Dynamic in vivo imaging of postimplantation mammalian embryos using whole embryo culture

Abstract

Due to the internal nature of mammalian development, much of the research performed is of a static nature and depends on interpolation between stages of development. This approach cannot explore the dynamic interactions that are essential for normal development. While roller culture overcomes the problem of inaccessibility of the embryo, the constant motion of the medium and embryos makes it impossible to observe and record development. We have developed a static mammalian culture system for imaging development of the mouse embryo. Using this technique, it is possible to sustain normal development for periods of 18-24 h. The success of the culture was evaluated based on the rate of embryo turning, heart rate, somite addition, and several gross morphological features. When this technique is combined with fluorescent markers, it is possible to follow the development of specific tissues or the movement of cells. To highlight some of the strengths of this approach, we present time-lapse movies of embryonic turning, somite addition, closure of the neural tube, and fluorescent imaging of blood circulation in the yolk sac and embryo.

Using a histone yellow fluorescent protein fusion for tagging and tracking endothelial cells in ES cells and mice

“…For time-lapse imaging, samples were cultured in a standard tissue culture incubator or on a microscope stage under conditions promoting the culture of mouse embryos (50% rat serum: 50% DMEM buffered with bicarbonate as described by Jones et al, 2002) and maintained under physiological conditions in a closed temperature-controlled, humidified, and oxygenated (95% air, 5% CO 2 ) chamber (Solent Sci, Portsmouth, UK, or custom made). For cytoplasmic counterstaining, samples were incubated for 10 min in Cell Tracker Orange (Molecular Probes, C-2927) at a dilution of 1:500 in dissecting or culture medium.…”

Maturation and enucleation of primitive erythroblasts during mouse embryogenesis is accompanied by changes in cell-surface antigen expression

“…These transgenic mouse lines have been useful in demonstrating a function for visceral endoderm signals in hematopoietic and endothelial development. 27,28 The human ⑀-globin::KGFP transgenic line has permitted dynamic in vivo imaging of the developing yolk sac and circulatory system 29 and quantitative analysis of hemodynamic changes during early embryogenesis. 30 Here, we have used this line to track quantitatively the differentiation and maturation of circulating EryPs during embryogenesis.…”

Multimodal imaging of mouse development: Tools for the postgenomic era

“…In these experiments, embryos were imaged for only minutes up to a few hours. Later, it was shown that robust development of early postimplantation embryos from 5.5-9.5 dpc can be supported on the microscope stage for 18 -30 hr using rich medium and limited evaporation (Jones et al, 2002(Jones et al, , 2004aSrinivas et al, 2004), conditions similar to those required for successful roller culture (Tam, 1998). Early postimplantation embryos are relatively transparent, and many of the dramatic changes that occur in the body plan can be readily imaged.…”

“…Live embryo imaging has also been used to study blood flow and dynamic changes in shear stress during vascular remodeling (Jones et al, 2002(Jones et al, , 2004b. Primitive erythroblasts labeled with GFP under the control of the Epsilon-globin promoter (Dyer et al, 2001) have been visualized in cultured 7.5-10.5 dpc embryos using confocal microscopy, providing a simple means to study early circulation and vessel formation (Jones et al, 2002;Fig.…”

“…Primitive erythroblasts labeled with GFP under the control of the Epsilon-globin promoter (Dyer et al, 2001) have been visualized in cultured 7.5-10.5 dpc embryos using confocal microscopy, providing a simple means to study early circulation and vessel formation (Jones et al, 2002;Fig. 3).…”