Miyata Takaki, Kawaguchi Ayano, Saito Kanako, Kuramochi Hiroshi, Ogawa Masaharu
Laboratory for Cell Culture Development, Advanced Technology Development Center, Brain Science Institute, RIKEN, Wako, Saitama, Japan.
J Neurosci Res. 2002 Sep 15;69(6):861-8. doi: 10.1002/jnr.10335.
Slice culture combined with the use of fluorescent dyes and/or the introduction of fluorescent protein genes provides live and three-dimensional information on cytogenetic and histogenetic events at the level of the individual cell. Using slices prepared from midembryonic mouse cerebral wall tissue upon which fine DiI crystals were placed on the pial or ventricular surface, we recently found that dividing progenitor cells do not lose their pia-connected (basal) processes and that the processes are inherited by daughter cells, including neurons (Miyata et al. [2001] Neuron 31:727-741). To understand more fully the biological significance of this inheritance process, the fate of each daughter cell should be monitored over a culture period extended long enough to allow a neuron to migrate up to the cortex or for a progenitor to proceed to the next round of division. Exposure of slices to 40%, instead of 20%, O(2) significantly improved their overall thickening, cell production, and layer formation and also provided better spatial resolution by preventing the loss of transparency that accompanies cell death.
切片培养结合荧光染料的使用和/或荧光蛋白基因的导入，可在单个细胞水平上提供有关细胞遗传学和组织遗传学事件的实时三维信息。利用从中胚层小鼠脑壁组织制备的切片，在软脑膜或脑室表面放置精细的DiI晶体，我们最近发现，正在分裂的祖细胞不会失去其与软脑膜相连的（基底）突起，并且这些突起会被包括神经元在内的子细胞继承（Miyata等人，[2001]《神经元》31:727 - 741）。为了更全面地理解这种继承过程的生物学意义，应在足够长的培养期内监测每个子细胞的命运，以使神经元迁移到皮质或使祖细胞进入下一轮分裂。将切片暴露于40%的氧气（而不是20%）中，显著改善了它们的整体增厚、细胞生成和层形成，并且通过防止伴随细胞死亡而出现的透明度丧失，还提供了更好的空间分辨率。
