327between quail and chick cells can be used to determine the origin of various cell
types in orthotopic xenografts (Le Douarin 1973 ). Cells in the chick blastodisc are
too small to reliably inject lineage tracing dyes, but the lipophilic dyes DiI and DiO
have been used to heritably label the cell membranes in particular embryonic regions
(Hatada and Stern 1994 ). Both methods have been applied to fate map the chicken
blastoderm (Hatada and Stern 1994 ; Callebaut et al. 1996 ). For the purposes of
building a fate map, the pre-gastrula stage blastodisc (EGK Stage XII), is organized
in regions defined by their distance from the PMZ, using Koller’s sickle as a refer-
ence point. The studies agree that the germ layers arise from three distinct regions
of the blastodisc. Cells that form the gut and other endodermal organs are restricted
to a pie-shaped sector next to the Koller’s sickle, cells at the opposite end of the disc
generate ectodermal tissues, and mesodermal progenitors arise from the middle
region (Fig. 7.5c). A detailed view of the fate maps shows considerable overlap
between the territories. Ectodermal precursors can be found throughout the entire
blastodisc, whereas mesodermal progenitors are excluded from the anterior 20
% epiblast. Precursors of all three germ layers can be found juxtaposed in the pos-
terior region.
7.4.4 The Mammalian Fate Map
Fate mapping mammalian embryos presented a greater technical challenge, since
they develop internally. The first fate maps of mouse embryos were made by inject-
ing blastomeres from the ICM of one blastocyst into the ICM of another, genetically
distinct embryo (Gardner and Rossant 1979 ). The resulting chimeras showed that
cells in the ICM contribute to all three germ layers of the fetus, as well as some extra-
embryonic tissues. Fate maps at later stages used a strategy conceptually identical to
the quail-chick chimeras used to map the avian blastodisc. In these experiments,
grafted mouse epiblast tissue was labeled either by incorporation of radioactive thy-
midine or by treatment with a Wheat Germ Agglutinin-gold conjugate, which binds
to cell surfaces (Beddington 1982 ; Copp et al. 1986 ; Tam and Beddington 1987 ; Tam
1989 ). Labeled explants were orthotopically transplanted into unlabeled host
embryos and grown in culture. Fate maps drawn from these experiments showed that
all regions of the epiblast contribute to the three germ layers until late in gastrulation,
when regions become specialized to contribute to different tissues. A fate map of the
pre-gastrulation stage epiblast was made at single resolution by injecting single cells
of epiblast with an HRP lineage tracer, and culturing the embryos until mid-gastrula-
tion (Lawson et al. 1991 ). Precursors of the three germ layers are organized in broad
territories, with progenitors of the definitive endoderm restricted to the posterior
most region of the cup, while the anterior is populated only by ectodermal precursors
(Fig. 7.5d). Precursors of all three germ layers are intermingled in the posterior,
while ectodermal and mesodermal precursors are juxtaposed in the central region of
the epiblast. A significant portion of the epiblast also generates extraembryonic
mesodermal tissues, such as the amnion and chorion.
7 Establishment of the Vertebrate Germ Layers