/. Embryol. exp. Morph. Vol. 21, 2, pp. 309-29, April 1969
Printed in Great Britain
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Effects of (3-mercaptoethanol on the fine structure
of the neural plate cells of the chick embryo
By PAUL-EMIL MESSIER 1
From the Laboratoire de Morphologie Animate,
Universite Libre de Bruxelles
In the course of his studies on mitosis in sea-urchin eggs, Mazia (1958 a, b)
first used mercaptoethanol (HS-CH2-CH2OH) in biological research. This
strongly reducing sulphydryl reagent penetrates easily into living cells and is
relatively non-toxic. In an extensive series of studies Brachet and coworkers,
using this substance, have illustrated its remarkable inhibitory effects on
regeneration in a large variety of biological specimens, including nucleate or
anucleate fragments of Acetabularia mediterranea, cephalic regions of Planaria,
the tail bud of tadpoles (Brachet, 1959) and bud regeneration in Hydra (DescotilsHeernu, Quertier & Brachet, 1961). Moreover, light-microscope and biochemical
studies have shown that mercaptoethanol is a powerful inhibitor of neurulation,
both in amphibians (Brachet, 1959,1963; Brachet et al. 1961; Malpoix, Quertier
& Brachet, 1963; Pohl & Quertier, 1963) and chick embryos (Pohl & Brachet,
1962; Pohl & Quertier, 1963).
The purpose of the present work is to give an account of the ultrastructural
changes undergone by the neural plate cells of the chick embryo when neurulation is inhibited with mercaptoethanol. As a basis of comparison normal
embryos are studied. Their age ranges from stage 5 (Hamilton, 1952), when the
neural plate is a completely flat cellular leaflet, to stage 11, at which time the
neural tube, now composed of cylindrical-shaped cells, has closed. In the course
of this investigation special attention was given to cytoplasmic microtubules.
The occurrence of these structures in differentiating motor neuroblasts of the
normal chick embryo has been discussed at length in Lyser's (1968) recent
work. However, the youngest embryos she described already had a closed
neural tube. Therefore, cytoplasmic microtubules will be considered in connexion
with the early morphogenetic movements of neurulation.
MATERIALS AND METHODS
In our investigations of normal untreated specimens chick embryos were
either explanted at stages 5-11 of Hamilton's table (1952) and immediately
1
Author's address: Departement d'Anatomie, Faculte de Medecine, Universite de
Montreal, C.P. 6128, Montreal, P.Q., Canada.
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fixed, or explanted at stage 5 and cultivated at 38 °C up to stage l i o n Spratt's
medium (1950). Three ml of this medium were poured into a watch glass and the
latter seated on damp cotton-wool in a Petri dish.
Embryos to be treated were explanted at stage 7+ or 8 - (2 or 3 somites)
and cultivated for 7-5 h at 38 °C on Spratt's medium, but with 0-01M mercaptoethanol added. Control embryos, developing on standard media, were cultivated, fixed and embedded at the same time as the treated ones.
The observations reported here are based on the study of twenty normal
embryos and fifty treated ones.
All embryos were fixed for 90 min in 1-25% phosphate-buffered glutaraldehyde (Dossel, 1966, personal communication). After dehydration in ethanol
solutions, the specimens were embedded in Araldite. To ensure that all embryos
would be sectioned transversely, they were embedded in flat capsules made of
aluminium foil. After polymerization the foil was discarded and the embryos,
which could easily be seen in detail with the binocular microscope, were
prepared for microtomy. The tissue blocks were handled so that the sections
would come from the middle region of the neural plate for stages 5 and 6
(i.e. half-way between the outer tip of the cephalic region and Hansen's node)
and from the region immediately in front of the first somites for stages 7-11.
Moreover, to obtain longitudinal sections of this region, a small piece of two
embryos of stage 11 was cut out from the first-somite level and re-embedded.
The thin sections, obtained with an LKB Ultrotome, were stained at room
temperature, first with 0-5 % uranyl acetate in 50 °C ethanol for 1 h, and then
with lead citrate according to Reynolds (1963) for 20 min. They were examined
in an A.E.I, electron microscope.
OBSERVATIONS
Normal untreated embryos
The material to be described here was obtained either from embryos that were
immersed in fixative immediately after they were separated from the yolk or
from embryos that were fixed only after having been incubated for varying
lengths of time on unmodified (see Materials and Methods) culture media. Since
no ultrastructural differences could be detected between embryos of either source,
they will be considered together. Only those structures that are morphologically
altered in the mercaptoethanol-treated embryos will be dealt with in the following description of the normal embryos. Considerable attention is given to
microtubules throughout the following description, since their occurrence will
be discussed at the end of this work.
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The scale line in the figures represents 1/t, with the exception of Fig. 5 A, where it
represents 0-1/4.
Fig. 1. Stage 5 (head process). (A) Transverse section through the primitive streak
region. Most cells are round and show irregular contours. An impressive degree
of free space prevails between cells; cytoplasmic processes are seen (c). The upper
cell has its longer axis parallel to the groove, x 8200. (B) Enlarged portion of the
preceding figure. Note that microtubules (m) are in an orientation parallel to that
of the cell's longer axis. At the bottom right, part of a mitochondria is seen. It
shows well-formed cristae traversing a fine granular matrix. Endoplasmic reticulum (r) is scant and few ribosomes are attached to its membranes. Glycogen is
seenO). x 41000.
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Stages 5 and 6 {head process and head fold)
Primitive streak region
At this early stage of embryonic development, a series of embryos were cut
transversely in the primitive streak region where, excepting the groove, a flat
leaflet of cells occurs and where no definite rolling-up forces are yet involved.
Most cells in this region are small, round-shaped and show irregular contours
(Fig. 1A). In such cells microtubules appear orientated at random (Fig. 5 A).
Exceptionally wide intercellular spaces occur between the cells making up this
leaflet and short cytoplasmic finger-like processes project into them (Fig. 1 A).
Such processes, though very few in number, are sometimes seen protruding
from the otherwise entirely smooth apical membrane (Fig. 5 A). A few elongated
cells with their long axis perpendicular to the groove can be observed. Others,
definitely less numerous, have their long axis parallel to the groove (Fig. 1 A).
In such a case, microtubules are seen in a direction parallel to that of the cell's
longer axis (Fig. 1A, B).
Throughout the cytoplasm of these cells, numerous mitochondria, with a
random distribution, are observed. According to their orientation in the
sections, they appear as small round-shaped or oblong bodies. They generally
show well-formed cristae traversing a fine granular matrix (Fig. 1 A).
The endoplasmic reticulum is remarkably scanty. It appears in the form of
short, closely applied membranes sparsely studded with ribosomes (Fig. 1B).
Neural plate region
The rather flat neural plate of the embryos at the head-process and head-fold
stages is composed of a pseudostratified epithelium, the cells of which show a
definite tendency towards the elongate form (Fig. 2 A). The nuclei show the same
tendency and have their long axis parallel to that of the cell which is perpendicular to the groove. However, as shown in Fig. 2 A, not all nuclei conform to this
general pattern. The nuclear ground substance of all the nuclei is made up of
homogeneous dispersed fine particulate material (Figs. 2 A, 5D); denser
granular areas of chromatin are also seen. The nucleolus is prominent (Fig. 5D).
Again the cells are separated by wide intercellular spaces into which slender
Fig. 2. (A) Stage 5 (head process): transverse section through the neural plate
region. Cells here show a definite tendency towards the elongate form. Most nuclei
also tend to be elongated. The nuclear ground substance of nuclei is made up of
homogeneously dispersed fine particulate material (see also Fig. ID). Free
intercellular space is evident. The groove occupies the top of the figure, x 4000.
(B) Stage 7 (one somite): longitudinal section of a centriole of a neural plate cell.
Centriolar cylinders are in close association with microtubules. The pericentriolar
vesicle on the right is seen to constrict at one end and to go on in the form of closely
parallel membranes (arrow). The axis of this centriole was parallel to that of the
cell and the microtubules were directed towards the nucleus, x 55000.
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cytoplasmic projections are seen (Figs. 2 A, 3 A). Such projections, short and
finger-like, often protrude from the apical membrane of the cells bordering
the groove (Fig. 2A); occasionally they are swollen and empty of cytoplasmic
material. In these transverse sections of neural plates most micro-tubules
appear to be orientated parallel to the cell's longer axis (Fig. 3B), but a
few microtubules are seen in perfect cross-section. They are generally, though
not necessarily (Fig. 3 A), confined to the apical region of the cells and often to
the vicinity of desmosomes (Fig. 3B).
These cells show numerous mitochondria. They appear as round or elongate
bodies (Fig. 3 A). When thin and elongate, they lie parallel to the cell's longer
axis (Fig. 3 A). They show typical cristae and a dense fine particulate matrix.
Profiles of endoplasmic reticulum are only slightly more abundant than in
sections of the primitive streak region. Indeed, at the neural plate level the still
scarce endoplasmic reticulum is observed in the form of short, closely applied
membranes imperfectly studded with ribosomes (Fig. 3). On the other hand, the
Golgi complex is an outstanding feature of these cells. It is always well developed
and usually occurs in the perinuclear area (Fig. 3 A). It is composed of long flat
cisternae, the content of which is moderately opaque to electrons. Other
elongate but shorter cisternae are slightly distended (Fig. 3A). The whole
system of flat and slightly dilated cisternae is constantly orientated parallel to
the cell's longer axis. Small vesicles, sometimes with densely staining contents,
also make up part of the apparatus (Fig. 3 A).
Stage 7 (one somite)
Thin sections of the neural plate cut transversely in front of the first somite in
the stage 7 embryos show features comparable to those observed on sections
from the mid-region of the neural plate at the head-process and head-fold
stages. However, the apical membranes of the cells bordering the groove are
becoming more and more indented and many short cytoplasmic projections are
visible.
Fig. 3. Stage 5 (head process). (A) Transverse section through neural plate. Mitochondria are highly elongated; their longer axis parallels that of the cell. They
show typical cristae and a dense fine particulate matrix. A well-developed Golgi
shows its cisternae system typically orientated in a direction parallel to that of the
cell's longer axis. Profiles of endoplasmic reticulum (r) are only slightly more
abundant than they were in the primitive streak region. It is in the form of short,
closely applied membranes, imperfectly studded with ribosomes. Some microtubules (arrows) are seen in cross-section in this middle region of a cell. Free
intercellular space and cytoplasmic process (c) are seen, x 41000. (B) Transverse
section through neural plate. In these cells, definitely sectioned longitudinally,
some microtubules are seen in perfect cross-section (single arrows). These are
generally confined to the apical region of the cells and often to the vicinity of
desmosomes. Other microtubules (double arrows) are seen longitudinally orientated,
x 30000.
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Again, in elongated cells, microtubules are observed. Most of the microtubules encountered are longitudinally orientated. However, some, usually
confined to the apical region of the cell, are seen in cross-section.
Among these embryos one neural plate cell sectioned longitudinally exhibited
a centriole, the units of which were in close association with microtubules. As
shown on Fig. 2B, some of the centriole cylinders lose their density towards one
end of the organelle and are seen in continuity with microtubules. One pericentriolar vesicle, constricted at one end, tapers off into closely parallel membranes highly reminiscent of microtubule structure.
This centriole was situated apically and orientated in a direction parallel to
that of the cell's longer axis. Microtubules were continuous to the centriolar end
facing the nucleus, towards which they were directed.
In embryos at this stage cells of the mesoderm and of the ectoderm were also
studied. In both types microtubules were seen.
Stage 9 (six somites)
At this stage the neural plate has rolled up considerably, and in sections cut
in front of the first somite it appears as a U-shaped open neural tube, the wall
of which is a pseudostratified columnar epithelium.
When these highly elongated cells are sectioned longitudinally, many microtubules are seen parallel to the cells' long axis (Fig. 4). However, a few microtubules do not conform to this general pattern.
The other cytoplasmic organelles of the six-somite embryos show features
comparable to those already described in earlier stages.
Stage 11 (thirteen somites)
The embryos described in this section represent the true controls.
Embryos sectioned in front of the first somites of the thirteen somite embryo
show a closed neural tube, the wall of which consists of pseudostratified
columnar epithelium. The amount of free space between the cells is considerably
reduced. The apical membranes bordering the neurocoel are quite irregular,
showing many indentations and short finger-like projections (Fig. 5B).
The outer edge of the neural tube is smooth and regular; the cells are aligned
along a regular base line and possess a continuous basement membrane
(Fig. 5C).
In these cells cut longitudinally, microtubules nearly always show a longitudinal orientation. Only very exceptionally were there a few microtubules,
generally towards the cellular apex, seen in oblique or cross-section. Two embryos
of this stage were cut out of the tissue blocks and re-embedded so as to produce
sections perfectly transverse to the cell's longer axis. In these sections all microtubules appeared in cross-section and their distribution seemed rather homogeneous throughout the cell (Fig. 6).
Effects of mereapt oethanol
Fig. 4. Stage 9 (six somites). Transverse section through the neural plate, showing
part of several cells. Note the abundance of microtubules. They are orientated in
a direction parallel to that of the cells' long axis, x 41000.
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319
The nuclei are highly elongated in the direction of the cell's longer axis. Their
nuclear ground substance appeared as a homogeneously dispersed fine particulate material and the chromatin as irregular, dense granular material around
the periphery of the nucleus (Fig. 7 A). The nucleolus is prominent.
In the neural tube cells of these embryos many mitochondria were observed.
They are abundant towards either end (apical-basal) of the cell and less numerous
in the immediate vicinity of the nucleus. In sections some appeared as small
round-shaped bodies, others as elongated organelles (Fig. 5B). They showed
characteristic cristae traversing a moderately granular matrix.
The endoplasmic reticulum, as in younger embryos, was scanty. It appeared
throughout the cytoplasm in the form of small cisternae, the membranes of
which were partly studded with ribosomes (Fig. 6).
Treated embryos
Embryos explanted at stages 7+ or 8 - of Hamilton's table (1952) and
treated for 7-5 h with 0 0 1 M mercaptoethanol have their morphogenetic
movements inhibited. As shown by Pohl & Brachet (1962), under such conditions
the closure of the nervous system is completely and permanently blocked.
Transverse sections of the embryos in front of the first somite show a wide,
open nervous system, still appearing in the form of a plate. The pseudostratified columnar epithelium making up the neural plate is composed of
closely packed cells between which very little, or no, free space is seen (Fig. 8).
The apical end of the cells bordering the groove is very irregular, highly
indented and exhibits slender finger-like projections, larger cytoplasmic projections and often apical blebs (Figs. 8, 9A). Cytoplasmic blebs, usually containing ribosomes and often endoplasmic reticulum and even mitochondria, are
seen to protrude freely into the groove. Such puffs or apical blebs of neighbouring
Fig. 5. (A) Stage 5 (head process). Primitive streak region. In the same cell some
microtubules are sectioned transversely (single arrows), others obliquely, while
a last group appear longitudinally orientated in the thickness of the section (double
arrows). In the upper left-hand corner note short finger-like process protruding in
the groove, x 55000. (B) Stage 11 (thirteen somites). Transverse section through
the neural plate. Part of many elongated cells may be seen. The apical membranes
bordering the neurocoel are irregular, showing indentation and short finger-like
projections. The amount of free space between cells has considerably reduced,
x 6000. (C) Stage 11 (thirteen somites). Cross-section through the neural plate.
The outer edge of the neural tube is smooth and regular cells have their base in line
and are covered by a continuous basement membrane, x 6000. (D) Stage 5 (head
process). Cross-section through the neural plate. Part of a nucleus is shown. The
nuclear ground substance is made up of homogeneously dispersed fine particulate
material, and denser areas of chromatin are seen (double arrows). The nucleolus is
prominent and characterized by its close-meshed nucleolonema (single arrows),
x 20000.
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cells may come close together but have never shown specialized zones of cellular
contact such as desmosomes.
In some cells, immediately beneath the apical membrane, a 0-5/6 thick region
of fine fibrillar material containing no other cytoplasmic inclusions was occasionally observed (Fig. 9B).
In this material the nuclei tend to be elongated. All have their longer axis
parallel to that of the cell. Most of the nuclei assume a regular outline, though
a few (as seen on Fig. 8) have irregular contours. Compared to the nuclei of the
normal embryos, all here have a more or less washed-out appearance. Interestingly, the more pronounced this appearance, the more regular is the
outline of the nucleus. In these nuclei the ground substance is in the form of a
delicate, fibrillar material (Fig. 7B). Chromatin is observed as small areas of
moderately dense granular material freely dispersed throughout the nucleus,
irregularly disposed around its periphery or surrounding the nucleolus (Fig. 7B).
The latter, prominent and highly contrasted, has a ribbon-like appearance at
low magnification (Fig. 8) and reveals very dense granular material when viewed
at higher magnification (Fig. 7B). It encloses many small rounded areas whose
density compares with that of the ground substance.
In these embryos the outer edge of the neural plate always showed a festooned
appearance, each festoon corresponding to the basal end of one cell (Fig. 8).
A basement membrane, with some interruptions, follows the irregularities of the
basal end of these cells.
In mercaptoethanol-treated embryos microtubules are abundant. They
usually lie parallel to the long axis of the cell; occasionally some, usually
situated at the apical end of the cell, are seen to be perpendicularly orientated to
that axis (Fig. 9 A). At high magnification their morphology seems comparable to
that observed in control embryos.
All the mitochondria in these cells are morphologically altered after mercaptoethanol treatment. They are still numerous, generally distributed apically and
basally to the nucleus, but they now assume a round shape and their diameter
ranges from 0-5/6 to approximately 2JLI (Fig. 8). Usually a variable number of
cristae persist, but they are often considerably reduced in number and size,
remaining only as small remnants attached to the mitochondrial membrane.
The matrix is low in density and is filled with moderately dense amorphous
material (Figs. 8, 9).
The endoplasmic reticulum is seen in the form of very long flat cisternae, the
Fig. 6. Stage 11 (thirteen somites). Longitudinal section through the neural plate.
These cells sectioned transversely through their longer axis show fifty crosssections of microtubules dispersed homogeneously throughout the cells. The
endoplasmic reticulum is scant, its membranes are imperfectly studded with
ribosomes. x 55000.
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membranes of which are studded with ribosomes. These cisternae are found
throughout the cytoplasm and are particularly abundant in the apical region of
the cell (Figs. 8, 9B).
DISCUSSION
The only other mention of the presence of microtubules in the cells of the
nervous system of the chick embryo is that of Lyser (1968), where their occurrence
in neuroblasts of the closed neural tube is pointed out and discussed.
In the present work they are seen as a constant feature as early as the headprocess stage and even in cells of the primitive streak region. Moreover, microtubules are by no means confined to these cells, for some have been seen in
mesoderm and ectoderm cells.
The study of the temporal changes in cell shape, from the early cuboidal cells
of the primitive streak of the head-process stage to the columnar ones of the
closed neural tube, has demonstrated a concomitant change in the general
scheme of microtubular orientation. Indeed, in the youngest embryos studied
(stages 5-6, primitive streak region) microtubules appear orientated at random
(Fig. 5 A). At the same stages in the neural-plate region fewer microtubules are
orientated at random and most tend to lie parallel to the long axis of the cell
(Fig. 3 A). Later in development (stages 7 and 11) fewer and fewer microtubules
were found to deviate from this longitudinal orientation.
Mercaptoethanol provided little information on the possible role(s) of microtubules. After treatment, microtubules were numerous and morphologically
unchanged. The only possible discrepancy lies in the observation of a greater
number of microtubules having an orientation parallel to the groove (Fig. 9 A),
but this finding was so sporadic that it cannot be the sole explanation of the
morphostatic effect of mercaptoethanol. On the other hand, mercaptoethanol is
reputed to interfere with the assembly of precursor protein molecules into
ordered structures such as microtubules (Gavin & Frankel, 1966; Wade &
Satir, 1968), and this would offer a possible means through which this substance
could act in blocking neurulation. Mercaptoethanol, which, from our experiments, does not seem to destroy already formed microtubules, could, as
suggested by Wade & Satir (1968), react with free -SH groups of the newly
synthesized component protein, thus preventing microtubule polymerization.
Fig. 7. (A) Stage 11 (thirteen somites). Transverse section through the neural plate
of a normal embryo. Parts of two nuclei are seen. The nuclear ground substance is
in the form of homogeneously dispersed fine paniculate material and chromatin is
observed as denser granular material disposed around the nucleus (single arrows).
The prominent nucleolus is characterized by its close-meshed nucleolonema (double
arrows), x 27 500. (B) Treated embryo. Transverse section through the neural
plate. Part of a nucleus is shown. The ground substance is in the form of a faint,
delicate, fibrillar material. Chromatin is observed as small areas of moderately
dense granular material freely dispersed throughout the nucleus. The nucleolus
shows very dense granular material in its core, x 27500.
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Even though microtubules were not morphologically altered in the treated
embryos, the possibility of their being involved in neurulation must be retained,
for it is conceivable that the changes induced by mercaptoethanol are not
detectable with the techniques presently used. More work, preferably concerned
directly with microtubules, is needed before one can safely conclude about their
role in cell deformation.
After mercaptoethanol treatment there is a complete disappearance of the
free spaces that were observed between cells in the normal embryos throughout
the time sequence studied (Figs. 1, 8). It would be tempting to explain this
discrepancy on the basis of Jurand & Tuft's (1961) observation of a disturbance
in water distribution after mercaptoethanol treatment. Certainly the appearance
of the basal regions of the cells, the rather turgid aspect of some apical ends and
the general swollen aspect of mitochondria are other evidences supporting this
view of cellular water imbalance. It remains unclear, however, whether the
primary effect of mercaptoethanol is on mechanisms responsible for the distribution of water (Tuft, 1961) or, as advocated by Brachet, on the mechanisms
responsible for morphogenetic movements. Indeed it is well known that osmosis
alone cannot account for mitochondrial changes in volume and many facts show
that their enzymic state is involved in the maintenance of their morphology
(Klein & Neff, 1960; also see Ernster & Lindberg, 1958). Obviously, much more
work will be needed before one can discover the type of enzymic or energetic
imbalance which is introduced by the action of mercaptoethanol and it would
be interesting to know if the lack (or decrease) of ATP, which is reputed to induce
mitochondrial swelling (see Ernster & Lindberg, 1958), is absent in mercaptoethanol-treated chick embryos as Brachet et al. (1961) have shown it to be in
amphibian eggs after a similar treatment. Furthermore, notwithstanding the
profound ultrastructural changes observed in our treated embryos, energyrequiring membrane synthesis is enhanced, as illustrated by the appearance of
the endoplasmic reticulum.
Finally, we are inclined to agree with Brachet, Decroly & Quertier (1963) and
feel it necessary to repeat that, taking into consideration (1) the very diverse
biochemical effects of mercaptoethanol (Brachet et al. 1961, 1963), (2) the
experimental results obtained by Malpoix et al. (1963) with explants, (3) the
incompatibility between Tuft's view and the inhibitory effects of mercaptoethanol on regeneration (see Introduction) and (4) evidence given here, it seems
Fig. 8. Treated embryos. Transverse section through the neural plate. No free space
is observed between these cells. The apical end of the cells bordering the groove is
very irregular, highly indented and exhibits slender finger-like projections (/),
larger cytoplasmic projections (single arrow) and apical blebs (double arrows). The
outer edge of the neural plate shows a festooned appearance, each corresponding
to the basal end of one cell. Nuclei have a washed-out appearance. Numerous
mitochondria are seen: most cristae are erased, x 4000.
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unlikely that water disturbance is the sole cause of the inhibition of morphogenetic movements following treatment with mercaptoethanol.
To conclude, it may be said that mercaptoethanol has profound effects on the
ultrastructure of the neural plate cells of the chick embryo. The modified general
cellular outline and the swollen aspects of mitochondria it causes are suggestive
of a water imbalance that could initiate mitochondrial swelling, then modify
ATP production and therefore prevent the contraction of a fibrous protein,
such as that proposed by Brachet and coworkers (Brachet et al 1961) as an
active force in neurulation.
SUMMARY
The effects of mercaptoethanol on the neural plate cells of stage 7 + and 8 —
chick embryos have been studied with the electron microscope. The ultrastructure
of the neural plate cells in normal embryos ranging from head process to
stage 11 were also studied. Moreover, at the head-process stage cells of the
primitive streak region were studied.
In normal embryos we found that: (1) wide intercellular spaces occur between
cells in younger stages; they are progressively reduced as embryos grow older;
(2) microtubules are present at all stages; (3) in younger stages microtubules
appeared orientated at random and in time more and more come to lie in a
direction parallel to the long axis of the cell; (4) some microtubules, mostly
apically situated, are always orientated in a direction parallel to the groove (or
the neurocoel) but their number decreases in time; (5) at the head-process stage
mitochondria in the cells of the primitive streak region are numerous and small,
whereas those of the neural plate cells, from that stage on, are numerous and
highly elongated; (6) at the head-process stage the Golgi apparatus is inconspicuous in the cells of the primitive streak region while it is highly developed in
the neural plate cells; (7) the endoplasmic reticulum remains scant throughout the
time sequence studied.
In the treated embryos we found that: (1) no free space exists between cells;
(2) the basal end of the cells is festooned and the apical region has a turgid
aspect; (3) microtubules are present and sporadically groups of them are
Fig. 9. Treated embryos. Transverse section through the neural plate. (A) Apical
part of four neural plate cells. Beyond the terminal bars turgid cytoplasmic puffs, or
blebs are seen to protrude freely in the groove. Such blebs of neighbouring cells
may come close together (upper right) but no specialized zone of cellular contact is
seen. In one cell numerous microtubules are shown (double arrows) in line with the
cell's longer axis. In another cell microtubules are perpendicularly orientated (single
arrows) to the cell's longer axis. Mitochondria (M) show absence of cristae. x 13 700.
(B) Apical part of a neural plate cell. Beneath the apical membrane (upper left
corner) a 0-5 /* thick zone of fine fibrillar material is seen. Endoplasmic reticulum (r)
is in the form of very long flat cisternae, the membranes of which are studded with
ribosomes. x 41000.
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orientated in a direction parallel to the groove; (4) mitochondria are swollen and
most cristae have disappeared; (5) definitely more endoplasmic reticulum
membranes are present; (6) nuclei have a washed-out appearance.
The meaning of these results is discussed.
RESUME
Effets du P-mercaptoethanol sur les ultrastructures des cellules de la
plaque neurale d" embryons de poulet
Les effets du mercaptoethanol sur les cellules de la plaque neurale d'embryons
de poulet aux stades 7+ et 8— ont ete etudies au microscope electronique.
L'ultrastructure des cellules de la plaque neurale d'embryons normaux depuis
le stade 'processus cephalique' jusqu'au stade 11 est aussi analysee. De plus,
les cellules de la ligne primitive ont ete etudiees au stade 'processus cephalique'.
Chez les embryons normaux, il a ete observe que: (1) Aux stades jeunes, les
cellules sont separees par de grands espaces intercellulaires; ceux-ci sont
progressivement reduits au cours du developpement. (2) Des microtubules sont
presents a tous les stades. (3) Aux stades jeunes, les microtubules sont orientes
au hasard; au cours du developpement il s'en trouve de plus en plus, orientes
suivant le grand axe de la cellule. (4) Dans la region apicale des cellules il y a
toujours des microtubules orientes parallelement a la goutiere neurale (ou au
neurocele) mais leur nombre decroit au cours du developpement. (5) Au stade
'processus cephalique', les mitochondries dans les cellules de la region de la
goutiere primitive sont petites et nombreuses. Par contre, dans les cellules de
la plaque neurale, depuis ce stade (proc. ceph.) et pour le reste du developpement,
les mitochondries sont nombreuses et tres allongees. (6) Au stade 'processus
cephalique' l'appareil de Golgi des cellules de la region de la goutiere primitive
est a peine developpe alors qu'il est tres developpe dans les cellules de la plaque
neurale. (7) Tout au long des stades etudies le reticulum endoplasmique reste
peu developpe.
Chez les embryons traites, il a ete observe que: (1) II n'existe pas d'espace
intercellulaire. (2) La partie basale des cellules est 'festonnee' et leur region
apicale est boursouflee. (3) Les microtubules sont presents et, sporadiquement,
il s'en trouve des faisceaux orientes parallelement a la goutiere. (4) Les mitochondries sont gonflees et leurs cretes sont disparus. (5) Le reticulum endoplasmique est plus abondant que chez les temoins. (6) Les noyaux sont vides de
leur contenu.
We wish to express our gratitude to Professor J. Brachet, who welcomed us in his Laboratory and suggested this line of work. We also wish to thank Dr P. Van Gansen for making
available to us the facilities of her E.M. section and Dr P. Malpoix for reading and improving
the English composition of this text. The author was the recipient of a NATO post-doctorate
fellowship.
Effects of mercaptoethanol
329
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{Manuscript received 5 September 1968)