/ . Embryo/, exp. Morph. Vol. 25, 2, pp. 263-276, 1971
Printed in Great Britain
263
Structure and developmental
tendency of the dorsal marginal zone in the
early amphibian gastrula
ByNOBUSHIGE IKUSHIMA1 AND SETSUKO MARUYAMA1
From the Biological Laboratory, Kansai Medical School
SUMMARY
The peripheral surface of a fertilized, uncleaved egg is subdivided through cleavage and
is allotted to constituent cells. This is called the primary surface. In an early morula a
constituent cell has two kinds of surfaces: the primary surface, and the secondary surface,
which does not participate in forming the periphery of the embryo. Electron-microscopic
observations showed structural differences between the two surfaces.
When the dorsal marginal zone of an early gastrula of Hynobius nebulosus is excised and
immersed in Feldman's solution, the piece can easily be separated into two layers: the outer
layer, whose constituent cells are given a share of the primary surface, and the inner layer,
whose constituent cells are completely covered only by the secondary surface.
Both an explanted piece of the outer layer and an intact double-layered piece show three
kinds of movement: spreading, convergence followed by stretching, and spherical thickening.
The inner layer is kinetically very inert, showing slight spreading and thickening.
An explanted piece of the outer layer differentiates into axial mesodermal structures,
while the inner layer does not.
When a piece of either the inner or the outer layer is implanted in the blastocoel of another
gastrula, it induces deuterencephalic and spino-caudal structures and seems to differentiate
into axial mesodermal structures.
Differences of kinetic properties and differentiation are considered to result from the
fact that the outer layer has the primary surface, while the inner layer does not.
Functional effects of the primary surface on the movement of tissues and differentiation
are discussed.
INTRODUCTION
It is well known that during gastrulation the dorsal marginal zone of an
early amphibian gastrula executes conspicuous movements of convergence
towards the median line and of stretching in the antero-posterior direction
(Vogt, 1922, 1929). A similar pattern of movements is also shown in this area
even when it is isolated from a gastrula (Ikushima, 1958, 1961). It has been
inferred that these movements of tissues occur owing to reshuffling or rearrangement of constituent cells (Willier, Weiss & Hamburger, 1955; Waddington,
1962); but the mechanism of the reshuffling and the rearrangement has not
yet been clearly demonstrated.
1
Authors'' address: Biological Laboratory, Kansai Medical School, Hirakata-City, Osaka,
Japan.
264
N. IKUSHIMA AND S. MARUYAMA
During experiments on dissociation of embryonic tissues, we found that
a piece taken from the dorsal marginal zone of an early gastrula of Hynobius
nebulosus could easily be separated into outer and inner layers after a short
treatment in dissociating agents. Moreover, we found that in the inner layer
movement and differentiation of the tissue was not necessarily similar to that
in the outer layer. The present experiments were attempted to investigate
differences of developmental tendencies of the two layers.
MATERIALS AND METHODS
Embryos of Hynobius nebulosus and Triturus pyrrhogaster were used. The
separation of the dorsal marginal zone into two layers was possible only in the
embryos of the former. The tissues were separated or dissociated in Feldman's
solution (Feldman, 1955). A square piece was isolated from the dorsal marginal
zone of an early gastrula (Harrison's stage 10, 1952). When the piece was
immersed in Feldman's solution the cells began to disaggregate, especially
at the corners of the piece, within 5-10 min. In these pieces the outer cell
layer could easily be peeled off from the inner cell layer with a hair loop.
Generally, the former seemed to be composed of a single layer of cells and
the latter of two or more layers of cells. Immediately after the separation
the two layers were removed from Feldman's solution to full strength Holtfreter's
solution.
In order to investigate the kinetic properties of the tissue the following
explantation experiments were performed. A square piece taken from the
dorsal marginal zone was cut into two parts along its median line. The two
parts were immediately combined again into one in which their inner surfaces
were placed in close contact with each other (Fig. 1). Within 12 h the two
parts were completely fused together, resulting in a single rectangular-shaped
explant. In a similar way rectangular explants were prepared from square
pieces of the outer and inner cell layers respectively. The kinetic properties of
the tissues were investigated by observing the change in configuration of these
rectangular explants during the first 48 h of cultivation.
To investigate the differentiating capacity, the intact square piece, and the
square pieces of the outer layer and of the inner layer, were cultured in full
strength Holtfreter's solution. To investigate the inducing capacity, they were
implanted into the blastocoel of another gastrula of the same age. Operations
and cultivation of these embryonic tissues were always performed in glass
dishes lined with 1-5% agar-agar. After cultivation for 3-4 weeks tissues were
fixed with Bouin's fixative, embedded in paraffin wax, sectioned at 10/6 and
then stained with Mayer's haemalaum and eosin. In some cases they were
fixed with OsO4 after cultivation for 24 h, embedded in Epon, sectioned at
10/* on an LKB Ultrotome and stained with toluidine blue (Trump,
Smuckler & Benditt, 1961).
Development of dorsal marginal zone
265
For electron-microscopic observations, specimens were fixed for 30 min in
ice-cold 1 % OsO4 in phosphate buffer at pH 8-0. After fixation they were
dehydrated rapidly in a graded series of ethanol and embedded in methacrylate.
Thin sections were cut with an LKB Ultrotome. Observations were carried
out with a Hitachi H-7 microscope, and micrographs were taken at an original
magnification of 2000-8000.
Fig. 1. Schematic representation of the method of explantation to investigate the
kinetic properties of the dorsal marginal zone. (A, B) Excision of a square piece
from the dorsal marginal zone of an early gastrula. (C) The piece is cut into
two parts along the median line. (D) The two parts are recombined.
RESULTS
1. Preliminary experiments
When morulae of H. nebulosus and T. pyrrhogaster were immersed in Feld-
raan's solution they dissociated into discrete cells within 30-60 min. As shown
in Fig. 2B the surface of a cell derived from the animal polar region is composed
of two parts - a heavily pigmented part and the remaining paler part. The
pigmented part is undoubtedly identical to the peripheral surface of the fertilized,
uncleaved egg. In the present report this part of the surface is provisionally
named 'primary surface', and the remaining part, the paler part in the case
of the animal polar cells, is named 'secondary surface'. As a result of cleavage
the primary surface is subdivided and allotted to every one of the constituent
cells of a morula. In the dorsal marginal zone of an early gastrula, lightmicroscopic observations showed that the surface of a cell in the outer layer
seemed to be composed of both primary and secondary surfaces, whereas in
the inner layer the surface seemed to be composed exclusively of secondary
surface.
During the present dissociation experiments, it was frequently observed that
the border of the primary surface began to constrict a cell (Fig. 3) or the primary
surface area shrank suddenly. These effects were most clearly shown in discrete
cells derived from a morula, and were observed not only in Feldman's solution
but also after they were removed into Holtfreter's solution. Although the
factors which make the primary surface area shrink have not yet been clearly
266
N. IKUSHIMA AND S. MARUYAMA
PSr
Fig. 2. (A) An intact morula of Hynobius nebulosus. (B) A morula dissociated
into discrete cells 45 min after it was immersed in Feldman's solution.
Fig. 3. Discrete cells which are constricted at the border of the primary surface.
The cells are derived from a morula of Triturus pyrrhogaster.
Fig. 4. Electron micrograph of a section of the surface area of a discrete cell from
a morula of T. pyrrhogaster.
Abbreviations: ASt, alveolar stratum; bl, borderline of the primary surface; PSr,
primary surface; PSt, cytoplasmic stratum containing pigment granules; SSr,
secondary surface.
Development of dorsal marginal zone
267
demonstrated, these observations seem to indicate that structural differences
exist between the two surfaces. In order to examine this possibility, discrete
cells from amorulaof T. pyrrhogasterwere investigated electron-microscopically.
In Fig. 4 a cross-section of the surface area of a discrete cell in which the
primary surface shrank markedly is shown. At the periphery of the primary
surface area a specific stratum is found just beneath the deeply folded plasma
membrane. This stratum shows a fine alveolar structure, and is mostly about
0-5/t thick; at the margin the thickness reaches 1 fi or more. Just inside this
stratum there is a cytoplasmic stratum about 5 pi thick containing an abundance
of pigment granules and hardly any yolk platelets. These two kinds of strata
do not seem to shrink independently but to be combined, forming a unitary
layer. This layer very probably represents the microscopically named 'primary
surface', and is presumably identical to the 'cortex' defined by Pasteels (1964)
and to the 'cortical layer' of Balinsky (1965). On the other hand, in the
secondary surface these two kinds of strata are hardly recognizable electronmicroscopically. From these results it seems beyond doubt that structural
differences exist between the primary and secondary surfaces.
2. Kinetic properties
(a) Combined explant of both the outer and the inner layers (Fig. 1). The
cross-section of this explant 12 h after combination (Fig. 5) reveals that a
continuous cell layer encircles an empty lumen. From this figure it may be
considered that the fusion of the two parts is finally accomplished through
lateral adhesion of the cut edges to each other and not by adhesion of the
inner surface.
Within the first 24 h of cultivation it could easily be seen that the lower
one-third of the explant became thick and spherical, and the middle one-third
became narrow in a lateral direction and stretched along the antero-posterior
direction of the original embryo. The upper one-third remained unchanged
in some specimens and wrinkled or spread out in the others. In the following
24 h these three kinds of movements of tissues became more and more marked.
(b) Combined explant of the outer layer. Immediately after the explants were
removed from the dissociating to the culture medium the constituent cells
became closely aggregated and the explant diminished slightly in size. Within
the first 24 h of cultivation the upper part became wrinkled and spread out, the
middle part narrowed and stretched and the lower part became thick and
spherical. In the following 24 h the spreading of the upper part and the narrowing
and the stretching of the middle part became more and more marked, but the
spherical thickening of the lower part hardly increased its thickness. Consequently, it may be stated that the change of shape in these explants is quite
similar to that in the preceding case (a).
(c) Combined explant of the inner layer. When the explants were removed
from the dissociating to the culture medium they diminished slightly in size,
l8
EM B 25
268
N. IKUSHIMA AND S. MARUYAMA
and their corners became rounded. During the first 24 h of cultivation the
lower part became slightly thicker, the upper part remained almost unchanged
and the middle part did not narrow or stretch at all or show any other type
of movement. In the following 24 h the explants spread out to some extent.
The spreading occurred not only in the upper part but also in the middle and
the lower part. These are shown in Fig. 6. The narrowing and the accompanying
stretching were never observed in any region of the explant during the first
48 h or in the following days of cultivation.
Fig. 6
Fig. 5. A cross-section of a combined explant of both the outer and the inner layer
12 h after the two parts were combined into one. An epoxy section stained with
toluidine blue, n, Nucleus.
Fig. 6. Change of configuration of the combined explant of the inner layer.
(A) 3 h after the two parts were combined; (B) 24 h after combination; (C) 48 h
after combination. The line in the figure represents 100 /i.
3. Differentiating capacity
(a) Differentiation of the unseparated explants. Immediately after the beginning of cultivation they began to curl up, so that the original peripheral surface
of the pieces, presumably identical to the primary surface, completely covered
the mass of cells. After 4 weeks' cultivation fifteen specimens were available.
Among them eleven remained a solid mass till the end of cultivation, while
the other four finally developed into large vesicles. Among the four vesicular
explants only a small mass of neural tissue and mesenchyme were found in
one specimen, and a small amount of muscle, pronephric tubules and lateral
Development of dorsal marginal zone
269
plate occurred without neural tissue in the other three. In the eleven specimens
remaining as solid masses, notochord, muscle, deuterencephalon and spinal
cord were always found (Fig. 7), and in one specimen an archencephalic
structure with an eye was also found.
(b) Differentiation of the explants of the outer layer. As soon as the explants
were removed from the dissociating to the culture medium they diminished
slightly in size and began to curl up, so that the cell mass was completely
covered by the primary surface. After 4 weeks' cultivation fourteen specimens
were available. Combinations of differentiated tissues appearing in these
explants are summarized in Table 1. In six specimens notochord, muscle,
deuterencephalon and spinal cord were found (Fig. 8). In two specimens an
abundance of muscle and ear vesicles occurred without notochord. In the
remaining cases neural or sensory structures were not found; in four specimens
notochord and muscle occurred without neural tissue and in the other two
only mesenchyme and mesenteric tissue were found in a large epithelial vesicle.
Table 1. Combination of tissues differentiated in explants
of the outer layer and of the inner layer
Combination of tissues
differentiated
Notochord + muscle
Notochord + muscle + neural tissue
Muscle + neural tissue
Mesenchyme
Mesenchyme + neural tissue
Undifferentiated tissue + neural
tissue
Total
Explants
of outer
layer
Explants
of inner
layer
4
6
2
2
0
0
0
0
0
0
3
10
14
13
(c) Differentiation of the explants of the inner layer. When the explants were
removed from the dissociating to the culture medium they diminished in size
and became slightly rounded. The curling up of the pieces was not so marked
as in the previous two cases. After 4 weeks' cultivation thirteen specimens
were available. Among them three specimens developed into large vesicles.
They were composed of a very small mass of neural tissue and mesenchyme,
and cartilage in one specimen (Fig. 9). The remaining ten were solid masses
composed of a large amount of undifferentiated tissue and a small amount
of neural tissue (Fig. 10); cartilage was found in two of them. These results are
summarized in Table 1.
4. Inducing capacity
(a) Implantation of the outer layer. During 3-4 weeks' cultivation of the
hosts, secondary embryonic structures were produced generally in their midventral region. Microscopic observation revealed that in sixteen specimens out
18-2
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N. IKUSHIMA AND S. MARUYAMA
nt
2nd emb
2nd emb
Development of dorsal marginal zone
271
of the twenty available deuterencephalon and spinal cord were always found
accompanied by well-differentiated notochord and muscle in the secondary
embryos (Fig. 11). In one specimen a large amount of neural tissue and four
ear vesicles were found without mesodermal organs. In two specimens only
a small amount of muscle and an abundance of mesenchyme were found
without notochord and neural tissue, and in the remaining specimen only
a small quantity of muscle and a large number of blood cells were found.
(b) Implantation of the inner layer. During 3-4 weeks' cultivation the secondary
embryonic structures were found generally in the mid-ventral region of the
hosts. Microscopic observations revealed that in twelve specimens out of the
seventeen available, deuterencephalon and spinal cord occurred with welldifferentiated notochord and muscle (Fig. 12). In three specimens, only mesenchyme and a small amount of muscle were found without notochord and neural
tissue. In the remaining two specimens archencephalic structures with eyes
occurred, one formed antero-ventrally and the other mid-ventrally.
The results of the implantation experiments reveal that the implanted pieces
of the outer and the inner layer both induce deuterencephalic and spino-caudal
structures in the hosts in the majority of cases. Although archencephalic
structures were induced only by the inner layer, they occurred in only two
specimens out of seventeen. From these results it seems difficult to point out
distinct differences in inducing capacity between the two layers.
On the other hand, it must be noted that well-differentiated notochord and
muscle are found in the secondary embryos which are induced by the pieces
of the inner layer. These pieces were not able to differentiate into axial mesodermal structures in the explantation experiments. However, in the case of
implantation experiments it seems probable that axial mesodermal structures
in the secondary embryos are derived from the implanted piece of the inner
layer.
Fig. 7. A section of an unseparated explant after 4 weeks' cultivation.
Fig. 8. A section of an explant of the outer layer after 4 weeks' cultivation.
Fig. 9. A section of an explant of the inner layer after 4 weeks' cultivation.
Fig. 10. A section of another explant of the inner layer after 4 weeks' cultivation.
Fig. 11. A section of an embryo in which a secondary embryo is induced by the
implanted piece of the outer layer.
Fig. 12. A section of an embryo in which a secondary embryo is induced by the
implanted piece of the inner layer.
Abbreviations: ca, Cartilage; e, ear vesicle; m, muscle; ne, neural tissue; nt,
notochord; und. a mass of undifferentiated tissue. The line in each figure represents
200/*.
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N. IKUSHIMA AND S. MARUYAMA
DISCUSSION
The role of the primary surface on the development
of the dorsal marginal zone
The present experiments show that when explanted pieces of the inner layer
are cultured in saline, they do not differentiate into axial mesodermal structures,
while pieces of the outer layer do. At first, this fact seems to indicate that the
intrinsic potential for development of the inner layer is quite different from
that of the outer layer. However, the results obtained in the implantation
experiments show that, like the outer layer, the inner layer can also differentiate
into axial mesodermal structures and can induce deuterencephalic and spinocaudal structures. These results do not necessarily support the above view;
another interpretation may be possible.
In the present explantation experiments the intact square explants and the
explants of the outer layer both begin to curl up immediately after the beginning
of cultivation. In consequence of the curling up, the mass of cells is completely
covered by the primary surface. In the case of the implantation experiments
there is no doubt that the implanted piece develops in the blastocoel which is
the interior of the primary surface. In contrast, the cells of the inner layer in
the explantation experiments are cultured without a covering of the primary
surface. The failure of notochord and muscle formation occurs exclusively
in this experimental situation. Looking through these results it seems likely
that the cells of the dorsal marginal zone of the early gastrula require a specific
environment in order to differentiate into axial mesodermal structures. This
environment is presumably produced when the aggregate of cells is covered
by the primary surface.
Next there arises the question of what sort of influence the covering of the
primary surface exerts upon the interior mass of cells. On this question a clear
solution cannot yet be given. However, as pointed out by Holtfreter (1943) and
ascertained by Loeffler & Johnston (1964), there seems no doubt that the outer
surface of the amphibian embryo has a specific trait which prevents or reduces
the entrance of water, electrolytes and other substances. From this, it may be
presumed that there is a specific environment within the permeability barrier,
caused by the protection from unfavourable external factors, or by the prevention of the escape of necessary substances from cells to the surrounding
medium, or by both. It seems likely that the cells of the dorsal marginal zone
can only realize their potency to differentiate into axial mesodermal tissue in
this specific environment, although its character has not yet been clearly
demonstrated.
In this report it is also shown that there are differences in kinetic properties
between the outer and the inner layer of the dorsal marginal zone. Generally
speaking, the outer layer seems to be kinetically active and the inner layer
inert. The most conspicuous difference is that the middle part of the former
Development of dorsal marginal zone
273
shows marked convergence and stretching, while the latter does not. At the
same time, it must also be noted that an intact piece composed of both the
outer and inner layers shows marked stretching as in the case of the outer
layer alone. From these facts it seems clear on the one hand that the outer
layer plays a leading role in the movement of the dorsal marginal zone, and
on the other hand that the inner layer, although seemingly inert kinetically, is
not rigid but stretches, conforming itself to the outer layer where the two
layers are in direct contact.
How do the cells of the outer layer play a leading role in the movement
of the dorsal marginal zone? In this report it is shown that the most marked
morphological difference between the two layers is that the cells of the outer
layer have a share of the primary surface, while those of the inner layer have
not. Consequently it seems possible to suppose that the primary surface is
responsible for the active movement of the dorsal marginal zone.
In order to verify this supposition it seems necessary to demonstrate first
that the primary surface has intrinsic tendencies to contract, to expand or to
stretch. On this point the results obtained by Curtis (1960) are relevant. He
demonstrates that the cortical material possesses morphogenetic properties,
and that these properties may be transferred with the cortical material even
when it is grafted. The cortical material used in his grafting experiments is
composed of a true cell surface at the periphery and a thin hyaline layer
containing mitochondria and pigment granules. Its thickness varies from 0-5
to 3-0 fi. The primary surface in the present experiments also contains a true
cell surface, a thin alveolar and a cytoplasmic stratum and pigment granules.
Its thickness as shown in Fig. 4 is about 5-6 [i. However, since this is undoubtedly
abnormal thickening owing to contraction, it seems reasonable to state that
the primary surface in the present report is identical to the cortical material
of Curtis's experiments. Consequently, it seems reasonable to consider that
the primary surface has intrinsic morphogenetic properties. Moreover, as shown
by Holtfreter (1943), when the viscosity of the surface layer is reduced by various
agents, the mass movements of gastrulation and neurulation are prevented
or altered and exogastrulae and other malformations are produced. This fact
may also support our present hypothesis.
On the problem of whether the primary surface
has a syncytial nature
As discussed above, it seems conceivable that the primary surface has
intrinsic morphogenetic properties. However, it seems also true that, in as
much as contraction or stretching of the primary surface occurs independently
in individual cells, these movements do not necessarily result in reshuffling or
rearrangement of cells in embryonic tissues. Thus it is difficult to understand
how independent movements of the primary surface in individual cells inevitably
result in a change of configuration of an embryonic tissue. The leading role
274
N. IKUSHIMA AND S. MARUYAMA
of the primary surface in movements of tissues could be most easily understood
if it is verified that the primary surfaces of individual cells are integrated into
a supercellular syncytial unit.
On this problem, Holtfreter (1943, 1948) has indicated that a special surface
layer named 'surface coat' exists at the periphery of amphibian eggs. According
to him, the spherical black pigment granules which are found at the surface
of the egg are contained in this coat, and this coat is divided up by the process
of cell division, but re-establishes a continuous envelope cementing together
the peripheral part of the blastomeres. However, recent electron-microscopic
observations do not necessarily support the existence of such a syncytial
envelope. These show that the pigment granules undoubtedly occur in the
cytoplasm which lies just inside the plasma membrane, and that no extracellular
cortical structures which correspond to Holtfreter's surface coat are to be
found (Wartenberg & Schmidt, 1961; Balinsky, 1965). Our present observations
also agree with these findings: that is, the primary surface which contains
the pigment granules is not an extracellular, but an intracellular structure.
So far as the primary surface is an intracellular structure, it must inevitably
be considered that the primary surface is not a syncytium but is composed of
discrete structures separated from each other by plasma membranes.
Thus there remains the difficult question of how the movements of discrete
primary surfaces give rise to mass movement of cells.
In our time-lapse microcinematography (N. Ikushima & S. Maruyama,
unpublished observations) it can be seen, as Holtfreter has pointed out, that
in cleaving eggs the pigmented surface which has once been divided by a cleavage
furrow behaves as if it re-establishes a continuous envelope. Moreover, it is
observed that the primary surface does not adhere to the primary or to the
secondary surface. Only the secondary surface adheres to that of another cell.
When dissociated discrete cells come in contact, adhesion always begins and
proceeds exclusively between their secondary surfaces. At the same time, it
seems worthy of notice that the adhesion of cells appears to be stabilized as
soon as the borders of the primary surfaces of respective cells come in contact
with each other. In other words, it appears that the border of the primary
surface of a cell has a particular adhesiveness to that of an adjacent cell. In
relation to this point, electron-microscopic studies in amphibian gastrulae show
that the cells composing the outer surface adhere to each other very closely
just at their distal ends and are probably joined by some cement substance
which is not easily broken (Balinsky, 1965). If it is true that cells of an early
embryo are connected with each other by cement substances which are secreted
outside cells just at the border of the primary surface, it may consequently
be conceivable that the intracellular, discrete primary surfaces are connected
with each other by cement substances across the plasma membranes. Of course
this does not mean that the primary surfaces are morphologically integrated
into a true syncytial layer, but it does indicate the possibility that discrete
Development of dorsal marginal zone
275
primary surfaces are functionally organized into a syncytial unit. On this
point, however, further studies would be necessary.
ZUSAMMENFASSUNG
Aufbau und Entwicklungstendenz der dorsalen Randzone in der friihen
Gastrula von Amphibien
Die Oberflache, die ein befruchtetes und ungeteiltes Ei umfasst, wird durch Zellteilung
abgeteilt und in die einzelnen Zellen ausgeteilt. Diese nennen wir eine primare Oberfiache.
In einer friihen Morula findet man die zweiartigen Oberflachen in einzelnen Zellen; die
eine ist die primare Oberfiache, und die andere, die sich nicht an dem Umfang des Keimes
beteiligt, ist die sekundare Oberfiache. Die strukturelle Verschiedenheit ist elektronenmikroskopisch zwischen zwei Oberflachen gezeigt.
Wenn ein Stuck aus der dorsalen Randzone isoliert und in die Feldmansche Losung
getaucht wird, kann es in die zwei Schichten geteilt werden. In der ausseren Schicht hat jede
Zelle die primare und sekundare Oberfiache. In der inneren Schicht hat jede Zelle nur die
sekundare Oberfiache.
Das Explantat der ausseren Schicht stellt die dreiartigen Gestaltungsbewegungen dar;
Ausbreitung, Konvergenz mit Streckung und sich Kugelung. Dieselbe Bewegungen finden
sich in gleicher Weise in dem normalen Explantat mit der ausseren und der inneren Schicht.
Das Explantat der inneren Schicht ist kinetisch sehr trage und stellt nur schwache Ausbreitung
und sich Kugelung dar.
In dem Explantat der ausseren Schicht werden die axialen, mesodermalen Strukturen
gebildet, aber gar nicht in dem Explantat der inneren Schicht.
Wenn ein Stuck der inneren Schicht ins Blastocol eingesteckt wird, induziert das Implantat
die deuterencephalen und spinalen Strukturen. Das Implantat wird gleichzeitig Chorda
und Somiten. Dieselbe Induktion und Differenzierung finden sich in gleicher Weise in der
Implantation der ausseren Schicht.
Die Verschiedenheit der Gestaltungsbewegungen und der Differenzierungsfahigkeiten
zwischen den ausseren und den inneren Schichten mag aus dem Grund geschehen, daB die
aussere Schichte die primare Oberflache, aber die innere Schicht sie nicht habe.
Funktion der primare Oberflache iiber die Gestaltungsbewegungen und die Differenzierungsfahigkeiten der dorsalen Randzone werden erortert.
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