1. Vertebrate Embryonic Patterning 1
Amphibian Gastrulation: Classical Observations & Experiments

2. Amphibian Development
Fertilization initiates cortical rotation
Cortical rotation relocalizes determinants
These determinants act to specify the dorsal-most regions of the embryo

3. Establishing Asymmetry
Localized mRNAs

4. Early Cleavages in Xenopus
gray crescent
Tiers A B C D

5. Fate Map of Xenopus Blastula

6. Primary Tissue Layers Ectoderm Mesoderm Endoderm skin, neural
notochord, somites, blood, kidney
Endoderm
gut, prechordal plate

7. Gastrulation Movements

8. Xenopus Gastrulation Movements

9. Gastrulation (again)

12. Xenopus
laevis

13. Classic Experiments Hans Spemann (1903)
Constriction of cleavage stage embryos & fertilized eggs
Demonstrated asymmetry in fertilized egg

14. Embryo Constriction Experiment

15. Egg Constriction Experiment
Constriction of the egg becomes the 1st cleavage plane. Constriction of egg has to bisect the gray crescent in order for twinning to occur. If division occur perpendicular to this, only ½ of the egg develops into an embryo.

16. Fate Map of Gray Crescent
dorsal blastopore lip
point where gastrulation begins
What is it about the gray crescent that makes it so important?

17. Spemann’s Specification Experiments
Presumptive neural plate ectoderm in the early gastrula was specified as ectoderm, but undetermined.
Later gastrula neuroectoderm was determined to a neural fate.

18. Dorsal Lip Transplantation

19. Perturbations of Axis Formation: Making Two Dorsal Axes

20. Cool Perturbations of Normal Development
Ventralization:
UV irradiation of vegetal pole shortly after fertilization
blocks cortical rotation by crosslinking cytoskeletal proteins
DAI Scale – rating of 0 to 5 of dorsal inhibition
0 is normal, 5 is completely ventralized
embryo develops as a lump of endoderm & ectoderm

21. More Cool Perturbations
Anterior/Dorsalization:
LiCl treatment
increases the amount of anterior/head tissue at the expense of posterior-dorsal mesoderm & ventral tissues
Posterior/Ventralization
Retinoic Acid treatment
decreases anterior/head structures

22. Implications
Strategies for cloning genes involved in dorsal/ventral specification
Subtractive library screens between dorsalized and ventralized embryos
Functional library screens
Rescue of ventralized embryos
Induction of secondary axis

23. Properties of Dorsal Lip Region
Autonomously specified
Dorsal lip never respecified by tissue at location of transplantation
Donor cells always contribute to notochord (mesoderm)
Secondary neural tube forms from recruited host cells
Donor lip cells organize host cells

24. Mesoderm Induced by Vegetal Cells

25. Nieuwkoop’s Experiment

26. Nieuwkoop’s Center Rescues Dorsal Axis in UV & Duplicates Axis in Wild-type

27. Inducing Centers in the Blastula
“Spemann”
(General)

28. Fate Map of Organizer and Nieuwkoop’s Center