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Medicine

基于蒙古药的大鼠慢性不可预测的轻度应激

Published: October 27, 2023
doi:
10.3791/65889
, , , , , , , , , , , ,
1Faculty of Humanities Education,Inner Mongolia Medical University, 2Faculty of Mongolian Medicine,Inner Mongolia Medical University, 3Faculty of Basic Medical,Inner Mongolia Medical University

Summary

该协议概述了基于蒙古医学理论的抑郁症慢性不可预测轻度压力 （CUMS） 模型，以及验证行为测试的方法。

Abstract

抑郁症是一种普遍存在的情感障碍，是导致全球残疾的主要原因。目前药物干预的局限性导致了这种疾病带来的巨大健康负担。迫切需要更深入地了解抑郁症的潜在机制，这使得具有转化潜力的临床前模型非常有价值。蒙古医学是传统医学的一个分支，认为疾病的发生与风、胆汁和痰液的平衡密切相关。在这项研究中，我们介绍了大鼠慢性不可预测的轻度应激（CUMS）方法的方案。在这个框架内，大鼠受到一系列波动的、温和的压力源的影响，以诱导出类似抑郁症的表型，模仿人类抑郁症的发病机制。该协议中采用的行为测定包括蔗糖偏好测试 （SPT），表明快感缺乏——抑郁症的核心症状;测量焦虑水平的旷场测试（OFT）;以及评估空间记忆和学习能力的莫里斯水迷宫测试（MWM）。CUMS 方法证明了诱导快感缺乏和导致长期行为缺陷的能力。此外，与其他旨在引发抑郁样行为的动物模型相比，该协议更符合蒙古医学理论。这种动物模型的开发和后续研究为蒙古医学领域未来的创新研究奠定了坚实的基础。

Introduction

重度抑郁症 （MDD） 是一种普遍存在的精神疾病，是全球第三大残疾原因，影响着超过 3 亿人1,2,3.值得注意的是，据估计，至少有一半的受影响个体没有得到充分的治疗4.鉴于这一差距，动物模型可以作为调查抑郁症病因的重要工具。迄今为止，存在 20 多种不同的抑郁症动物模型5.其中，Paul Winer 于 1987 年改进的慢性不可预测的轻度压力 （CUMS） 模型是最常用的6.CUMS 模型的前提是，将啮齿动物暴露于各种社会环境压力源会导致类似于焦虑、紧张和抑郁的症状。该方法涉及在几周内将动物暴露于各种轻度压力源中，最终导致一系列行为改变，包括快感缺乏和抑郁样行为7,8.这些变化伴随着内分泌和神经递质谱的变化，例如 5-HT 的减少9,10.这些结果与在被诊断患有MDD的人类中观察到的结果非常相似，从而验证了该模型的实用性。CUMS 模型因其在评估抗抑郁药方面的有效性而受到特别重视，表现出高水平的表面、结构和预测有效性11,12.与其他模型不同，CUMS 对长期服用单胺能抗抑郁药的影响敏感。例如，选择性血清素再摄取抑制剂 （SSRIs），如西酞普兰、帕罗西汀和氟西汀，已被证明可以预防和逆转慢性压力条件下的快感缺乏12,13.此外，新的速效抗抑郁药，如氯胺酮，也证明了这种模型的疗效14,15.相比之下，其他测试，如强迫游泳测试（FST）和尾部悬挂测试（TST）在模拟长期行为变化方面不太可靠，通常反映出对急性压力的适应，而不是持久的抑郁症状16.这些特征强调了CUMS模型在抑郁症研究中的稳健有效性。CUMS模型最显着的特征之一是快感缺乏症，该模型因其在经典研究中的高可靠性而得到认可 – 无法在日常活动中体验快乐或兴趣17,18.这种现象通常使用蔗糖偏好测试进行评估，许多抗抑郁药已被证明可以逆转蔗糖消耗减少。CUMS 文献中也常用其他几个指标，包括开场测试 （OFT），它评估自愿运动行为、探索倾向和紧张程度，从而衡量抑郁症的严重程度19.其他测试，如高架加迷宫 （EPM） 评估类似焦虑的行为，莫里斯水迷宫测试 （MWM） 检查认知功能20，FST 评估对负面情绪和行为绝望的易感性20.此外，影响人类的大多数压力源本质上都是社会性的。社会关系欠佳的个体，其特征是社会活动、网络和支持有限，患各种疾病的风险更高21,22.这也与啮齿动物有关，啮齿动物是群居的群居动物。例如，隔离饲养的大鼠表现出所谓的隔离综合征的特征，这种特征会引起社会压力并加速抑郁症的发作23.

蒙古医学是中医的一个重要分支，它认为疾病的发作是内在因素和外部因素之间的复杂相互作用。这些外部因素被称为四个辅助条件，包括气候变化、饮食、生活方式和突发事件，如感染、令人震惊的事件和心理障碍。疾病过程被概念化为三个元素（称为三种类型的同源体）与七种身体成分与四种辅助条件之间的持续相互作用24.蒙古医学认为，人体作为一个整体运作，由三个同源之间的相对平衡维持。这种平衡的破坏被认为是疾病的前兆24.鉴于动物实验在连接传统医学和现代医学方面发挥着关键作用，开发与蒙古医学领域研究相关的动物模型至关重要。因此，我们采用了 28 天隔离方法与 CUMS 相结合来模拟这些生理和心理压力源。我们选择了九个特定的不可预测的应激源，并试图通过蒙古医学的三同源理论来支持这种建模方法。建立稳健的动物模型是推进蒙古医学基础研究的基础，并将对其基础研究做出重大贡献。

Protocol

实验方案获得了内蒙古医科大学（YKD202301172）动物实验护理伦理委员会的批准，并遵守了美国国立卫生研究院的动物护理和伦理指南。我们动物中心的许可证号是NO.110324230102364187。获得 24 只雄性 Sprague-Dawley （SD） 大鼠，每只 8 周龄 （200 g ± 20 g），并将其饲养在温度为 22°C ± 2°C，湿度为 55% ± 15% 的受控环境中。用啮齿动物维持饲料喂养大鼠，用玉米芯喂养饲料和纯净水作为垫料。在实验前，将?…

Representative Results

CUMS诱导的大鼠抑郁模型中的行为测试结果为了证实 CUMS 程序在诱导抑郁样行为方面的功效，进行了操作检查。雄性Sprague-Dawley（SD）大鼠被随机分配到MOD或CON组，为期4周，如步骤1.2.3所述。随后，处死大鼠，并完全解剖其海马体，使用酶联免疫吸附测定（ELISA）28评估5-HT，这是一种与抑郁症病理生理学密切相关的神经递质10。 <p class="jove_content"…

Discussion

抑郁症是一种精神障碍，其特征是情绪低落、缺乏快乐和精力减少等症状30.在抑郁症研究领域，建立可靠的动物模型对于推进治疗干预至关重要。在各种动物模型中，CUMS模型因其高信度、效度以及与人类抑郁症31特征的一致性而特别值得关注。它非常适合在各种环境中模拟低水平压力源在长时间内的累积效应。在这项研究中，我们采用了步骤 1.2.3 和表 2</…

Disclosures

The authors have nothing to disclose.

Acknowledgements

我们感谢中国内蒙古医科大学蒙古医学院提供的仪器和实验室，本研究得到了必要的仪器和实验室设施的支持。本研究获得国家自然科学基金（Grant No. 81760762）和内蒙古医科大学项目（Grant No.YKD2022MS074）和内蒙古自治区高等教育科研项目（批准号）NJZY22661）内蒙古自治区中蒙医学重点实验室开放基金项目（批准号）MYX2023-K07）。

Materials

1.5 mL centrifuge tube service Biotechnology Co., Ltd EP-150-M
1000 µL Pipette service Biotechnology Co., Ltd IC021198160223
10 µL pipette tip service Biotechnology Co., Ltd IC012395160823
10 µL pipette tip service Biotechnology Co., Ltd TP-10
1250 µL pipette tip service Biotechnology Co., Ltd TP-1250
2 mL centrifuge tube service Biotechnology Co., Ltd EP-200-M
200 µL pipette tip service Biotechnology Co., Ltd TP-200
200 µL pipette tip service Biotechnology Co., Ltd IC021029160323
300 µL Multi-Channel Pipette service Biotechnology Co., Ltd IC091006161022
50 µL Pipette service Biotechnology Co., Ltd DS35110
Automatic plate washing machine rayto Life Sciences Co., Ltd RT-3100
Benchtop High-Speed Freezing Centrifuge dalong construction Co., Ltd D3024R
electronic balance Mettler Toledo International Trade (Shanghai) Co., Ltd ME203E/02
Electrothermal blast drying oven Labotery Experimental Instrument Equipment Co., Ltd GEL-70
Enzyme Label Detector BioTeK Co., Ltd Epoch
High Speed Tissue Grinder service Biotechnology Co., Ltd KZ-Equation 2-F
Horizontal Freezer Mellow Group Co., Ltd BCD-318AT
Laboratory Ultrapure Water Machine Jinan Aiken Environmental Protection Technology Co., Ltd   AK-RO-C2
Morris water maze video trail analysing system  Tai Meng Tech Co., Ltd WMT-200
Rat 5-HT ELISA Kit Lian Ke bio Co., Ltd,China 96T/48T
SPF grade Sprague Dawley (SD) rats SPF (Beijing) Biotechnology Co  SCXK(JING)2019-0010
Sprague Dawley rats Beijing Biotechnology Co., Ltd, China  SCXK (JING) 2019-0010
Vertical Refrigerated Display Cabinet Xingx Group Co., Ltd LSC-316C
video tracking system Tai Meng Tech Co., Ltd ZH-ZFT
vortex mixer Servicebio technology Co., Ltd MV-100
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Keywords: Chronic Unpredictable Mild StressCUMSMongolian MedicineDepressionAnhedoniaOpen Field TestMorris Water MazeAnimal ModelBehavioral Assays
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Xin, H., Su, S., Wu, R., Wei, L., Su, N., Qi, L., Wu, R., A, R., Tong, L., Wang, W., Zhang, J., Hu, R., Li, L. Chronic Unpredictable Mild Stress in Rats based on the Mongolian medicine. J. Vis. Exp. (200), e65889, doi:10.3791/65889 (2023).
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