诱导小鼠抑郁行为的慢性固定性压力协议
1Department of Anatomy and Convergence Medical Sciences, Institute of Health Sciences, Bio Anti-aging Medical Research Center,Gyeongsang National University Medical School, 2Department of Biological Sciences,Pusan National University, 3Department of Physiology and Convergence Medical Sciences, Institute of Health Sciences,Gyeongsang National University Medical School
Summary
本文提供了一个简化和标准化的协议,用于诱导长期固定小鼠的抑郁样行为使用约束剂。此外,还解释了验证抑郁症诱导的行为和生理技术。
Abstract
抑郁症尚未完全了解,但各种因果关系已经报告。最近,抑郁症的患病率有所上升。然而,治疗抑郁症或研究抑郁症是稀缺的。因此,本文提出了运动限制引起的抑郁小鼠模型。慢性轻度压力 (CMS) 是一种众所周知的诱导抑郁行为的技术。然而,它需要一个复杂的程序,由各种轻度应力的组合组成。相反,慢性固定压力 (CIS) 是一种易于获取的慢性应激模型,从约束模型修改,该模型通过限制运动在一定时期内限制运动来诱发抑郁行为。为了评估抑郁样的行为,本实验中结合蔗糖偏好测试(SPT)、尾部悬浮试验(TST)和ELISA测定来测量应力标记皮质酮水平。所述协议说明了CIS的诱导和评价行为和生理因素的变化,以验证抑郁症。
Introduction
重度抑郁症(MDD)是全世界精神残疾的主要原因,其发病率增长快于预期。2001年,世界卫生组织预测,到2020年,MDD将成为世界上第二大常见疾病。然而,它已经是第二最常见的在2013年1。此外,目前的抗抑郁药有许多局限性,包括延迟效果,耐药性,复发,和各种副作用2,3。因此,研究人员必须开发更有效的抗抑郁药。然而,MDD的模糊病理生理学对新型抗抑郁药的发展构成了障碍。
长期压力是MDD的主要危险因素。它可以诱发下丘脑-垂体-肾上腺(HPA)轴功能障碍,这也与MDD病因4,5相关。如前所述,HPA 轴在压力引起的精神病理生理学(包括抑郁和焦虑障碍)中起关键作用,通过增加皮质酮水平6、7、8、9.许多动物模型都基于 HPA 轴的持续激活,在 MDD4患者中观察到。此外,由慢性压力和皮下注射的糖皮质激素引起的高糖皮质激素会导致抑郁行为以及神经细胞死亡、神经元过程萎缩和啮齿动物大脑中成人神经发生减少10,11.与抑郁症相关的另一个重要大脑区域是前额叶皮质 (mPFC)。mPFC在控制大脑分区域(如下丘脑和杏仁核)中起着至关重要的作用,控制情绪行为和压力反应8,9。例如,背膜mPFC的病变诱发HPA轴功能障碍和增强皮质酮分泌由于约束应力12,13。最近的一项研究还表明,反复约束压力增加皮质酮水平,可以通过谷氨酰胺补充谷氨酰胺在mPFC9神经元和星形细胞之间的谷氨酸-谷氨酰胺循环来降低。
Katz14提出了第一个用于研究MDD病因的慢性应激范式。Willner等人随后根据Katz的发现提出了一个慢性轻度应激(CMS)模型。他们证实,该模型具有预测有效性,通过观察抗抑郁药恢复CMS引起的类人无一样的行为15,16。通常,CMS 模型由各种轻度应力的组合组成,例如轻微噪音、笼子倾斜、湿床上用品、改变的暗箱循环、笼摇、强迫游泳和社会失败。CMS模型被研究者广泛使用;然而,该模型的可复制性较差,而且时间和能效都很低。因此,对诱导抑郁行为和生理分析以评估抑郁症的标准化和简化方案的需求日益增长。与CMS模型相比,慢性固定应力(CIS;也称为慢性约束应激)模型更简单、更高效;因此,CIS模型可以广泛用于慢性应激研究17,18,19,20,21,22,23, 24.此外,CIS可用于雄性小鼠和雌性小鼠,以发展抑郁症行为25,26。在CIS期间,动物被放置在一个适合身体大小的圆柱体中,每天1-8小时,为期2周或4周9,27,28。其中,每天2小时的克制应激状态,持续2周,足以引起小鼠9、28的抑郁行为,疼痛最小。在约束条件下,血液皮质酮水平迅速上升9,28,29。几项研究表明,CIS模型具有预测有效性,证实CIS引起的抑郁症状症状通过抗抑郁药19,20,30,31恢复。在这里,我们报告CIS的详细程序,以及一些在小鼠CIS后的行为和生理结果。
Protocol
所有实验规程和动物护理均按照庆尚国立大学动物研究大学动物护理委员会(GLA-100917-M0093)的指导原则进行。 1. 材料 小 鼠 在产后第7周使用重达22-24克的C57BL/6菌株背景的雄性。在实验前在繁殖室中活动1周。注:所有小鼠都是从实验室动物公司购买的。 在12小时的明/暗周期(早上6点亮灯)下,将小鼠单独在温度控制的活体(22-24°C)中,并可正常使用实验室的会?…
Representative Results
在代表性实验中,所有数据均从每组6-8只小鼠中获取。具有代表性的材料和将鼠标自愿插入约束器的方法如图1所示。 为了在CIS诱导后进行行为测试和血液采样,小鼠接受了如图2A中概述的实验程序。如图2和图3所示,CIS能很好地诱导抑郁样的行为,并释放压力标记皮质酮。此外,这些指标通过?…
Discussion
大脑的复杂性和MDD的异质性使得创建完全重现这种状况的动物模型变得十分困难。许多研究人员已经克服了这一困难,使用内皮内皮型的方法32,其中阿内多尼亚(缺乏兴趣奖励刺激)和绝望被认为是进化保护,在动物模型中可量化的行为,这同样出现在抑郁症患者中33。在本文中,我们提出了一种方法,其中独联体足以引起一种绝望和绝望,表明独联体和MDD之间的翻译相关…
Disclosures
The authors have nothing to disclose.
Acknowledgements
这项研究得到了韩国国家科学研究基金会(NRF)基础科学研究计划的支持,该基金会由韩国教育部资助(NRF-2015R1A5A2008833和NRF-2016R1D1A3B03934279)和卫生科学基金会(IHS)资助GNU-2016-02)在庆尚国立大学。
Materials
| 1 ml disposable syringes | Sungshim Medical | P000CFDO | |
| Balance | A&D Company | FX-2000i | |
| Ball nozzle | Jeung Do B&P | JD-C-88 | |
| CCTV camera | KOCOM | KCB-381 | |
| Corticosterone ELISA kits | Cayman Chemical | ||
| Digital lux meter | TES | TES-1330A | |
| Ethovision XT 7.1 | Noldus Information Technology | ||
| Isoflurane | HANA PHARM CO., LTD. | Ifran solution | |
| Mice | Koatech | C57BL/6 strain | |
| Restrainer | Dae-jong Instrument Industry | DJ-428 | |
| Saccharose (sucrose) | DAEJUNG | 7501-4400 | |
| Small animal isoflurane anaesthetic system | Summit | ||
| Acrylic bar | The apparatus was made in the lab for TST test | ||
| Tail suspension box | The apparatus was made in the lab | ||
| Timer | Electronics Tomorrow | TL-2530 | |
| Water bottle | Jeung Do B&P | JD-C-79 |
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Cite This Article
Son, H., Yang, J. H., Kim, H. J., Lee, D. K. A Chronic Immobilization Stress Protocol for Inducing Depression-Like Behavior in Mice. J. Vis. Exp. (147), e59546, doi:10.3791/59546 (2019).