Presented is a concise battery of mouse neonatal motor tests. Using these tests, neonatal motor deficits can be demonstrated in a variety of neonatal motor disorders. By having a standardized set of tests, results from different studies can be compared, allowing for better and accurate reporting between groups.
As the sheer number of transgenic mice strains grow and rodent models of pediatric disease increase, there is an expanding need for a comprehensive, standardized battery of neonatal mouse motor tests. These tests can validate injury or disease models, determine treatment efficacy and/or assess motor behaviors in new transgenic strains. This paper presents a series of neonatal motor tests to evaluate general motor function, including ambulation, hindlimb foot angle, surface righting, negative geotaxis, front- and hindlimb suspension, grasping reflex, four limb grip strength and cliff aversion. Mice between the ages of post-natal day 2 to 14 can be used. In addition, these tests can be used for a wide range of neurological and neuromuscular pathologies, including cerebral palsy, hypoxic-ischemic encephalopathy, traumatic brain injury, spinal cord injury, neurodegenerative diseases, and neuromuscular disorders. These tests can also be used to determine the effects of pharmacological agents, as well as other types of therapeutic interventions. In this paper, motor deficits were evaluated in a novel neonatal mouse model of cerebral palsy that combines hypoxia, ischemia and inflammation. Forty-eight hours after injury, five tests out of the nine showed significant motor deficits: ambulation, hindlimb angle, hindlimb suspension, four limb grip strength, and grasping reflex. These tests revealed weakness in the hindlimbs, as well as fine motor skills such as grasping, which are similar to the motor deficits seen in human cerebral palsy patients.
开发儿童受伤或利用啮齿类动物疾病的新车型往往是困难的,因为大鼠和小鼠的惊人的能力,迅速从神经损伤恢复。因此,为了验证任何新的儿科疾病模型,深入研究细胞和分子的变化必须去手牵手与行为的结果。在许多方面,功能行为恢复可能比在治疗或翻译相关性方面潜在的细胞的变化更为重要。随着研究人员更多地了解在成人和新生儿损伤,很显然,他们的反应是非常不同的,两者不能外推。例如,新生小鼠显示不同水平的神经生长因子,脑源性神经营养因子,神经营养因子-3和脊髓损伤后1,2胶质细胞源性神经营养因子。此外,有新生儿中风后3,D显著血脑屏障泄漏emonstrate皮层神经元重排周围神经损伤后4,并有脊髓损伤和缺血缺氧5,6延迟或减缓星形胶质细胞增生。因此,平移儿科研究使用发育等效模型和这些模型是为两个蜂窝/分子变化和年龄适当行为测试评价是很重要的。
脑瘫(CP)是一种运动障碍,影响3:1000名活产儿每年(NIH)。带CP的儿童显示出一系列的症状和共病的条件下,根据疾病的严重程度。难度与运动和协调是最常见的迹象,在达到马达发育标志延迟沿。其他症状包括肌张力异常(增加或减少),降低的精细动作技能,行走困难,流口水过多和吞咽,和语音延迟(NIH)。 CP的根本原因被认为是在芽前或围产期期间,或最多一年产后缺乏氧和/或血液流向脑部的。此外,炎症现在认为是在CP的发展的重要组成部分。
大多数CP案件与周围的脑室,被称为脑室周围白质软化症(PVL)脑白质损伤有关。这种神经标志表明,最初的侮辱导致CP期间大脑发育的时期,少突胶质细胞最容易受到侮辱发生。快速少突胶质细胞的生长在一个人的期间,也期间时少突胶质细胞是最容易受到伤害,是24之间 – 周32妊娠。在啮齿动物,等效周期是产后天7月2日至7日,并且是当CP在这个模型中被诱导。
这是用于进行此处所述的测试CP的新生小鼠模型结合缺氧和局部缺血与炎症创建一个注射URY,更好地模仿神经退行性疾病在人类CP看到。该模型解决一些在CP的其他动物模型中观察到的大的缺点,缺乏鲜明运动缺陷类似于人类CP患者,以及不同的白质损伤。通过使用相同的模型协作者以前的研究已经表明,在加入炎症增强脑白质损伤,从而更好地模仿在儿童看到具有CP 8的PVL。在以前的数据的基础上,提出了以评估在运动行为变化动物年龄的新生儿电机测试的综合电池。
使用动物模型来研究人类疾病是否存在人类和啮齿动物之间,并且所执行的行为测试有直接关系到人类的症状的细胞和分子响应之间的重叠是唯一相关。之一的儿科疾病的研究的主要问题是,许多研究者使用成年啮齿动物创建模型,以及成年啮齿动物行为的评价,在不考虑发育差异可能对疾病过程重要。由于这些问题,重要的是在儿科疾病使用研究不仅适当调整的发育时间点( 例如,人的中枢神经系统发育,在28 – 51周内,相当于一个产后第2天-第7天的啮齿动物)7,而且行为测试,将审查适当的运动,感觉或反射性发育行为。因此,因为每个新的新生儿疾病模型开发,它必须经过严格的测试,以确保蜂窝和行为反应将提供啮齿类动物和人类之间最合适的翻译资料。
脑瘫是一种运动障碍,它持续到成年。许多当今脑瘫模型的一个问题是缺乏可重复的,标准化的电机测试,可以在儿科患者看到的赤字相关的。在这种新的模式,它结合缺氧,缺血和炎症的新生小鼠,运动行为,用具体的新生小鼠试验电池进行评估。为了减小主体和增加定量报告,几个测试已被修改,以包括非常具体的,但容易评估,可以标准化的措施。此外,前端和后肢评价可以单独执行,并且左/右差异可被确定。测试该电池是特异于新生小鼠最多两个周龄。
这种CP模型演示行走困难(行走,后肢足角),以及肢体特异性无力(4肢悬浮,后肢悬挂),和赤字发育反射(抓握反射)。尽管在本研究中检测只有一个时间点,这些缺陷可以随时间被跟踪。
有测试其他电池可以在新生儿中使用,如测试的福克斯的电池或发育里程碑15的Heyser的评价。然而,这些测试的新生儿进行比较的成人,其响应可能是不一样的,因为新生儿仍在发展。福克斯的电池和Heyser的Assement测试依赖于观测主观信息用二分(是或否)的评估,而不是客观的数据(角度,姿势的基础上强度等 )。由于这些测试的主观性,许多科学家已经适应,添加或删除的标准,从而使他们的结果无比给他人,并限制ING在对于特定疾病或病症建立基线赤字方面的数据的有用性。通过建立一组标准化电机测试是定性和专为测试新生儿,从单个研究小组结果能够精确和可靠地报告和比较。
The authors have nothing to disclose.
We would like to thank everyone at Shriners Hospital Pediatric Research Center, in particular Dr. Mickey Seltzer, of whom without his support, this work would not have been funded. In addition, we would like to thank Isha Srivastava, who contributed to early data collection and Amy He, who helped with the figures. This study was funded by Shriners Hospitals for Children. No funding source played a role in experimental design or decision to submit the paper for publication.
C57BL/6 mice | Charles River Laboratories | STRAIN CODE: 027 | C57BL/6NCrl is the exact strain we use |
Anesthesia Dish, PYREX™ Crystallizing Dish | Corning Life Sciences Glass | 3140125 | Capacity: 25.03 oz. (740mL); Dia. x H: 4.92 x 2.55 in. (125 x 65mm). However, any small round glass container will work. A 2 cup capacity pyrex food storage bowl with flat bottom will also work and is much cheaper (Pyrex model number: 6017399). |
Covered lead ring | Fisher Scientific | S90139C | Lead ring for stablizing flasks in a water bath. It is used inside the anesthesia dish. |
Scalpel Blade #11 | World Precision Instrucments, Inc. | 500240 | |
Small Vessel Cauterizer | Fine Science Tools | 18000-00 | |
Micro Hook | Fine Science Tools | 10064-14 | |
Vetbond Suture Glue | 3M | 1469SB | n-butyl cyanoacrylate adhesive |
Lipopolysaccharide | Sigma Life Science | L4391 | Lipopolysaccaride from e.coli 0111:B4, gamma irradiated |
12×12 inch opaque box | Acrylic Display Manufacturing: A division of Piasa Plastics | C4022 | Colored Acrylic 5-Sided Cube, 3/16" Colored Acrylic, 12"W x 12"D x 12"H; http://www.acrylicdisplaymfg.com/html/cubes_19.html |
Camera/camcorder | JVC | GC-PX100BUS | Any camcorder that works well in low light and can be imported and edited. We use the JVC GC-PX100 Full HD Everio Camcorder. |
Covidien Tendersorb™ Underpads | Kendall Healthcare Products Co | 7174 | |
WypAll L40 | Kimberly-Clark Professional | 5600 | Any surface with moderate grip will do |
Surface at 45 degree incline | We use a cardboard box. | ||
Thin wire from a pipe cleaner | Creatology | M10314420 | Any pipe cleaner from any craft store will work. |
50mL conical tube | Falcon | 352070 | |
Fiberglass Screen Wire | New York Wire www.lowes.com | 14436 | Any supplier can be used as long as their screen is 16×16 or 18×16 |
Razor blade | Fisherbrand | 12-640 | A wooden stick applicator or wooden part of a cotton-tipped swab will also work. |
OPTIX 24-in x 4-ft x 0.22-in Clear Acrylic Sheet to make Clear Acrylic Walkway | PLASKOLITE INC | 1AG2196A | Clear acrylic (1/8" thick) with sides and a top to limit exploration. We bought a sheet of acrylic from a local hardware store and had them cut it to size. (2) 2"x2"; (3) 2"x 18"; (1) 2"x15.5"; (1) 2"x3". Using clear tape, tape all sides together, with the 15.5" piece on top. Tape the 3" piece to the end of the 15.5" piece to create a flap/entryway for the mice. Alternatively, part or all of the walkway can be glued together, and only taping on the top pieces. This design will allow for the walkway to be opened for easy cleaning. |
Protractor | Westscott | ACM14371 |