Summary

电子 von Frey 与小鼠鬼脸量表相关联,以评估 埃文西锥虫感染小鼠的异常性疼痛和疼痛

Published: July 19, 2024
doi:

Summary

在这里,我们提出了一种方案,用于评估感染 埃文西锥虫的小鼠的伤害感受,使用电子von Frey装置作为工具,测量后爪和内脏的机械阈值。

Abstract

传染病的发病机制仍然是一个需要研究的复杂领域。在家畜中可以观察到几种临床症状的病程,例如异常性疼痛和疼痛。然而,对其途径的了解和正确的治疗需要对照实验,其中许多实验使用实验动物。测量后爪和内脏机械阈值的变化是观察啮齿动物疼痛感知变化的有用技术。戒断反应可以首先在基线测试中测量,从而更好地控制实验组。后续测试可以在诱导感染并将药物添加到方案中后进行。使用与使用面部量表相关的电子 von Frey 设备来观察疼痛样变化,可以进行简单、精确和一致的评估,以评估小鼠的异常性疼痛和疼痛。因此,使用本方法治疗 埃文西锥 虫感染的实验代表了评估实验室感染动物异常性疼痛和疼痛的有用方法,可应用于家畜动物的常规治疗。

Introduction

埃文西锥虫是该疾病的病原体,在南美洲称为“surra”或“mal-das-cadeiras”1,2。它通常影响马和牛,但也影响野生动物,通过噬血蝙蝠、塔巴尼德和口蝇叮咬传播 1,3。Surra 是家畜的一种主要疾病,如果没有正确的治疗,它可能会致命,表现出非特异性临床症状,如贫血、食欲不振和体重减轻、肌肉无力和流产,这些症状可能因宿主和地理区域而异 2,4,5,6。

在疾病过程中,受感染动物的异常性疼痛和疼痛的表达仍然是一个新话题2,7。了解这些体征背后的发病机制的冲动是通过在经典的锥虫杀虫方案中添加有效的镇痛药物来改进和完善目前使用的治疗方法的重要一步 5,6。在这种情况下,使用小鼠模型复制疾病的可能性代表了一个优势,因为小鼠可以很容易地保持在实验室的受控环境中,因此,产生的结果比使用牲畜的田间实验更一致。

在大量实验中,通常使用电子 von Frey (EvF) 设备来评估异常性疼痛 2,8,9。该装置用于评估组织对机械刺激的敏感性:一旦该装置接触动物的爪子,通过连接到该装置的 20-200 μL 尖端,记录动物收回爪子的力。

在这些实验中,啮齿动物通常被用作标准动物,大多数结果都外推到其他物种,因为大多数研究的疾病来自其他物种,在这些物种中很难进行对照研究。此外,异常性疼痛和疼痛密切相关。使用特定的面部量表来评估感染小鼠的疼痛起着重要作用,作为确认佐剂,以确认 T. evansi 感染过程中疼痛的存在 2,10

在该协议中,我们展示了一种新的模型来评估实验感染 T. evansi 的小鼠的异常性疼痛和疼痛,显示出变量之间的高度相关性,因此被证明是一个强大的模型。此外,它需要少数研究人员来执行整个过程,从而减少了实验过程中人为干预的机会。它还使研究者能够在急性病程中复制目标疾病,并在实验设计中添加几种不同的治疗药物,从而在短时间内获得一致的结果。

Protocol

所有实验均使用10周龄的成年雌性瑞士小鼠(35-55g)进行。将动物饲养在聚砜笼(每个笼3-5只动物)中,房间温度受控(21±1°C),12小时光照/ 12小时黑暗循环,标准实验室食物和 水随意饮水。在每次测试中,每组分配10只动物,以证明药物治疗的一致效果。实验设计由圣卡塔琳娜州立大学(CEUA)伦理委员会提交并批准(协议编号:6019201123)。所有动物实验均符合ARRIVE指南(动物研究: <em…

Representative Results

通过电子 von Frey 装置在右后爪和内脏组织上评估的 T. evansi 感染导致的机械阈值降低根据先前与可用 T. evansi 样品2 相关的数据,该实验进行了 5 天。感染小鼠在感染后第3天开始显示出右后爪机械阈值的显着差异,并且在感染后2天内与未感染的小鼠相比,仍然显着低于对照组(图1A)。腹部触觉敏感性的类似结果显示,感染小?…

Discussion

在涉及受感染动物的实验中,一个关键步骤是控制寄生虫血症水平。不同菌株的 T. evansi 在小鼠中可能以不同的方式表现,导致从急性感染到慢性感染 2,4,5,6此外,感染剂量的变化可能会减少或延长小鼠的存活时间。因此,建议在实验前获得生存曲线,以确定实验的正确周期,并防止因意外的…

Disclosures

The authors have nothing to disclose.

Acknowledgements

作者特别感谢圣卡塔琳娜州立大学的财政支持,感谢动物和太空的药理学实验室,以及血液寄生虫和载体生物化学实验室的冷冻保存血液样本,这些血液样本感染了这些实验中使用的 埃文西锥虫

Materials

26 G 1/2" needle coupled to insulin syringe TKL 80288090100 Used to infund solutions on laboratory animals
Accessories for von Frey analgesimeter INSIGHT EFF 303 Containment box with support for digital analgesimeter assessment
D-(+)-Glucose SIGMA-ALDRICH G7021 A monosaccharide which is the main source of energy in the form of ATP for living organisms
Digital analgesimeter INSIGHT von Frey Wi-Fi The von Frey Wi-Fi is a portable device used to assess tissue sensitivity to mechanical stimuli
Gilson type 10 µL polypropylene tip CRALPLAST 18261 Polypropylene to be used on eletronic von Frey apparatus, recommended for hind paw allodynia assessment
Laboratory water bath BEING INSTRUMENT BW-22P Used to heat liquid and semi-solid substances contained in appropriate recipients to specific temperature
Phosphate buffered saline SIGMA-ALDRICH 806552 A balanced salt solution buffer used for a variety of cell culture applications
Swiss mice (Mus musculus) from both gender UFSC Swiss Webster Laboratory animals used for controlled experiments
Trypanosoma evansi cryiopreserved sample UDESC Sample used to infect all mice, ceded by the Hemoparasites and Vectors Biochemistry Laboratory
Universal type 10 µL polypropylene tip CRALPLAST 18171 Polypropylene to be used on eletronic von Frey apparatus, recommended for visceral allodynia assessment

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Cite This Article
Cipriani, D. S., Borges, G. K., Miletti, L. C., Bastos-Pereira, A. L. Electronic von Frey associated with the Mouse Grimace Scale to Assess Allodynia and Pain in Trypanosoma evansi-Infected Mice. J. Vis. Exp. (209), e65743, doi:10.3791/65743 (2024).

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