DEVELOPMENTAL BIOLOGY, 8, 186-204 ( 1963)

Ultrastructural Differentiations during Formation of the

Blastoderm in the Drosophila melanogaster Embryo1

ANTHONY P. MAHOWALD

Department ofBiology, The .lohns Hopkins University,

Baltimore 18, Maryland, and
Woodstock College, Woodstock, Maryland

Accepted August 3, 1963

INTRODUCTION

The ultrastructure of the cleavage furrows during the transition

from the syncytial blastema to the cellular blastoderm in the Drosoph-
ila embryo has been recently described (Mahowald, 1963). During
the 30 minutes needed for this transition, other modifications of the
cytoplasmic ultrastructure occur. In this paper these changes will be
described and the evidence found for differential localization of cyto-
plasmic organelles will be presented.

MATERIALS AND METHODS

The wild-type stocks of Drosophila melanogaster, the preparatory

techniques for collection of embryos, and the methods of fixation used
are the same as those described in a previous report (Mahowald,
1963). OsO,, I%, buffered at pH 7.8-8.0 with Verona1 acetate and with
Zetterqvist’s balanced salts added, was the most successful fixative.
For unknown reasons, fine structure was not preserved if sucrose was
added to the osmium fixative instead of salts. Although differences
between osmium and permanganate fixation were noted (see below),
most of the electron microscopy was done on sections of osmium-fixed
embryos because of the difficulties found in attempting to remove the
vitelline membrane after permanganate fixation. Unless this membrane

1Portions of this work are taken from a thesis presented in partial fulfillment
of the requirements of a Doctor of Philosophy degree at The Johns Hopkins
University. The author was a National Science Foundation Predoctoral Fellow at
this time. Part of the research was supported by research grant GB-309 to
Woodstock College from the National Science Foundation.
186
 ULTRASTRUCTURE OF DROSOPHILA BLASTODERM 187

is removed, fixation is poor. Embryos were usually embedded in

Araldite. Sections were mounted on 200-mesh grids, coated with a
thin carbon film, or on SjGstrand-type grids also coated with a carbon
film. Karnovsky’s alkalized lead stain (1961) was used to increase
contrast in most cases, although other types of lead stains and satu-
rated uranyl acetate were also used. Mitochondrial counts were made
on electron micrographs printed at about X20,000. Full details of
materials and methods used can be found in a previous report
( Mahowald, 1963).
OBSERVATIONS
General Fine Structure
At fertilization, yolk and cytoplasmic components are evenly dis-
tributed throughout the egg of Drosophila except for the narrow strip
of pcriplasm at the periphery. During the early cleavage divisions the
nuclei are found in protoplasmic islands scattered throughout the
central part of the embryo. At the time of the 8th and 9th cleavages
most of the nuclei migrate into the periplasm and during the next
three divisions a nearly complete separation of yolk from the cytoplasm
occurs (Fig. 1). A few nuclei remain behind in the yolk (often
called vitellophages or yolk nuclei); their function remains unknown.
The pole cells form at this time. Three more synchronous divisions of
the cleavage nuclei occur at the periphery during the syncytial blas-
tema stage, Immediately after the last blastema division, cell mem-
branes form and nucleoli appear for the first time. These pre-
blastoderm stages have been thoroughly studied cytologically and
the literature reviewed by Sonnenblick (1950). Some observations on
the fine structure have been previously described (Mahowald, 1963;
Okada and Waddington, 1959).
At the end of the blastema divisions, the spherical nuclei are only
34 ,J,in diameter and are located about 1 ,J.from the surface. During
the formation of the cellular blastoderm they increase in size mltiI they
are about 4-5 ,L wide and 10-15 ,J.long (Fig. 1). Nucleoli first appear
at the outer edge of nuclei as small bodies (Fig. 2). Before the cell
furrows have advanced more than a fourth of the length of the
nuclei, full-sized nucleoli are evident. Their fine structure is typical
and usually a nucleolonema-like structure can be found (Fig. 14).
The nuclear envelope possessestypical annuli at all stages studied
(Figs. 2 and 15). The outer membrane bears dense particles, 130-
188 ANTHONY P. MAHOWALII

FIG. 1. Section (0.5 p thick), stained with toluidine blue, showing the
cytology of the Drosophila embryo at the blastoderm stage. The central region is
mostly yolk and a few yolk nuclei are visible. Around the yolk is the blastoderm
with the nuclei at the periphery. Cell furrows are at the base of the nuclei.
Fragments of the vitelline membrane which were not removed during fixation still
adhere to the edge (arrow) and distort that region of the embryo. Araldite em-
bedding. Magnification: x 380.
 ~JL,THASTRUCTURE OF DROSOPHILA BLASTODERM 189

150 h in size, which probably correspond to ribosomes. Tangential

sections of the envelope show that the ribosomes are arranged in
spirals or parallel rows (\Yatson, 1959).
The centrides arc always located within an area that is virtually
free of ribosomes and which has a fine granular ultrastructure (Fig.
II). This area is probably identical with the centrosome of classical
cytology (\Vilson, 1928). The centriole is a short cylinder, 160 m,u in
diameter but only 150-175 n-q. in length compared to the typical
length of X30-.500 m/i, (de Harven and Bernhard, 1956) (Fig. 11 ).
The centriole is also atypical in that it has a central tubule 15 111;~
in diameter and appears to have radial “spokes” in addition to the
usual nine sets of cylinders at the periphery (Figs. 3 and 3). Gull
( 1961) occasionally found similar structures in the centriole of the
snail Vioipanrs. Two fully formed centrioles were never found ad-
jacent to each other, but in serial sections structures resembling pro-
centrioles (Bernhard and de Harven, 1960; Gall, 1961) were found
(Fig. 3). Since this stage is immediately after the rapid blastema
divisions (a nuclear division every 10 minutes at 25”C), replication
of centrioles probably occurs later than in other embryos (Mazia
et al., 1960).
The clztloplasnlic reticulum (ER) is present in the preblastoderm
stages of Drosoplzilu embryos in the form of long flattened cisternae,
frequently branched, with ribosomes attached (Fig. 5). Most of the
hasophilia of the cytoplasm, however, is due to numerous unattached
ribosomes which are usually in clusters rather than scattered evenly
in the cytoplasm (Fig. 11).
Amzzllate lanudlae (Swift, 1956) are rarely found in the cytoplasm
of the syncytial blastema, but during blastoderm formation local
differentiations of the flattened cisternae of ER (Figs. 6 and 7)
appear in the blastoderm adjacent to the yolk. These differentiations,

FIG. 2. Low magnification picture of the snrfacc of the cLmbr\o at it time

when the furrows (F) are beginning to form. Sphrricnl nucleoli (Nzr ) are prrs-
ent. The surface of the embryo has manv projcctiony throughout thr stage
studirtl. Araldite cmbcdding, lead stain. hZngnification: x 17,000.
Fx. 3. Cross section of a centriolc showing the 9 0utc.r sets of cylinders
and thta ct~ntral cylinder with radial “spokrs.” Amlditc, cmbcclcling, lead stain.
Magnificntion: X 50,000.
FIG. 4. IIigh magnification of cmtriole. Aralditt ~mlxdding, lrxl stain.
Magnification: x 96,000.
 ANTHONY P. MAHOWALD

FIG. 5. Survey picture of the cytoplasm below the nucleus at a stage when
the furrows (F) are approaching the yolk. Long lamellar ER is present, ex-
tracted lipid droplets ( L), mitochondria, and many dense bodies ( db ) similar to
those described by Karasaki ( 1959 ). Araldite embedding, lead stain. hlagnifica-
tion: X 17,000.
FIG. 6. Local differentiations within a long cisterna of ER, which have the
 ULTRASTRUCTURE OF DROSOPHILA BLASTODERM 191

which have the characteristics of annulate lamellae, may he of the

same ER cisterna folded back on itself (Fig. 6) or of adjacent
cisternae (Fig. 7). In the embryos studied during formation of the
ventral furrow [the LYminute period after completion of blas-
toderm formation ( Sonnenblick, 1950) 1, the annulate lamellae were
larger (Fig. S), having up to 5 lamellae in one stack. Annulate
lamellae have not been found near the nuclear envelope of the
blastoderm nuclei, but these local differentiations of the ER are
found adjacent to the nuclear envelope of yolk nuclei (Fig. 9). In
this location also adjacent lamellae may be differentiations of the
same cisterna, folded back on itself, or of different cisternae. Fewer
annulate lamellae are found adjacent to yolk nuclei during the
blastoderm stage in comparison to earlier stages, and, in contrast, it
is during the blastoderm stage that these structures are first found
in the cytoplasm of the blastoderm cells.
Clusters of agranular ocsicles nrzd rnembmnes (Fig. 6) are present
throughout the cytoplasm. Continuities between these membranes
and the granular ER are common during the late stage of blastoderm
formation but not during early stages. These clusters of agranular
vesicles assume a fine structure more typical of Golgi (two or three
flattened agranular cisternae surrounded by agranular vesicles) dur-
ing late stages of blastoderm formation, especially above the nucleus
(Figs. 11 and 12).
Many mitochondria are present, about 10% showing an oblong
profile; elongate profiles are rarely found, Wessel (1960) has shown
that even if the mitochondria were threadlike, their profiles in thin
sections would usually be oval or round, both because the plane of
section would rarelv cut the mitochondrion lengthwise and because a
threadlike mitochondrion would probably be cut more than once.
Because of the scarcity of long profiles in thin sections, mitochonclria
of the blastoderm are probably not elongate. In preliminary observa-
tions of embryos shortly after the completion of blastoderm forma-
tion, long profiles and branched mitochondria are readily found. This
probably indicates a change in mitocbondrial morphology as embrvo-
genesisproceeds.

nppearance of annulatc lamcllac. A Golgi region (G) is also present, as well as

lipid droplets ( L), mitochondria, d a tlmse bodv. Aralditc cmlxdcling, lend
stain. hlngnification: X 17,000.
192 ANTHONY P. MAHOWALD

FIG. 7. Differentiations within three separate cisternae of ER, which have

the appearance of annulate lamellae. Araldite embedding, lead stain. Magnifica-
tion: X 33,000.
 ULTRASTRUCTURE OF DROSOPHILA BLASTODERM 193

Desrnosomes form very soon after the initial stages of cell furrow
formation. By the end of blastoderm formation, they are very com-
mon above the nucleus, usually located on each side of interdigitating
projections of adjacent cells. They have not been found below the
nucleus, even adjacent to interdigitations. Since the developmental
period studied here is less than 30 minutes long, these organrlles
obviously can differentiate very rapidly.

As tllc nucleus moves away from the surface, a distinct cytoplusmic

region appears between tlic nucleus and the outer plasma mcmbranc.
Thr cytoplasm of this area probably originated from below tlLc
nucleus and flowed past it as the nucleus moved from the surface.
Fine-structllre changes appear very rapidly in this newly segregated
cytoplasmic area which results in the formation of a complex of
agranular membranes, ribosomes, granular KR, and mitochondria.
This supranuclcar complex was first noted in permangaiintc-tixcc1it[,-~x~~l
material, where the membranous features can bc seen (Fig. 10).
Long, irregularly arranged double membranes, usually in groups of
two or thrcr flattened cisternac, are found above the nuclens. One cud
of the group of cisternae joins to a diffuse area where membranes are
difficult to detect.
In osmium-fixed material the distinction between agranular and
granular ER is evident in the supranuclear complex (Figs. 11 and
13). Clusters of agranular vesicles, occasionally with a typical Golgi
appearance (Figs. 11 and 12), are found scattered in the cytoplasm.
Granular tubules and cisternae are frequently found to be continuous
with the agranular membranes. Some of the agranular membranes are
not in clusters, but rather in a loose meshwork of tubules in the
cytoplasm (Fig. 11). The results after permanganate fixation con-
sistently indicated the presence of more long profiles of ER than

FIG. 8. Two groups of annulatc lamellac, one outside the hlastoclerm at

the cdgc of the yolk (Y), the other within the blastoderm cells ( BC ). Con-
tinuitics with the ER nrr not as evident nt this stage (formation of the wntral
furrow), Araldite cmbcdding, lead stain. Magnification: x 17,000.
FIG. 9. Annulate lam&r structures at the edge of a yolk nucleus at the
wcond l&sterna division. The nuclear envelope (NE) is not clearly visible
hecause of the plane of section. Araldite cmbcdding, lead stain. hlagnification:
x 35,000.
 ANTHONY P. MAHOWALD

FIG. 10. Survey picture of the complex of membranes present above the
nuclei of the ventral blastoderm at the end of blastoderm formation. Arrows
indicate the diffuse area mentioned in the text. The vitelline membrane (VM)
is present at the outer edge of embryo. 2% KMnOa fixation, Epon embedding.
Magnification: x 13,000.

were found after osmium fixation. This difference is similar to the

results of Rosenbluth (1963), who found distinct differences in the
preservation of membranes by these two fixatives. Neither the reason
for the difference nor which fixative preserves the structure of the
living state of the membranes best is known.
It is difficult to establish with certainty the sequence of events that
lead to the formation of this new cytoplasmic complex, but at least
some of the consistent features can be mentioned. The nuclear
envelope is frequently considered as a possible source of ER (Porter,
1961a). Continuities between the outer nuclear membrane and the
ER were commonly found only during the early stages when the
nuclei are enlarging; only occasionally were they found in later
stages of blastoderm formation. These continuities at the early stage
could possibly contribute to the formation of the complex, but it is at
 ULTRASTRUCTURE OF DROSOPHILA BLASTODERM 195

F’K:. 11. Osmium-fix4 embryo at thr end of blastodcrm formation on the

ventral side showing the differentiation of the cytoplnsmic membranes above the
nuclei. Flattened Golgi vcsiclrs (G) arc evident at thr top of the figure, while
clusters of agranular vesicles are found ahove the centriole (ce) in the lower
center of figure. Many agranular tulmlcs are found in the upper right quarter
of the figure. The arrows indicate placrs where continuities of agranular and
granular ER are eqxcially evident. Unattached ribosomes are usually present in
clusters. The m&us is located at the base of the figure with the crntrosome
present in an indentation. The periphery of the embryo is about 1 y above the
tap of the figure. Araldite emhcdding, uranyl acetate stain. Magnification:
x 20.000.
196 ANTHONY P. MAHOWALD

FIG. 12. An early stage in the differentiation of the supranuclear complex,

showing the Golgi (G) and tubular ER. Araldite embedding, lead stain. Magni-
fication: x 23,000.
 ULTRASTRUCTURE OF DROSOPHILA HL.AETODERli 197

least equally probable that they are a part of nuclear memlxxne

growth during the stage of nuclear expansion. It would be very
difficult to decide between these alternatives in thin sections. Another
possible site for the origin of the new cytoplasmic membranes is the
Colgi. Since the Golgi are prominent in the region above the nucleus
an d since there are continuities bctnwn them and both the agranular
and granular ER (Fig. ll), it is quite possible that much of the
complex arises from the Golgi bv the frequentlv postulated method of
vesicular budding.
Clusters of granular ER (Figs. 14 and 1s) are regnlarly found
adjacent to the nucleus. or projecting radiallv from the nuclear enve-

FIG. 14. Localization of ER adjacent to the nuclear membrane in the same

region whew the n~deolus is locatrd. The embryo was fixed during an early
stage of complex formation. Araltlitr rmbcdding, Irntl stain. Magnification:
x21,000.
FIG. 15. Cluster of ER adjacent to the blastodcrm nucleus during an ctarly
stage in formation of the supranudear complcs. Th(b arro\V points to an annulus
of nuclwr envelope. Mcthncrl\-late embedding, uranvl a&ate stain. hlagnifica-
tion: X 38,000.

lope but without contacting it. The nucleolus is frequently located in

this region of the nucleus. Thr ER probably does not originate from

FIG. 13. The cytoplasmic complex in the mid-dorsal region of thr embryo
during the ventral furrow stage. No cistrrnae appear to be present, but only a
mass of granular and agranular tubules near the outer edge of the embryo.
Aralclitc embedding, lead stain. Magnification: x 17,000.
198 ANTHONY P. MAHOWALD

the outer nuclear membrane since no continuities between the

nuclear membrane and the ER are found at this time.
At the stage when the furrows have progressed below the nucleus,
differences can be noticed, in the supranuclear complex, between the
mid-dorsal and mid-ventral regions of the embryo. On the ventral
side, the tubules give rise to cisternae (Figs. 10 and 11) which be-
come long and pronounced by the end of blastoderm formation. IIow-
ever on the dorsal side, the complex of agranular and granular tubules
increases in size, but only rarely are cisternae found. This difference is
still evident when the supranuclear complex is examined in embryos
that have completed blastoderm formation (Fig. 13).

Difierential Localization of Mitochondria

Since the mid-dorsal and mid-ventral regions of the blastoderm
have distinct embryonic fates, the former giving rise to extraembryonic
membranes and the hypoderm, the latter to the mesoderm (Poulson,
1950), a more detailed examination of these two regions was made.
Although the more rapid growth of cell furrows on the ventral side
first seen with the light microscope (Mahowald, 1963) was confirmed,
and although the supranuclear complex attained a different “degree”
of differentiation, few additional differences could be clearly demon-
strated. In Table 1 data accumulated on the distribution of mito-

1’,4BLE 1
DISTRIBUTION OF MIT~CHONDRIA ABOVE NUCLEUS

hlid-ventral Rlid-dorsal

Kumber Number
Per per
Age Embryo Area (~2) 100 82 Area (!2) 100 pz

Cleavage furrow at middle of nucleus a 231 38.4 234 46.5

I) 243 45.7 237 36.8
Cleavage furrow at base of nucleus 253 67.9 “23 61.4
Fl 259 49.8 242 38.2
Cleavage furrow next to yolk e 549 60.6 301 36.8
f 674 56.0 676 37.4
g 343 59.0 350 38.8

chondria in the supranuclear region are itemized. Attempts were

made to obtain similar quantitative data for the distribution both of
mitochondria and of lipid droplets in the regions below the nuclei,
but the lack of clear “landmarks” rendered it impossible to obtain
 ULTRASTRUCTURE OF DROSOPHILA BLASTODERhI 199

consistent results. The edge of the embryo and the top of the nucleus
provided easily recognized boundaries for the data of Table 1. These
data are for three periods during the formation of the cellular blasto-
derm: the first, when the cleavage furrows are at the level of the
middle of the nuclei (15 minutes before the completion of blastoderm
formation); the second, when the furrows are at the base of the
nuclei (10 minutes before the completion); and the last stage, which
corresponds to the completion of blastoderm formation. No clear
difference in the number of mitochondria per 100 p2 was found be-
tween the dorsal and ventral regions at the first two periods studied,
but at the end of blastoderm formation 1.6 times as many mito-
chondria were found in the supranuclear region of the ventral side.

DISCUSSION
The sequence of events in the formation of cell membrane furrows
has been previously presented (Mahowald, 1963). During the transi-
tion from the syncytial blastema to the cellular blastoderm, the segre-
gation of yolk particulates from cytoplasm is completed and a series
of cytoplasmic changes occur. The reason for terminating the study at
the end of blastoderm formation is that cell movements of embryo-
genesis immediately ensue which result in frequent changes in cell
shape, thus rendering a study of this type overly difficult.
Annulate lamellae have been frequently described in oocytes and
spermatids, and only occasionally in other tissues (Porter, 1961b).
ROSS (1962) recently found these structures during only one stage of
the development of the mammalian adrenal gland, a finding indicative
of the transient nature of the organelle. In Drosophila they have
previously been noted in oogenesis (King and Devine, 1958; Maho-
wald, unpublished observations) and in pole cells (Mahowald, 1962).
The origin of the annulate lamellae has usually been attributed to the
nuclear envelope, and it has been postulated to function as a
mechanism of transfer of genetic information from nucleus to cyto-
plasm (Swift, 1956). In the DrosophiZa embryo, the lamellae first
appear as localized differentiations of the ER adjacent to the nuclear
envelope of yolk nuclei. This observation is consistent with the pro-
posed site of origin in other organisms (Swift, 1956). Since the
lamellae found in the cytoplasm after the completion of blastoderm
formation are larger (Fig. 8) than those found in the cytoplasm during
blastoderm formation (Figs. 6 and 7) and since at both times they
are not near any nuclear envelope, they can probably increase in
200 ANTHONY P. MAHOWALD

size away from a nuclear envelope. Waddington (1962) also con-

cluded that during oogenesis in DrosophiZa the lamellar stacks in-
crease in size without participation of the nuclear envelope. Poulson
(1950) and Poulson and Waterhouse (1960) have shown that the
yolk nuclei contribute to the formation of the embryonic gut. The
evidence presented here suggests that they also contribute an RNA
containing organelle (Rebhun, 1956) to the blastoderm cells. It is
unfortunate that at present the functional importance of the an-
nulate lamellae is not known, so that the significance of this proposed
contribution of the yolk nuclei might be determined.
The rapid appearance of new ER has been noted in many systems,
and this has been correlated with new synthetic activity occurring at
the same time (Hay, 1958; Slautterback and Fawcett, 1959). Con-
tinuities between the various types of membrane systems, i.e., between
agranular and granular ER and both of these with Golgi vesicles,
have been frequently noted in other cell types. This has been the basis
for the theory that all cytoplasmic membranes are differentiations of
the same system (Porter, 1961b). Observations on successively older
Drosophila embryos indicate that in the formation of the supranuclear
complex, granular ER may originate from the nuclear envelope dur-
ing the early stages of blastoderm formation and during later stages
from agranular vesicles and tubules, possibly of Golgi origin. Occa-
sional configurations of ER adjacent to the nuclear envelope during
later stages (Figs. 14 and 15) indicate a further relationship with the
nucleus, but this could be simply a chance occurrence.
Mitochondria are very common in the supranuclear region, and
frequency counts indicate 1.6 times as many mitochondria on the
mid-ventral as on the mid-dorsal region at the completion of blasto-
derm formation. Since the mid-ventral region invaginates and forms
the primitive mesoderm of the embryo immediately after the com-
pletion of blastoderm formation, this higher frequency of mito-
chondria as well as the greater “degree” of differentiation of the
supranuclear complex might be expected because of the active role
the mesoderm plays in subsequent embryogenesis (Poulson, 1950).

Embryological 1mplication.s
Two recent reviews of the evidence for the mosaic character of
the dipteran embryo (Anderson, 1962; Counce, 1961) have concluded
that it is not as determinative as previously thought, i.e., from the
 ULTRASTRUCTURE OF DROSOPHILA BLASTODERM 201

moment of fertilization. Yajima (1960) has been able to demonstrate

in Chironomus dorsuZis (a lower dipteran) a high degree of regulation
during cleavage stages. He interprets the results of his centriiugation
experiments to mean that regional determination becomes established
at the start of the blastema stage, i.e., when nuclei first appear in the
periplasm, whereas complete cellular determination only occurs at the
time of the formation of the cellular blastoderm. Idris (1959) has
shown by means of ligature experiments that in C&X pipicns (also a
lower dipteral?) the developmental fates are fixed before the syn-
cytial blastema stage, but that prior to this time some regulation can
occur. In higher dipterans there is less clear evidence for regulation,
but results from experiments with ultrasonics on Drosopllilu embryos
( Counce, 1961) and results of constriction experiments in Calliphora
( Nitschmann, referred to bv Counce, 1961) have been intcrprctcd as
indicating regulation at pr;blastema stages. Recently Hathaway and
Srlman (1961) have approached the question of mosaicism ani fates
in Drosophila by means of micro-spot irradiation with UV light, and
they concluded that the embryo is mosaic at the 1st hour of develop-
ment (6th cleavage). However, since the embryos showed excessive
sensitivity after irradiation at 1 hour, frecluently not surviving to a
stage when llistogencsis is occurring, it is difficult to conclude from
the evidence given whether mosaicism is complete at this stage. hlore
experimentation is needed to clarify the exact time when the
Drosophilu embryo becomes determinate,
Since there is general agreement that the DmsophiZu embryo is
mosaic at the blastoderm stage, attempts were made to find variations
in fine structure at earlier stages when possibly this mosaicism is
not complete. However, except for the polar region (1Iahowald,
1962)? no clearly distinct areas were found. In the early embryo
various special organelles are found in the periplasm, e.g., F-granules
(Okada and Waddington, 1959), dense multivesicular bodies (Rlaho-
wald, 1982), but these have disappeared before the time of blasto-
derm formation. There is at present no evidence that they have
specific localizations or that they are important in embryological
determination.
At the blastoderm stage when the nuclei are for the first time
segregated into individual cells, changes rapidly occur in the cyto-
plasm which are characteristic of differentiating cells: the nucleolus
forms for the first time, indicating new nuclear or genetic activity;
202 ANTHONY P. MAHOWALD

besides the lamellar ER which was present throughout the early

stages of embryogenesis, a new cytoplasmic system of ER is estab-
lished, presumably for synthesis of different enzymes or cell products
needed at this stage of embryogenesis; and finally, since the nuclei
are no longer in a syncytium, the cellular differentiations which occur
are established in individual cells, not in a common cytoplasm. In
the two regions studied most extensively for differences in fine struc-
ture, i.e., the mid-dorsal and mid-ventral, differences were found both
in the distribution of mitochondria and in the degree of differentia-
tion of the complex of agranular and granular ER. More extensive
comparisons of regions of the blastoderm may disclose other distinc-
tive differentiations of ultrastructure, thus indicating a variety of
specializations for different cell types.

SUMMARY

The ultrastructure of the Drosophila melanogaster embryo is

described during the transition from the syncytial to the cellular
blastoderm. The centriole is unusually short, only X50-175 rnp in
length, and has a central cylinder and radial “spokes.” Although two
centrioles have not been found together, a structure resembling a pro-
centriole is frequently found with the centriole. Differentiations of
cisternae of ER with the characteristics of annulate lamellae are found
in the blastoderm adjacent to the yolk. Some evidence suggests that
these differentiations first form adjacent to yolk nuclei during the
blastema and early blastoderm stages and that subsequently they are
incorporated into the blastoderm cells.
A new supranuclear cytoplasmic region is formed in blastoderm
cells after the nuclei have moved from the surface. Within this area a
cytoplasmic complex forms consisting of Golgi, agranular tubules, and
granular tubules and cisternae. The possible origins of the new ER
are discussed. Definite differences in fine structure, both in the num-
ber of mitochondria and in the type of differentiation of the supra-
nuclear complex, were found between the mid-dorsal and mid-ventral
regions of the embryo. The differentiation of the supranuclear com-
plex is discussed in relation to the establishment of the determinative
nature of the embryo and to cellular differentiation.
The author wishes to thank Dr. T. R. F. Wright and Dr. M. Beer for their
assistance during the course of the work, and also Dr. Wright and Dr. II.
Ursprung for their aid in the preparation of the manuscript.
 ULTRASTRUCTURE OF DROSOPHILA BLASTODERM 203

REFERENCES
ANDERSON, II. A. ( 1962). Th e e p’g1 enetics of the larva in Diptcra. Acta 2001.
(Stockholm) 43, 221-228.
BEHNHAHD, W., and nrz HARvEN, E. ( 1960). L’ultrastructure du centriole et
d’autrrs ClGments cle l’appareil achromaticlue. Intern. Kong. Elekt7onenmikro-
skopic 4 Berlin 1958 Verhcdl. 2, 217-227.
COUNCE, S. J. (1961). The analysis of insect emhrvogencxis. Ann. Rcu. Entomb.
6, 295-312.
DE HAWEN, E., and BERNHAHU, W. ( 1956). &ude ;IIX microscope ClwtronirIue
clc l’ultrastructure du cmtriolr chcz lrs vert&r&s. 2. Zellforsch. Mikroskop.
Anut. 45, 378-398.
GALL, J. C. ( 1961 ). Crntriole replication, a study of spc~rmntogcwsis in the
snail Vizjip(irI(.F. J. Biopltys. Bioc&w. Cytol. 10, 163-193.
HAITIAWAY, D. S., and SELAIAN, C. C. (1961). Certain aspects of cell lineage
and morphogenesis studied in embryos of Drosophila melnnogoster with an
ultra-violet micro-beam. J. Embryol. Erptl. Morphol. 9, 310-325.
HAY, E,. D. (1958). The fine structure of l&sterna cells and differentiating
cariilage cells in regenrru~ing limb of Antblystoma larvae. J. Biophys. Bioclicm.
Cytol. 4, 583-592.
IDIUS, B. E. M. (1959). Die Enhvicklung im geschniirten Ei von Culer pipiens
L. (Diptera). Arch. Entwicklungsmech. Organ. 152, 230-262.
KARASAKI, S. (1959). Electron microscopic studies on the cytoplasmic struc-
tures of ectoderm cells of the Triturm embryo during the early phase of
differentiation. Embryologin 4, 247-272.
KARNOVSKY, M. J. ( 1961). Simple methods for “staining with lead” at high
pH in electron microscopy. J. Biophys. Biochem. Cytol. 11, 729-732.
KING, R. C., and DE~INZ, R. L. ( 1958). Oogenesis in adult Drosophila meluno-
guster. VII. The sub-microscopic morphology of the ovary. Grou;th 22, 299-
326.
MAHOWALD, A. P. ( 1962). Fine structure of pole ~11s and polar granu1r.s in
Drosophila n&noguster. J. Erptl. Zool. 151, 201-216.
MAHOWALD, A. P. ( 1963). Electron microscopy of the formation of the> c~llulnr
blastoderm in Drosophila melnnoguster. Exptl. Cell Research in press.
MAZIA, D., IIAHHIS, I’., and BIBHING, T. ( 1960). The multiplicity of mitotic
center5 ant1 the time-course of their duplication and separation. J. Biophy~.
Biochem. Cytol. 7, l-20.
OKADA, lil., and WADDINGTCX, C. H. (1959). The submicroscopic structure of
the Drosophila egg. J. Emhryol. Exptl. Morphol. 7, 583-597.
POHTER, K. R. ( 1961a ), The cndoplasmic reticulum: some current interpretations
of its form and lunctions. 111 “Biological Structure and Function” (T. \V.
Goodwin and 0. Linclberg, eds. ), Vol. 1, pp. 127-155. Academic Press,
New York.
PORTER, K. R. ( 196lb). The grouncl substance: observations from clcctron
microscopy. 171 “The Cell” (J. Brachct and A. E. hlirskp, rds. ), Vol. 2, pp.
621-675. Academic Press, New York.
POULSON, D. F. ( 1950). IIistogcmcsis, orgnnogenc~sis, and differentiation in the
204 ANTHONY P. MAHOWALD

embryo of Drosophila melanogaster Meigen. In “Biology of Drosophila” ( M.

Demerec, ed.), pp. 168-274. Wiley, New York.
POULSON, D. F., and WATERHOUSE, D. F. ( 1960). Experimental studies on pole
cells and midgut differentiation in Diptera. Austrcllian J. Biol. Sci. 13, 541-567.
REBHUN, L. I. (1956). Electron microscopy of basophilic structures of some
invertebrate oocytes. I. Periodic lamellae and the nuclear envelope. J. Biopilys.
Biochem. Cytol. 2, 93-104.
ROSENBLUTH, J. ( 1963). Contrast between osmium-fixed and permnnganate-fixed
toad spinal ganglia. J. Cell Biol. 186, 143-157.
ROSS, M. H. (1962). Annulate lamellae in the adrenal cortex of the fetal rat.
J. Ultrastruct. Res. 7, 373-382.
SLAUTTERBACK, D. B., and FAWCETT, D. W. (1959). The development of the
cnidoblasts of Hydra, an electron microscope study of cell differentiation. J.
Biophys. Biochem. Cytol. 5, 441-552.
SONNENBLICK, B. P. (1950). The early embryology of Drosophila melunogaster.
In “Biology of Drosophila” (M. D emerec, ed.), pp. 62-167. Wiley, New York.
SWIFT, II. (1956). The fine structure of annulate lamellae. J. Biophys. Biochem.
Cytol. 2, Suppl., 415-418.
WADDINGTON, C. H. (1962). “N ew Patterns in Genetics and Development.”
Columbia Univ. Press, New York.
WATSON, M. L. ( 1959). Further observations on nuclear envelope of animal cells.
J. Biophys. Biochem. Cytol. 6, 147-156.
WESSEL, W. (1960). tiber die Form der Mitochondrien in Elecktronenmikro-
skopishen Bilden. Z. Zellforsch. Mikroskop. Anut. 52, 712-714.
WILSON, E. B. ( 1928). “The Cell in Development and Heredity.” Macmillan,
New York.
YAJIMA, H. (1960). Studies on embryonic determination of the harlequin fly,
Chironomus dorsalis. I. Effects of centrifugation and of its combination with
constriction and puncturing. J. Embryol. Exptl. Morphol. 8, 198-215.