sheet Microscopy

使用MRI，组织清除和光片显微镜对完整的新生小鼠大脑进行全脑单细胞成像和分析

Tissue clearing followed by light-sheet microscopy (LSFM) enables cellular-resolution imaging of intact brain structure, allowing quantitative analysis of ...