Özet

创伤性脑损伤诱导受控皮质冲击的母模型

Published: August 16, 2019
doi:

Özet

在这里,我们描述了通过开头控制皮质撞击诱导鼠外创伤性脑损伤的协议。

Abstract

疾病控制和伤害预防中心估计,美国每年有近200万人遭受创伤性脑损伤(TBI)。事实上,TBI是造成所有与伤害有关的死亡率超过三分之一的因素。然而,TBI病理生理学的细胞和分子机制却知之甚少。因此,能够复制与人类患者TBI相关的损伤机制的TBI临床前模型是一项关键的研究需要。TBI 的受控皮质冲击 (CCI) 模型利用机械装置直接影响暴露的皮层。虽然没有一种模型能够全面概括人类患者中TBI的不同损伤模式和异质性,但CCI能够诱导广泛的临床应用TBI。此外,CCI 易于标准化,使调查人员能够跨实验和跨调查组比较结果。以下协议详细介绍了在 TBI 的鼠模型中使用市售影响装置的严重 CCI。

Introduction

美国疾病控制和预防中心估计,每年约有200万美国人遭受创伤性脑损伤(TBI)。事实上,TBI在美国所有与伤害有关的死亡中,超过30%的死亡率,每年医疗费用接近800亿美元,每人每年在严重的TBI3、4、5中生存近400万美元。TBI的影响突出表现在其幸存者遭受的重大长期神经认知和神经精神并发症与行为,认知和运动损伤的阴险开始称为慢性创伤性脑病(CTE)6,7,8,9,10.即使是亚临床脑震荡事件——那些不导致临床症状的影响——也可能导致长期神经功能障碍11、12。

研究TBI的动物模型从1800年末开始被采用。在 20 世纪 80 年代,开发了一种用于建模 TBI 的气动冲击器。这种方法现在被称为受控皮质撞击(CCI)14。CCI的控制和可重复性使研究人员将模型调整为啮齿动物15。我们的实验室使用这种模型,通过市售的冲击器和电子驱动装置16、17诱导TBI。该模型能够根据所使用的生物力学参数产生广泛的临床适用的TBI状态。在我们的实验室中引起严重损伤后,TBI大脑的由性评价显示,显著的叶边皮质和海马损失以及反向水肿和变形。此外,CCI产生一致的运动和认知功能损伤,通过行为测定18测量。CCI 的限制包括颅切除术和获取冲击器和驱动装置的费用。

TBI的其他几种模型存在,并在文献中得到了很好的建立,包括横向流体打击模型、降重模型和爆炸伤害模型19,20,21。虽然每种型号各有其独特优势,但其主要缺点是混合损伤、高死亡率和缺乏标准化,分别为22。此外,这些模型都没有提供 CCI 的精度、精度和可重复性。通过调整生物力学参数输入到驱动装置中,CCI 模型允许调查人员精确控制损伤的大小、损伤的深度以及施加到大脑的动能。这使得研究者能够将TBI的整个光谱应用于大脑的特定区域。它还允许从实验到实验的最大可重复性。

Protocol

所有程序都通过了西北大学机构动物护理和使用委员会的批准。C57BL/6小鼠是从杰克逊实验室购买的,并组在西北大学比较医学中心(伊利诺伊州芝加哥)的屏障设施中。所有动物都被安置在12/12小时的光/暗周期,免费获得食物和水。 1. 诱导麻醉 用氯胺酮(125毫克/千克)和木兰素(10毫克/千克)在腹内注射给小鼠麻醉。 2. 每15分钟监测一次生?…

Representative Results

冲击器直接安装在立体框架上,分辨率高达 10 μm,用于控制冲击点、深度和穿透力。使用的电磁力可以产生1.5~6米/s的撞击速度。这使得在整个临床相关TBI范围内具有无与伦比的精度和可重复性。调查人员可以进行PILOT实验,改变损伤参数,如撞击器尖端尺寸、撞击速度和撞击深度,以确定最能产生所需伤害程度的参数。该协议描述了一个严重的TBI到左腹腔位区域,通过执行5毫米颅切除术2毫米左?…

Discussion

有几个步骤对于应用可靠和一致的伤害至关重要。首先,小鼠必须到达外科麻醉的深平面,以确保在颅切除术期间没有运动。虽然许多麻醉方案可用于诱导啮齿动物的一般麻醉,但引起呼吸抑郁的麻醉剂,如吸入麻醉剂,在与严重的TBI结合时,可能导致呼吸停止。本协议使用氯胺酮(125毫克/千克)和木兰胺(10毫克/千克)注射在腹内。这种药物组合在5分钟内产生麻醉的手术平面,持续时间约为30…

Açıklamalar

The authors have nothing to disclose.

Acknowledgements

这项工作得到了美国国家卫生研究院GM117341助学金和美国外科医生学会C.詹姆斯·卡里科研究奖学金的资助。

Materials

AnaSed Injection Xylazine Sterile Solution LLOYD, Inc. 5939911020
Buprenorphine SR Lab 0.5mg/mL Zoopharm-Wildlife Pharmaceuticals USA BSRLAB0.5-182012
High Speed Rotary Micromotor KiT0 Foredom Electric Company K.1070
Imapact one for Stereotaxix CCI Leica Biosystems Nussloch GmbH 39463920
Ketathesia Ketamine HCl Injection USP Henry Schein, Inc 56344
Mouse Specific Stereotaxic Base Leica Biosystems Nussloch GmbH 39462980
Trephines for Micro Drill Fine Science Tools, Inc 18004-50

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Bu Makaleden Alıntı Yapın
Schwulst, S. J., Islam, M. B. Murine Model of Controlled Cortical Impact for the Induction of Traumatic Brain Injury. J. Vis. Exp. (150), e60027, doi:10.3791/60027 (2019).

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