The invention of the pilidium larva in an otherwise perfectly good spiralian phylum Nemertea

One of the most remarkable larval types among spiralians, and invertebrates in general, is the planktotrophic pilidium. The pilidium is found in a single clade of nemerteans, called the Pilidiophora, and appears to be an innovation of this group. All other nemerteans have either planktotrophic or lecithotrophic juvenile-like planuliform larvae or have direct development. The invention of the pilidium larva is associated with the formation of an extensive blastocoel that supports the delicate larval frame and elaborate ciliary band. Perhaps the most striking characteristic of the pilidium is the way the juvenile worm develops inside the larva from a series of isolated rudiments, called the imaginal discs. The paired cephalic discs, cerebral organ discs, and trunk discs originate as invaginations of larval epidermis and subsequently grow and fuse around the larval gut to form the juvenile. The fully formed juvenile ruptures the larval body and, more often than not, devours the larva during catastrophic metamorphosis. This review is an attempt to examine the pilidium in the context of recent data on development of non-pilidiophoran nemerteans, and speculate about the evolution of pilidial larval development. The author emphasizes the difference between the planuliform larvae of Palaeonemerteans and Hoplonemerteans, and suggest a new name for the hoplonemertean larvae--the decidula.     

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.     

Vestigial prototroch in a basal nemertean, Carinoma tremaphoros (Nemertea; Palaeonemertea)

Nemerteans have been alleged to belong to a protostome clade called the Trochozoa that includes mollusks, annelids, sipunculids, echiurids, and kamptozoans and is characterized by, among other things, the trochophore larva. The trochophore possesses a prototroch, a preoral belt of specialized ciliary cells, derived from the trochoblast cells. Nemertea is the only trochozoan phylum for which presence of the trochophore larva possessing a prototroch had never been shown. However, so little is known about nemertean larval development that comparing it with development of other trochozoans is difficult. Development in the nemertean clade Pilidiophora is via a highly specialized planktonic larva, the pilidium, and most of the larval body is lost during a drastic metamorphosis. Other nemerteans (hoplonemerteans and palaeonemerteans) lack a pilidium, and their development is direct, forming either an encapsulated or planktonic "planuliform" larva, producing a juvenile without a dramatic change in body plan. We show that early in the development of a member of a basal nemertean assemblage, the palaeonemertean Carinoma tremaphoros, large squamous cells cover the entire larval surface except for the apical and posterior regions. Although apical and posterior cells continue to divide, the large surface cells cleavage arrest and form a contorted preoral belt. Based on its position, cell lineage, and fate, we suggest that this belt corresponds to the prototroch of other trochozoans. Lack of differential ciliation obscures the presence of the prototroch in Carinoma, but differentiation of the trochoblasts is clearly manifested in their permanent cleavage arrest and ultimate degenerative fate. Our results allow a meaningful comparison between the development of nemerteans and other trochozoans. We review previous hypotheses of the evolution of nemertean development and suggest that a trochophore-like larva is plesiomorphic for nemerteans while a pilidium type of development with drastic metamorphosis is derived.     

Expression profiling of Drosophila imaginal discs

Background  

In the Drosophila larva, imaginal discs are programmed to produce adult structures at metamorphosis. Although their fate is precisely determined, these organs remain largely undifferentiated in the larva. To identify genes that establish and express the different states of determination in discs and larval tissues, we used DNA microarrays to analyze mRNAs isolated from single imaginal discs.     

Loss and gain of the juvenile rudiment and metamorphic competence during starvation and feeding of bryozoan larvae

SUMMARY In many animals, larval structures and juvenile rudiments develop independently. One advantage of this independence is that juvenile rudiments can be expended as a nutrient reserve or for energy conservation. When bryozoan cyphonautes larvae were starved, structures required for settlement and metamorphosis shrank. When the larvae were again fed, these structures grew back. Starvation reduced the size of both the internal sac, a rudiment of postlarval juvenile structures, and the pyriform organ, which functions in sensing and crawling on the substratum at settlement. In contrast, starvation affected neither the size of the larval shell nor the lengths of the ciliary bands used in swimming and feeding. Starved larvae that had reduced the pyriform organ and internal sac did not metamorphose in response to stimuli from a laminarian alga. The laminarian alga did stimulate metamorphosis of the same larvae after renewed feeding, when the larvae had regrown these structures. Thus starved larvae expended body parts needed for settlement and metamorphosis when food was scarce while retaining structures for feeding, swimming, and defense. Starved larvae thereby retained the capacity to regrow structures needed for settlement and metamorphosis when they again encountered food. Advantages from expendable juvenile rudiments may enhance selection for their being developmentally distinct from structures for larval swimming and feeding.     

Invasive cell behavior during Drosophila imaginal disc eversion is mediated by the JNK signaling cascade

Drosophila imaginal discs are monolayered epithelial invaginations that grow during larval stages and evert at metamorphosis to assemble the adult exoskeleton. They consist of columnar cells, forming the imaginal epithelium, as well as squamous cells, which constitute the peripodial epithelium and stalk (PS). Here, we uncover a new morphogenetic/cellular mechanism for disc eversion. We show that imaginal discs evert by apposing their peripodial side to the larval epidermis and through the invasion of the larval epidermis by PS cells, which undergo a pseudo-epithelial-mesenchymal transition (PEMT). As a consequence, the PS/larval bilayer is perforated and the imaginal epithelia protrude, a process reminiscent of other developmental events, such as epithelial perforation in chordates. When eversion is completed, PS cells localize to the leading front, heading disc expansion. We found that the JNK pathway is necessary for PS/larval cells apposition, the PEMT, and the motile activity of leading front cells.     

The peculiar nemertean larva pilidium recurvatum belongs to Riserius sp., a basal heteronemertean that eats Carcinonemertes errans,a hoplonemertean parasite of Dungeness crab

A typical nemertean pilidium larva resembles a hat with ear flaps. But one type, called pilidium recurvatum, looks more like a sock, swimming heel first. This distinctive larva was discovered in 1883 off the coast of Rhode Island and subsequently found in plankton samples from other parts of the world. Despite the long time since discovery, and its significance in discussions of larval evolution, this larva remained unidentified even to the family level. We collected pilidium recurvatum larvae from plankton samples in Coos Bay, OR, and identified them as belonging to the heteronemertean genus Riserius based on juvenile morphology and DNA sequence data. Phylogenetic analysis suggests that two distinct types of pilidium recurvatum from Oregon represent two new species within this currently monotypic genus. We describe the morphology of pilidium recurvatum using confocal microscopy and compare it to that of the typical pilidium, discussing possible implications for larval feeding. We also report our surprising discovery that juveniles of Riserius sp. from Oregon prey on another nemertean, Carcinonemertes errans, an egg predator of Cancer magister (Dungeness crab), a commercially important species. We speculate that the species‐level diversity and geographic distribution of Riserius may be much greater than currently appreciated.     

What are and what are not imaginal discs: reevaluation of some basic concepts (Insecta, Holometabola).

Some general aspects of the concept of imaginal discs in the Holometabola are reevaluated. Their monolayer character and continuity with the surrounding epidermis are confirmed. Studies on the imaginal discs of the silkworm (Bombyx mori) and data from the literature show that the discs and their peripodial cells produce cuticle during larval life, as well as at metamorphosis. In B. mori it is demonstrated that adult and larval antennae are produced by the same cells or their progeny. The results also suggest that segments of the typically three-segmented larval antenna of Holometabola are not scape, pedicel, and one-segmented flagellum; at least segments 2 and 3 are of flagellar origin. Based on these and some additional facts it is argued that: (1) No larval organs are "replaced" at metamorphosis, but strict "sequential homology" is always maintained. (2) Imaginal discs are not undifferentiated structures destined to form the adult after larval breakdown, cannot be unambiguously defined, and do not represent qualitatively different epidermal structures. Classical imaginal discs (invaginated and present also in pre-final larval instars) arose several times independently and were not present in the larvae of ancestral Holometabola. (3) Since the disc cells are not undifferentiated and "embryonic" (if these words have a defined meaning at all), it is unreasonable to expect that the processes taking place in discs at metamorphosis would differ fundamentally from those occurring in other diploid metamorphosing epidermal cells.     

Development Capacities of Mixed Normal and Neoplastic 1(3)gl Imaginal Discs and Neuroblast Tissue in Drosophila hydei

The pleiotropic mutant lethal(3)giant larvae [l(3)gl] of Drosophila hydei exhibits among other anatomical defects, hypertrophy of the larval brain and imaginal discs. Both hypertrophic tissues when transplanted into wild-type female flies behave as fast growing and lethal neoplasms. Implanted into mature wild-type larvae they fail to metamorphose. When l(3)gl neoplastic brain tissue or imaginal discs were mixed with normal imaginal discs, cultured in vivo in the abdomen of adult females and transplanted into mature wild-type larvae, the following results were obtained. The invasive l(3)gl brain neoplasm, while fatal for adult hosts, had no effect on the metamorphosis of normal imaginal disc tissue. On the other hand, the noninvasive l(3)gl imaginal disc neoplasms when mixed with normal imaginal disc tissue inhibited its development and metamorphosis in the wild-type host. This inhibitory effect was not observed when the tissues were injected as separate implants into the same host.     

Imaginal discs regulate developmental timing in Drosophila melanogaster

The regulation of body size in animals involves mechanisms that terminate growth. In holometabolous insects growth ends at the onset of metamorphosis and is contingent on their reaching a critical size in the final larval instar. Despite the importance of critical size in regulating final body size, the developmental mechanisms regulating critical size are poorly understood. Here we demonstrate that the developing adult organs, called imaginal discs, are a regulator of critical size in larval Drosophila. We show that damage to, or slow growth of, the imaginal discs is sufficient to retard metamorphosis both by increasing critical size and extending the period between attainment of critical size and metamorphosis. Nevertheless, larvae with damaged and slow growing discs metamorphose at the same size as wild-type larvae. In contrast, complete removal of all imaginal tissue has no effect on critical size. These data indicate that both attainment of critical size and the timely onset of metamorphosis are regulated by the imaginal discs in Drosophila, and suggest that the termination of growth is coordinated among growing tissues to ensure that all organs attain a characteristic final size.     

The morphostatic actions of juvenile hormone

The maintenance of "status quo" in larvae by juvenile hormone (JH) involves both the programming of ecdysteroid-dependent synthesis during the molt and the suppression of morphogenetic growth during the intermolt. The latter morphostatic action does not require ecdysteroids, and has been studied in the formation of imaginal discs in Manduca sexta. Preultimate larval instars have both invaginated discs and imaginal primordia, both of which grow isomorphically with the larva. In the last instar, the young discs/primordia initiate the morphogenesis and patterning that results in a mature disc. JH suppresses both the initiation and progression of the signaling that transforms immature discs or primordia into a fully patterned imaginal disc. This transformation normally occurs in the context of the rapid growth of the last larval stage, and nutrient-dependent factors appear to be able to override the JH suppression. The morphostatic action of JH may have been important for the evolution of the larval stage. Studies on embryos of basal, hemimetabolous insects show that their premature exposure to JH can truncate patterning programs and cause precocious tissue maturation, factors essential for organizing a novel larval form. This suppression of embryonic patterning then results in embryonic fields that remain dormant as long as JH is present. These are the primordia that can transform into imaginal discs once JH disappears in preparation for metamorphosis.     

The role of nutrition in creation of the eye imaginal disc and initiation of metamorphosis in Manduca sexta

With the exception of the wing imaginal discs, the imaginal discs of Manduca sexta are not formed until early in the final larval instar. An early step in the development of these late-forming imaginal discs from the imaginal primordia appears to be an irreversible commitment to form pupal cuticle at the next molt. Similar to pupal commitment in other tissues at later stages, activation of broad expression is correlated with pupal commitment in the adult eye primordia. Feeding is required during the final larval instar for activation of broad expression in the eye primordia, and dietary sugar is the specific nutritional cue required. Dietary protein is also necessary during this time to initiate the proliferative program and growth of the eye imaginal disc. Although the hemolymph titer of juvenile hormone normally decreases to low levels early in the final larval instar, eye disc development begins even if the juvenile hormone titer is artificially maintained at high levels. Instead, creation of the late-forming imaginal discs in Manduca appears to be controlled by unidentified endocrine factors whose activation is regulated by the nutritional state of the animal.     

Implantation Serine Proteinases heterodimerize and are critical in hatching and implantation

Background

Metamorphosis is a complex, highly conserved and strictly regulated development process that involves the programmed cell death of obsolete larval organs. Here we show a novel functional role for the aspartic proteinase cathepsin D during insect metamorphosis.

Results

Cathepsin D of the silkworm Bombyx mori (BmCatD) was ecdysone-induced, differentially and spatially expressed in the larval fat body of the final instar and in the larval gut of pupal stage, and its expression led to programmed cell death. Furthermore, BmCatD was highly induced in the fat body of baculovirus-infected B. mori larvae, suggesting that this gene is involved in the induction of metamorphosis of host insects infected with baculovirus. RNA interference (RNAi)-mediated BmCatD knock-down inhibited programmed cell death of the larval fat body, resulting in the arrest of larval-pupal transformation. BmCatD RNAi also inhibited the programmed cell death of larval gut during pupal stage.

Conclusion

Based on these results, we concluded that BmCatD is critically involved in the programmed cell death of the larval fat body and larval gut in silkworm metamorphosis.     

Cellular basis of the dynamic behavior of the imaginal thoracic discs during Drosophila metamorphosis

The eversion, migration, spreading, and fusion of the thoracic imaginal discs during metamorphosis of Drosophila are described using timed whole-mount preparations and several molecular markers. The leading edge of the migrating disc epithelia consists of two groups of cells, stalk cells (S cells) and specialized imaginal cells (I cells), that both express the gene puckered. With this and other markers, opening of the stalk, eversion of the discs, migration of the leading edges, and fusion of the imaginal epithelia can be visualized in detail. Fusion is initiated by S cells that migrate over the larval epithelium and constitute a bridge between two imaginal epithelia. S cells are subsequently lost and imaginal fusion is mediated by the I cells that remain at the site of fusion. The possible cellular basis of this process is discussed. Fusion along the dorsal midline of the notum from the mesothoracic wing discs occurs earlier than that of the prothoracic and metathoracic discs, which remain in a lateral position. For a relatively long period (30 h) the mesothoracic epithelium becomes attached to the head and abdomen, causing a temporary local discontinuity of the order of segments. Later the pro- and metathoracic discs intercalate between head and mesothorax and between abdomen and mesothorax, respectively, to reestablish the normal order.     

Transcriptional signature of accessory cells in the lateral line, using the Tnk1bp1:EGFP transgenic zebrafish line

Background

A metamorphic life-history is present in the majority of animal phyla. This developmental mode is particularly prominent among marine invertebrates with a bentho-planktonic life cycle, where a pelagic larval form transforms into a benthic adult. Metamorphic competence (the stage at which a larva is capable to undergo the metamorphic transformation and settlement) is an important adaptation both ecologically and physiologically. The competence period maintains the larval state until suitable settlement sites are encountered, at which point the larvae settle in response to settlement cues. The mechanistic basis for metamorphosis (the morphogenetic transition from a larva to a juvenile including settlement), i.e. the molecular and cellular processes underlying metamorphosis in marine invertebrate species, is poorly understood. Histamine (HA), a neurotransmitter used for various physiological and developmental functions among animals, has a critical role in sea urchin fertilization and in the induction of metamorphosis. Here we test the premise that HA functions as a developmental modulator of metamorphic competence in the sea urchin Strongylocentrotus purpuratus.

Results

Our results provide strong evidence that HA leads to the acquisition of metamorphic competence in S. purpuratus larvae. Pharmacological analysis of several HA receptor antagonists and an inhibitor of HA synthesis indicates a function of HA in metamorphic competence as well as programmed cell death (PCD) during arm retraction. Furthermore we identified an extensive network of histaminergic neurons in pre-metamorphic and metamorphically competent larvae. Analysis of this network throughout larval development indicates that the maturation of specific neuronal clusters correlates with the acquisition of metamorphic competence. Moreover, histamine receptor antagonist treatment leads to the induction of caspase mediated apoptosis in competent larvae.

Conclusions

We conclude that HA is a modulator of metamorphic competence in S. purpuratus development and hypothesize that HA may have played an important role in the evolution of settlement strategies in echinoids. Our findings provide novel insights into the evolution of HA signalling and its function in one of the most important and widespread life history transitions in the animal kingdom - metamorphosis.     

Larval growth and perimetamorphosis in the echinoid <Emphasis Type="Italic">Echinocardium cordatum</Emphasis> (Echinodermata): the spatangoid way to become a sea urchin

The prejuvenile development of Echinocardium cordatum (Echinoidea) was investigated by means of scanning electron, confocal and light microscopes, aiming to illustrate the early life history of a spatangoid representative and to compare it with the other major echinoid groups. During the larval development of E. cordatum, two periods follow one another. The first one takes 12 days; it ends with the formation of a complete echinopluteus with twelve elongated larval arms. The second lasts from 3 to 12 days; it is entirely devoted to the building of the echinid rudiment and ends with the acquisition of larval competence. No appendage other than arms develops at the larva’s outer surface. Competent larvae are demersal. They settle onto the substratum and test it for suitability using the five rudiment podia that protrude through the vestibule opening. Metamorphosis is a rapid event that lasts less than an hour. The rudiment does not everse and its spines and podia actively tear up the larval epidermis which is progressively covered by the growing vestibular epidermis. The resulting postlarva is short-lived and morphologically similar to both the late rudiment and the early juvenile, which, however, is exotrophic. Late rudiments in E. cordatum show basic spatangoid features being bilaterally symmetric and having clavulae and sphaeridia. More importantly, they already have the convex shape and the appendage cover of early juveniles. Metamorphosis in E. cordatum appears to be less complex, i.e. no rudiment is everted, and more complete, since, in contrast to “regular” echinoids, no transitory appendages are seen. Metamorphosis/development of E. cordatum, thus, is closer to that of clypeasteroids, since the rudiment of the latter already bears juvenile definitive appendages, when everted during metamorphsis.     

Potential for paired vestibules in plutei (Echinodermata, Echinoidea)

Abstract. Echinoids are generally thought to develop only a single ectodermal invagination (V1), which expands into a vestibule and contributes to the formation of the juvenile rudiment on the left side of the larval body. However, in 2 echinoid species, Strongylocentrotus purpuratus and Dendraster excentricus, a second invagination (V2) was observed to form on the right side of the larval body exactly opposite V1. Formation of the invaginations was followed and characterized using light microscopy and antibodies directed against the S2amide echinoderm neuropeptide as a morphological tag. V2 consistently formed later than V1, and appeared capable of contributing to the formation of a second rudiment when it received the appropriate mesodermal influence from contact with a hydrocoel. Otherwise, in both species, V2 formed a skeletal element that has not been previously described in this location. Because of the location, shape, and timing of its formation, V2, like V1, appears to have the innate ability of the larval ectoderm to contribute to juvenile rudiment formation.     

Metamorphosis of imaginal discs of Drosophila melanogaster

Summary Imaginal discs ofDrosophila melanogaster larvae, 24–53 hrs after oviposition, were transplanted into mature immobile larval hosts. The transplants did not respond to the hormonal stimuli of metamorphosis, but instead completed their larval development. When reinjected into mature larval hosts, they now differentiated the full set of their presumptive imaginal structures. The process of acquiring competence for metamorphosis appears to be independent of the hormonal conditions.Supported by a credit of the Swiss National Foundation granted to Prof. Dr. E. Hadorn. I thank Dr. R. Nöthiger for his valuable criticism during this investigation.     

Primordial germ cells originate from the endodermal strand cells in the ascidian Ciona intestinalis

The origin of germ cells in the ascidian is still unknown. Previously, we cloned a vasa homologue (CiVH) of Ciona intestinalis from the cDNA library of ovarian tissue by polymerase chain reaction and showed that its expression was specific to germ cells in adult and juvenile gonads. In the present study, we prepared a monoclonal antibody against CiVH protein and traced the staining for this antibody from the middle tailbud stage to young adulthood. Results showed that positive cells are present in the endodermal strand in middle tailbud embryos and larvae. When the larval tail was absorbed into the trunk during metamorphosis, the CiVH-positive cells migrated from the debris of the tail into the developing gonad rudiment, and appeared to give rise to a primordial germ cell (PGC) in the young juvenile. The testis rudiment separated from the gonad rudiment, the remainder of which differentiated into the ovary. PGCs of the testis rudiment and the ovary rudiment differentiated into spermatogenic and oogenic cells, respectively. When the larval tail containing the antibody-positive cells was removed, the juveniles did not contain any CiVH-positive cells after metamorphosis, indicating that the PGCs in the juvenile originated from part of the larval tail. However, even in such juveniles, positive cells newly appeared in the gonad rudiment at a later stage. This observation suggests that a compensatory mechanism regulates germline formation in C. intestinalis.