Ultrastructure and formation of the body cavity lining in Phoronis muelleri (Phoronida, Lophophorata)

Among other characteristics a trimeric coelomic compartmentation consisting of an anterior protocoel, followed by a mesocoel and a posterior metacoel is traditionally believed to substantiate the sister-group relationship between Lophophorata and Deuterostomia, together forming the Radialia. As molecular data cannot support this hypothesis a reanalysis of the coelomic cavities in Phoronida is undertaken, because corresponding coelomic compartmentation is widely accepted to support the Radialia hypothesis. A coelomic cavity can be recognized on the ultrastructural level because its lining is a true epithelium with polarized cells interconnected by apical adherens junctions. This study reveals that neither in larval nor adult Phoronis muelleri (Phoronida) an anterior cavity with such a lining is present. What on the light microscopic level leads to the impression of a cavity inside the larval episphere, actually is an enlarged subepidermal extracellular matrix with an amorphous, presumably gel-like filling, into which several muscle cells are embedded. Larvae, thus, possess only one coelomic cavity, the large trunk coelom of the larva which is adopted in the adult organization. The second coelomic cavity of adult P. muelleri, the lophophore coelom, develops as a double-layer of epithelialized mesodermal cells at the base of the adult tentacle buds and becomes fluid filled during metamorphosis. Like the larval episphere, larval tentacles and most parts of the blastocoel are filled by an amorphous matrix. Reanalysis of the literature and comparison with Brachiopoda and Bryozoa allows the hypothesis that a protocoel is lacking in all Lophophorata, and that merely two unpaired coelomic cavities, one tentacle and one trunk coelom, can be assumed for the ground pattern of this taxon. These results do not provide further evidence for the Radialia hypothesis, but also do not contradict it. Accepted: 28 August 2000     

The Development of the Brachiopod Crania (Neocrania) anomala (O. F. Müller)and its Phylogenetic Significance

The freely spawned eggs of Crania go through radial cleavage, embolic gastrulation, and the posteroventral part of the archenteron forms mesoderm through modified enterocoely. The blastopore closes in the posterior end of the larva. The ciliated, lecithotrophic larva has four pairs of coelomic pouches and three pairs of dorsal setal bundles. At metamorphosis, the larva curls ventrally by contraction of a pair of midventral muscles, which are extensions of the first pair of coelomic sacs; the larva attaches by the epithelium just behind the closed blastopore. The brachial valve is secreted by the middle part of the dorsal epithelium and the pedicle valve is secreted by the attachment epithelium. The second pair of coelomic sacs develop small attachment areas at the edge of the dorsal valve and become the lophophore coelom (mesocoel); the third pair of coelomic sacs become the body coelom (metacoel) with the adductor muscles. The posterior position of the closing blastopore is characteristic of deuterostomes. The ventral curving of the settling larva and the formation of both valves from dorsal epithelial areas indicate that the brachiopods have a very short ventral side as opposed to the phoronids. It is concluded that both groups have originated from a creeping ancestor with a straight gut.     

Fine structure of the epistome in Phoronis ovalis: significance for the coelomic organization in Phoronida

Abstract. The hypothesis of a common ancestry of the lophophorate taxa Brachiopoda, Bryozoa, Phoronida, and the Deuterostomia can be traced back to the late 19th century when Masterman recognized a tripartite organization of the body consisting of pro-, meso-, and metasome, along with coelomic body cavities in each compartment, as characteristic for Echinodermata, Pterobranchia, Phoronida, and Brachiopoda. This idea became quite popular under the name "archicoelomate" concept. The organization of the phoronids, and especially of their transparent actinotroch larva, has for a long time been used as a touchstone for the validity of this concept. As a coelomic lining can reliably be recognized only on the ultrastructural level, this technique has been applied for adults of Phoronis ovalis , which is assumed to be a sister species to all other phoronids. Phoronis ovalis contains only two coelomic compartments, a posterior coelom inside the trunk (metasoma), occupying the space between the trunk epidermis and the digestive epithelium, and an anterior lophophoral coelom inside and basal to the tentacular crown (mesosoma). There is no coelomic cavity inside the epistome (prosoma). This part of the body is filled with myoepithelial cells, which are continuous with the epithelial lining of the lophophore cavity. These cells form a lumenless bilayer and possess long, tiny myofilamentous processes, which are completely embedded in an extracellular matrix. A comparison with data on P. muelleri shows that there is no need to assume three different coelomic cavities in Phoronida, in contrast to the predictions of the archicoelomate concept. At least for this taxon, a correspondence to the situation in deuterostomes can hardly be found.     

Ultrastructure of the coelom in the brachiopod Lingula anatina

The organization of the coelomic system and the ultrastructure of the coelomic lining are used in phylogenetic analysis to establish the relationships between major taxa. Investigation of the anatomy and ultrastructure of the coelomic system in brachiopods, which are poorly studied, can provide answers to fundamental questions about the evolution of the coelom in coelomic bilaterians. In the current study, the organization of the coelom of the lophophore in the brachiopod Lingula anatina was investigated using semithin sectioning, 3D reconstruction, and transmission electron microscopy. The lophophore of L. anatina contains two main compartments: the preoral coelom and the lophophoral coelom. The lining of the preoral coelom consists of ciliated cells. The lophophoral coelom is subdivided into paired coelomic sacs: the large and small sinuses (= canals). The lining of the lophophoral coelom varies in structure and includes monociliate myoepithelium, alternating epithelial and myoepithelial cells, specialized peritoneum and muscle cells, and podocyte‐like cells. Connections between cells of the coelomic lining are provided by adherens junctions, tight‐like junctions, septate junctions, adhesive junctions, and direct cytoplasmic bridges. The structure of the coelomic lining varies greatly in both of the main stems of the Bilateria, that is, in the Protostomia and Deuterostomia. Because of this great variety, the structure of the coelomic lining cannot by itself be used in phylogenetic analysis. At the same time, the ciliated myoepithelium can be considered as the ancestral type of coelomic lining. The many different kinds of junctions between cells of the coelomic lining may help coordinate the functioning of epithelial cells and muscle cells.     

The phylogenetic position of Brachiopoda—a comparison of morphological and molecular data

Analyses of rRNA and rDNA among Metazoa result in a hypothesis of a sistergroup relationship of Brachiopoda and certain spiralian taxa, whereas analyses of morphological data imply that Brachiopoda show affinities to Deuterostomia within the Radialia. Regarding Brachiopoda as a derived spiralian taxon must be followed by a reinterpretation of the evolution of distinct brachiopod morphological characters—like cleavage pattern, coelom or larva. The experimental insertion of a monophyletic taxon consisting of Brachiopoda and Phoronida into a widely accepted phylogenetic tree of Spiralia leads to the hypothesis that at least trimeric organization, mesosomal tentacular apparatus and heterogeneously assembled metanephridia are products of convergent evolution in Brachiopoda plus Phoronida and Deuterostomia. The hypothesis of a radialian nature of Brachiopoda and Phoronida, as implied by morphological data, remains as the most parsimonious possibility to explain the evolution of seven regarded characters (cleavage pattern, larva, tentacular apparatus, coelom, metameric segmentation, metanephridia and chaetae) in Brachiopoda. Due to the conflicting results of both methods a hitherto undetected systematical problem is discussed possibly hindering data comparability. If the course of evolution can principally be inferred from the information preserved in recent and fossil animals, the results should be congruent in the analyses of both, molecular and morphological data.     

The pericardium in the deuterostome <Emphasis Type="Italic">Saccoglossus kowalevskii</Emphasis> (Enteropneusta) develops from the ectoderm via schizocoely

The origin of mesoderm and coelomic compartments has traditionally been given high value for phylogenetic considerations of animal relationships. Two main modes have been distinguished, associated with the two main groups of animals: schizocoely with protostomes and enterocoely with deuterostomes. During enterocoely, coelomic compartments are formed from the endoderm. Here, we show that the pericardium of the deuterostome Saccoglossus kowalevskii, an enteropneust, is ontogenetically derived from the ectoderm and develops by schizocoely. The pericardium develops from a solid cluster of epidermis cells situated underneath the ectodermal nerve net above the basement membrane of the epidermis. The undifferentiated cells are interconnected by spot desmosomes, become separated from the epidermis and develop a central cavity. Pericardial cells become epithelial, by developing apical adherens junctions, a single apical cilium and basal striated myofibres. The differentiated pericardium possesses a cavity and surrounds a central blood vessel, the heart, situated in the basal extracellular matrix. The pericardium is an integral part of the anterior excretory complex, and comparisons to other deuterostomes indicate that pericardia are homologous despite differing ontogenies. Original data generated for the present study are deposited on MorphDBase ().     

Development of head and trunk mesoderm in the dogfish,Scyliorhinus torazame: I. Embryology and morphology of the head cavities and related structures

Vertebrate head segmentation has attracted the attention of comparative and evolutionary morphologists for centuries, given its importance for understanding the developmental body plan of vertebrates and its evolutionary origin. In particular, the segmentation of the mesoderm is central to the problem. The shark embryo has provided a canonical morphological scheme of the head, with its epithelialized coelomic cavities (head cavities), which have often been regarded as head somites. To understand the evolutionary significance of the head cavities, the embryonic development of the mesoderm was investigated at the morphological and histological levels in the shark, Scyliorhinus torazame. Unlike somites and some enterocoelic mesodermal components in other vertebrates, the head cavities in S. torazame appeared as irregular cyst(s) in the originally unsegmented mesenchymal head mesoderm, and not via segmentation of an undivided coelom. The mandibular cavity appeared first in the paraxial part of the mandibular mesoderm, followed by the hyoid cavity, and the premandibular cavity was the last to form. The prechordal plate was recognized as a rhomboid roof of the preoral gut, continuous with the rostral notochord, and was divided anteroposteriorly into two parts by the growth of the hypothalamic primordium. Of those, the posterior part was likely to differentiate into the premandibular cavity, and the anterior part disappeared later. The head cavities and somites in the trunk exhibited significant differences, in terms of histological appearance and timing of differentiation. The mandibular cavity developed a rostral process secondarily; its homology to the anterior cavity reported in some elasmobranch embryos is discussed.     

Mesodermal and axial determinants contribute to mesoderm regionalization in <Emphasis Type="Italic">Bufo arenarum</Emphasis> embryos

The existence of mesodermal determinants in the equator of Bufo arenarum embryos has been previously demonstrated. In this work, their role in dorso-ventral regionalization of mesoderm was studied by transferring the determinants to animal blastomeres. The transfer was performed by cleavage reorientation and cytoplasmic microinjection. Forced inclination during early cleavage caused deviation of the third cleavage plane and annexation of equatorial cytoplasm into animal quartets. Animal blastomeres from embryos oriented with the dorsal side up, incorporated ventro-equatorial cytoplasm and formed blood cells, mesenchyme, and coelomic epithelium. In contrast, animal blastomeres from embryos oriented with the ventral side up, acquired dorso-equatorial cytoplasm and developed notochord, somites, mesenchyme, coelomic epithelium and nervous tissue. In order to investigate if this dorso-ventral differentiation pattern responds to an interaction of mesodermal and axial factors, isolated 8-cell-stage animal quartets were microinjected with subcortical cytoplasm from: (a) the ventro-equatorial region of synchronous embryos; (b) the vegetal pole of uncleaved eggs; (c) a combination of both cytoplasms. As expected, the implanted ventro-equatorial cytoplasm promoted ventral mesoderm differentiation. Conversely, the joint transfer of ventro-equatorial cytoplasm and vegetal pole cytoplasm behaved as the dorso-equatorial cytoplasm, promoting dorso-lateral mesoderm and neural formation. Thus, mesoderm regionalization in B. arenarum embryos seems to be caused by a concurrent action of both mesodermal and axial determinants.     

Nephridial development and body cavity formation in <Emphasis Type="Italic">Artemia salina</Emphasis> (Crustacea: Branchiopoda): no evidence for any transitory coelom

The Ecdysozoa-hypothesis on the origin of arthropods questions the homology of segmentation in arthropods, onychophorans, and annelids. The implication of convergent gain of metamery in these groups seems to conflict particularly with the correspondence in the development of serial coelomic cavities and metanephridia. Ultrastructural studies of the mesoderm development in Onychophora revealed that main correspondence with the state in annelids concerns the involvement of epithelial lining cells of the embryonic coelomic cavities in the formation of the visceral and somatic musculature. The significance of this correspondence, however, remained unclear as comparable data on the state in arthropods were still missing. Developmental studies on selected representatives covering all major arthropod subgroups aim to fill in this gap. Data were raised by a combination of transmission electron microscopy and fluorescent stainings of the muscular system and nuclei for the anostracan crustacean Artemia salina. In this species, putative transitory coelomic cavities proved to be absent in all trunk segments. In the second antennal and second maxillary segments small, compact nephridial anlagen develop into a sacculus and excretory duct. The sacculus originates from the terminal cells of the nephridial duct, which is formed in advance. The lumen of the sacculus is inconspicuous in its earliest functional stage and later enlarges to a bulb; it accordingly represents no remnant of any primarily large coelomic cavity. The muscular system is entirely formed prior to and independent of coelomic or nephridial anlagen. Visceral and somatic mesoderm already separate in the caudal body region. Transitory segmental clusters of mesodermal cells are composed of somatic cells only and accordingly represent no “somites”. Our observations overall do not provide any support for the homology of coelomic cavities in annelids and arthropods.     

Ectopic mesodermal expression promoted by the eight-cell stage Bufo arenarum subequatorial cytoplasm

Mesodermal determinants were investigated by cytoplasmic transfer and blastomere isolation in the eight-cell stage of Bufo arenarum. Their existence was confirmed by assaying the subequatorial cytoplasm’s ability to respecify the developmental potency of animal quartets. The gray subequatorial cytoplasm, but not animal cytoplasm, is able to divert the ectodermal fate of animal quartets to several mesodermal components. The source of the transplanted cytoplasm was important in determining the category of the resulting structures. Ventral subequatorial cytoplasm from ventrovegetal blastomeres generated ventral derivatives, namely erythrocytes and mesenchyma. Dorsal subequatorial cytoplasm from dorsovegetal blastomeres produced dorsolateral derivatives, such as notochord, muscle, nephric tubules, and coelomic epithelium, including mesenchyma. On the other hand, transfer of vegetal pole cytoplasm to animal quartets resulted in the formation of groups of endoderm-like cells dispersed among epidermal cells. However, the presence of such cells did not cause any mesodermal induction. The present findings suggest the existence of cytoplasmic information responsible for mesodermal specification. The alternative hypothesis that animal blastomeres become mesoderm due to vegetal induction is questioned. Received: 9 October 1998 / Accepted: 10 March 1999     

Anatomy of the trunk mesoderm in tunicates: homology considerations and phylogenetic interpretation

The tadpole stage of tunicates has played a pivotal role in understanding chordate evolution. While the organization of the mesoderm has been given high importance in comparative anatomical studies of Bilateria, this morphological character remains largely unexplored in tunicate tadpoles. For larvae of the phlebobranch ascidian Ciona intestinalis, the presence of two mesodermal pockets had been claimed, raising the possibility that paired coelomes are present in the larval ascidian. Using computer assisted 3D-reconstructions based on complete series of 1 μm-sections analyzed by light microscopy complemented by TEM-investigation of selected regions a comparative anatomical study of tadpole stages from four major tunicate clades, Aplousobranchiata, Phlebobranchiata, Stolidobranchiata, and Appendicularia is presented. In the aplousobranch Clavelina lepadiformis numerous mesodermal cells are found throughout the entire trunk plus the unpaired ventral rudiment of the pericardium. In the phlebobranch Ascidia interrupta, massive mesodermal components occur in the posterior trunk, whereas more anteriorly situated mesoderm consists of loose streaks of cells or isolated cells. This is also the case in the stolidobranch ascidians Herdmania momus and Styela plicata. In the stolidobranch Molgula occidentalis and the appendicularian Oikopleura dioica the anterior trunk is entirely devoid of mesodermal cells. TEM-investigation revealed that all mesodermal structures in the trunk of tunicate tadpoles were mesenchymal with the exception of a ventral portion of the mesoderm in C. lepadiformis, which probably corresponds to the developing pericardium, and the differentiated pericardium of the juvenile O. dioica. Thus no evidence for paired coelomic cavities in Tunicata was found. Outgroup comparison suggests that the reduction of paired coelomic cavities is an apomorphic trait of Tunicata. Within Tunicata a stepwise evolutionary reduction of the anterior larval mesenchyme is documented.     

Two essential processes in the formation of a dorsal axis during gastrulation ofCynops embryo

The isolated upper marginal zone from the initial stage ofCynops gastrulation is not yet determined to form the dorsal axis mesoderm: notochord and muscle. In this experiment, we will indicate where the dorsal mesoderm-inducing activity is localized in the very early gastrula, and what is an important event for specification of the dorsal axis mesoderm during gastrulation. Recombination experiments showed that dorsal mesoderm-inducing activity was localized definitively in the endodermal epithelium (EE) of the lower marginal zone, with a dorso-ventral gradient; and the EE itself differentiated into endodermal tissues, mainly pharyngeal endoderm. Nevertheless, when dorsal EE alone was transplanted into the ventral region, a secondary axis with dorsal mesoderm was barely formed. However, when dorsal EE was transplanted with the bottle cells which by themselves were incapable of mesoderm induction, a second axis with well-developed dorsal mesoderm was observed. When the animal half with the lower marginal zone was rotated 180° and recombined with the vegetal half, most of the rotated embryos formed only one dorsal axis at the primary blastopore side. The present results suggest that there are at least two essential processes in dorsal axis formation: mesoderm induction of the upper marginal zone by endodermal epithelium of the lower marginal zone, and dorsalization of the upper dorsal marginal zone evoked during involution.     

Urodeles remove mesoderm from the superficial layer by subduction through a bilateral primitive streak

Urodeles begin gastrulation with much of their presumptive mesoderm in the superficial cell layer, all of which must move into the deep layers during development. We studied the morphogenesis of superficial mesoderm in the urodeles Ambystoma maculatum, Ambystoma mexicanum, and Taricha granulosa. In all three species, somitic, lateral, and ventral mesoderm move into the deep layer during gastrulation, ingressing through a "bilateral primitive streak" just inside the blastopore. The mesodermal epithelium appears to slide under the endodermal epithelium by a mechanism we term "subduction." Subduction removes the large expanse of superficial presumptive somitic and lateral-ventral mesoderm that initially separates the sub-blastoporal endoderm from the notochord, leaving the endoderm bounding the still epithelial notochord along the gastrocoel roof. Subduction may be a common feature of urodele gastrulation, differing in this regard from anurans. Subducting cells constrict their apices and become bottle-shaped as they approach the junction of the mesodermal and endodermal epithelia. Subducting bottle cells endocytose apical membrane and withdraw the tight junctional component cingulin from the contracting circumferential tight junctions. Either in conjunction with or immediately after subducting, the mesodermal cells undergo an epithelial-to-mesenchymal transition. The mechanism by which epithelial cells release their apical junctions to become mesenchymal, without disrupting the integrity of the epithelium, remains mysterious, but this system should prove useful in understanding this process in a developmental context.     

Organization of the epistome in <Emphasis Type="Italic">Phoronopsis harmeri</Emphasis> (Phoronida) and consideration of the coelomic organization in Phoronida

Results provided by modern TEM methods indicate the existence of the lophophoral and trunk coelomes but not of the preoral coelom in Phoronida. In the present work, the epistome in Phoronopsis harmeri was studied by histological and ultrastructural methods. Two kinds of cells were found in the frontal epidermis: supporting and glandular. The coelomic compartment is shown to be inside the epistome. This compartment has a complex shape, consists of a central part and two lateral branches, and contacts the lophophoral coelom, forming two complete dissepiments on the lateral sides and a partition with many holes in the center. TEM reveals that some portions of the incomplete partition are organized like a mesentery, with the two layers of cells separated by ECM. The myoepithelial cells of the coelomic lining form the circular and radial musculature of the epistome. Numerous amoebocytes occur in the coelom lumen. The tip of the epistome and its dorso-lateral parts lack a coelomic cavity and are occupied by ECM and muscle cells. The fine structure of the T-shaped vessel is described, and its localization inside lophophoral coelom is demonstrated. We assert that the cavity inside the epistome is the preoral coelom corresponding to the true preoral coelom of the larva of this species. Proving this assertion will require additional study of metamorphosis in this species. To clarify the patterns of coelom organization in phoronids, we discuss the bipartite coelomic system in Phoronis and the tripartite coelomic system in Phoronopsis.     

Mesodermal Gene Expression in the Acoel Isodiametra pulchra Indicates a Low Number of Mesodermal Cell Types and the Endomesodermal Origin of the Gonads

Acoelomorphs are bilaterally symmetric small marine worms that lack a coelom and possess a digestive system with a single opening. Two alternative phylogenetic positions of this group within the animal tree are currently debated. In one view, Acoelomorpha is the sister group to all remaining Bilateria and as such, is a morphologically simple stepping stone in bilaterian evolution. In the other, the group is a lineage within the Deuterostomia, and therefore, has derived a simple morphology from a more complex ancestor. Acoels and the closely related Nemertodermatida and Xenoturbellida, which together form the Acoelomorpha, possess a very limited number of cell types. To further investigate the diversity and origin of mesodermal cell types we describe the expression pattern of 12 orthologs of bilaterian mesodermal markers including Six1/2, Twist, FoxC, GATA4/5/6, in the acoel Isodiametra pulchra. All the genes are expressed in stem cells (neoblasts), gonads, and at least subsets of the acoel musculature. Most are expressed in endomesodermal compartments of I. pulchra developing embryos similar to what has been described in cnidarians. Our molecular evidence indicates a very limited number of mesodermal cell types and suggests an endomesodermal origin of the gonads and the stem cell system. We discuss our results in light of the two prevailing phylogenetic positions of Acoelomorpha.     

An SEM study of cellular morphology,contact, and arrangement,as related to gastrulation inXenopus laevis

Summary Cellular morphology, contact, and arrangement in the late blastula and in various stages of gastrulation ofXenopus were examined by SEM of specimens dissected after fixation or fractured in amyl acetate. The prospective ectoderm of the blastocoel roof consists of several layers of interdigitating cells connected by numerous small protrusions which may function in the decrease in number of cell layers observed during ectodermal epiboly. During gastrulation, prospective mesoderm is regionally differentiated by cellular morphology and arrangement into preinvolution mesoderm, the mesodermal involution zone, and involuted mesoderm. The involuted anterodorsal (head), lateral, and ventral mesoderm consists of a stream of loosely-packed, irregularly shaped cells having large extensions of the cell body attached locally to other cells by small protrusions. Involuted posterodorsal mesoderm (chordamesoderm) consists of elongated cells arranged in palisade fashion and connected by similar protrusions. Involuted mesodermal cells in all regions are attached to the overlying prospective ectodermal cells by numerous small protrusions along the entire interface between the two cell layers. Suprablastoporal endodermal cells involute as an epithelial sheet, changing in shape in the process, to form the roof of the archenteron. Bottle cell morphology, arrangement, and position with respect to the mesodermal cell stream is described. Evidence presented here and elsewhere suggests that involution of mesoderm and of the archenteron roof inXenopus is dependent primarily upon the relative movement of the mesodermal cell stream and of the overlying ectoderm.     

Coelom formation: binary decision of the lateral plate mesoderm is controlled by the ectoderm.

Most triploblastic animals including vertebrates have a coelomic cavity that separates the outer and inner components of the body. The coelom is lined by two different tissue components, somatopleure and splanchnopleure, which are derived from the lateral plate region. Thus, the coelom is constructed as a result of a binary decision during early specification of the lateral plate. In this report we studied the molecular mechanisms of this binary decision. We first demonstrate that the splitting of the lateral plate into the two cell sheets progresses in an anteroposterior order and this progression is not coordinated with that of the somitic segmentation. By a series of embryological manipulations we found that young splanchnic mesoderm is still competent to be respecified as somatic mesoderm, and the ectoderm overlying the lateral plate is sufficient for this redirection. The lateral ectoderm is also required for maintenance of the somatic character of the mesoderm. Thus, the ectoderm plays at least two roles in the early subdivision of the lateral plate: specification and maintenance of the somatic mesoderm. We also show that the latter interactions are mediated by BMP molecules that are localized in the lateral ectoderm. Evolutionary aspects of the coelom formation are also considered.