Van Hoeymissen Evelien, Philippaert Koenraad, Vennekens Rudi, Vriens Joris, Held Katharina
Laboratory of Endometrium, Endometriosis and Reproductive Medicine, Department of Development and Regeneration, KU Leuven; Laboratory of Ion Channel Research, VIB-KU Leuven Center for Brain and Disease Research, Leuven, Belgium and Department of Molecular Medicine, KU Leuven.
Laboratory of Ion Channel Research, VIB-KU Leuven Center for Brain and Disease Research, Leuven, Belgium and Department of Molecular Medicine, KU Leuven.
J Vis Exp. 2020 Sep 22(163). doi: 10.3791/61753.
The hippocampus is a highly organized structure in the brain that is a part of the limbic system and is involved in memory formation and consolidation as well as the manifestation of severe brain disorders, including Alzheimer's disease and epilepsy. The hippocampus receives a high degree of intra- and inter-connectivity, securing a proper communication with internal and external brain structures. This connectivity is accomplished via different informational flows in the form of fiber pathways. Brain slices are a frequently used methodology when exploring neurophysiological functions of the hippocampus. Hippocampal brain slices can be used for several different applications, including electrophysiological recordings, light microscopic measurements as well as several molecular biological and histochemical techniques. Therefore, brain slices represent an ideal model system to assess protein functions, to investigate pathophysiological processes involved in neurological disorders as well as for drug discovery purposes. There exist several different ways of slice preparations. Brain slice preparations with a vibratome allow a better preservation of the tissue structure and guarantee a sufficient oxygen supply during slicing, which present advantages over the traditional use of a tissue chopper. Moreover, different cutting planes can be applied for vibratome brain slice preparations. Here, a detailed protocol for a successful preparation of vibratome-cut horizontal hippocampal slices of mouse brains is provided. In contrast to other slice preparations, horizontal slicing allows to keep the fibers of the hippocampal input path (perforant path) in a fully intact state within a slice, which facilitates the investigation of entorhinal-hippocampal interactions. Here, we provide a thorough protocol for the dissection, extraction, and acute horizontal slicing of the murine brain, and discuss challenges and potential pitfalls of this technique. Finally, we will show some examples for the use of brain slices in further applications.
海马体是大脑中一个高度有组织的结构,是边缘系统的一部分,参与记忆的形成和巩固,以及包括阿尔茨海默病和癫痫在内的严重脑部疾病的表现。海马体具有高度的内部和相互连接性,确保与大脑内部和外部结构进行适当的通信。这种连接是通过纤维通路形式的不同信息流来实现的。在探索海马体的神经生理功能时,脑片是一种常用的方法。海马体脑片可用于多种不同的应用,包括电生理记录、光学显微镜测量以及多种分子生物学和组织化学技术。因此,脑片是评估蛋白质功能、研究神经疾病所涉及的病理生理过程以及用于药物发现目的的理想模型系统。切片制备有几种不同的方法。使用振动切片机进行脑片制备能够更好地保存组织结构,并在切片过程中保证充足的氧气供应,这比传统使用组织切片机具有优势。此外,振动切片机脑片制备可以应用不同的切割平面。在此,提供了一个成功制备小鼠脑振动切片水平海马体切片的详细方案。与其他切片制备方法不同,水平切片能够使海马体输入路径(穿通路径)的纤维在切片内保持完全完整的状态,这便于研究内嗅 - 海马体相互作用。在此,我们提供了一个关于小鼠脑解剖、提取和急性水平切片的详尽方案,并讨论了该技术的挑战和潜在陷阱。最后,我们将展示一些脑片在进一步应用中的实例。