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Development 120, 135-141 (1994)
Printed in Great Britain  The Company of Biologists Limited 1994
Interactions between primordial germ cells play a role in their migration in
mouse embryos
Miranda Gomperts*, Martin Garcia-Castro, Chris Wylie and Janet Heasman
Wellcome/CRC Institute and Department of Zoology, University of Cambridge, Tennis Court Road, Cambridge CB2 1QR, UK
*Corresponding author
SUMMARY
Primordial germ cells (PGCs) are the founder cell population of the gametes which form during the sexually mature
stage of the life cycle. In the mouse, they arise early in
embryogenesis, first becoming visible in the extraembryonic mesoderm, posterior to the primitive streak, at 7.5
days post coitum (d.p.c.). They subsequently become incorporated into the epithelium of the hind gut, from which
they emigrate (9.5 d.p.c.) and move first into the dorsal
mesentery (10.5 d.p.c.), and then into the genital ridges that
lie on the dorsal body wall (11.5 d.p.c.). We have used
confocal microscopy to study PGCs stained with an
antibody that reacts with a carbohydrate antigen (StageSpecific Embryonic Antigen-1, SSEA-1) carried on the
PGC surface. This allows the study of the whole PGC
surface, at different stages of their migration.
The appearance of PGCs in tissue sections has given rise
to the conventional view that they migrate as individuals,
each arriving in turn at the genital ridge. In this paper, we
show that PGCs leave the hind gut independently, but then
extend long (up to 40 µm) processes, with which they link
up to each other to form extensive networks. During the
10.5-11.5 d.p.c. period, these networks of PGCs aggregate
into groups of tightly apposed cells in the genital ridges. As
this occurs, their processes are lost, and their appearance
suggests they are now non-motile. Furthermore, we find
that PGCs taken from the dorsal mesentery at 10.5 d.p.c.
perform the same sequence of movements in culture. At
first they are actively locomotory. They become linked to
other PGCs via long processes and form clusters of nonmotile cells. This aggregation together into closely apposed
masses may be an important component of PGC migration
from the gut into the genital ridges and would also allow
signalling interactions between PGCs.
We show also that the first PGCs to emigrate from the
hind gut, between 9 and 9.5 d.p.c., do so directly into the
area where the genital ridges will form. This suggests that
an adhesive interaction between these ‘pioneer germ cells’
and the target tissue may play a role in the localisation of
PGCs into the genital ridges as they aggregate.
INTRODUCTION
genital ridges by 11.5 d.p.c. Their morphology in sectioned
material (Clark and Eddy, 1975) and their behaviour in culture
(Donovan et al., 1986) suggest that PGCs actively migrate
from the hindgut to the genital ridges.
PGCs isolated before, during, or after migration have distinctive adhesive properties in vitro (Donovan et al., 1986;
ffrench-Constant et al., 1991). Primordial germ cells isolated
during the 8.5-10.5 d.p.c. period adhere to certain feeder cell
monolayers (Donovan et al., 1986), and to purified extracellular matrix molecules (ffrench-Constant et al., 1991; de Felici
and Dolci, 1989). Moreover, it has been found that they are
invasive (Stott and Wylie, 1986). Little is known, however,
about their interactions with surrounding somatic cells during
migration in vivo. An understanding of the changes in cellular
associations of primordial germ cells, therefore, may shed light
on the mechanism by which these cells are directed to the
genital ridges.
During migration, primordial germ cells express on their cell
surfaces the Stage Specific Embryonic Antigen 1 (SSEA1, Fox
et al., 1981), a trisaccharide of the form galactose [β1-4]N-
Primordial germ cells (PGCs) are the founder population of the
gametes. They are set aside at early stages of embryonic development, and later join the somatic tissue of the gonad by a combination of passive displacement and active migration (see
Wylie and Heasman, 1993 for review). In the mouse, PGCs
can first be identified at the late gastrula stage (7.5 days post
coitum, d.p.c.) due to their expression of the enzyme alkaline
phosphatase (Ginsberg et al., 1990). This useful marker of
mouse primordial germ cells allows their subsequent distribution to be followed in tissue sections. At 8.5 d.p.c., the PGC
population is localized to the base of the developing allantois
and, by 9.5 d.p.c., they are in the epithelial lining of the
hindgut. Between 9 and 9.5 d.p.c., they emigrate from the
hindgut into its dorsal mesentery (10.5 d.p.c.) and from there
into the genital ridges on the dorsal wall of the abdomen.
During this period, PGC numbers increase from less than 100
to 25,000, indicating an approximate cell cycle time of 16-17
hours (Tam and Snow, 1981). Most PGCs have colonized the
Key words: primordial germ cell, migration, cell adhesion, mouse
germ cell
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M. Gomperts and others
acetylglucosamine[α1-3]fucose (Gooi et al., 1981). We have
used the monoclonal antibody, TG1 (Beverley et al., 1980),
which recognises SSEA1 (see Donovan et al., 1987), to
examine the 3-dimensional morphology of primordial germ
cells in situ at different stages of migration. Using confocal
microscopy to analyse the whole-mount samples, we have
examined both the time and pattern of primordial germ cell
emigration from the hindgut as well as their principal cellular
associations. Their appearance in histological sections suggests
that germ cells migrate to the genital ridges independently of
each other and use cues from the surrounding somatic cells and
extracellular matrix to guide them. A surprising fact to emerge
from our work is that after their emigration from the gut, primordial germ cells extend long processes, which they use to
associate with each other. The sequence of events seen by
confocal microscopy between 9.5 and 11.5 d.p.c. suggests that
PGC:PGC aggregation plays a role in the accumulation of
these cells in the genital ridges. This interpretation is supported
by the observation that primordial germ cells isolated from
embryos during this same period and cultured in vitro, behave
similarly. We also show that the first PGCs to emigrate from
the hind gut do so directly into the region of the genital ridges.
This suggests that an interaction between these ‘pioneer’ germ
cells and the target tissue may be involved in the localisation
of PGCs into the genital ridges as they aggregate.
MATERIALS AND METHODS
Whole-mount immunohistochemistry
Embryos were obtained from MF1 mice on 9-11 d.p.c. of timed pregnancies (day 0 is the day on which a vaginal plug is found). The yolk
sacs were removed and the embryos fixed in freshly prepared
paraformaldehyde (4% in PBS) for 2 hours at 4°C. To remove excess
fixative, the samples were washed in several changes of PBS. At this
stage, embryos were dehydrated into 100% methanol and were either
stored at −20°C or embedded in PEDS wax for sectioning. Embryos
at 10.5 d.p.c. or older were rehydrated to PBS and dissected further
to expose primordial germ cell-containing tissues. To increase the
access of the antibody to the tissues, the samples were incubated for
15 minutes at room temperature in PBS containing 2 mg/ml BSA,
0.1% Triton X-100, 0.02% sodium azide (PBTA). Non-specific
antibody-binding sites were blocked by incubating the samples in
PBTA containing 10% goat serum (PBTAS). The embryos were then
incubated overnight at 4°C in an undiluted supernatant of the mouse
hybridoma line TG1, which secretes a monoclonal antibody that reacts
with SSEA1 (a gift from Peter Beverley). The samples were washed
hourly for 5-6 hours in PBTA at room temperature before incubation
overnight at 4°C with the secondary antibody, a fluorescein-conjugated goat anti-mouse Ig (Nordic) diluted 1/50 in PBTA. After
extensive washing, the embryos were dehydrated through a
methanol:PBS series to 100% methanol. All incubations to this point
in the procedure were performed on a rotating platform. The embryos
were cleared in benzoyl alcohol:benzoyl benzoate (1:2) and mounted
in this mixture on cavity slides for viewing by confocal microscopy.
The number of PGCs attached to each other was counted by
following individual PGCs through z-series images in 0.5 µm steps.
The standard error of the percentage (SE%) was calculated from the
data according to the formula (pq/n)G. This can be used to assign confidence limits for the percentages such that there is a 68% chance that
the population % lies between ±1SE% of the sample %, a 95% chance
that the population lies between ±2SE% of the sample and a 99%
chance that the population lies between ±3SE% of the sample %.
Thick (20 µm) sections of wax-embedded embryos were dewaxed
in acetone, blocked in PBS containing 10% goat serum/0.02% sodium
azide (PSA) and stained as described for the whole mounts except that
the antibody incubations were performed at room temperature for 3
hours and washes were with PSA. The sections were mounted in
aqueous mounting medium containing an anti-quench agent (90%
glycerol, 10% water, 100 mg/ml DABCO triethylenediamine
(Sigma)).
All samples were analysed using a BioRad scanning-laser confocal
microscope (MRC 600).
In vitro culture of primordial germ cells
The primordial germ cell-containing tissues (the urogenital ridges and
the hindgut mesenteries) of 10.5 d.p.c. mouse embryos were dissected
away from other tissues in Ca2+-Mg2+-free PBS. The tissue fragments
were triturated repeatedly in a small volume of PBS to give a singlecell suspension. The cells were then washed in DMEM supplemented
with 10% FCS, glutamine (4 mM), penicillin and streptomycin. They
were pelleted and resuspended in the same medium before plating out
onto a preformed STO cell monolayer (prepared as described previously (Donovan et al., 1986)) on poly-D-lysine-coated chamber slides
(Lab-tek). The samples were fixed for 15 minutes at room temperature in 4% paraformaldehyde at various intervals after plating. They
were stained, mounted and viewed essentially as described above for
the sections except that there was no preblocking step, and washes
and second antibody dilutions were in PBS.
Vital staining of primordial germ cells for aggregation
assays
Primordial germ cells were isolated as described above and the singlecell suspension divided in two. Half the cells were washed into
DMEM supplemented as described above. The remainder of the cells
were labelled using the PKH26 Red Fluorescent General Cell Linker
Kit (Sigma) according to the manufacturers instructions. All the cells
treated in this way became labelled. The labelled cells were fractionated by Percoll (Pharmacia) centrifugation to eliminate labelled
somatic cells. The labelled cell sample (250 µl) was added to an equal
volume of 65% Percoll in PB1 containing 2.6% BSA (Barton et al.,
1993) and then laid onto a cushion of 65% Percoll (300 µl) in an
Eppendorf tube. 300 µl of 25% and then 200 µl of 20% Percoll were
gently applied on top. The sample was then centrifuged at 270 g for
20 minutes. An 800 µl sample, consisting predominantly of somatic
cells was removed from the top of the gradient and discarded. The
remaining primordial germ cell-enriched fraction (enriched to 25%)
was washed in DMEM plus supplements and then mixed back with
the unlabelled cells before plating onto a STO cell monolayer. The
samples were fixed and stained for SSEA1 as described above.
RESULTS
10.5 d.p.c. germ cells interact with each other to
form large networks
A low-power confocal image of a 10.5 d.p.c. mouse embryo
fragment stained for SSEA1 is shown in the mid-sagittal plane
in Fig. 1A. In this focal plane, the germ cells can be seen concentrated in the dorsal part of the hindgut mesentery although
a few germ cells are scattered more ventrally. The gut lumen
is also stained by this antibody and it can be seen looping away
from the dorsal body wall of the embryo. The targets of germ
cell migration, the genital ridges, are not in the plane of focus
in this image.
We have optically sectioned (0.5 µm sections) labelled
embryos at higher magnifications and find that germ cells
extend an array of processes from their surfaces. These
processes appear to link the germ cells together (Fig. 1B,D).
Mouse PGC migration
137
Fig. 1. (A) Confocal image of a 10.5 d.p.c. mouse embryo fragment
stained for SSEA1 showing the localisation of primordial germ cells.
The anterior end of the embryo is to the right-hand side of the image.
The ventral body wall of the embryo has been dissected away to
reveal the primordial germ cell-containing tissues. Black open
triangles, primordial germ cells; white arrow heads, hindgut lumen;
m, hindgut mesentery. Scale bar, 500 µm. (B) Four confocal images
cells from a 10.5 d.p.c. mouse embryo stained for SSEA1. Each
image is separated from the last by 3 µm in the Z axis. In frame 1
three apparently separate cells are visible. By moving 3 µm out of
the plane of the page, however (frame 2), it can be seen that cell ‘x’
has a long process with which it associates with cell ‘y’ (frame 3). In
frame four it can be seen that cell ‘y’ touches cell ‘z’. Scale bar, 20
µm. (C,D) Confocal images of 10.5 d.p.c. mouse embryos stained for
SSEA1, showing two kinds of germ cell process. (C) A lamellipodialike process with microspikes protruding from its surface (scale bar,
10 µm). (D) Two cells associating with each other via a long
filopodia-like process (scale bar, 20 µm).
We have examined the complete surfaces of 70 individual primordial germ cells in the mesenteries of six 10.5 d.p.c.
embryos in ‘Z series’ images and have found that 90%
(1SE%=3.6) are linked either via side-by-side associations or
via fine processes. The processes vary in length from 3 µm
(microspike-like, Fig. 1C) to 40 µm (filopodia-like, Fig. 1D)
and are of apparently random orientation. We describe germ
cells associated in this way as being ‘networked’.
Network formation is established after the germ
cells emerge from the hindgut endoderm, and leads
to their aggregation in the genital ridges
We have compared the appearance of primordial germ cells in
a temporal series of whole mounts beginning at 9 d.p.c. (there
are no antibody markers for younger primordial germ cells) in
order to determine when the germ cell network is established
and what happens after it forms. A confocal image of primordial germ cells in the hindgut endoderm of a 9 d.p.c. mouse
embryo is shown in Fig. 2A. At this stage, the hindgut, in
which the germ cells are embedded, is next to the dorsal aorta.
Although some of the germ cells have an elongate appearance
suggestive of motile cells, few associate with each other at this
stage. Fig. 2B,C shows slightly later embryos (9.25-9.5 d.p.c.)
where we have found germ cells with processes projecting out
of the gut endoderm into the surrounding tissue, close to the
site where the genital ridges will develop. These ‘pioneer’
germ cells may therefore contact the gonadal region as early
as 9 d.p.c. By 9.5 d.p.c., most germ cells have migrated out
from the epithelial lining of the gut and it is at this time that
networks first become evident (Fig. 2D). We examined
complete Z-series images from 243 individual primordial germ
cells in nine embryos, and found that 35% (1SE%=3.0) are
networked by 9.5 d.p.c.
Fig. 3A shows a genital ridge and its associated
mesonephros from an 11.5 d.p.c. mouse embryo. The germ
cells at this stage are now within the genital ridges. The appearance of these cells is quite distinct from those seen at 10.5
d.p.c. PGCs from 11.5 d.p.c. embryos are present as large
aggregates of rounded cells, which associate with maximal
contact of their surfaces and show few or no processes (Fig.
3B). Comparison of the 9.5, 10.5, and 11.5 d.p.c. images shows
that the germ cell networks seen at 10.5 d.p.c. are part of a
sequence of events whereby germ cells emerge from the hind
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M. Gomperts and others
Fig. 2. Confocal images from 9-9.5 d.p.c. embryos stained for
SSEA1 (A) Primordial germ cells in the hindgut endoderm of a 9
d.p.c. mouse embryo. Scale bar, 100 µm (B) A 9.25-9.5 d.p.c. mouse
embryo showing primordial germ cells with process projecting out of
the hindgut endoderm. Scale bar, 100 µm (C) A 50 µm thick section
through a similar aged embryo to that shown in B, with a single
primordial germ cell sending a process out of the gut endoderm into
a site adjacent to the developing mesonephros. The gut lumen is out
of the plane of focus and appears as a blur in this image as does the
end of the primordial germ cell process. Scale bar, 40 µm.
(D) Confocal image of a 9.5 d.p.c. mouse embryo showing germ
cells coming together into a network. The plane of focus is such that
the hindgut lumen clearly visible. Scale bar, 40 µm. Open triangles
(either black or white) indicate primordial germ cells. Black
arrowheads indicate the hindgut lumen. White arrowheads indicate
the periphery of the hindgut endoderm. da, dorsal aorta; s, somite,
mn, developing mesonephros.
Fig. 3. Confocal images from an 11 d.p.c. mouse embryo stained for
SSEA1 (A) A genital ridge and associated mesonephros. The
primordial germ cells are packed into the tissues of the developing
gonad. Scale bar, 100 µm. (B) Rounded up primordial germ cells,
which have a non-motile appearance and which directly associate
with each other, maximizing their cell surface contacts. They are
quite distinct from the primordial germ cells of younger embryos.
Scale bar, 10 µm.
gut separately and come together into closely apposed masses
in the genital ridges.
10.5 d.p.c. germ cells form networks and aggregate
in vitro
The interactions between PGCs could occur passively or
actively. They are undergoing cell division during migration
and so connected cells might be siblings that remain adherent
to one another. Indeed, networks could be formed passively by
Mouse PGC migration
139
Fig. 4. The hindgut mesentery and associated mesonephroi
were dissected from embryos at 10.5 d.p.c. After
disaggregation, the cells were plated onto a STO cell
monolayer at a density of 8-10 embryo equivalents per well.
At intervals of 2 (A, scale bar, 50 µm), 24 (B-D, scale bars,
10, 10 and 25 µm, respectively) and 48 (E, scale bar, 10 µm)
hours, the cells were fixed and stained for SSEA1. Examples
of the conformations of the cells found at these times are
shown.
initially adherent siblings being partially pulled apart by the
extension of the mesentery, thus forming the networks seen at
10.5 d.p.c. Alternatively, a single PGC may actively extend
processes until it encounters another PGC and then selectively
adhere to it. To determine the involvement of morphological
movements in network formation, we isolated PGCs from 10.5
d.p.c. embryos, disaggregated them mechanically and plated
them onto a substratum of a confluent monolayer of irradiated
STO fibroblasts. The cultures were fixed and the germ cells
stained for SSEA1, 2 hours, 24 hours and 48 hours after
plating. Examples of the cell conformations are shown in Fig.
4. We find that during the first 2 hours in culture the PGCs
adhere to the STO cell monolayer, flatten out and start to
extend processes. After 24 hours in culture, some of them have
formed networks. Processes extending up to 50 µm, and indis-
tinguishable morphologically from those seen in the embryo,
have been observed between PGCs in culture. By 48 hours,
however, clusters of at least 8 PGCs are apparent in which they
are closely apposed and rounded up, identical in appearance to
those found in 11.5 d.p.c. embryos. In a separate experiment,
in which the density of the cells seeded was halved so that we
could assess the proportion of cells associating with time, we
found in four replicate samples that, after 2 hours in culture,
15.1% (1SEM=1.7) of PGCs were networked. After 24 hours,
this number increased to 65% (1SEM=1.94) and, by 48 hours,
the number of PGCs in networks had risen to 86.25%
(1SEM=1.8) in the culture. Thus progressively more PGCs are
found in clusters than as single cells attached to STO cells, suggesting that PGCs may preferentially adhere to each other.
Clusters of up to 7 cells were observed in this experiment. Such
140
M. Gomperts and others
from unlabelled germ cells. The cultures were fixed at 24 hours
and the germ cells stained for SSEA1. Fig. 5 shows that, in
some cases, aggregates of PGCs forming in culture contain
both labelled and unlabelled cells, and therefore cannot have
arisen by cell division alone.
DISCUSSION
Fig. 5. Germ cell-containing tissue from 10.5 d.p.c. embryos was
isolated as described in the legend to Fig. 4. Half the disaggregated
cells were labelled with a rhodamine-conjugated dye (PKH26). The
cells from the labelled sample were enriched for germ cells and then
mixed with the unlabelled sample before plating onto a STO cell
monolayer. The cultures were fixed and stained for SSEA1 after 24
hours. Examples of ‘mixed’ clusters are shown. PGC1 is both
PKH26-positive (red) and SSEA1-positive (green), PGC2 is PKH26negative but SSEA1-positive, and PGC3 is an aggregate of both
PKH26-positive and -negative PGCs.
clusters have also been documented previously and shown to
increase in size with time (Godin et al., 1991). The fact that
clusters of PGCs form in vitro suggests that morphological
movements within the embryo are not essential for network
formation.
To distinguish between the roles of mitosis and cell aggregation in cluster formation, we labelled a proportion of the cells
with a rhodamine-conjugated dye before plating them onto the
STO monolayer. The labelled cells were subjected to a Percoll
gradient enrichment procedure to eliminate excess somatic
cells, since this makes it difficult to distinguish the labelled
The object of this work was to identify structures in the embryo
with which primordial germ cells interact, and analyse changes
in cell shape during migration using confocal microscopy. The
advent of the confocal microscope has enabled three-dimensional analysis to be performed, and with associated real time
imaging techniques, analysis of migratory cells in living tissues
is now possible in some organisms. Clearly, such experiments
rely on the availability of suitable markers and, in this respect,
SSEA1 is particularly useful as it is distributed over the entire
PGC surface. An obvious and surprising observation is that
progressive germ cell-germ cell association occurs during the
2-day period of migration from the hind gut to the genital
ridges. As they emigrate from the hindgut, primordial germ
cells are not generally associated with each other. 1 day later,
however, when the germ cells are in the mesentery, they are
connected by long processes. 1 day later still, they are aggregated into clusters of closely apposed cells in the genital ridges.
This sequence of events also occurs when 10.5 d.p.c. primordial germ cells are cultured in vitro, suggesting that it is not
simply due to activity of the surrounding tissues. It is well documented that primordial germ cells actively divide during their
migration (see Wylie and Heasman, 1993 for review). It may
be argued, therefore, that clusters of PGCs represent sibling
cells. We have demonstrated, however, by a differential
staining procedure, that cell aggregation does play a role in
germ cell cluster formation.
Another significant observation is that the first primordial
germ cells to emigrate from the gut do so when there is no
dorsal mesentery. The genital ridges begin to form medial to
the mesonephros as small thickenings on the dorsal abdominal
wall at 9.5 d.p.c. (see Clark and Eddy, 1975). Thus the first primordial germ cells to emigrate from the hindgut extend
processes into a site very close to, or actually in, the developing genital ridge.
Our observations raise two important points concerning
primordial germ cell migration, neither of which is apparent
by examination of the cells by conventional microscopic
methods. Firstly, germ cells do not migrate independently of
each other, but instead form an extensive network of
connected cells. Secondly, the progression of events seen in
vivo and in vitro suggests that PGC-PGC adhesion may play
a role in their accumulation in the genital ridges. Previous
work on primordial germ cells and other migratory cell types
has focused on heterotypic attachments between the
migratory cells and the cells and extracellular matrix surrounding them (Donovan et al., 1986; de Felici and Dolci,
1989; ffrench-Constant et al., 1991; Hynes and Lander,
1992). Our work, however, demonstrates the occurrence of
interactions between the migratory cells themselves. Does the
process of germ cell-germ cell aggregation result in their
accumulation in the genital ridges? And if so, how? One
explanation is that the primordial germ cells leaving the gut
Mouse PGC migration
endoderm first, during 9-9.5 d.p.c., enter directly into the site
where the genital ridges will develop and anchor themselves
to the target tissue. As later primordial germ cells emerge
from the gut, they migrate in the elongating mesentery and
extend long processes which attach to other primordial germ
cells. Aggregation would then draw the primordial germ cells
into the genital ridges.
This model of primordial germ cell migration is supported
by their behaviour when isolated during the migratory phase
and cultured on feeder cells. The primordial germ cells adhere,
spread and move on the surfaces of the embryo fibroblasts.
During this period, they associate with each other, such that
after 2 days in culture, only a small proportion are found as
single cells. The majority are found in aggregates of closely
apposed and apparently non-motile cells. Thus, although primordial germ cells stick to somatic cells and use them as a substratum for migration, it appears that they may adhere preferentially to each other.
Furthermore, these observations raise the possibility that
germ cell-germ cell contacts (rather than germ cell-somatic cell
contacts) play a role in switching off the migratory phenotype.
Signalling interactions between PGCs would be allowed by
their close apposition, and may play an important role in the
migratory and/or proliferative behaviour of these cells.
It will be important to identify the adhesion molecules
mediating germ cell-somatic cell adhesion and germ cell-germ
cell adhesion. There are a number of candidate molecules. Ncadherin, for instance, has been reported to be present on
chicken germ cells (Hatta et al., 1987) but so far cadherins have
not been identified on mouse germ cells. Xenopus germ cells
express a cell surface glycolipid during the period when they
leave the gut, a factor which bears a carbohydrate moiety that
has been found to play a role in cell-cell adhesion during the
blastula stage (Turner et al., 1992). SSEA1 itself has been
implicated in cell-cell adhesion in the early mouse embryo
(Bird and Kimber, 1984) and thus may play a role in homotypic
germ cell-germ cell adhesion. SSEA1 is involved in compaction at the morula stage, during which cells maximize their
contacts forming a tight ball of cells. This process has a close
resemblance to that which we see during primordial germ cell
migration.
In summary, these observations are incompatible with the
view, suggested by their appearance in tissue sections, that
PGCs migrate as individuals, and arrive at the genital ridges in
turn, like runners finishing a race. Instead, the principal
movements seem to involve PGCs interacting with each other,
first via long processes, and then by aggregation. It will be
important to identify the molecules involved in this homotypic
adhesion event, since we would predict them to be critically
important in PGC migration.
141
We are grateful to the Wellcome Trust for financial support for this
work. We also thank Aaron Crawford, Julie Cooke and Colin Sharpe
for useful discussions, constructive criticisms and proof reading.
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(Accepted 21 September 1993)