Trunk Ectoderm Research Articles

Retinoic acid signaling and neurogenic niche regulation in the developing peripheral nervous system of the cephalochordate amphioxus.

The retinoic acid (RA) signaling pathway regulates axial patterning and neurogenesis in the developing central nervous system (CNS) of chordates, but little is known about its roles during peripheral nervous system (PNS) formation and about how these roles might have evolved. This study assesses the requirement of RA signaling for establishing a functional PNS in the cephalochordate amphioxus, the best available stand-in for the ancestral chordate condition. Pharmacological manipulation of RA signaling levels during embryogenesis reduces the ability of amphioxus larvae to respond to sensory stimulation and alters the number and distribution of ectodermal sensory neurons (ESNs) in a stage- and context-dependent manner. Using gene expression assays combined with immunohistochemistry, we show that this is because RA signaling specifically acts on a small population of soxb1c-expressing ESN progenitors, which form a neurogenic niche in the trunk ectoderm, to modulate ESN production during elongation of the larval body. Our findings reveal an important role for RA signaling in regulating neurogenic niche activity in the larval amphioxus PNS. Although only few studies have addressed this issue so far, comparable RA signaling functions have been reported for neurogenic niches in the CNS and in certain neurogenic placode derivatives of vertebrates. Accordingly, the here-described mechanism is likely a conserved feature of chordate embryonic and adult neural development.

Decoupling brain from nerve cord development in the annelid Capitella teleta: Insights into the evolution of nervous systems

In the deuterostomes and ecdysozoans that have been studied (e.g. chordates and insects), neural fate specification relies on signaling from surrounding cells. However, very little is known about mechanisms of neural specification in the third major bilaterian clade, spiralians. Using blastomere isolation in the annelid Capitella teleta, a spiralian, we studied to what extent extrinsic versus intrinsic signals are involved in early neural specification of the brain and ventral nerve cord. For the first time in any bilaterian, we found that brain neural ectoderm is autonomously specified. This occurs in the daughters of first-quartet micromeres, which also generate anterior neural ectoderm in other spiralians. In contrast, isolation of the animal cap, including the 2d micromere, which makes the trunk ectoderm and ventral nerve cord, blocked ventral nerve cord formation. When the animal cap was isolated with the 2D macromere, the resulting partial larvae had a ventral nerve cord. These data suggest that extrinsic signals from second-quartet macromeres or their daughters, which form mesoderm and endoderm, are required for nerve cord specification in C. teleta and that the 2D macromere or its daughters are sufficient to provide the inductive signal. We propose that autonomous specification of anterior neural ectoderm evolved in spiralians in order to enable them to quickly respond to environmental cues encountered by swimming larvae in the water column. In contrast, a variety of signaling pathways could have been co-opted to conditionally specify the nerve cord. This flexibility of nerve cord development may be linked to the large diversity of trunk nervous systems present in Spiralia.

A comprehensive fate map by intracellular injection of identified blastomeres in the marine polychaete Capitella teleta

BackgroundThe polychaete annelid Capitella teleta (formerly Capitella sp. I) develops by spiral cleavage and has been the focus of several recent developmental studies aided by a fully sequenced genome. Fate mapping in polychaetes has lagged behind other spiralian taxa, because of technical limitations.ResultsTo generate a modern fate map for C. teleta, we injected 1,1'-dioctadecyl-3,3,3'3'-tetramethylindocarbocyanine perchlorate (DiI) into individual identified blastomeres through fourth-quartet micromere formation. Confocal laser scanning microscopy at single-cell resolution was used to characterize blastomere fates during larval stages. Our results corroborate previous observations from classic studies, and show a number of similarities with other spiralian fate maps, including unique and stereotypic fates for individual blastomeres, presence of four discrete body domains arising from the A, B, C and D cell quadrants, generation of anterior ectoderm from first quartet micromeres, and contributions to trunk ectoderm and ventral nerve cord by the 2d somatoblast. Of particular interest are several instances in which the C. teleta fate map deviates from other spiralian fate maps. For example, we identified four to seven distinct origins of mesoderm, all ectomesodermal. In addition, the left and right mesodermal bands arise from 3d and 3c, respectively, whereas 4d generates a small number of trunk muscle cells, the primordial germ cells and the anus. We identified a complex set of blastomere contributions to the posterior gut in C. teleta, which establishes the most complete map of posterior gut territories to date.ConclusionsOur detailed cellular descriptions reveal previously underappreciated complexity in the ontogenetic contributions to several spiralian larval tissues, including the mesoderm, nervous system and gut. The formation of the mesodermal bands by 3c and 3d is in stark contrast to other spiralians, in which 4d generates the mesodermal bands. The results of this study provide a framework for future phylogenetic comparisons and functional analyses of cell-fate specification.

Convergence of a head-field selector Otx2 and Notch signaling: a mechanism for lens specification

Xenopus is ideal for systematic decoding of cis-regulatory networks because its evolutionary position among vertebrates allows one to combine comparative genomics with efficient transgenic technology in one system. Here, we have identified and analyzed the major enhancer of FoxE3 (Lens1), a gene essential for lens formation that is activated in the presumptive lens ectoderm (PLE) when commitment to the lens fate occurs. Deletion and mutation analyses of the enhancer based on comparison of Xenopus and mammalian sequences and in vitro and in vivo binding assays identified two essential transcriptional regulators: Otx2, a homeodomain protein expressed broadly in head ectoderm including the PLE, and Su(H), a nuclear signal transducer of Notch signaling. A Notch ligand, Delta2, is expressed in the optic vesicle adjacent to the PLE, and inhibition of its activity led to loss, or severe reduction, of FoxE3 expression followed by failure of placode formation. Ectopic activation of Notch signaling induced FoxE3 expression within head ectoderm expressing Otx2, and additional misexpression of Otx2 in trunk ectoderm extended the Notch-induced FoxE3 expression posteriorly. These data provide the first direct evidence of the involvement of Notch signaling in lens induction. The obligate integration of inputs of a field-selector (Otx2) and localized signaling (Notch) within target cis-regulatory elements might be a general mechanism of organ-field specification in vertebrates (as it is in Drosophila). This concept is also consistent with classical embryological studies of many organ systems involving a ;multiple-step induction'.

Expression of hunchback during trunk segmentation in the branchiopod crustacean Artemia franciscana

Comparative studies have shown that some aspects of segmentation are widely conserved among arthropods. Yet, it is still unclear whether the molecular prepatterns that are required for segmentation in Drosophila are likely to be similarly conserved in other arthropod groups. Homologues of the Drosophila gap genes, like hunchback, show regionally restricted expression patterns during the early phases of segmentation in diverse insects, but their expression patterns in other arthropod groups are not yet known. Here, we report the cloning of a hunchback orthologue from the crustacean Artemia franciscana and its expression during the formation of trunk segments. Artemia hunchback is expressed in a series of segmental stripes that correspond to individual thoracic/trunk, genital, and postgenital segments. However, this expression is not associated with the segmenting ectoderm but is restricted to mesodermal cells that associate with the ectoderm in a regular metameric pattern. All cells in the early segmental mesoderm appear to express hunchback. Later, mesodermal expression fades, and a complex expression pattern appears in the central nervous system (CNS), which is comparable to hunchback expression in the CNS of insects. No regionally restricted expression, reminiscent of gap gene expression, is observed during trunk segmentation. These patterns suggest that the expression patterns of hunchback in the mesoderm and in the CNS are likely to be ancient and conserved among crustaceans and insects. In contrast, we find no evidence for a conserved role of hunchback in axial patterning in the trunk ectoderm.

Expression of a SoxB and a Wnt2/13 gene during the development of the mollusc Patella vulgata.

We cloned and analysed the expression of a SoxB gene ( PvuSoxB) in the marine mollusc, Patella vulgata. Like its orthologues in deuterostomes, after an early broad ectodermal distribution, PvuSoxB expression only persists in cells competent to form neural structures. In the post-gastrulation larva, PvuSoxB is expressed in the prospective neuroectoderm in the head and in the trunk. No expression can be seen dorsally, around the mouth and the anus, or along the ventral midline. We also report the expression of a Wnt2/13 orthologue ( PvuWnt2) in Patella. After gastrulation, PvuWnt2 is expressed in the posterior part of the mouth, along the ventral midline and around the anus. This expression seems to be complementary to that of PvuSoxB in the trunk. We suggest the existence of a fundamental subdivision of the Patella trunk ectoderm into midline (mouth, midline, anus) and more lateral structures.

Surface ectoderm is necessary for the morphogenesis of somites

The paraxial mesoderm of the neck and trunk of mouse embryos undergoes extensive morphogenesis in forming somites. Paraxial mesoderm is divided into segments, it elongates along its anterior posterior axis, and its cells organize into epithelia. Experiments were performed to determine if these processes are autonomous to the mesoderm that gives rise to the somites. Presomitic mesoderm at the tailbud stage was cultured in the presence and absence of its adjacent tissues. Somite segmentation occurred in the absence of neural tube, notochord, gut and surface ectoderm, and occurred in posterior fragments in the absence of anterior presomitic mesoderm. Mesodermal expression of Dll1 and Notch1, genes with roles in segmentation, was largely independent of other tissues, consistent with autonomous segmentation. However, surface ectoderm was found to be necessary for elongation of the mesoderm along the anterior–posterior axis and for somite epithelialization. To determine if there is specificity in the interaction between ectoderm and mesoderm, ectoderm from different sources was recombined with presomitic mesoderm. Surface ectoderm from only certain parts of the embryo supported somite epithelialization and elongation. Somite epithelialization induced by ectoderm was correlated with expression of the basic-helix-loop-helix gene Paraxis in the mesoderm. This is consistent with the genetically defined requirement for Paraxis in somite epithelialization. However, trunk ectoderm was able to induce somite epithelialization in the absence of strong Paraxis expression. We conclude that somitogenesis consists of autonomous segmentation patterned by Notch signaling and nonautonomous induction of elongation and epithelialization by surface ectoderm.

Induction of the epibranchial placodes.

The cranial sensory ganglia, in contrast to those of the trunk, have a dual embryonic origin arising from both neurogenic placodes and neural crest. Neurogenic placodes are focal thickenings of ectoderm, found exclusively in the head of vertebrate embryos. These structures can be split into two groups based on the positions that they occupy within the embryo, dorsolateral and epibranchial. The dorsolateral placodes develop alongside the central nervous system, while the epibranchial placodes are located close to the top of the clefts between the branchial arches. Importantly, previous studies have shown that the neurogenic placodes form under the influence of the surrounding cranial tissues. In this paper, we have analysed the nature of the inductive signal underlying the formation of the epibranchial placodes. We find that epibranchial placodes do not require neural crest for their induction, but rather that it is the pharyngeal endoderm that is the source of the inductive signal. We also find that, while cranial ectoderm is competent to respond to this inductive signal, trunk ectoderm is not. We have further identified the signalling molecule Bmp7 as the mediator of this inductive interaction. This molecule is expressed in a manner consistent with it playing such a role and, when added to ectoderm explants, it will promote the formation of epibranchial neuronal cells. Moreover, the Bmp7 antagonist follstatin will block the ability of pharyngeal endoderm to induce placodal neuronal cells, demonstrating that Bmp7 is required for this inductive interaction. This work answers the long standing question regarding the induction of the epibranchial placodes, and represents the first elucidation of an inductive mechanism, and a molecular effector, underlying the formation of any primary sensory neurons in higher vertebrates.

The pattern of protein and glycoprotein synthesis in presumptive lens and non-lens ectoderm of the chicken embryo.

Lens induction is a classic example of the tissue interactions that lead to cell specialization during early vertebrate development. Previous studies have shown that a large region of head ectoderm, but not trunk ectoderm, of 36 h (stage 10) chicken embryos retains the potential to form lenses and synthesize the protein δ-crystallin under some conditions. We have used polyacrylamide gel electrophoresis and fluorography to examine protein and glycoprotein synthesis in presumptive lens ectoderm and presumptive dorsal (trunk) epidermis to look for differentiation markers for these two regions prior to the appearance of δ-crystallin at 50 h. Although nearly all of the proteins incorporating3H-leucine were shared by presumptive lens ectoderm and trunk ectoderm, these two regions showed more dramatic differences in the incorporation of3H-sugars into glycoproteins. when non-lens head ectoderm that has a capacity for lens formation in vitro was labeled, a hybrid pattern of glycoprotein synthesis was discovered: glycoproteins found in either presumptive lens ectoderm or trunk ectoderm were oftentimes also found in other head ectoderm. Therefore, molecular markers have been identified for three regions of ectoderm committed to different fates (lens and skin), well before features of terminal differentiation begin to appear in the lens.

Immunocytochemical analysis of embryonic compartmentation with a monoclonal antibody against a cytokeratin-related antigen

Mab 113F4, a monoclonal antibody recognizing an antigen in the outer synaptic layer of the chick neural retina, also recognizes an antigen appearing in all three germ layers of the gastrulating chick embryo. However, as neurulation proceeds, the antigen is down-regulated in three distinct patterns. First, the antigen is lost specifically from those trunk ectodermal cells destined to form the neural plate and, later, the neural tube. It remains absent from any neural derivative until day 13 when it appears in the outer synaptic layer of the neural retina, coincident with synaptogenesis in this region. Second, the entirety of the head ectoderm loses this antigen as the head lifts off the blastoderm. This down-regulation is followed later by a similar loss of antigen expression in the trunk ectoderm. Third, expression in the mesoderm becomes limited to the lateral plate and extraembryonic epithelia. Endodermal derivatives continue to express the antigen throughout development. Antigen 113F4 is localized within the cytoplasm and is organized in a fibrillar pattern. The intracellular localization of this antigen and its characteristic spatio-temporal tissue distribution are consistent with the antigen being a cytokeratin or cytokeratin-related antigen. The changes in tissue distribution suggest a possible role in tissue modelling in response to inductive interactions during development.

Isolation of a chick cytokeratin cDNA clone indicative of regional specialization in early embryonic ectoderm

During early vertebrate development, a series of inductive tissue interactions appear to be involved in establishing regional specializations that are eventually elaborated in the basic body plan of the embryo. These early inductive interactions are particularly difficult to study because they often occur in the absence of any associated morphological changes. In the chick embryo, the regional subdivision of the early ectoderm is evidenced by a marked lens-forming bias in the head ectoderm, which is absent from the presumptive dorsal epidermis of the trunk region. This striking divergence in developmental state is present long before any differentiation into lens or epidermal phenotypes can be detected. As a strategy for isolating genes whose differential expression might be a reflection of this regional subdivision, a cDNA library was prepared from early embryos and screened for differential hybridization to radiolabelled probes prepared from head ectoderm and trunk ectoderm. Two related cDNA clones were isolated that hybridize to transcripts present at much higher levels in trunk ectoderm than in head ectoderm. Sequence analysis of one of these clones revealed a high degree of similarity to members of the type II subfamily of intermediate filament cytokeratins. This clone (pCKse1) was used to examine cytokeratin gene expression in ectodermal tissues. A large increase in the level of CKse1 transcripts was found to take place in trunk ectoderm, approximately coordinate with neurulation, contrasting sharply with the much lower levels detected in head ectoderm and neural ectoderm at all stages tested. These results indicate that differential cytokeratin gene expression can occur within a contiguous layer of simple embryonic epithelia, and that this expression pattern coincides closely to the subdivision of the early ectoderm into regions with distinct developmental potencies. This type of regulation has not been described previously for members of the cytokeratin gene family.

Diferenciación de Cristalinos en Explantes de Embrión de Polio en Ausencia de Hipoblasto y Vesícula Optica1

In vitro studies of lens formation in chick embryo have suggested the action of two factors leading the lens induction in the cephalic ectoderm in the absence of optic vesicle: preliminary instructive specific stimulus (homotypic endo-mesoderm) and permissive unspecific stimulus (heterotypic mesenchymes). In order to detect the true capacities of tissues that exert this influence in the cultural condition, series of in vitro experiments were planned. Exclusion experiments: explants including presumptive lens ectoderm were cultured previous to a progressive exclusion of adjacent tissues to trigger lens formation (endoderm, mesoderm and neural tissue), from stage 1 to 7 of HAMBURGER/HAMILTON. Recombinant experiments: Recombinations of caudal epiblast with cephalic hypoblast from blastoderms stages 3, 4 and 5; and recombinations of cardiac mesoderm stage 7 with trunk ectoderm stage 11, were cultured in close association. Lens and lentoids were formed in the presumptive lens ectoderm, even when endoderm, neural tissue and optic vesicle were excluded, but always in presence of subjacent mesoderm. Observation of cephalic epiblast after to be separated mechanically from the underlying tissues showed that the presumptive cardiac mesoderm remains in contact with the epiblast. Beside the cardiac area was capable of forming lens bodies in contact with the trunk ectoderm. It was concluded that the cardiac mesoderm is able to exert a instructive specific stimulus and a permissive unspecific stimulus during in vitro lens formation.

Transfilter Lens Induction in Avian Embryo

The directive influence of the optic vesicle during lens induction of 2-day chick embryos was studied in vitro. Trunk ectoderm was chosen for the responding tissue. This uncommitted ectoderm formed distinct lentoid bodies when grown together with the optic vesicle. Crystallin synthesis was demonstrated in the lentoids with fluorescein labelled antiserum. By interposing filters of different thicknesses and pore sizes between the interacting tissues, three questions were put forward: (1) Whether the intimate contact between the interactants was essential for induction, (2) how far the inductive influence of the optic vesicle extended, and (3) what was the smallest pore through which the inductive influence could penetrate. The inductive influence reached across a Millipore filter with a thickness of 100 micrometer. It also penetrated a dialyzer membrane, allowing passage of molecules with molecular weights (MW) of less than 12,000 daltons. It was concluded that the directive, inductive signal(s) passing from the optic vesicle, diffused in the extracellular space as far as 100 micrometer and had a molecular weight of less than 12,000 daltons.

Permissive and directive interactions in lens induction

The interactive events leading to lens formation and the developmental potentialities of the presumptive lens ectoderm were examined in vitro. The presumptive lens ectoderm of both mouse and chick embryos was capable of forming a lens even when isolated from the optic vesicle before the two tissues reach the stage of close association. This lens-forming bias can be released with favourable culture conditions and by various heterotypic mesenchymes. The same permissive, unspecific conditions or heterotypic tissues failed to trigger lens formation in trunk ectoderm. The directive effect of the optic vesicle was demonstrated in experiments where it was grown in contact with the trunk ectoderm. The latter developed distinct lentoid bodies synthesizing lens proteins. The origin of the lentoid was confirmed in interspecies combination of chick and quail tissues. It is concluded that lens formation is governed by a series of interactive events consisting of both directive and permissive influences.

Chondrogenesis in chick embryo somites grafted with adjacent and heterologous tissues

ABSTRACT A variety of heterologous tissues have been tested for the ability to promote cartilage differentiation in isolated chick-embryo somites, using a modified chorioallantoic grafting technique. Of the 12 tissues tested only 3- and 4-day embryonic ectoderm promoted somite chondrogenesis in somites that fail to chondrify when grafted in isolation. This activity of ectoderm was evident in grafts of somites isolated with adjacent ectoderm, and in grafts of somites recombined with ectoderm derived from several sources. Four-day embryonic limbbud ectoderm, including the apical ridge, was capable of promoting somite chondrogenesis, but to no greater extent than dorsal trunk ectoderm of the same age. It is suggested that the ability of embryonic ectoderm to promote cartilage differentiation in isolated somites is associated with its ability to synthesize basement membrane material (sulphated glycosaminoglycans and collagen), in association with adjacent somite mesoderm.

The development of Neomenia carinata Tullberg (Mollusca Aplacophora)

The development of <i>Neomenia carinata</i> Tullberg (Mollusca Aplacophora)