Pulse labeling and long-term tracing of newborn neurons in the adult subgranular zone
Pulse labeling and long-term tracing of newborn neurons in the adult subgranular zone
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DOI：
10.1038/cr.2010.141
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发表时间：
2011-02-01
期刊：
影响因子：
44.1
通讯作者：
Xiong, Zhi-Qi
中科院分区：
文献类型：
--
作者：
Cheng, Xuewen;Li, Yang;Xiong, Zhi-Qi
过去几十年的研究表明，成人大脑产生神经祖细胞，这些细胞增殖并分化为新生神经元，这些新生神经元整合到现有的回路中。然而，由于标记和操纵神经祖细胞和新生神经元的可用方法的限制，新产生的神经元的详细分化过程和潜在机制在很大程度上是未知的。在这项研究中，我们设计了一个严格控制的，非侵入性的系统的基础上Cre/loxP重组，以实现长期跟踪和遗传操作的成年神经元在体内。在该系统中，他莫昔芬诱导型重组酶CreER（T2）由基于BAC的双皮质素（DCX，新生神经元的标志物）启动子驱动。通过将Cre细胞系与报告基因小鼠杂交，我们发现齿状回（DG）中的新生神经元可以被他莫昔芬诱导的黄色荧光蛋白（YFP）表达选择性脉冲标记。YFP阳性神经元通过与细胞类型特异性标记物的共免疫染色来鉴定，并通过电生理记录来表征。此外，这些神经元的迁移分析表明，这些标记的神经元大多数迁移到颗粒细胞层的内部。此外，新生颗粒神经元内分子层的棘状生长呈倒U型动态变化，而外分子层的棘状生长呈楔形变化。我们的转基因工具提供了一种有效的方法来选择性地标记和操纵成年小鼠DG中的新生神经元。
Research over the past decades has demonstrated that adult brain produces neural progenitor cells which proliferate and differentiate to newborn neurons that integrate into the existing circuit. However, detailed differentiation processes and underlying mechanisms of newly generated neurons are largely unknown due to the limitation of available methods for labeling and manipulating neural progenitor cells and newborn neurons. In this study, we designed a tightly controlled, noninvasive system based on Cre/loxP recombination to achieve long-term tracing and genetic manipulation of adult neurons in vivo. In this system, tamoxifen-inducible recombinase, CreER(T2), was driven by BAC-based promoter of doublecortin (DCX, a marker of newborn neurons). By crossing this Cre line with reporter mouse, we found that newborn neurons in the dentate gyrus (DG) could be selectively pulse-labeled by tamoxifen-induced expression of yellow fluorescent protein (YFP). YFP-positive neurons were identified by coimmunostaining with cell type-specific markers and characterized by electrophysiological recording. Furthermore, analysis of the migration of these neurons showed that the majority of these labeled neurons migrated to the inner part of granule cell layer. Moreover, spine growth of inner molecular layer of newborn granule neurons takes a dynamic pattern of invert U-shape, in contrast to the wedge-shaped change in the outer molecular layer. Our transgenic tool provides an efficient way to selectively label and manipulate newborn neuron in adult mouse DG.