Regionalisation of segment-forming capacities during early embryogenesis inDrosophila melanogaster

Summary Transverse fragmentation of the egg ofDrosophila melanogaster results in the formation of partial larvae. Anterior and posterior egg fragments develop the respective partial larval patterns. The partial patterns do not add up to the complete pattern.Fragmentation near the middle of the egg during early cleavage causes a gap of 3–4 segments on average in the larva. This gap is reduced to 2 segments on average if operations are performed at the early syncytial blastoderm stage. Fragmentation near the pole regions from early cleavage stages onwards causes a gap of only 2 larval segments on average. When the egg is fragmented at the columnar cellular blastoderm stage or later, the gap at all positions amounts to the size of one segment or less. A gap is also found after incomplete fragmentation, when the ooplasmic bridge between both egg parts was constricted beyond a certain limit.A specific shift of the segment-forming capacities along the egg axis is observed from syncytial blastoderm stages onwards.After partial longitudinal fragmentation no additional structures are observed. In general, the partial transverse patterns add up to the complete pattern, but minor structures like single denticles are missing near the fragmentation site.The results are discussed with respect to current concepts of segment pattern formation during early embryogenesis in dipterans.     

A morphological and developmental study of Drosophila embryos ligated during nuclear multiplication

In the Drosophila embryo, determination is established at the cellular blastoderm and a mosaic type development is observed after this time. Before the blastoderm stage, however, development is not of the mosaic type, as ligation during the nuclear multiplication stage causes a change in the spatial organization of the larval pattern. An aberration in determination leads to an increase in segment size, an increase in the number of cells per segment, and a decrease in segment number. This abnormal determination of blastoderm cells has also been demonstrated experimentally by marking corresponding regions of the blastoderm in ligated (posterior fragments only) and nonligated embryos. When the blastoderms of nonligated and ligated embryos are punctured at the same site, ligated embryos produce larvae with damage in segments posterior to the segments damaged in larvae from nonligated embryos. Ultrastructurally, no abnormalities were observed in the plasma membrane at the time of ligation or later in blastoderm cells which formed in the ligation area of these embryos. Evidence from this study, as well as other sources, indicates that determination of segmentation is under maternal control.     

Determination During Early Embryogenesis in Drosophila melanogaster

Ligation of developing embryos of Drosophila melanogaster wasperformed at three different stages of nuclear multiplicationand at the cellular blastoderm stage. Egg fragments of variablesizes are able to continue development up to the hatching stage.Partial embryos differentiate larval structures, anterior fragmentsforming larval head and posterior fragments larval abdominalstructures. These fragments differentiate a variable numberof the twelve larval cuticular bands formed by intact embryos.We found that ligation at cellular blastoderm can lead to anteriorand posterior fragments which differentiate together all thetwelve bands, indicating that at this stage the embryo developsthese patterns in a mosaic fashion. Ligation of younger embryosprevents the differentiation of some intermediate larval cuticularbands, while the terminal ones are consistently differentiated.The number and position of the deleted bands is correlated withthe time and position of ligation. This indicates that the mosaicpattern present in the egg at blastoderm is not fully formedat earlier stages in development.     

Adult abnormalities resulting from a localized blastoderm defect in a maternal-effect mutant of Drosophila.

In the mutant mat(3)3 of Drosophila melanogaster, there is a temperature-sensitive maternal effect on blastoderm formation. When oogenesis occurs in homozygous mat(3)3 females at the fully restrictive temperature of 29°C, the embryonic progeny form a defective cellular blastoderm in which cells are either completely or partially missing from a posterior-dorsal region, and the embryos die before hatching. Transplantation tests for the presence in the embryos of primordial imaginal cells capable of developing into adult structures showed a relatively high yield of eye and antenna structures, an intermediate yield of labium structures, and low or zero yields of wing, haltere, and leg structures. These results are consistent with the fate mapping of the primordial imaginal cells by analysis of gynandromorph mosaics; the eye and antenna map in the fully cellular region of the mutant blastoderm, the labium near the border of the defective region, and the wing, haltere, and legs within the defective region. When oogenesis oocurs at a lower temperature, the lethal maternal effect in mat(3)3 is reversed, but there is a nonlethal effect on larval and adult progeny of the mat(3)3 females. Many of the adults are missing one or more cuticular structures, usually a leg, haltere, or abdominal segment, and many of the larvae are missing the corresponding imaginal discs from which the thoracic structures are derived. These selective effects on imaginal development appear to be caused by maternally induced blastoderm defects that are less extensive at the lower temperature of oogenesis.     

The potentialities of transplanted early gastrula nuclei ofDrosophila melanogaster. Production of their imago descendants by germ-line transplantation

Summary Wild-type nuclei, taken out of cells from five regions of early gastrula embryos, were implanted singly into unfertilizedy w sn ³lz^50e eggs ofDrosophila melanogaster. The different types of nuclei initiated development with nearly equal frequencies of about 60%. 2.9% of the 1073 nuclear transfers developed as far as one of the three larval instars, and one reached the pupal stage.All individuals showed stage-specific patterns of defect. Most of these abnormalities were probably due to some inevitable damage caused by the implantation procedure such as disarrangement of the internal egg morphology and loss of peripheral egg substance. The proportions of individuals arrested at different embryonic and larval stages were similar for the five nuclear groups.Fertile imagos, descendants of all five types of donor nuclei, were produced via germ-line mosaics in two ways: (1) Pole cells of nuclear-transplant blastoderm stages were implanted into the pole cell region of host blastoderm eggs. (2) Gonads were taken from nuclear-transplant larvae and implanted into host larvae. In both cases gametes developed from the transplants as could be recognized from the genotypes of their progeny. By means of suitable crosses it was possible to get clones of flies whose large chromosomes were descended from the chromosomes of only one transplanted nucleus, that is, each clone was the descendant of one somatic nucleus. The data presented show that the nuclei remain omnipotent until the early gastrula stage.     

Fortpflanzung und entwicklung vonAnemonia sulcata (Anthozoa,Actiniaria) II. Frühentwicklung,Blastula und Gastrula

Early developmental stages of theAnemonia germ are characterized by asynchronously dividing nuclei and an extreme delay of blastomere differentiation. The nuclei migrate to the periphery, whereas nutritive substances remain in the interior. Following this stage, the appearance of cell boundaries results in the formation of the blastoderm and the simultaneous division of the yolk into many fragments. Most of them are exclusively filled with reserve material; only very few contain zooxanthellae or nuclei. On the embryo's surface, conically shaped bundles of long microvilli are obvious. They appear to be less regularly arranged than the spines of oocytes before insemination. Pigment granules that have originated from fusing Golgi vesicles are crowded peripherally in the blastoderm cells. In the nucleoplasm single annulate lamellae that seem to be cut off from the nuclear envelope can frequently be found. There is no further cellular differentiation until gastrulation is completed. Though yolk-containing cellular fragments occupy nearly the whole blastocoel, entoderm formation occurs by invagination. Ultrastructural observations provide evidence of the existence of interstices between entoderm cells that allow all nutritive substances to pass gradually into the gastric cavity. In the region of the blastoporus there are cellular processes enveloping reserve material. Presumably, these observations indicate a so-called filtration of nutritive yolk (Korschelt & Heider, 1909) that might represent an additional mode for the transfer of yolk-containing cellular fragments from blastocoel into gastrocoel.     

Immunohistochemical observations of the endocrine pancreas during metamorphosis of the sea lamprey,Petromyzon marinus L.

Summary Light-microscopic immunohistochemistry was used to localize insulin- and somatostatin-immunoreactive cells within developing endocrine pancreatic tissue of metamorphosing lampreys, Petromyzon marinus. The extrahepatic common bile duct and a portion of the intrahepatic bile duct develop into the caudal portion of the endocrine pancreas. The cranial pancreas is composed of follicles originating in the intestinal and diverticular epithelia, thus following the method of formation of pancreatic follicles from gut epithelium in larvae. In both the cranial and caudal portions, and in an intermediate cord of isolated follicles which connect these two major masses, insulin-immunoreactive cells appear first and are followed by cells showing somatostatin-immunoreactivity. In all stages of metamorphosis individual endocrine cells demonstrate immunoreactivity to a single hormone. Biliary atresia in lamprey may have some adaptive significance in providing cells that produce a caudal endocrine pancreas.Supported by NSERC of Canada grant No. A5945 and MRC of Canada grant No. MA8629 to JHY     

Developmental potencies of nuclei from cleavage,preblastoderm, and syncytial blastoderm transplanted into unfertilized eggs ofDrosophila melanogaster

Summary Wild-type nuclei from eggs ofDrosophila melanogaster at various developmental stages and from different regions of the egg—cleavage nuclei, pole nuclei from preblastoderm, and lateral nuclei from syncytial blastoderm—were singly implanted into unfertilizedy w sn ³ lz _50e eggs to determine their developmental potencies.All three types of transplanted nuclei were almost equally effective in initiating development of unfertilized eggs. Development was arrested in one of five critieal embryonic stages or in one of the three larval instars. The frequency of individuals reaching a distinct stage was approximately the same for all three types of donor nuclei. The stage-specific pattern of defects was independent of the type of nucleus transplanted.The deviations from normal development were broadly similar to those seen in controls developing from fertilized eggs which had only been punctured or into which cytoplasm had been injected. Many defective embryos also occurred in these control experiments. These and other observations indicate that a large proportion of irregularly developed individuals found after nuclear transfer can be ascribed to loss of egg material, disturbances in the internal organization of the egg during nuclear implantation, and the difficulty the implanted nucleus has in adjusting to the autonomous processes within the egg, such as the formation and migration of cytoplasmic islands.Some of the defective embryos and larvae originating from nuclear transfer were implanted into adult hosts. After culture for 14 days the early embryonic stages had formed several larval structures, and the late embryonic and larval stages had developed all larval organs. The proliferated imaginal primordia of thesein vivo cultured embryos and larvae, as well as the imaginal disks of the third instar larva, were then implanted into larval hosts with which they passed through metamorphosis and differentiated into imaginal structures. All three types of donor nuclei were capable of producing all adult structures derivedin situ from imaginal disks. The phenotype of these structures waswild-type, thus demonstrating their origin from the transplanted nuclei.The problem as to why not all transplanted nuclei initiated development, and why development after nuclear transplantation was arrested at the third larval instar, at the latest, is discussed.This article is dedicated to Professor Friedrich Seidel on the occasion of his 75th birthday.     

Early development of the kelp fly,Coelopa frigida (Diptera). II. Morphology of cleavage and blastoderm formation

Cleavage and blastoderm formation in Coelopa frigida are extremely rapid developmental processes. In short (6–7 minutes) successive cell cycles, nuclei multiply and spread out through the egg. The movement seems to be aided by endoplasmic vesicles and cisternae which are in direct contact with the nuclear membrane. The first cells to separate from the egg plasmodium in early superficial cleavage stages are the pole cells. Precursor material from multivesicular bodies forms the pole cell membranes. The primary nuclei from the posterior pole region are removed from the blastoderm by the pole cell segregation. Blastoderm nuclei from the regions adjacent to the posterior pole migrate into the residual periplasm after pole cell segregation has been completed and constitute the blastoderm nuclei in that region of the egg. Nucleoli are not revealed during internal cleavage. They appear in pole cells shortly after their segregation. The generation time of the blastoderm nuclei increases after the twelfth cleavage. Concurrently, nucleoli form in the blastoderm nuclei and permanent cell membranes separate individual blastoderm cells. After blastoderm cells have been separated from each other, they remain in contact with the interior yolk sac by means of cytoplasmic canals. This contact is maintained at least during the early phases of blastokinesis. Observations on nuclear migration and rapid membrane formation are discussed as examples of protein assembly from subunits as an alternative to de novo protein synthesis in early stages of development.     

A comparative ontogenetic study of the tetraodontiform caudal complex

Konstantinidis P. and Johnson, G. D. 2012. A comparative ontogenetic study of the tetraodontiform caudal complex. —Acta Zoologica (Stockholm) 93 : 98–114. Interpretation of the caudal complex of adult Tetraodontiformes has proven problematic because of the consolidation of the component elements. Here, we show that an ontogenetic approach offers considerable elucidation of the homology of the caudal complex, resulting in a new understanding of the grundplan of these fishes. The reductions of structures of the caudal complex are interpreted in a phylogenetic context. The caudal skeleton of larval triacanthodids resembles that of many adult percomorphs; however, during subsequent development epural 3 disappears, while epural 2 is reduced so that it can hardly be distinguished from the uroneural remnants. Juvenile triacanthids have an epural 2 that is lost in ontogeny, and the cartilaginous parhypural becomes integrated into the large hypural plate. In ostraciids and diodontids, the parhypural is absent throughout development. The hypural plates of adult balistids, monacanthids and tetraodontids have a conspicuous diastema between the dorsal and ventral portions. However, in early stages of the former two, the dorsal and ventral portions are continuous in cartilage proximally and remain fused in the adults. In tetraodontids, the two hypurals are separate from their initial appearance in cartilage and never fuse, raising the question of homology of the individual hypurals among the different families.     

Cranial muscle development in frogs with different developmental modes: Direct development versus biphasic development

Normal development in anurans includes a free swimming larva that goes through metamorphosis to develop into the adult frog. We have investigated cranial muscle development and adult cranial muscle morphology in three different anuran species. Xenopus laevis is obligate aquatic throughout lifetime, Rana (Lithobates) pipiens has an aquatic larvae and a terrestrial adult form, and Eleutherodactylus coqui has direct developing juveniles that hatch from eggs deposited on leaves (terrestrial). The adult morphology shows hardly any differences between the investigated species. Cranial muscle development of E. coqui shows many similarities and only few differences to the development of Rana (Lithobates) and Xenopus. The differences are missing muscles of the branchial arches (which disappear during metamorphosis of biphasic anurans) and a few heterochronic changes. The development of the mandibular arch (adductor mandibulae) and hyoid arch (depressor mandibulae) muscles is similar to that observed in Xenopus and Rana (Lithobates), although the first appearance of these muscles displays a midmetamorphic pattern in E. coqui. We show that the mix of characters observed in E. coqui indicates that the larval stage is not completely lost even without a free swimming larval stage. Cryptic metamorphosis is the process in which morphological changes in the larva/embryo take place that are not as obvious as in normal metamorphosing anurans with a clear biphasic lifestyle. During cryptic metamorphosis, a normal adult frog develops, indicating that the majority of developmental mechanisms towards the functional adult cranial muscles are preserved. J. Morphol. 275:398–413, 2014. © 2013 Wiley Periodicals, Inc.     

Localized defects of blastoderm formation in maternal effect mutants of Drosophila

In the three maternal effect lethal mutant strains of D. melanogaster described in this report, the homozygous mutant females produce defective eggs that cannot support normal embryonic development. The embryos from these eggs begin to develop for the first 2 hr after fertilization in an apparently normal way, forming a blastula containing a cluster of pole cells at the posterior end and a layer of syncytial blastoderm nuclei. During the subsequent transition from a syncytial to a cellular blastoderm, cell formation in the blastoderm is either partially or totally blocked. In mutant mat(3)1 no blastoderm cells are formed, indicating that there are separate genetic controls for pole cells and blastoderm cells. The other two mutants form an incomplete cellular blastoderm in which certain regions of the blastoderm remain noncellular. The noncellular region in mutant mat(3)3 is on the posterior-dorsal surface, covering about 30% of the total blastoderm. In mutant mat(3)6 blastoderm cells are formed only at the anterior and posterior ends, separated by a noncellular region that covers about 70% of the total blastoderm. The selective effects on blastoderm cell formation in the three mutants emphasize the importance of components present in the egg before fertilization for the transition from a syncytial to a cellular blastoderm.The genes defective in the three mutants are essential only for oogenesis and not for any other period of development, as indicated by a strict dependence of the lethal phenotypes on the maternal genotypes. Heterozygous embryos from the eggs of homozygous mutant females die, whereas homozygous mutant embryos from the eggs of heterozygous females develop into viable adults.One of the mutants, mat(3)3, has a temperature-sensitive phenotype. Homozygous mat(3)3 females maintained at a restrictive temperature of 29°C show the lethal maternal effect. However, at a permissive temperature of 20°C the females produce viable adult progeny. The temperature-sensitive period in mat(3)3 females occurs during the last 12 hr of oogenesis, consistent with the maternal effect phenotype of the mutant.     

Expression of laminin and of a laminin-related antigen during early development of Drosophila melanogaster

This paper reports the characterization of two immunologically related proteins that may be involved in cell adhesion during Drosophila development. These proteins, laminin chain A and a 240K component, share the epitope recognized by monoclonal antibody RD3 (Mab RD3). The two antigens show different developmental expression profiles. Laminin is detected only from 6 to 8 h of development onwards; its concentration increases during embryogenesis to reach steady-state value in larvae, pupae and adult flies. By contrast, the 240K antigen, not found in oocytes, is present before blastoderm stages; its concentration increases during gastrulation, decreases at the end of organogenesis and the antigen is no longer detected in third instar larvae. Light and electron microscope immunolocalization in imaginal discs indicates that laminin is distributed apically in the lumen and basally in the basal membrane that surrounds the nonevaginated disc. During morphogenesis laminin is detected at the basal side of the evaginating part of the disc epithelium. Immunolocalization on paraffin sections of early embryos suggests that the 240K antigen is related to (1) cell formation and polarization in association with cytoskeleton components, (2) establishment of cell-extracellular substratum interactions during the blastoderm cell sheet organization and (3) basement membrane deposition during embryonic germ cell layer segregation. This 240K protein is poorly or not glycosylated, is resistant to chondroitinase ABC and collagenase and appears therefore as a new extracellular component that might be specifically involved in early processes of morphogenesis.     

aproctous,a locus that is necessary for the development of the proctodeum in Drosophila embryos,encodes a homolog of the vertebrate Brachyury gene

The proctodeum of the Drosophila embryo originates from the posterior end of the blastoderm and forms the hindgut. By enhancer-trap mutagenesis, using a P-element-lacZ vector, we identified a mutation that caused degeneration of the proctodeum during shortening of the germ band and named it aproctous (apro). Expression of the lacZ reporter gene, which was assumed to represent expression of the apro gene, began at the cellular blastoderm stage in a ring that encompassed about 10–15% of the egg's length (EL) and included the future proctodeum, anal pads, and posterior-most part of the visceral mesoderm. In later stages, strong expression of lacZ was detected in the developing hindgut and anal pads. Expression continued in the anal pads and epithelium of the hindgut of larvae; the epithelium of the hindgut of the adult fly also expressed lacZ. The spatial patterns of the expression of lacZ in various mutants suggested that the embryonic expression of apro was regulated predominantly by two gap genes, tailless (tll) and huckebein (hkb): tll is necessary for the activation of apro, while hkb suppressed the expression of apro in the region posterior to 10% EL. Cloning and sequencing of the apro cDNA revealed that apro was identical to the T-related gene (Trg) that is a Drosophila homolog of the vertebrate Brachyury gene. apro appears to play a key role in the development of tissues derived from the proctodeum.     

Analysis of morphogenetic movements in the development of the notum anlage of Drosophila melanogaster

Summary A comparison of the morphogenetic maps of the notum anlage of Drosophila melanogaster derived from the gynandromorph data and mosaics induced by somatic crossing-over during the first instar larval stage revealed that practically no major morphogenetic movements occur in the development of the anlage between the blastoderm and first instar larval stages and the adult stage. By comparing the morphogenetic map derived from gynandromorphs and the fate map derived from data on the transplantation of fragments of the mature wing imaginal disc, it was observed that no major morphogenetic movements occur in the notum anlage between the stages of the allocation of the disc and the mature disc. The results are consistent with the observations of other authors concerning the larval development of eye-antenna, wing and leg discs.     

Osteological development of the lumpfish,inimicus japonicus (pisces: Synanceiidae)

The osteological development of the synanceiidInimicus japonicus, was described on the basis of five larvae and four juveniles (4.2–10.1 mm BL) reared in the laboratory, and two wild adult specimens. All bones, except for the basisphenoid, were formed in all larvae and juveniles, but fusions between the uppermost actinost and scapula, upper caudal plate and urostyle, and third preural centrum and hemal spine were not completed by 10.1 mm BL. Following comparison with the adult condition, a rod-like ossified bone without a tooth plate on the upper branchial arch of larvae and juveniles was considered homologous with the second pharyngobranchial. The number of epurals and length of the neural spine on the second preural centrum varied (unrelated to growth) and it is inferred thatJ. japonicus shows intraspecific variations in these bones.     

Hox C6 expression during development and regeneration of forelimbs in larval Notophthalmus viridescens

 A central theme concerning the epimorphic regenerative potential of urodele amphibian appendages is that limb regeneration in the adult parallels larval limb development. Results of previous research have led to the suggestion that homeobox containing genes are ”re-expressed” during the epimorphic regeneration of forelimbs of adult Notophthalmus viridescens in patterns which retrace larval limb development. However, to date no literature exists concerning expression patterns of any homeobox containing genes during larval development of this species. The lack of such information has been a hindrance in exploring the similarities as well as differences which exist between limb regeneration in adults and limb development in larvae. Here we report the first such results of the localization of Hox C6 (formerly, NvHBox-1) in developing and regenerating forelimbs of N. viridescens larvae as demonstrated by whole-mount in situ hybridization. Inasmuch as the pattern of Hox C6 expression is similar in developing forelimb buds of larvae and epimorphically regenerating forelimb blastemata of both adults and larvae, our results support the paradigm that epimorphic regeneration in adult newts parallels larval forelimb development. However, in contrast with observations which document the presence of Hox C6 in both intact, as well as regenerating hindlimbs and tails of adult newts, our results reveal no such Hox C6 expression during larval development of hindlimbs or the tail. As such, our findings indicate that critical differences in larval hindlimb and tail development versus adult expression patterns of this gene in these two appendages may be due primarily to differences in gene regulation as opposed to gene function. Thus, the apparent ability of urodeles to regulate genes in such a highly co-ordinated fashion so as to replace lost, differentiated, appendicular structures in adult animals may assist, at least in part, in better elucidating the phenomenon of epimorphic regeneration. Received: 6 November 1998 / Accepted: 12 December 1998     

Sauropod haemal arches: morphotypes,new classification and phylogenetic aspects

Sauropod haemal arches are caudal bony structures that have been traditionally incorporated into two different types observed in different anatomical views: Y-shaped (anterior view) and forked (lateral view). This research proposes a classification combining information observed in anterior and lateral views. Four types of ‘Y-shaped’ and six types of ‘V-shaped’ chevrons were recognised. Complete chevron series in some eusauropods allows the comparison of topological equivalent structures along the tail and also among taxa. A basal titanosaur from Argentina exhibits mid-caudal chevron morphology in which more than one cranial and caudal process is present, arising from the distal blades as well as from the proximal rami, a condition not seen before in a sauropod dinosaur. The morphological variability seen in sauropod chevrons along the tail is in close relationship with the development and distribution of muscle. caudofemoralis longus, as seen in extant crocodiles and as previously proposed for non-avian theropods and for caudal centra and transverse processes of sauropod dinosaurs. Two new characters related to middle chevrons are proposed here, in which the transitional morphology is described.