Serotonergic and FMRFamidergic nervous systems in gymnolaemate bryozoan larvae

A growing body of data from nervous systems of marine invertebrate larvae provides an ideal background for comparisons among higher taxa. The currently available data from Bryozoa, however, do not allow for a consistent hypothesis of an ancestral state for this taxon, which would be necessary for phylogenetic inferences. The larval nervous systems of the four gymnolaemate species Flustrellidra hispida, Bugula fulva, Alcyonidium gelatinosum, and Bowerbankia gracilis are examined by means of antibody staining against the neurotransmitters serotonin and FMRFamide, as well as against acetylated α-tubulin. Despite considerable variation, a comparison reveals a common pattern of the distribution of serotonin. The neurotransmitter is found in at least two cells in the apical organ as well as in paired axial and lateral nerves emerging from a central nerve nodule. A ring nerve is present below the corona and at least two serotonergic cells are found between the corona cells. Serotonergic coronal cells might represent unique bryozoan features, whereas the remaining elements show resemblance to the situation found in most spiralian taxa. The data do not provide support for a closer relationship of Bryozoa to Phoronida or Brachiopoda.     

Inferring larval type in fossil bryozoans

Pachut, J.F. & Fisherkeller, P. 2010: Inferring larval type in fossil bryozoans. Lethaia, Vol. 43, pp. 396–410. Larval type in extinct organisms might be recognizable because larvae of living marine invertebrates are approximately of the same size as the initial post‐larval organism. Two larval types typically occur. Planktotrophic larvae feed on other members of the plankton, potentially prolonging their larval existence and producing broad geographic distributions. Conversely, lecithotrophic larvae feed on yolk supplied by the fertilized egg, often settle quickly after release, and display more restricted distributions. However, some lecithotrophic bryozoans undergo embryonic fission forming multiple, small, polyembryonic larvae. The relationship between post‐larval size and larval type was evaluated in bryozoans by comparing the size of the ancestrula, the founding individual of a colony, to the sizes of extant planktotrophic, lecithotrophic and polyembryonic lecithotrophic larvae and ancestrulae. The sizes of larvae and ancestrulae in extant lecithotrophic and planktotrophic cheilostome (gymnolaemate) species are statistically the same. They are, however, statistically larger than the polyembryonic larvae of extant cyclostomes (stenolaemates). In turn, the sizes of cyclostome larvae are indistinguishable from the ancestrulae of extant and fossil cyclostomes, the ancestrulae of other fossil stenolaemate species measured from the literature, and the ancestrulae of three of four genera from North American Cincinnatian strata. Ancestrulae of a fourth genus, Dekayia, are the same size as cyclostome ancestrulae but are statistically smaller than the ancestrulae of other stenolaemates. With few exceptions, stenolaemates have statistically smaller larvae and ancestrulae than both lecithotrophic and planktotrophic cheilostomes. We infer that the sizes of fossil ancestrulae permit the discrimination of taxa that had polyembryonic lecithotrophic larvae from those possessing other larval types. This inference is strengthened, in several cases, by the co‐occurrence of brood chambers (gynozooecia) and restricted palaeobiogeographic distributions. The presence of cyclostomes in Early Ordovician strata suggests that polyembryony may have been acquired during the initial radiation of Class Stenolaemata. Polyembryony appears to be a monophyletic trait, but confirmation requires the demonstration that species of several stenolaemate suborders lacking skeletally expressed brood chambers possessed polyembryonic larvae. □Ancestrulae, evolution, fossil bryozoans, gynozooecia, larvae.     

Adrenoceptor Compounds Prevent the Settlement of Marine Invertebrate Larvae: Balanus amphitrite (Cirripedia), Bugula neritina (Bryozoa) and Hydroides elegans (Polychaeta)

The effects of the neurotransmitter blockers idazoxan and phentolamine on the larval settlement of three marine invertebrate species belonging to three different phyla were investigated by using in vitro concentration-response bioassays. Since neurotransmitters are known to influence metamorphic transitions in invertebrate larvae, neurotransmitter blockers were tested to evaluate their sublethal effects on larvae. The α-adrenergic antagonists idazoxan and phentolamine inhibited settlement of Balanus amphitrite (Cirripedia), Bugula neritina (Bryozoa) larvae, and larvae of the polychaete Hydroides elegans (Polychaeta) in a concentration- and taxon-dependent manner. At concentrations of 10^?3 M of both agents, larvae of all three species became immobile and subsequently died within 24 h. While cumulative settlement rates were observed after 48 h for B. amphitrite and H. elegans, and after 5 h for B. neritina, > 90% of the larvae that settled did so within 24 h for the first two species and within 1 h for B. neritina. The tendency of the hydrophobic idazoxan and phentolamine to accumulate at solid surfaces most probably contributes to their successful inhibition of larval settlement. This ability makes them particularly attractive as candidates for the development of slow-release carriers in antifouling paints.     

Structure and occurrence of cyphonautes larvae (bryozoa,ectoprocta)

We have studied larvae of the freshwater ctenostome Hislopia malayensis with scanning electron microscopy (SEM), confocal laser scanning microscopy (CLSM), and LM of serial sections. Some additional observations on larvae of M. membranacea using SEM and CLSM are also reported. The overall configuration of muscles, nerves, and cilia of the two larvae are identical. However, the larva of H. malayensis is much smaller than that of M. membranacea, which may explain most of the differences observed. Although all major nerves and muscle strands are present in H. malayensis, they are generally composed of fewer fibers. The H. malayensis larva lacks the anterior and posterior intervalve cilia. Its pyriform organ is unciliated with only a small central depression. The adhesive epithelium is not invaginated as an adhesive sac and lacks the large muscles interpreted as adhesive sac muscles in the M. membranacea larva. The velum carries two rows of ciliated cells, though the lower “row” consists of only one or two cells. Both rows of ciliated cells are innervated by nerves, which have not been detected in the M. membranacea larva. The ciliated ridge of H. malayensis lacks the frontal cilia. The planktotrophic cyphonautes larvae in a number of ctenostome clades and in the “basal” cheilostome clade Malacostega (and probably in the earliest cheilostomes) support the idea that the cyphonautes larva is the ancestral larval type of the Eurystomata. It may even represent the ancestral larval type of the bryozoans (= ectoprocts). J. Morphol. 271:1094‐1109, 2010. © 2010 Wiley‐Liss, Inc.     

Ultrastructure of mesoderm formation and development in Membranipora membranacea (Bryozoa: Gymnolaemata)

Mesoderm origin in Bryozoa is largely unknown. In this study, embryonic and early larval stages of Membranipora membranacea, a bryozoan exhibiting a planktotrophic cyphonautes larva, are investigated using mainly ultrastructural techniques. Shortly after the onset of gastrulation, an ectodermal cell, which is situated centrally at the prospective anterior pole of the larva, can be recognized by its constricted apical surface and enlarged basal part. It is also distinct from other ectodermal cells by the composition of its cytoplasm. In later stages, it has left the epidermis, lost its epithelial character, and is situated subepithelially, between the basal sides of the ectodermal and endodermal sheets. A blastocoelic cavity is not present at this stage. This cell divides and gives rise to a group of cells forming a muscular and neuronal strand at the anterior side of the larva. The majority of the larval musculature originates from this ingression. Despite this evidence for an ectodermal origin, additional sources of mesoderm can so far not be excluded. The literature on mesoderm origin in Bryozoa is reviewed and the results are compared to known data from other metazoan taxa.     

Potent cytotoxins produced by a microbial symbiont protect host larvae from predation

Larvae of the sessile marine invertebrate Bugula neritina (Bryozoa) are protected by an effective chemical defense. From the larvae, we isolated three bryostatin-class macrocyclic polyketides, including the novel bryostatin 20, that deterred feeding by a common planktivorous fish that co-occurs with B. neritina. A unique bacterial symbiont of B. neritina, Endobugula sertula, was hypothesized as the putative source of the bryostatins. We show that: (1) bryostatins are concentrated in B. neritina larvae and protect them against predation by fish; (2) the adults are not defended by bryostatins; and (3) E. sertula produces bryostatins. This study represents the first example from the marine environment of a microbial symbiont producing an anti-predator defense for its host and, in this case, specifically for the hosts larval stage, which is exceptionally vulnerable to predators.Electronic Supplementary Material Supplementary material is available in the online version of this article at     

Development of the musculature in the limpet Patella (Mollusca, Patellogastropoda)

 Whole-mount technique using fluorescent-labelled phalloidin for actin staining and confocal laser scanning microscopy as well as semi-thin serial sectioning, scanning and transmission electron microscopy were applied to investigate the ontogeny of the various muscular systems during larval development in the limpets Patella vulgata L. and P. caerulea L. In contrast to earlier studies, which described a single or two larval shell muscles, the pretorsional trochophore-like larva shows no less than four different muscle systems, namely the asymmetrical main head/foot larval retractor muscle, an accessory larval retractor with distinct insertion area, a circular prototroch/velar system, and a plexus-like pedal muscle system. In both Patella species only posttorsional larvae are able to retract into the shell and to close the aperture by means of the operculum. Shortly after torsion the two adult shell muscles originate independently in lateral positions, starting with two fine muscle fibres which insert at the operculum and laterally at the shell. During late larval development the main larval retractor and the accessory larval retractor become reduced and the velar muscle system is shed. In contrast, the paired adult shell muscles and the pedal muscle plexus increase in volume, and a new mantle musculature, the tentacular muscle system, and the buccal musculature arise. Because the adult shell muscles are entirely independent from the various larval muscular systems, several current hypotheses on the ontogeny and phylogeny of the early gastropod muscle system have to be reconsidered. Received: 23 June 1998 / Accepted: 25 November 1998     

Myogenesis in two polyclad platyhelminths with indirect development,Pseudoceros canadensis and Stylostomum sanjuania

SUMMARY Myogenesis of two representatives of Platyhelminthes, Stylostomum sanjuania and Pseudoceros canadensis, was followed from egg deposition until well‐differentiated free‐swimming larval stages, using F‐actin staining and confocal laserscanning microscopy. Zonulae adhaerentes are the only structures to stain before 50% of development between egg deposition and hatching in S. sanjuania, and before 67% of development in P. canadenis. Subsequently, irregular fibers appear in the embryo, followed by a helicoid muscle close to the apical pole. Three longitudinal muscle pairs form, of which the dorsal pair remains more pronounced than the others. Gradually, new muscles form by branching or from double‐stranded muscle zones adjacent to existing muscles. This results in an elaborate muscular bodywall that consists of a single helicoid muscle as well as multiple circular and longitudinal muscles. Diverse retractor muscles insert at the sphincter muscles around the stomodeum. The overall arrangement and formation mode of the larval musculature appears very similar in both species, although only P. canadensis has a primary circular muscle posterior to the helicoid muscle. Muscle formation in the apical region of the embryo precedes that at the abapical pole and the primary longitudinal muscles form slightly later than the primary circular muscles. Myogenesis and larval myoanatomy appears highly conserved among polyclad flatworms, but differs significantly from that of other trochozoan clades. Our data suggest that the larval muscular ground pattern of polyclad larvae comprises a bodywall consisting of a helicoid muscle, circular and longitudinal muscles, several retractor muscles, and sphincter muscles around the stomodeum.     

Neuromuscular development in Novocrania anomala: evidence for the presence of serotonin and a spiralian-like apical organ in lecithotrophic brachiopod larvae

SUMMARY The phylogenetic position of Brachiopoda remains unsettled, and only few recent data on brachiopod organogenesis are currently available. In order to contribute data to questions concerning brachiopod ontogeny and evolution we investigated nervous and muscle system development in the craniiform (inarticulate) brachiopod Novocrania anomala . Larvae of this species are lecithotrophic and have a bilobed body with three pairs of dorsal setal bundles that emerge from the posterior lobe. Fully developed larvae exhibit a network of setae pouch muscles as well as medioventral longitudinal and transversal muscles. After settlement, the anterior and posterior adductor muscles and delicate mantle retractor muscles begin to form. Comparison of the larval muscular system of Novocrania anomala with that of rhynchonelliform (articulate) brachiopod larvae shows that the former has a much simpler muscular organization. The first signal of serotonin-like immunoreactivity appears in fully developed Novocrania anomala larvae, which have an apical organ that consists of four flask-shaped cells and two ventral neurites. These ventral neurites do not stain positively for the axonal marker α-tubulin in the larval stages. In the juveniles, the nervous system stained by α-tubulin is characterized by two ventral neurite bundles with three commissures. Our data are the first direct proof for the presence of an immunoreactive neurotransmitter in lecithotrophic brachiopod larvae and demonstrate the existence of flask-shaped serotonergic cells in the brachiopod larval apical organ, thus significantly increasing the probability that this cell type was part of the bauplan of the larvae of the last common lophotrochozoan ancestor.     

Adrenoceptor compounds prevent the settlement of marine invertebrate larvae: Balanus amphitrite (Cirripedia), Bugula neritina (Bryozoa) and Hydroides elegans (Polychaeta)

The effects of the neurotransmitter blockers idazoxan and phentolamine on the larval settlement of three marine invertebrate species belonging to three different phyla were investigated by using in vitro concentration-response bioassays. Since neurotransmitters are known to influence metamorphic transitions in invertebrate larvae, neurotransmitter blockers were tested to evaluated their sublethal effects on larvae. The alpha-adrenergic antagonists idazoxan and phentolamine inhibited settlement of Balanus amphitrite (Cirripedia), Bugula neritina (Bryozoa) larvae, and larvae of the polychaete Hydroides elegans (Polychaeta) in a concentration-and taxon-dependent manner. At concentrations of 10(-3) M of both agents, larvae of all three species became immobile and subsequently died within 24 h. While cumulative settlement rates were observed after 48 h for B. amphitrite and H. elegans, and after 5 h for B. neritina, >90% of the larvae that settled did so within 24 h for the first two species and within 1 h for B. neritina. The tendency of the hydrophobic idazoxan and phentolamine to accumulate at solid surfaces most probably contributes to their successful inhibition of larval settlement. This ability makes them particularly attractive as candidates for the development of slow-release carriers in antifouling paints.     

Morphogenesis and phenotypic divergence in two developmental morphs of Streblospio benedicti (Annelida,Spionidae)

Abstract. The morphology of marine invertebrate larvae is strongly correlated with egg size and larval feeding mode. Planktotrophic larvae typically have suites of morphological traits that support a planktonic, feeding life style, while lecithotrophic larvae often have larger, yolkier bodies, and in some cases, a reduced expression of larval traits. Poecilogonous species provide interesting cases for the analysis of early morphogenesis, as two morphs of larvae are produced by a single species. We compared morphogenesis in planktotrophic and lecithotrophic morphs of the poecilogonous annelid Streblospio benedicti from the trochophore stage through metamorphosis, using observations of individuals that were observed alive, with scanning electron microscopy, or in serial sections. Offspring of alternate developmental morphs of this species are well known to have divergent morphologies in terms of size, yolk content, and the presence of larval bristles. We found that some phenotypic differences between morphs occur as traits that are present in only one morph (e.g., larval bristles, bacillary cells on the prostomium and pygidium), but that much of the phenotypic divergence is based on heterochronic changes in the differentiation of shared traits (e.g., gut and coelom). Tissue and organ development are compared in both morphs in terms of their structure and ontogenetic change throughout early development and metamorphosis.     

Reversible anti-settlement activity against Amphibalanus (=Balanus) amphitrite,Bugula neritina,and Hydroides elegans by a nontoxic pharmaceutical compound,mizolastine

Mizolastine, an antihistamine pharmaceutical, was found to significantly inhibit larval settlement of the barnacle Amphibalanus (=Balanus) amphitrite, the bryozoan Bugula neritina, and the polychaete Hydroides elegans with EC₅₀ values of 4.2, 11.2, and 4.1 µg ml^?1, respectively. No toxicity against the larvae of these three species was observed at the concentration range tested during incubations with mizolastine. To determine whether the anti-settlement activity of mizolastine is reversible, recovery bioassays using these three species were conducted. More than 70% of the larvae that had been exposed for 4 h to mizolastine at concentrations four-fold greater than their respective EC₅₀ values completed normal metamorphosis. The results of the recovery bioassay provide evidence that the anti-settlement effect of mizolastine is reversible in addition to being nontoxic. The anti-settlement activities of several intermediates of the synthesis process of mizolastine were also examined. One of the intermediates, 2-chloro-1-(4-fluorobenzyl)-1H-benzo[d]imidazole, inhibited larval settlement and metamorphosis with low toxicity. These results may improve the understanding of the key functional group responsible for the anti-settlement activity of mizolastine.     

The fate of the larval epidermis in the Desor-larva of <Emphasis Type="Italic">Lineus viridis</Emphasis> (Pilidiophora,Nemertea) displays a historically constrained functional shift from planktotrophy to lecithotrophy

Pilidiophora constitutes a clade of nemerteans characterized by a peculiar larval type, the pilidium. A characteristic of this larva is the transitory epidermis in which the juvenile develops from imaginal discs. The primary function of this larval envelope is assumed to be feeding and dispersal. When juvenile development is complete, the larval epidermis is ruptured and swallowed by the juvenile. According to recent cladistic and molecular analyses of the Nemertea, the intracapsular Desor-larva of the sibling species Lineus viridis and L. ruber is thought to have evolved from a pelagic pilidium. The general course of development has been demonstrated to be similar to that of the pilidium, in which the juvenile forms from imaginal discs under the larval epidermis. The two Lineus species, however, differ in their mode of larval feeding: L. ruber being ootrophic and L. viridis being lecithotrophic. In order to elucidate the transition from the planktotrophic pilidum to lecithotrophic development, I studied the early cleavage and metamorphosis from intracapsular Desor-larva to juvenile stages in L. viridis from the island of Sylt, using light microscopical, electron microscopical, and fluorescent staining methods. Due to the specific cleavage pattern with equally sized 1st quartet animal blastomeres and vegetal blastomeres in L. viridis, the larval epidermis later contains a considerable amount of the yolk reserve. During metamorphosis, the larval epidermis is ingested by the juvenile thus displaying behavior similar to that of the pilidium larva. In contrast to the pilidium, the function of the larval epidermis of the Desor-larva has shifted from feeding and dispersal to direct food supply. Thus, the development of L. viridis is a perfect example for strong historical constraints that prevent ancestral larval structures from being lost.     

Some Like It Fat: Comparative Ultrastructure of the Embryo in Two Demosponges of the Genus Mycale (Order Poecilosclerida) from Antarctica and the Caribbean

During embryogenesis, organisms with lecithotrophic indirect development usually accumulate large quantities of energetic reserves in the form of yolk that are necessary for larval survival. Since all sponges have lecithotrophic development, yolk formation is an ineludible step of their embryogenesis. Sponge yolk platelets have a wide range of morphological forms, from entirely lipid or protein platelets to a combined platelet showing both lipids and proteins and even glycogen. So far, there are no comparative studies on the nature and content of yolk in congeneric species of sponges inhabiting contrasting environments, which could have putative effects on the larval adaptation to environmental conditions. Here, we have taken advantage of the worldwide distribution of the sponge genus Mycale, in order to compare the embryogenesis and yolk formation in two species inhabiting contrasting latitudinal areas: M. acerata from Antarctic waters and M. laevis from the Caribbean. We have compared their brooded embryos and larvae using scanning and transmission electron microscopy, and calculated their energetic signatures based on the nature of their yolk. While the general morphological feature of embryos and larvae of both species were very similar, the main difference resided in the yolk nature. The Antarctic species, M. acerata, showed exclusively lipid yolk, whereas the Caribbean species, M. laevis, showed combined platelets of lipids and proteins and less frequently protein yolk platelets. The larvae of M. acerata were estimated to possess a two-fold energetic signature compared to that of M. laevis, which may have important ecological implications for their survival and for maintaining large population densities in the cold waters of the Southern Ocean.     

Comparative morphology of pond,stream and phytotelm‐dwelling tadpoles of the South American Redbelly Toads (Anura: Bufonidae: Melanophryniscus)

We present a comprehensive review of larval morphology in the Neotropical toad genus Melanophryniscus. The taxa studied included 23 species with representatives of recognized phenetic groups and different larval ecomorphological guilds: pond, stream, and phytotelm‐dwelling tadpoles. Their external morphology variation is congruent with current phenetic arrangement based on adult features, but also reflects the habitat where larvae develop. Lotic tadpoles (i.e. M. tumifrons group and M. krauczuki) in general exhibit a more depressed body, a longer tail with lower fins, and larger oral discs than lentic forms (i.e. M. stelzneri group, M. moreirae, M. sanmartini, and M. langonei). Despite their peculiar, confined microhabitat, phytotelm larvae do not diverge markedly from non‐arboreal species. The distinctive features of all species are the presence of a pineal end organ and the placement of the intestinal reversal point at the left of the abdomen in typical larval stages. The buccal cavity and musculoskeletal anatomy are quite conserved between species, yet some characteristics differ from those of other bufonids. The presence of one pair of subhyoid muscles is apparently an exclusive trait of Melanophryniscus among Bufonidae. © 2014 The Linnean Society of London, Biological Journal of the Linnean Society, 2014, 112 , 417–441.     

The bioenergetic fuel for non-feeding larval development in an endemic palaemonid shrimp from the Iberian Peninsula,Palaemonetes zariquieyi

Palaemonetes zariquieyi, an endemic palaemonid species of shrimp that lives in freshwater and brackish coastal habitats in eastern Spain, shows an abbreviated, non-feeding larval development comprising only three zoeal stages. To identify the endogenous bioenergetic fuel that allows for food-independent development from hatching to metamorphosis, larvae were reared under controlled laboratory conditions, and ontogenetic changes in dry weight (W), elemental (CHN), and lipid composition (total lipids, principal lipid classes, and fatty acids [FA]) were quantified at the onset of each zoeal stage and in the first juvenile. Values of W, C, and H per larva and per mass unit of W decreased throughout the time of larval development, while the N content showed only a weak decline (suggesting strong lipid but only little protein degradation). Correspondingly, directly measured values of total lipids (both in μg/larva and in % of W) decreased gradually, with neutral lipids (NL) consistently remaining the predominant and most strongly used fraction; sterol esters and waxes were not detected. In contrast to the NL, the fraction of polar lipids (PL) per larva remained stable and, as a consequence, tended to increase as a percentage of total lipids. Likewise, other important lipid fractions such as free FA and cholesterol remained stable throughout the time of larval development. Among the FA, palmitic (16:0), oleic (18:1n–9), linoleic (18:2n–6), and eicosapentaenoic (20:5n–3) acid were predominant, showing a significant decrease during larval development; stearic (18:0), vaccenic (18:1n–7), and arachidonic acid (20:4n–6) were found only in small amounts. Our results indicate that the lecithotrophic development of P. zariquieyi is primarily fuelled by the utilization of lipids (especially triacylglycerides and other NL), which is reflected by a decreasing carbon content. Proteins and PL, by contrast, are preserved as structurally indispensable components (nerve and muscle tissues, cell membranes). The abbreviated and non-feeding mode of larval development of P. zariquieyi may have an adaptive value in land-locked freshwater habitats, where planktonic food limitation is likely to occur. The patterns of reserve utilization are similar to those previously observed in other palaemonid shrimps and various other groups of decapod crustaceans with lecithotrophic larvae. This suggests a multiple convergent evolution of bioenergetic traits allowing for reproduction in food-limited aquatic environments.