Vertebrate Embryonic Patterning 1 Amphibian Gastrulation: Classical Observations & Experiments
Amphibian Development Fertilization initiates cortical rotation Cortical rotation relocalizes determinants These determinants act to specify the dorsal-most regions of the embryo
Establishing Asymmetry Localized mRNAs
Early Cleavages in Xenopus gray crescent Tiers A B C D
Fate Map of Xenopus Blastula
Primary Tissue Layers Ectoderm Mesoderm Endoderm skin, neural notochord, somites, blood, kidney Endoderm gut, prechordal plate
Gastrulation Movements
Xenopus Gastrulation Movements
Gastrulation (again)
Xenopus laevis
Classic Experiments Hans Spemann (1903) Constriction of cleavage stage embryos & fertilized eggs Demonstrated asymmetry in fertilized egg
Embryo Constriction Experiment
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.
Fate Map of Gray Crescent dorsal blastopore lip point where gastrulation begins What is it about the gray crescent that makes it so important?
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.
Dorsal Lip Transplantation
Perturbations of Axis Formation: Making Two Dorsal Axes
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
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
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
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
Mesoderm Induced by Vegetal Cells
Nieuwkoop’s Experiment
Nieuwkoop’s Center Rescues Dorsal Axis in UV & Duplicates Axis in Wild-type
Inducing Centers in the Blastula “Spemann” (General)
Fate Map of Organizer and Nieuwkoop’s Center