在多发性硬化症的小鼠模型测量进行性神经功能残疾
Summary
An optimized testing protocol is presented in this paper for the Rotarod performance test, used for measuring progressive neurological disability in TMEV-infected mice.
Abstract
After intracerebral infection with the Theiler’s Murine Encephalomyelitis Virus (TMEV), susceptible SJL mice develop a chronic-progressive demyelinating disease, with clinical features similar to the progressive forms of multiple sclerosis (MS). The mice show progressive disability with loss of motor and sensory functions, which can be assessed with multiple apparatuses and protocols. Among them, the Rotarod performance test is a very common behavioral test, its advantage being that it provides objective measurements, but it is often used assuming that it is straightforward and simple. In contrast to visual scoring systems used in some models of MS, which are highly subjective, the Rotarod test generates an objective, measurable, continuous variable (i.e., length of time), allowing almost perfect inter-rater concordances. However, inter-laboratory reliability is only achieved if the various testing parameters are replicated. In this manuscript, recommendations of specific testing parameters, such as size, speed, and acceleration of the rod; amount of training given to the animals; and data processing, are presented for the Rotarod test.
Introduction
Theiler鼠脑脊髓炎病毒（TMEV）是一种嗜神经单链RNA病毒持续感染的鼠的中枢神经系统（CNS）。在易感小鼠中，感染TMEV导致免疫介导的慢性进行性脱髓鞘疾病，称为TMEV诱导脱髓鞘疾病（TMEV-IDD）。小鼠的实验性感染发生的疾病过程类似，在多发性硬化症（MS）的渐进形式看到。 TMEV-IDD的特征在于两个不同的阶段：急性期和慢性期。急性期是一种温和的，通常亚临床性脑炎1,2。第二，慢性期，开始约一个月感染后，由一个缓慢进展残疾特征在于脱髓鞘，炎症和轴索损害1,2。在小鼠中观察到的疲软与痉挛，偶尔严重的强直性痉挛有关。
由于目前还没有medicatioNS改善逐步残疾患者，研究人员特别是TMEV，国际直拨电话，它代表了监测疾病修饰药物对疾病进展的影响，最佳的动物模型所吸引。然而，在小鼠中，以及在MS患者，残疾进展的监测需要在延长的时间周期的连续的临床观察。在小鼠中，对于残疾进展的长期监测可与旋转杆的性能测试来完成。
在转棒性能测试是一种标准的行为测试，用于评估运动相关的功能，例如协调性，平衡，和疲劳在啮齿类动物。小鼠必须保持平衡的转弯杆，这是在连续加速旋转;落入时间延迟从该杆被记录。与神经功能障碍的动物不能长留在旋转杆作为对照，他们当转速超过正常脱落的电机容量。更多的神经功能缺损动物们，他们越早脱落杆，以及更短的时间延迟。
旋转试验在传统的视觉评分系统的优点在于，它产生一个目标，可测量的可变时间延迟该最终可用于统计分析以量化疗法和实验程序3的效果。
在免疫学（LONI）中的达特茅斯实验室，将小鼠进行的适配协议，在那里它们被TMEV感染前测试，以便与机器使他们熟悉并评估其正常的“基线”平衡协调和电机控制4， 5。一旦基线建立和小鼠被感染TMEV，它们过了一段数月测定一次或每周两次。实际测试协议为期150天的平均值，从而允许进行评估平衡，协调和电机控制的降幅比脱髓鞘疾病的全过程。
几百TMEV，IDD和假治疗的小鼠迄今已测试达特茅斯神经功能障碍。这些小鼠接受了各种免疫调节治疗方法，但没有药剂已发现可有效地改善残疾进展6,7。本文章及相关协议描述了如何通过定性TMEV-IDD小鼠显示进行性神经损害。特别是，该协议提供的认为是一般适合学习使用旋转试验TMEV-IDD小鼠神经失具体的测试参数的建议。此过程提供对照评估一个基线（1）该小鼠模型来进行性MS和（2）它的实用性，用于测试旨在治疗进行性神经疾病如MS疗法的相关性。显然，转棒性能测试和当前优化测试参数和协议不仅在该TMEV-IDD小鼠模型检测进行性神经障碍是有用的，但是，可以在中神经变性疾病的其他病毒诱导的和/或遗传小鼠模型揭示损伤是有用的。
Protocol
Representative Results
Discussion
尽管有一些限制，所述旋转杆的性能测试代表用于评估TMEV-IDD运动功能和功能障碍，以及药理干预对残疾进展中的小鼠的影响的重要工具。
旋转试验是在1957年首次描述为在啮齿类动物11测量神经功能缺损的工具。鼠害必须走在旋转的杆，随着转速，并尽量避免坠落到地面。落入延迟被记录并用作定量终点运动功能：越神经功能缺损的动物有，越早它们脱落杆。此测试?…
Disclosures
The authors have nothing to disclose.
Acknowledgements
The authors thank the staff of the Center for Comparative Medicine and Research (CCMR) at Dartmouth for their expert care of the mice used for these studies. The authors also acknowledge Emily Clough for her excellent administrative support.
Materials
| Mice SJL/JCrHsd 4 to 6 week old | Envigo | #052 | |
| TMEV virus stock | |||
| Isoflurane vaporizer | Harvard Apparatus | #340471 | |
| Insulin Syringes U- 100 29g x 0.5cc | BD | #328203 | |
| Rotamex-5 4 Lane Rota-Rod for Mice with RS-232 and Software | Columbus Instruments | #0890M |
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