419Building on his seminal discovery of mesoderm induction (Nieuwkoop 1969 ),
Nieuwkoop and colleagues began a series of studies that defined the basic parameters
underlying the induction of urodele PGCs. First, they cultured the primitive ectoderm
of early axolotl gastrulae with vegetal blastomeres from either the ventral or dorsal
side of the embryo (Boterenbrood and Nieuwkoop 1973 ). When associated with ven-
tral cells, the primitive ectoderm elicited a robust induction of PGCs, along with
somatic ventral mesodermal derivatives, such as blood and mesonephric tissue. Dorsal
vegetal blastomeres induced a very different response, resulting in the production of
dorsal axial structures, notably including notochord. These studies represent the first
demonstration that patterning of the mesodermal mantle in amphibians occurs in
response to signals from the presumptive endoderm. Equally provocative, they
prompted the hypothesis that PGCs are typical mesoderm derivatives, formed similar
to somatic cells, blurring the fundamental distinction between germ cells and somatic
cells that had been a fixture in the germ cell literature until that time. Nonetheless,
these experiments could not exclude the possibility that PGCs are derived exclusively
from cells in the animal cap that harbor germinal granules which might predispose
them towards germ line development. Sutasurja and Nieuwkoop ( 1974 ) addressed
this possibility. They dissected animal caps into discreet regions, and provided these
with equivalent inducing stimuli. Their data showed that all regions within the animal
moiety were capable of producing PGCs. Interestingly, from the data in this study the
authors concluded that PGCs and somatic cells are derived from the same population
of precursors, which in modern parlance is the functional definition for cells in the
pluripotent ground state (Nichols and Smith 2009 ).
The hypothesis that PGCs might be derived from typical cells in the primitive ecto-
derm was difficult to reconcile with work from experimental models that contain germ
plasm. However, Ikenishi and Nieuwkoop ( 1978 ) were unable to detect germ plasm in
the PGCs of tailbud stage (stage 23) embryos. Germ–cell specific material, called nuage,
which has some features of germinal granules, was first observed in PGCs in larval stage
embryos (stage 40), as PGCs begin to approach the presumptive gonad. While Ikenishi
and Nieuwkoop proposed that that their findings negate the likelihood of predetermined
germ cells in urodeles, other workers considered the evidence insufficient to exclude a
role for early acting germ cell determinants (Michael 1984 ; Smith et al. 1983 ). Therefore,
the issue of whether or not urodele embryos contain germ plasm, or related material,
was considered unresolved by the time the twentieth century came to an end. However,
a related larger issue, which influenced contemporary interpretations of data up to that
period, also remained unresolved: Could closely related species evolve divergent mech-
anisms to regulate cell fate specification in early development?
8.5.3 PGC Induction Is a Conserved Basal Trait of Vertebrates
The issue of whether urodeles and anurans employ shared, or divergent, modes of
PGC specification led Johnson et al. ( 2001 ) to revisit germ line development in
axolotls. They asked if axolotl oocytes contain germ plasm, and demonstrated that
8 Mechanisms of Vertebrate Germ Cell Determination