Acute dissociation for analyses of NMDA receptor function in cortical neurons during aging
Acute dissociation for analyses of NMDA receptor function in cortical neurons during aging
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DOI：
10.1016/s0165-0270(03)00196-1
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发表时间：
2003-10-15
影响因子：
3
通讯作者：
Magnusson, KR
中科院分区：
文献类型：
--
作者：
Kuehl-Kovarik, MC;Partin, KM;Magnusson, KR
本研究旨在确定是否在单个皮层神经元水平上可检测到NMDA ε 2（NR2B）亚基的功能年龄差异。使用蛋白酶K和胰蛋白酶的组合，然后手动研磨，从年轻的成年人，中年人，或老年小鼠的额叶和前额叶皮质急性分离神经元。过夜培养后，使用膜片钳电生理学和快速灌注获得对300 μ M NMDA的全细胞反应，有或没有有效的NR2B拮抗剂艾芬地尔。从所有年龄段的动物中分离健康的相亮皮质神经元。细胞直径和电容在年龄之间是一致的。我们能够进行动力学分析的NMDA诱发的反应，并表现出显着增加的失活率随着年龄的增长。此外，我们观察到高浓度的艾芬地尔对老年动物的NMDA诱发反应的显着影响。因此，这种方法对于从年轻和年老动物的大脑中分离神经元是理想的，并且为在单个细胞水平上进行功能分析提供了强大的工具。(C)2003 Elsevier B.V.保留所有权利。
The present study was designed to determine if functional age differences in the NMDA epsilon2 (NR2B) subunit were detectable at the level of individual cortical neurons. Neurons were acutely dissociated from the frontal and prefrontal cortices of young adult, middle-aged, or old mice, using a combination of proteinase K and trypsin followed by manual trituration. After overnight culture, patch-clamp electrophysiology and rapid perfusion were used to obtain whole-cell responses to 300 muM NMDA, with or without the potent NR2B antagonist ifenprodil. Healthy, phase-bright cortical neurons were isolated from animals of all ages. Cell diameter and capacitance was consistent between ages. We were able to perform kinetic analyses of the NMDA-evoked response, and demonstrated a significant increase in the rate of deactivation with increasing age. In addition, we observed a significant effect of high-concentration ifenprodil on the NMDA-evoked response in old animals. Thus, this method is ideal for the dissociation of neurons from the brain of both young and old animals, and offers a powerful tool for functional analysis at the level of the individual cell. (C) 2003 Elsevier B.V. All rights reserved.