Kawaue Takumi, Sagou Ken, Kiyonari Hiroshi, Ota Kumiko, Okamoto Mayumi, Shinoda Tomoyasu, Kawaguchi Ayano, Miyata Takaki
Department of Anatomy and Cell Biology, Nagoya University Graduate School of Medicine, 65 Tsurumai, Showa-ku, Nagoya, 466-8550, Japan.
Dev Growth Differ. 2014 May;56(4):293-304. doi: 10.1111/dgd.12131. Epub 2014 Apr 9.
To achieve highly sensitive and comprehensive assessment of the morphology and dynamics of cells committed to the neuronal lineage in mammalian brain primordia, we generated two transgenic mouse lines expressing a destabilized (d4) Venus controlled by regulatory elements of the Neurogenin2 (Neurog2) or Gadd45g gene. In mid-embryonic neocortical walls, expression of Neurog2-d4Venus mostly overlapped with that of Neurog2 protein, with a slightly (1 h) delayed onset. Although Neurog2-d4Venus and Gadd45g-d4Venus mice exhibited very similar labeling patterns in the ventricular zone (VZ), in Gadd45g-d4Venus mice cells could be visualized in more basal areas containing fully differentiated neurons, where Neurog2-d4Venus fluorescence was absent. Time-lapse monitoring revealed that most d4Venus(+) cells in the VZ had processes extending to the apical surface; many of these cells eventually retracted their apical process and migrated basally to the subventricular zone, where neurons, as well as the intermediate neurogenic progenitors that undergo terminal neuron-producing division, could be live-monitored by d4Venus fluorescence. Some d4Venus(+) VZ cells instead underwent nuclear migration to the apical surface, where they divided to generate two d4Venus(+) daughter cells, suggesting that the symmetric terminal division that gives rise to neuron pairs at the apical surface can be reliably live-monitored. Similar lineage-committed cells were observed in other developing neural regions including retina, spinal cord, and cerebellum, as well as in regions of the peripheral nervous system such as dorsal root ganglia. These mouse lines will be useful for elucidating the cellular and molecular mechanisms underlying development of the mammalian nervous system.
为了对哺乳动物脑原基中神经元谱系细胞的形态和动态进行高度敏感且全面的评估,我们构建了两种转基因小鼠品系,它们表达由Neurogenin2(Neurog2)或Gadd45g基因的调控元件控制的不稳定型(d4)金星荧光蛋白。在胚胎中期的新皮质壁中,Neurog2-d4Venus的表达大多与Neurog2蛋白的表达重叠,只是起始时间稍有延迟(1小时)。虽然Neurog2-d4Venus和Gadd45g-d4Venus小鼠在脑室区(VZ)表现出非常相似的标记模式,但在Gadd45g-d4Venus小鼠中,在含有完全分化神经元的更基底部区域可以看到细胞,而这些区域没有Neurog2-d4Venus荧光。延时监测显示,VZ中大多数d4Venus(+)细胞有延伸至顶端表面的突起;其中许多细胞最终缩回其顶端突起并向基底迁移至室下区,在那里可以通过d4Venus荧光对神经元以及经历产生终末神经元的分裂的中间神经源性祖细胞进行实时监测。一些d4Venus(+) VZ细胞反而进行核迁移至顶端表面,在那里它们分裂产生两个d4Venus(+)子细胞,这表明在顶端表面产生神经元对的对称终末分裂可以可靠地进行实时监测。在包括视网膜、脊髓和小脑在内的其他发育中的神经区域以及背根神经节等周围神经系统区域也观察到了类似的谱系定向细胞。这些小鼠品系将有助于阐明哺乳动物神经系统发育的细胞和分子机制。