Summary

视神经横断:一个成人在中枢神经系统的神经元凋亡模型

Published: May 12, 2011
doi:

Summary

视神经横断是一个成人中枢神经系统损伤的广泛使用的模型。视网膜神经节细胞的90%(视网膜神经节细胞),其轴突完全横断(干切断)干切断后14天之内死亡。这种模式很容易经得起实验操作和高度重现。

Abstract

视网膜神经节细胞(RGC的)是中枢神经系统的神经元,输出的视觉信息从视网膜到大脑,通过视神经。视神经眼轨道内可以访问,并完全切断(axotomized),切割研资局在整个人口的轴突。视神经横断重现模型在1-4成人中枢神经系统的细胞凋亡的神经细胞死亡。这种模式是特别有吸引力,因为眼睛的玻璃体腔行为作为药物输送到视网膜的胶囊,允许通过人工注射的实验操作。通过玻璃体液体化学品的扩散,确保他们的行为后,整个研资局人口。此外,视网膜神经节细胞可以选择性地转用短干扰RNA(siRNAs)的质粒或病毒载体的视神经5-7或注入自己的目标,上丘8向量的切端。这使研究人员能够学习所需的神经元群的细胞凋亡的机制,没有其他旁观者神经元或神经胶质细胞周围的混杂影响。另外一个好处是与细胞的存活,可在伤后量化的易用性和准确性。视网膜是一个平面,分层组织和RGC的是在最里层,神经节细胞层的本地化。随着时间的推移,可跟踪的视网膜神经节细胞的生存,申请时间干切断视神经切开结束了荧光示踪(3%荧光金),或上丘(研资局的目标)前一周注入示踪剂干切断。示踪逆行运输,标签研资局在整个人口。由于神经节细胞层是单层(一个细胞厚),研资局的密度可量化的扁平式的组织中,无体视需要。视神经横断导致的90%受伤的视网膜神经节细胞的凋亡,14天以内 postaxotomy 9-11。研资局凋亡有一个特点的时间进程,使细胞死亡是推迟3-4天postaxotomy后,这些细胞迅速退化。这提供了一个针对在细胞凋亡有关的途径的实验操作的时间窗口。

Protocol

1。外科技术使用无菌技术操作,并按照您的特定机构的动物使用协议,应进行实验。活体组织接触到的工具和材料(解决方案,测试物质,示踪剂,针等)必须是无菌的,以防止感染动物福利和研究的潜在的负面影响和不利影响。 2。麻醉老鼠将被麻醉的兽医异氟醚蒸发器系统。使用医疗级氧气在0.8 L / min的异氟醚气体蒸发的速度。放置在附加麻醉中的动…

Discussion

这个手术过程中有许多变化,在本议定书中的几个步骤是没有必要的。它是只需要收回,覆盖在视神经以获得神经肌肉。然而,这个结果在一个非常有限的工作空间周围神经的横断关键的最后阶段更加困难。在某些情况下是可取的转染细胞和视神经残端回缩所有眼外肌,泪腺是有利于在这种情况下所提供的增加的存取空间。

在协议中最重要的步骤是步骤4.3-4.6。重要的是,不?…

Declarações

The authors have nothing to disclose.

Acknowledgements

PDK是支持由CIHR经营授予(澳门币86523)

Materials

Material Name Tipo Company Catalogue Number Comment
Stereotaxic Frame   Stoelting, Kopf, WPI    
Rat Gas Mask   Stoelting, Kopf, WPI    
Anesthesia System   VetEquip 901806  
Isoflurane (PrAErrane)   Baxter Corp DIN 02225875  
Surgical Microscope   WPI, Zeiss, Leica    
Fluorogold -(Hydroxystilbamidine bis(methanesulfonate)   Sigma 39286  
Gelfoam   Pharmacia & Upjohn    
Tears Naturale P.M.   Alcon    
Proviodine   Medline MDS093945H  
Vannas spring scissors   Fine Science Tools 15000-00  
Fine tip Dumont forceps   Fine Science Tools 11252-00  
Micro surgical hook   Fine Science Tools 10062-12  
Eye dressing serrated forceps   Fine Science Tools 11152-10  
Dumont #7b sharp curved serrated forceps   Fine Science Tools 11270-20  
Cauterizer   Fine Science Tools 18010-00  

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Magharious, M. M., D’Onofrio, P. M., Koeberle, P. D. Optic Nerve Transection: A Model of Adult Neuron Apoptosis in the Central Nervous System. J. Vis. Exp. (51), e2241, doi:10.3791/2241 (2011).

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