Fig. 1. Diagrams of experimental manipulations of axolotl gastrulae. (A) A tilted dorsal vegetal view of an intact axolotl embryo at stage 10 reveals the state of the bottle cell apices (arrows) at the time of microsurgical manipulation. (B) Lateral view of the axolotl early gastrula fate map according to Pasteels (Pasteels, 1942). Dorsal is right. An, animal; bp, blastopore; ec, ectoderm; en, endoderm; lp, lateral plate mesoderm; nc, notochord; ne, neurectoderm; pcm, prechordal mesoderm; pe, pharyngeal endoderm; s, somitic mesoderm. (C) Dorsal view of the axolotl early gastrula fate map (Pasteels, 1942). Colors indicate tissues as in B. The broken black lines divide the spherical embryo at 30° intervals, both in latitude and longitude. The broken white line indicates the region of the explants shown in Ea, Eb and Ec, extending from 45° (the level of the blastopore is bp) to 110° (the animal extent of the axial mesoderm), and 30° bilaterally from the midline. (D) Keller sandwiches were constructed from paired dorsal marginal zones and allowed to develop intact until the hatchling larval stage (a). Keller endomesodermal isolates were generated (b) by removing the converging and extending endomesoderm (pink and blue regions below broken line) at early (b1) and later (b2) stages of gastrulation. (Ea) Single dorsal lip explants were trimmed to the size of the presumptive axial mesoderm (pink) and pharyngeal endoderm (blue) according to published fate maps. The predicted endodermal and mesodermal regions were then separated microsurgically (see Materials and Methods section for details). (Eb) Keller sandwiches were constructed, as in D, trimmed to the size of the future axial mesoderm and pharyngeal endoderm, and subdivided into suprablastoporal endoderm (blue) and more animal axial mesoderm (pink). (c) Fused dorsal lip explants were created by first trimming Keller sandwiches, and then subdividing them into even thirds. The most animal axial mesoderm and suprablastoporal endoderm were either raised separately (c1) or allowed to fuse (c2). The intermediate portions, containing the imprecise border between endoderm and mesoderm, were also raised separately (c1,c2).