Fig. 4. The dorsal mesoderm of Zoep;ace embryos undergoes massive cell death at the end of gastrulation. (A-D) Lateral views (tail region) of live embryos at 30 hours of development stained with acridine orange. Dead cells stain green. The genotype, which is inferred from the phenotype and statistical analysis is indicated at the top right. (E,F) Views from the dorsoposterior region of live embryos at the shield stage (the inferred genotype is indicated top right). (G,H) Confocal sections of live embryos injected at the tail-bud stage with Annexin V Alexa 488 15 minutes before inspection (the inferred genotype is indicated top right). (I-M) Live Zoep/ace embryos inspected with Nomarski optics. (I) Focus on the dorsal (axial) and dorso-lateral hypoblast at tailbud stage, presented as three overlapping panels because of the curvature of the underlying yolk ball. Axially, notochord does not form; instead there is a large mass of dying cells that have morphology typical of apoptotic cells. Paraxially (left), 100-160 µm away from the axis, dead cell bodies of various sizes are also found (boxed region; enlarged in K) next to live cells of typical migrating (converging) non-axial hypoblast cell morphology (black arrowheads). (J) 4-somite stage, parasagittal view (caudal to the lower right) through the neuroectoderm, somitic mesoderm, yolk syncytial layer and yolk core. The inset shows the yolk syncytial layer and yolk portion appearing at a shallower focus. Both focal planes reveal numerous cell corpses (arrows) in the yolk syncytial layer, some of which abut and penetrate the underlying yolk core. (K-L) Higher-magnification views of cell corpses. (K) Dead cell bodies in the paraxial/lateral hypoblast at tailbud stage, from panel I inset. (L) Cell corpses (arrows) at 4-somite stage inside the yolk syncytial layer, one of them (left) indenting the underlying yolk mass. A yolk syncytial layer nucleus (white arrowhead), seen here slightly out of focus behind a dead cell body, is in focus in panel M. Abbreviations: n, neuroectoderm; sm, somitic mesoderm; y, yolk core; ysl, yolk syncytial layer. Scale bar: 20 µm.