Fate mapping the avian epiblast with focal injections of a fluorescent-histochemical marker: ectodermal derivatives.
Fate mapping the avian epiblast with focal injections of a fluorescent-histochemical marker: ectodermal derivatives.
复制标题
通过局部注射荧光组织化学标记物（外胚层衍生物）来绘制鸟类外胚层的命运图。
DOI：
10.1002/jez.1402550309
复制
发表时间：
1990
期刊：
影响因子：
--
通讯作者：
Sheard,P
中科院分区：
文献类型：
--
作者：
Schoenwolf,GC;Sheard,P
描述了一种用于禽胚外胚层命运定位的显微注射技术。它包括用荧光组织化学标记物注射培养胚皮的外胚层，在活体胚皮注射时检测罗丹明荧光，并在注射后24小时收集的同一胚胎的组织学切片上检测辣根过氧化物酶活性。我们的研究结果表明，这一过程常规地标记细胞，从而确定它们的命运，并构建未来的命运图。两个这样的图谱是关于外胚层的外胚层衍生物的：一个是关于Hensen淋巴结进展的晚期（3c到4阶段），另一个是关于淋巴结退化的早期阶段（4 +到5阶段）。这些新地图有六个重要特征。首先，他们表明，无论淋巴结是前进还是后退，平坦的神经板向Hensen淋巴结中心延伸至少300 μm，向两侧延伸300 μm，向尾侧延伸1mm。其次，它们证实了我们之前基于鹌鹑/小鸡嵌合体的命运映射研究。也就是说，它们表明外胚层的结前中线区域形成了前脑的基底和视神经泡的腹外侧部分以及MHP细胞（即，主要是包含在弯曲神经板的中间铰链点内的楔形神经上皮细胞）；相反，结旁区和结后区贡献L细胞（即，主要是构成神经板外侧的纺锤形神经上皮细胞）。第三，他们揭示了MHP细胞的第二个来源，Hensen淋巴结，验证了先前基于氚化胸腺嘧啶标记的其他研究。第四，与其他基于活体染色、碳标记和绒毛膜尿囊移植的研究相反，但与我们之前基于鹌鹑/小鸡嵌合的研究一致，他们证明，在扁平神经板阶段，促进神经管的四个颅足部分支（即前脑、中脑、后脑和脊髓）的细胞尚未在空间上彼此分离。尽管更多的颅神经板细胞倾向于形成更多的颅分支，更多的尾侧细胞倾向于形成更多的尾侧分支。因此，单次注射常规标记多个神经管细分。讨论了我们目前的结果与别人以前的结果不一致的可能原因。第五，他们认为，在平面神经板阶段，特别是在尾端结后区域，贡献于表面外胚层和神经板的细胞在空间上还没有完全分离。第六，他们描绘了这些位点的位置。因此，显微注射技术为鸟类外胚层，特别是神经板及其分支提供了重要的新信息。
A microinjection technique is described for fate mapping the epiblast of avian embryos. It consists of injecting the epiblast of cultured blastoderms with a fluorescent‐histochemical marker, examining rhodamine fluorescence at the time of injection in living blastoderms, and assaying for horseradish peroxidase activity in histological sections obtained from the same embryos collected 24 h postinjection. Our results demonstrate that this procedure routinely marks cells, allowing their fates to be determined and prospective fate maps to be constructed. Two such maps are presented for ectodermal derivatives of the epiblast: one for late stages of Hensen's node progression (stages 3c through 4) and one for early stages of node regression (stages 4 + through 5). These new maps have six significant features. First, they show that regardless of whether the node is progressing or regressing, the flat neural plate extends at least 300 μm cranial to, 300 μm bilateral to and 1 mm caudal to the center of Hensen's node. Second, they confirm our previous fate mapping studies based on quail/chick chimeras. Namely, they show that the prenodal midline region of the epiblast forms the floor of the forebrain and the ventrolateral part of the optic vesicles as well as MHP cells (i.e., mainly wedge‐shaped neurepithelial cells contained within the median hinge point of the bending neural plate); in contrast, paranodal and postnodal regions contribute L cells (i.e., mainly spindle‐shaped neurepithelial cells constituting the lateral aspects of the neural plate). Third, they reveal a second source of MHP cells, Hensen's node, verifying previous studies of others based on tritiated thymidine labeling. Fourth, they demonstrate, in contrast to studies of others based on vital staining, carbon marking, and choriolallantoic grafting but in accordance with our previous studies based on quail/chick chimeras, that the cells contributing to the four craniocaudal subdivisions of the neural tube (i.e., forebrain, midbrain, hindbrain, and spinal cord) are not yet spatially segregated from one another at the flat neural plate stage, although more cranial neural plate cells tend to form more cranial subdivisions and more caudal cells tend to form more caudal subdivisions. Thus, single injections routinely mark multiple neural tube subdivisions. Probable reasons for the discrepancy between our present results and the previous results of others is discussed. Fifth, they suggest that cells contributing to the surface ectoderm and neural plate are not yet completely spatially segregated from one another at the flat neural plate stage, particularly in caudal postnodal regions. Sixth, they delineate the locations of the otic placodes. Therefore, the microinjection technique has contributed important new information about the avian epiblast and, in particular, about the neural plate and its subdivisions.