The attachment complex of brachiolaria larvae of the sea star <Emphasis Type="Italic">Asterias rubens</Emphasis> (Echinodermata): an ultrastructural and immunocytochemical study

The attachment complex of brachiolaria larvae of the asteroid Asterias rubens comprises three brachiolar arms and an adhesive disc located on the preoral lobe. The former are used in temporary attachment and sensory testing of the substratum, whereas the latter is used for permanent fixation to the substratum at the onset of metamorphosis. Brachiolar arms are hollow structures consisting of an extensible stem tipped by a crown of dome-like ciliated papillae. The papilla epidermis is composed of secretory cells (type A, B and C cells), non-secretory ciliated cells, neurosecretory-like cells and support cells. Type A and B secretory cells fill a large part of the papilla epidermis and are always closely associated. They presumably form a duo-gland adhesive system in which type A and B cells are respectively adhesive and de-adhesive in function. The adhesive disc is an epidermal structure mainly composed of secretory cells and support cells. Secretory cells produce the cement, which anchor the metamorphic larva to the substratum until the podia are developed. The relatedness between the composition of the adhesive material in the brachiolaria attachment complex and in the podia of adults was investigated by immunocytochemistry using antibodies raised against podial adhesive secretions of A. rubens. Type A secretory cells were the only immunolabelled cells indicating that their temporary adhesive shares common epitopes with the one of podia. The attachment pattern displayed by the individuals of A. rubens during the perimetamorphic period—temporary, permanent, temporary—is unique among marine non-vertebrate Metazoa.     

Mechanisms of rapid morphogenetic movements in the metamorphosis of the bryozoan Bugula neritina (cheilostomata,cellularioidea). I. Attachment to the substratum

The larval morphology, settlement behavior, and the rapid morphogenetic movements that occur during the first 60 sec of metamorphosis of the cellularioid cheilostome bryozoan Bugula neritina have been examined and analyzed by light and electron microscopy. The larva attaches to the substratum at the onset of metamorphosis by the eversion of the internal sac. At the same time, the coronal cilia reverse their direction of beat, spreading an adhesive secreted by the neck region of the everting sac over the metamorphosing larva. During attachment, the larva goes through several configurations that coincide with the sequential contraction and relaxation of certain larval muscles. Histological and ultrastructural evidence indicates that the neck and wall regions of the internal sac are everted by the contraction of the muscles in the equatorial plane of the larva at the same time that the roof region in pulled toward the larval equator by the contraction of the axial muscles. The subsequent relaxation of the axial muscles allows the roof region to be everted by the antagonistic force generated by the sustained contraction of the equatorial musculature. After the roof region attaches to the substratum, the apical disc is temporarily retracted by a second contraction of the axial muscles. The apical disc subsequently reextends as the axial muscles relax just before coronal involution. A comparison of the ontogenetic sequence of rapid morphogenetic movements in the metamorphoses of cheilostome and ctenostome bryozoans indicates that cellularioid cheilostomes have undergone peramorphosis in the aspect of development.     

Adaptations to benthic development: functional morphology of the attachment complex of the brachiolaria larva in the sea star Asterina gibbosa

The asteroid Asterina gibbosa lives all its life in close relation to the sea bottom. Indeed, this sea star possesses an entirely benthic, lecithotrophic development. The embryos adhere to the substratum due to particular properties of their jelly coat, and hatching occurs directly at the brachiolaria stage. Brachiolariae have a hypertrophied, bilobed attachment complex comprising two asymmetrical brachiolar arms and a central adhesive disc. This study aims at describing the ultrastructure of the attachment complex and possible adaptations, at the cellular level, to benthic development. Immediately after hatching, early brachiolariae attach by the arms. All along the anterior side of each arm, the epidermis encloses several cell types, such as secretory cells of two types (A and B), support cells, and sensory cells. Like their equivalents in planktotrophic larvae, type A and B secretory cells are presumably involved in a duo-glandular system in which the former are adhesive and the latter de-adhesive in function. Unlike what is observed in planktotrophic larvae, the sensory cells are unspecialized and presumably not involved in substratum testing. During the larval period, the brachiolar arms progressively increase in size and the adhesive disc becomes more prominent. At the onset of metamorphosis, brachiolariae cement themselves strongly to the substratum with the adhesive disc. The disc contains two main cell types, support cells and secretory cells, the latter being responsible for the cement release. During this metamorphosis, the brachiolar arms regress while post-metamorphic structures grow considerably, especially the tube feet, which take over the role of attachment to the substratum. The end of this period corresponds to the complete regression of the external larval structures, which also coincides with the opening of the mouth. This sequence of stages, each possessing its own adhesive strategy, is common to all asteroid species having a benthic development. In A. gibbosa, morphological adaptations to this mode of development include the hypertrophic growth of the attachment complex, its bilobed shape forming an almost completely adhesive sole, and the regression of the sensory equipment.     

Evaluation of the attachment strength of individuals of Asterina gibbosa (Asteroidea, Echinodermata) during the perimetamorphic period

A turbulent channel flow apparatus was used to determine the adhesion strength of the three perimetamorphic stages of the asteroid Asterina gibbosa, i.e. the brachiolaria larvae, the metamorphic individuals and the juveniles. The mean critical wall shear stresses (wall shear stress required to dislodge 50% of the attached individuals) necessary to detach larvae attached by the brachiolar arms (1.2 Pa) and juveniles attached by the tube feet (7.1 Pa) were one order of magnitude lower than the stress required to dislodge metamorphic individuals attached by the adhesive disc (41 Pa). This variability in adhesion strength reflects differences in the functioning of the adhesive organs for these different life stages of sea stars. Brachiolar arms and tube feet function as temporary adhesion organs, allowing repetitive cycles of attachment to and detachment from the substratum, whereas the adhesive disc is used only once, at the onset of metamorphosis, and is responsible for the strong attachment of the metamorphic individual, which can be described as permanent adhesion. The results confirm that the turbulent water channel apparatus is a powerful tool to investigate the adhesion mechanisms of minute organisms.     

Larval Attachment by Eversion of the Internal Sac in the Marine Bryozoan Bowerbankia gracilis (Ctenostomata: Vesicularioidea): A Muscle-Mediated Morphogenetic Movement

The larval morphology and settlement of the vesicularioid ctenostome bryozoan Bowerbankia gracilis has been investigated by light and electron microscopy in an attempt to elucidate the mechanism of attachment to the substratum at the onset of metamorphosis. The oral epithelium in the free-swimming larva is infolded to form a glandular internal sac at the oral pole. The internal sac is not specialized into distinct regions, but consists of a uniform, simple columnar epithelium filled with secretory granules. The hemispherical internal sac is underlain by a cup-shaped layer of undifferentiated cells that constitutes the polypide rudiment. The cupiform layer of undifferentiated cells is in turn embraced by a network of muscle fibers called the rete muscularis. At the onset of metamorphosis, the larva constricts oro-laterally and the internal sac is everted against the substratum. As the sac everts, the glandular cells secrete an adhesive that is wafted up over the metamorphosing larva by the reversed beating of the coronal cilia. At the same time, the cupiform layer of undifferentiated cells flattens in the plane of the oro-lateral constriction and doubles in thickness. The cells of the cupiform layer undergo a corresponding transformation from short columnar cells to flask-shaped cells that bulge into the glandular cells of the internal sac. The narrow ends of the flask-shaped cells abut the strongly contracted muscle fibers of the rete muscularis. It is hypothesized that the contraction of the muscle fibers of the rete muscularis is responsible for the change in shape of the undifferentiated cells and, consequently, for the eversion of the internal sac. On the basis of this study and a review of the literature, it is concluded that attachment to the substratum at the onset of metamorphosis typically is effected by the eversion of an internal sac in larvae of the ctenostome superfamily Vesicularioidea.     

Neuromuscular structure of the larva to early ancestrula stages of the cyclostome bryozoan Crisia eburnea

For the first time, the development of a cyclostome bryozoan has been studied with immunochemistry and confocal laser scanning microscopy, with emphasis on nerves and muscles. The larva is covered by multiciliated cells, which are latitudinally strongly elongated and show phalloidin-stained cell junctions. We hypothesize that these cells contract at metamorphosis and squeeze the apical invagination and the adhesive sac out. Ectodermal, longitudinal muscle cells extend from the cells of the inner, conical cuticularized part of the apical invagination to the lower part of the corona, around the adhesive sac pore. These muscles are retained in the ancestrula. Scattered monociliated nerve cells are interspersed between the coronal ciliary cells. An equatorial nerve in the larva disappears at metamorphosis. The central, conical part of the cuticle becomes the terminal membrane of the ancestrula, and the underlying ectodermal and mesodermal cell layers differentiate into the polypide bud, forming a deep narrow invagination, differentiating into vestibule–atrium, mouth ring and pharynx–stomach–rectum. Tentacles develop from the ring of cells around the mouth, and a small ganglion with four nerves innervating each of the tentacles develops at the anal side of the mouth. These new findings yield further support for previous homology statements of bryozoan larvae and development.     

Evaluation of the attachment strength of individuals of Asterina gibbosa (Asteroidea,Echinodermata) during the perimetamorphic period

Abstract

A turbulent channel flow apparatus was used to determine the adhesion strength of the three perimetamorphic stages of the asteroid Asterina gibbosa, i.e. the brachiolaria larvae, the metamorphic individuals and the juveniles. The mean critical wall shear stresses (wall shear stress required to dislodge 50% of the attached individuals) necessary to detach larvae attached by the brachiolar arms (1.2 Pa) and juveniles attached by the tube feet (7.1 Pa) were one order of magnitude lower than the stress required to dislodge metamorphic individuals attached by the adhesive disc (41 Pa). This variability in adhesion strength reflects differences in the functioning of the adhesive organs for these different life stages of sea stars. Brachiolar arms and tube feet function as temporary adhesion organs, allowing repetitive cycles of attachment to and detachment from the substratum, whereas the adhesive disc is used only once, at the onset of metamorphosis, and is responsible for the strong attachment of the metamorphic individual, which can be described as permanent adhesion. The results confirm that the turbulent water channel apparatus is a powerful tool to investigate the adhesion mechanisms of minute organisms.     

Adhesion and fusion of the extraembryonic epiblast

A tissue culture model system has been devised to examine the attachment, expansion, and fusion of epithelial cell sheets. A normal embryonic epithelial tissue, the extraembryonic epiblast of the chick, is isolated mechanically and cultured on its natural substratum, the vitelline membrane. This persistently migratory tissue has distinct adhesive and non-adhesive regions. A serum-free chemically-defined culture medium has been formulated that permits determination of the effects of individual growth and trophic factors. Attachment of transferred epiblasts is dependent upon the presence of mineralocorticoids in the medium. This suggests that fluid transport is required for the cell sheet to make its initial attachment to the culture substratum. Expansion of the cell sheet following attachment, and the fusion of epiblasts advancing toward each other, does not require the presence of mineralocorticoid. No exogenous adhesive glycoproteins are required for attachment, expansion, or fusion. Antibody localization shows that endogenous laminin is present on the attachment surface of the specialized adhesive edge region of the extraembryonic epiblast. Following fusion of confronted epiblasts into one coherent cell sheet, the laminin disappears. Throughout these studies the adhesive and non-adhesive regions of the epiblast are identified by their characteristic distributions of actin microfilaments, localized using rhodamine-phalloidin staining.     

Micro- and nanostructure of the adhesive material secreted by the tube feet of the sea star Asterias rubens

To attach to underwater surfaces, sea stars rely on adhesive secretions produced by specialised organs, the tube feet. Adhesion is temporary and tube feet can also voluntarily become detached. The adhesive material is produced by two types of adhesive secretory cells located in the epidermis of the tube foot disc, and is deposited between the disc surface and the substratum. After detachment, this material remains on the substratum as a footprint. Using LM, SEM, and AFM, we described the fine structure of footprints deposited on various substrata by individuals of Asterias rubens. Ultrastructure of the adhesive layer of attached tube feet was also investigated using TEM. Whatever the method used, the adhesive material appeared as made up of globular nanostructures forming a meshwork deposited on a thin homogeneous film. This appearance did not differ according to whether the footprints were fixed or not, and whether they were observed hydrated or dry. TEM observations suggest that type 2 adhesive cells would be responsible for the release of the material constituting the homogeneous film whereas type 1 adhesive cells would produce the material forming the meshwork. This reticulated pattern would originate from the arrangement of the adhesive cell secretory pores on the disc surface.     

Evidence for adhesive activity of the ventrolateral flange in Giardia lamblia

Giardia lamblia is an intestinal protozoan that inhabits the intestinal tract of man and other mammals by attaching to the mucosal surface via the contractile activity of an attachment organelle called the ventral adhesive disk. We have investigated the presence of other attachment mechanisms in G. lamblia trophozoites by using microfabricated substrates that sterically interfere with formation of the hypothesized "negative pressure" under the ventral adhesive disk that would mediate attachment to a substratum. Pillars measuring 1 microm high and 2 microm in diam. were constructed in microarrays with spacings smaller than the diameter of the ventral adhesive disk. Using high resolution field emission scanning electron microscopy, the attachment of trophozoites to the tops of pillars in the microfabricated substrates was investigated. Firm adhesion of trophozoites was observed to be mediated by direct attachment of the ventrolateral flange membrane to the tops of microfabricated pillars. Attachment to microfabricated surfaces was 16% of that observed for attachment mediated by the ventral adhesive disk (4.4 +/- 1.5 cells/100 micro2 micropillar surface vs. 25.9 +/- 3.1 cells/100 micro2 flat substrate, p < 0.0001) This is the first report of trophozoite adhesion to a substratum by a mechanism other than the direct attachment of the ventral adhesive disk, and provides experimental evidence that the ventrolateral flange may play a role in trophozoite adhesion. A hypothesis is presented describing how the adhesive nature of the ventrolateral flange might be involved in normal attachment of G. lamblia trophozoites to a substratum.     

Structure of the organs of attachment of brachiolaria larvae of Stichaster australis (Verrill) and Coscinasterias calamaria (Gray) (Echinodermata: Asteroidea)

The structure of the brachiolar arms and adhesive disk of the brachiolaria larvae of Stichaster australis (Verrill) and Coscinasterias calamaria (Gray) was determined from light microscopy and from scanning and transmission electron microscopy. The structure of these organs was very similar in both species.The brachiolar arms are comprised of a stem region terminating in a crown of adhesive papillae which are made up of a variety of secretory cell types. Principal among these are elongated cells producing very electron-dense secretory particles, which are released at the free cell surface attached to cilia. Secretory particles appear to be important in temporary attachment of the brachiolar arms to the substratum. Ciliary sense cells, possibly used in the recognition of specific substrata are located at the tip of adhesive papillae.The adhesive disk is comprised of large cells packed with secretory droplets and elongated intracellular fibres. In the attached adhesive disk, secretory droplets are lost, having formed the cement that attaches the disk to the substratum. It appears that adhesive papillae lateral to the adhesive disk hold the disk in position close to the substratum during secretion and hardening of the cement. The intracellular fibres are the principal anchoring structures running from microvilli (locked into the attachment cement) on the surface of the disk to the underlying connective tissue of the attachment stalk.     

Unexpected role of plasminogen activator inhibitor 1 in cell adhesion and detachment

Plasminogen activator inhibitor 1 (PAI-1) is the primary physiological inhibitor of plasminogen activation in vivo, and thus it is one of the main regulators of the fibrinolytic system. In this regard, individuals with elevated PAI-1 seem to have an increased risk for thrombotic disease, whereas those lacking the inhibitor develop a lifelong bleeding diathesis. Unexpectedly, recent observations demonstrate that cancer patients with high PAI-1 levels have a poor prognosis for survival. This correlation with metastatic disease may be related to the observation that high PAI-1 levels decrease the adhesive strength of cells for their substratum, and that this de-adhesive activity of PAI-1 is not related to its role as a protease inhibitor. Initial insights into potential mechanisms by which PAI-1 regulates the attachment, detachment, and migration of cells are addressed in this review.     

Structure of Adnate Colony Portions in Crisiidae (Bryozoa Cyclostomata)

The “rhizoidsrdquo; surrounding the base of the erect “colony” emanating from the ancestrula in the Crisiidae, especially the simple species Crisidia cornuta (L.), are a regular adnate system of autozooids. Each autozooid is composed of a proximal adnate part and a distal peristome (in some species kenozooids are possibly intercalated). The autozooid peristomes support erect branches identical in budding and structure with the branch emanating from the erect peristome of the ancestrula. Thus, the complete crisiid colony consists of an adnate system of ancestrula and autozooids, which form erect branches from their peristomes. The adnate zooid system is comparable in autozooid morphology and budding pattern with simple uniserial stomatoporids. The tentative hypothesis is proposed that the crisiid group has developed from primitive stomatoporids; the adnate zooid system of the stomatoporids apparently evolved peristomial budding to produce the erect colony branches characteristic of crisiids.     

Spinning activity of the spider Trogloneta granulum (Araneae, Mysmenidae): web, cocoon, cocoon handling behaviour, draglines and attachment discs

The spider species Trogloneta granulum, which in the wild lives inside scree slopes, builds three-dimensional orb webs. During egg-laying and egg sac building, the females stay with their dorsa down at the central part of the web. In this process, the hub is used as a platform. The threads of the hub are not incorporated into the silk cover of the egg sac. The silk wall of the egg sac is very thin, with all the silken threads constituting it having a uniform ultrastructure. The silk wall of the egg sac and the spinnerets are permanently linked by a dragline. Draglines produced by T. granulum affect the direction of movements of the female carrying its cocoon. Egg sacs are handled using draglines. The low number of piriform glands leads to the formation of very simple attachment discs, which fix the individual threads to the substratum. Thread bundles are attached to the substratum by means of accumulated attachment discs.     

On the morphology of antennular sensory and attachment organs in cypris larvae of the deep‐sea vent/seep barnacles,Ashinkailepas and Neoverruca

Barnacle cypris larvae show high morphological variation in the organs used in search of and attaching to a substratum. This variation may represent adaptation to the habitat of the species. Here, we studied SEM level morphologies of cypris antennular sensory and attachment organs in a deep‐sea vent endemic species (Neoverruca sp.) and a vent/seep inhabiting species (Ashinkailepas seepiophila). We compare them with three species from other environments. The antennular morphologies of Neoverruca sp. and A. seepiophila were similar, which is consistent with recent molecular studies showing a close relationship of the two species. The setation pattern of the antennules was very conservative among species from various environments. In contrast, striking differences were observed in the structure of the attachment organ (the third antennular segment). Neoverruca sp. and A. seepiophila had no velum or a skirt surrounding the attachment disc on the third segment, while other cirripede cyprids almost always have either of these structures. In addition, both cyprids of A. seepiophila and Neoverruca sp. had the attachment disc angled toward the substratum, whereas it faces distally in cyprids from hard bottom inhabiting barnacles. We suggest that both velum/skirt and the angle of the attachment disc play an important role, when the antennules are contacting the substratum during surface exploration. Differences in attachment organ structures may be highly adaptive, enabling cirripede species to enter new habitats during evolution. J. Morphol. 277:594–602, 2016. © 2016 Wiley Periodicals, Inc.     

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.     

Adhesive mechanisms in cephalopods: a review

Several genera of cephalopods (Nautilus, Sepia, Euprymna and Idiosepius) produce adhesive secretions, which are used for attachment to the substratum, for mating and to capture prey. These adhesive structures are located in different parts of the body, viz. in the digital tentacles (Nautilus), in the ventral surface of the mantle and fourth arm pair (Sepia), in the dorsal epidermis (Euprymna), or in the dorsal mantle side and partly on the fins (Idiosepius). Adhesion in Sepia is induced by suction of dermal structures on the mantle, while for Nautilus, Euprymna and Idiosepius adhesion is probably achieved by chemical substances. Histochemical studies indicate that in Nautilus and Idiosepius secretory cells that appear to be involved in adhesion stain for carbohydrates and protein, whilst in Euprymna only carbohydrates are detectable. De-adhesion is either achieved by muscle contraction of the tentacles and mantle (Nautilus and Sepia) or by secretion of substances (Euprymna). The de-adhesive mechanism used by Idiosepius remains unknown.     

Identification of two putative adhesive polypeptides in Caulerpa prolifera rhizoids using an adhesion model system

The rhizoid section of the green alga Caulerpa prolifera (Cp) is active in attaching the developing plant to the substratum. A model system for the study of the adhesion of Cp rhizoids has been developed and identification of two putative adhesive polypeptides of Caulerpa (Vn-Cp) was revealed by immunodetection. A method for fast induction of new rhizoids was established using blade-base cutting followed by a few days of incubation. The new rhizoids were gently enclosed between two cover glasses and incubated until firm attachment developed. While analyzing protein extracts, two ∼60–70 kDa polypeptides (Vn-Cp I and Vn-Cp II) were identified by immunodetection with monoclonal antibodies to human vitronectin (Vn). The relative concentration values of the Vn-Cp proteins increased significantly in the ‘cell-wall’ fraction of the attached rhizoids during the incubation period. However, Vn-Cp proteins were not detected in non-attached rhizoids. Furthermore, the Vn-Cp proteins were also detectable on glass substratum subsequent to attached rhizoid removal. The induction and accumulation of Vn-Cp proteins on the ‘cell-wall’ of Caulerpa rhizoids and the firm attachment of the rhizoids to the glass substratum during the incubation period suggest that Vn-Cp proteins play a significant role in adhesion, which may be similar to the function of vitronectin in other adhesion systems. Furthermore, the high accumulation of Vn-Cp proteins on the glass substratum during attachment of new rhizoids suggests that the Vn-Cp proteins are secreted to the extracellular matrix and directly connect rhizoids to the glass substratum as an intermediate compound. These unique properties of Cp make it an excellent model system for the establishment of high amounts of adhesive material for future research. This revised version was published online in August 2006 with corrections to the Cover Date.     

Organization of the flagellar apparatus and associate cytoplasmic microtubules in the quadriflagellate alga Polytomella agilis

The organization of microtubular systems in the quadriflagellate unicell Polytomella agilis has been reconstructed by electron microscopy of serial sections, and the overall arrangement confirmed by immunofluorescent staining using antiserum directed against chick brain tubulin. The basal bodies of the four flagella are shown to be linked in two pairs of short fibers. Light microscopy of swimming cells indicates that the flagella beat in two synchronous pairs, with each pair exhibiting a breast-stroke-like motion. Two structurally distinct flagellar rootlets, one consisting of four microtubules in a 3 over 1 pattern and the other of a striated fiber over two microtubules, terminate between adjacent basal bodies. These rootlets diverge from the basal body region and extend toward the cell posterior, passing just beneath the plasma membrane. Near the anterior part of the cell, all eight rootlets serve as attachment sites for large numbers of cytoplasmic microtubules which occur in a single row around the circumference of the cell and closely parallel the cell shape. It is suggested that the flagellar rootless may function in controlling the patterning and the direction of cytoplasmic microtubule assembly. The occurrence of similar rootlet structures in other flagellates is briefly reviewed.