223when the chick blastoderm thins to a single layer, forming the area pellucida epiblast.
Another set of studies showed that uterine eggs could be incubated in a variety of
orientations without rotation, or even hung from a chalaza without shells, and the
axis always formed along the gravitational axis with the posterior end uppermost
(Kochav and Eyal-Giladi 1971 ; Eyal-Giladi and Fabian 1980 ). Thus, it is the response
to gravity that is critical, not the stress of movement or effect of rotation per se.
Axial polarity and bilateral symmetry in the blastoderm is evident both morpho-
logically and molecularly prior to primitive streak formation. The future posterior
half of the embryo, in which the primitive streak forms, can first be distinguished by
the formation of a ridge of cells in the deep layer of the posterior area opaca,
Koller’s/Rauber’s sickle (Callebaut and Van Nueten 1994 ). Additionally, the hypo-
blast layer (analogous to the anterior visceral endoderm, see below) begins to form
in this region, coalescing from isolated hypoblast islands delaminating from the
epiblast in a posterior-to-anterior progression. The hypoblast is then replaced by the
endoblast (posterior visceral endoderm), derived from Koller’s sickle, in the same
progression (Stern 1990 ; Stern and Downs 2012 ). Molecular analyses have also
identified early differential gene expression in this region, including Gdf1 (alias
cVg1; Seleiro et al. 1996 ; Shah et al. 1997 ) in the PMZ of the epiblast and Goosecoid
(Gsc) in Koller’s sickle (Izpisúa Belmonte et al. 1993 ). Transplantation experiments
showed non-cell/tissue autonomous induction of ectopic axes, sparking parallels
between the PMZ and the amphibian Nieuwkoop center (see Sect. 6.3.5).
The mechanisms leading to these developmental events in the posterior are
unknown. The prevailing hypothesis for the initiation of this posterior polarity is the
differential exposure of presumptive areas to maternal cytoplasm during cleavage.
In the chicken egg, a particular cytoplasmic layer, the subgerminal ooplasm (gamma-
and delta-ooplasm) is contained in a central region below the blastodisc and overly-
ing the latebra and Nucleus of Pander (Callebaut 2005 ). With the blastoderm offset
from the animal pole by the inertia of the rotating egg (see above), this cytoplasm
would have more prolonged contact with cells arising in the upper (future posterior
end) of the embryo. The subgerminal cytoplasm may also be differentially inherited
by primordial germ cells, which form in response to cytoplasmic determinants (germ
plasm) in birds (Tsunekawa et al. 2000 ). It is thus possible that unknown axial deter-
minants might also be localized to this region, as in amphibians.
6.2.3.2 Early Polarization of the Mammalian Embryo
The evolution of implantation in therian mammals resulted in many changes in the
structure of the egg and early embryo, including a loss of yolk, the reemergence of
holoblastic cleavage and the early segregation of embryonic and extra embryonic
tissues, forming a preimplantation blastocyst (see Chap. 4 ). In light of these major
alterations in life history, it has long been of interest to determine the extent that
axes in the mammalian embryo are determined by cytoplasmic asymmetries as in
other vertebrates.
6 Vertebrate Axial Patterning: From Egg to Asymmetry