基于改进多平台法的大鼠中心疲劳模型
Summary
在这里, 我们提出了一个使用改进的多平台方法 (MMPM) 来引入大鼠中心疲劳模型的协议。
Abstract
本文采用改进的多平台法 (MMPM), 介绍了一种大鼠中心疲劳模型。多平台箱设计成一个水箱, 底部有狭窄的平台。模型鼠被投入了坦克并且在平台上站立了14小时 (18:00-8:00) 每天连续21天, 与空白控制小组设置为对比。建模结束时, 模型组大鼠表现出明显的疲劳状态。为了评估模型, 我们进行了几个行为测试, 包括开放场测试 (常), 高附加迷宫 (吸盘) 测试, 和详尽的游泳 (ES) 测试。结果表明: 模型大鼠焦虑、空间认知功能障碍、肌肉表现不佳、自愿活动减少, 证实了中枢性疲劳的诊断。中枢神经递质的变化也验证了结果。最后, 该模型成功地模拟了中心疲劳, 未来的研究与模型可以帮助揭示疾病的病理机制。
Introduction
疲劳是威胁人类健康的主要因素之一1。在过去的几十年里, 各种研究已经证明疲劳是外围触发的, 但集中驱动, 总是伴随着情感和认知障碍。意大利生理学家 a. Mosso 首先提出了 “中心疲劳2” 这个词。它一般被定义为有限的自愿活动和认知功能障碍, 由于中枢神经系统的冲动传输功能障碍 (CNS)3。与周围肌肉疲劳相比, 中枢性疲劳强调中枢神经系统的变化, 以及由此产生的情绪/行为紊乱, 包括抑郁、焦虑、认知障碍和记忆力减退。一项研究表明, 许多因素会导致中枢性疲劳, 其中过度的体力活动和精神压力是相当不可缺少的4。至于发病机制, 如色氨酸-犬尿氨路径假说5解释某些途径的变化;然而, 还需要更深入的研究来揭示中枢性疲劳的中枢-周边相关性。
由于中心疲劳的基本机制尚不清楚, 有效的动物模型对进一步的研究具有十分重要的意义。现有的疲劳模型大多是由过度运动引起的, 如跑步机6和负重游泳7, 很少关注心理因素。为了更好地模拟中心疲劳的发展, 我们小组开发了 MMPM 的大鼠模型。在建模过程中, 老鼠仍然站在狭窄的平台上, 在多个平台盒长时间, 包括部分睡眠时间。与过度运动模型不同, MMPM 模型采用部分睡眠剥夺作为心理因素, 考虑到复杂的中枢性疲劳发病机制。
对于模型评价, 我们使用经常和测验来确定焦虑情绪和自愿活动。采用 ES 试验对周围肌肉的性能进行测量。此外, 我们采取大鼠的大脑和检测多巴胺 (DA)/血清素 (5 HT) 的内容, 在两个 hypothalamuses 观察中枢神经递质的差异。
下面提出的协议旨在模拟在人类生活中常见的情况下, 反复的体力活动和缺乏睡眠导致的中枢疲劳。然而, 通过调整模型的持续时间, 它可以用于许多其他领域, 如睡眠观察和应力研究。在今后的研究中, 我们希望这一模型能帮助发现更多的中枢神经系统变化及其与周边体系的联系, 揭示中枢性疲劳的发病机制。
Protocol
所有的动物都是按照中国关于实验室动物的道德使用和护理的法律来维护的。 1. 预建模前准备 实验室准备 在实验前至少30分钟运行紫外线灯。 控制实验室温度在 25 3 摄氏度, 相对湿度约30% 左右。 打开实验室灯在6:00 和关闭在18:00 建立一个 12 h/12 h 光/暗循环。 多平台箱结构 构造一个不透明的?…
Representative Results
我们用 MMPM 描述了一种大鼠中心疲劳模型。24只大鼠随机分为对照组和模型组, 每组12只大鼠。该模型装置设计成一个水箱, 底部有狭窄的平台 (图 1)。模型鼠每天在平台上站立14小时, 包括部分睡眠时间, 21 天 (图 2)。 行为测试是在建模后进行的, 以评估大鼠的情绪和物理变化。通常的…
Discussion
MMPM 最初设计为睡眠剥夺9。老鼠被发射到水箱里, 平台固定在底部。由于对水的本能的恐惧, 老鼠仍然站在平台上, 没有睡眠发生。研究表明, 不同时间的睡眠剥夺导致大鼠行为和情绪的变化, 包括认知障碍10, 消极情绪11, 中心疲劳。一些研究人员证明, 采用单平台法 (SPM) 的慢性睡眠剥夺可诱发中枢性疲劳, 并有认知障碍和社会障碍12…
Disclosures
The authors have nothing to disclose.
Acknowledgements
这项工作得到北京自然科学基金 (7162124 号) 和北京中医药大学新奥基金会的支持。
Materials
| multiple platform sleep deprivation water tank | Customization,it is provided by the neuroimmunological laboratory of Beijing University of Chinese Medicine | 110cm x 60cm x 40cm. There are 15 plastic small platforms at the bottom. The small platform is 6.5cm in diameter and 8cm high | |
| Wistar rats | Beijing Weitong Lihua Experimental Animal Technology Company | license number SYXK (Beijing) 2016-0011 | Use 32 Wistar healthy male rats ,8 week old (200-210 g) |
| Agilent 1100LC high performance liquid chromatograph | Agilent | G1379A, G1311A, G1313A , G1316A | G1379A, G1311A type chromatographic pump, G1313A automatic sampler, G1316A column temperature box |
| DECADE II SDC electrochemical detector | Dutch ANTEC company | glassy carbon electrode, Ag/AgCl reference electrode, workstations (Clarity CHS) | |
| Biofuge Stratos high-speed refrigeration centrifuge | HERAEUS | ||
| VCX130 ultrasonic fracturing instrument | SONICS | ||
| ACS-ZEAS electronic scale | Phos technology development, Beijing. | The weight of the weighing rats can be accurate to 0.1g. | |
| Open Field Box | Customization,it is provided by the neuroimmunological laboratory of Beijing University of Chinese Medicine | wooden box of open field 100 cm by 100 cm x 40 cm, inside wall and bottom as the gray.The bottom is divided into 25 equal area squares, each of which is 20cm x 20cm, and the 16 grids along the outer wall are the external ones, and the other 9 grids are central.The camera is mounted above the median. | |
| Elevated Plus-maze | Beijing zhongshi dechuang technology development co. LTD. | The open arms and close arms of the cross are composed of 30cm x 5cm x 15cm, and the central area is 5cm x 5cm, with a camera mounted above the center and 45cm high. | |
| rat swimming bucket. | Zhenhua biological instrument equipment co., LTD. Anhui,China. | The volume of plastic drum is 70cm x 30cm x 110cm, which is used for swimming in rats. | |
| Thermometer | Shiya instrument co., LTD., changzhou,China. | Control water temperature | |
| Small water pump | Xincheng technology co., LTD., chengdu,China. | Used for water tank and swimming behavior. | |
| Ethovition3.0 behavioral software. | Nuldus,Netherlands | Measurement analysis of rat behavior videos. |
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Cite This Article
Zhang, W., Zhang, W., Dai, N., Han, C., Wu, F., Wang, X., Tan, L., Li, J., Li, F., Ren, Q. A Rat Model of Central Fatigue Using a Modified Multiple Platform Method. J. Vis. Exp. (138), e57362, doi:10.3791/57362 (2018).