The organization of the nervous system in Plathelminthes. The neuropeptide F-immunoreactive pattern in Catenulida,Macrostomida, Proseriata

The organization of the nervous system of Archilopsis unipunctata Promonotus schultzei and Paramonotus hamatus (Monocelididae, Proseriata) and Stenostomum leucops (Catenulida) and Microstomum lineare (Macrostomida) was studied by immunocytochemistry, using antibodies to the authentic flatworm neuropeptide F (NPF) (Moniezia expansa). The organization of the nervous system of the Monocelididae was compared to that of the nervous system of Bothriomolus balticus (Otoplanidae), a previously studied species of another family of the Proseriata. The results show that the main nerve cords (MCs), independent of lateral or ventral position in the Monocelididae and the Otoplanidae, correspond to each other. The study also confirms the status of the lateral cords as main cords (MCs) in S. leucops and M. lineare. Common for MCs in the members of the investigated taxa are the following features: MCs consist of many fibres, originate from the brain and are adjoined to 5-HT-positive neurons. In Monocelididae and Otoplanidae, the MCs additionally have the same type of contact to the pharyngeal nervous system. Also common for both proseriate families is the organization of the two lateral nerve cords, with weaker connections to the brain, and the pair of dorsal cords running above the brain. The organization of the minor cords differs. The Monocelididae have a pair of thin ventral cords forming a mirror image of the dorsal pair. Furthermore, an unpaired ventral medial cord connecting medial commissural cells was observed in P. schultzei. Marginal nerve cords, observed in Otoplanidae, are absent in Monocelididae. All minor nerve cords are closely connected to the peripheral nerve plexus. The postulated trends of condensation of plexal fibres to cords and/or the flexibility of the peripheral nerve plexus are discussed. In addition, the immunoreactivity (IR) pattern of NPF was compared to the IR patterns of the neuropeptide RFamide and the indoleamine, 5-HT (serotonin). Significant differences between the distribution of IR to NPF and to 5-HT occur. 5-HT-IR dominates in the submuscular and subepidermal plexuses. In the stomatogastric plexus of M. lineare, only peptidergic IR is observed in the intestinal nerve net. The distribution of NPF-IR in fibres and cells of the intestinal wall in M. lineare indicates a regulatory function for this peptide in the gut, while a relationship with ciliary and muscular locomotion is suggested for the 5-HT-IR occurring in the subepidermal and submuscular nerve, plexuses. In M. lineare, the study revealed an NPF- and RFamide-positive cell pair, marking the finished development of new zooids. This finding indicates that constancy of these cells is maintained in this asexually reproducing and regenerating species.     

Glyoxylic acid induced fluorescence in the nervous system of Gyratrix hermaphroditus (Kalyptorhynchia,Polycystididae)

Catecholamines (CAs) are found in the neuropile of the brain, in 3 pairs of longitudinal nerve cords, in the transverse ventral commissure, in anterior ventral and dorsal nerves, in two pharyngeal nerve rings and in 24 neurons in the nervous system of Gyratrix hermaphroditus. The CA distribution pattern in compared with those of other neuroactive substances. Homology of neurons in the family of Polycystididae and in Plathelminthes in general is discussed.     

The nervous system of <Emphasis Type="Italic">Convolutriloba</Emphasis> (Acoela) and its patterning during regeneration after asexual reproduction

We describe the serotonergic and cholinergic nervous system of the asexually reproducing acoel Convolutriloba longifissura Bartolomaeus & Balzer, 1997 by means of immunohistochemistry, conventional histochemistry and transmission electron microscopy. Immunocytochemical staining for serotonin revealed neurons in the brain, in a pair of ventral main longitudinal cords, in two pairs of smaller dorsal longitudinal nerve cords, and in a submuscular nerve net. The brain comprises a ventral-anterior commissure and a less intensely stained dorsal commissure joined together by connectives into a three-ringed scaffold from which the longitudinal nerves extend. We followed the regeneration of the serotonergic part of the nervous system up to the second day after fission. Within this time period, the offspring reestablished bilateral symmetry in the nervous system and developed full motor control. The presence of aminergic cell bodies associated with the main lateral nerve cords of C. longifissura shows that the acoelan nervous system is more similar to that of other platyhelminths (triclads, rhabditophorans) than previously assumed. The presence of serotonergic cell bodies along the main nerve cord correlates with the capacity for asexual reproduction via fissioning. We also describe the single fission mode of C. hastifera Winsor 1990, which brings the modes of asexual reproduction employed by members of the Convolutrilobinae to three.     

Embryonic development of the alimentary canal and ectodermal derivatives in the primitive moth,Neomicropteryx nipponensis issiki (Lepidoptera,Micropterygidae)

The formation of the alimentary canal, nervous system, and of other ectodermal derivatives in the embryo of the primitive moth, Neomicropteryx nipponensis Issiki, is described. The stomodaeum is formed from an invagination in the medioposterior portion of the protocephalon. The proctodaeum arises as an extension of the amnioproctodaeal cavity. The midgut epithelium orginates from anterior and posterior rudiments in blind ends of the stomodaeum and proctodaeum. The decondary dorsal organ is formed in developing midgut. The development of the brain is typical of insects. The ventral nerve cord originates in large part from neuroblasts arising in 3 gnathal, 3 thoracic, and 11 abdominal segments. Intrasegmental median cord cells probably differentiate into both ganglion cells and glial elements of the ventral nerve cord; intersegmental cells appear not to participate in the formation of the nervous system. The stomatogastric nervous system develops from three evaginations in the dorsal wall of the stomodaeum, and consists of the frontal, hypocerebral, and ventricular ganglia, the recurrent nerve, and corpora cardiaca. Five stemmata arise from the epidermis on each side of the head. Five pairs of ectodermal invaginations are formed in the cephalognathal region to produce the tentorium, mandibular apodemes, corpora allata, and silk glands. Prothoracic glands orginate in the prothorax. Mesothoracic spiracles shift anteriorly to the prothorax during development. Oenocytes arise in the first seven abdominal segments. Invaginated pleuropodia are formed in the first abdominal segment.     

Comparative analysis of the nervous systems in presumptive progenetic dinophilid and dorvilleid polychaetes (Annelida) by immunohistochemistry and cLSM

Entire nervous systems of the dinophilids Dinophilus (two species) and Trilobodrilus (three species) and the dorvilleids Parapodrilus psammophilus and Ophryotrocha gracilis (larva) were stained with antisera directed against serotonin, Phe‐Met‐Arg‐Phe‐NH₂ (FMRFamide) and acetylated α‐tubulin and analysed by confocal laser scanning microscopy (cLSM). Adult dinophilids and the dorvilleid larva exhibit the same structure of the ventral nerve cord, with two main nerves spaced far apart, one median and two paramedian nerves. A serotonergic plexus is situated between the paramedian nerve pair, above the ventral locomotory ciliary band. These similarities between adults and larva corroborate the presumed progenetic origin of dinophilids. However, since larval nervous systems of other polychaete taxa also seem to be organized in this way, this result cannot support the view that dinophilids originate from dorvilleids. In P.psammophilus the main nerve cords are widely separated only in the last segment, indicating that this pattern may be correlated with the absence of parapodia. The unpaired median nerve of dinophilids, P. psammophilus and many other polychaetes, is considered to be part of the basic annelid body plan. The ground pattern of the ventral paired dinophilid ganglia is represented by three (anterior, main, posterior) commissures, conserved in most of the ganglia in the Dinophilus species and mostly reduced to a main commissure in the Trilobodrilus species. The dinophilid species and P. psammophilus possess six pairs of ganglia indicating six trunk segments – in contrast to former views. The two rings behind the prostomium in both the dinophilid and the dorvilleid species contain one pair of ganglia only, corroborating the presumed homology of this peristomial region in the two taxa. The Dinophilus nervous system with 12 longitudinal nerves and three perpendicular nerve rings per segment resembles orthogonal nervous structures characteristic of platyhelminths.     

Muscle fibers in the central nervous system of nemerteans: Spatial organization and functional role

The system of muscle fibers associated with the brain and lateral nerve cords is present in all major groups of enoplan nemerteans. Unfortunately, very little is known about the functional role and spatial arrangement of these muscles of the central nervous system. This article examines the architecture of the musculature of the central nervous system in two species of monostiliferous nemerteans (Emplectonema gracile and Tetrastemma cf. candidum) using phalloidin staining and confocal microscopy. The article also briefly discusses the body‐wall musculature and the muscles of the cephalic region. In both species, the lateral nerve cords possess two pairs of cardinal muscles that run the length of the nerve cords and pass through the ventral cerebral ganglia. A system of peripheral muscles forms a meshwork around the lateral nerve cords in E. gracile. The actin‐rich processes that ramify within the nerve cords in E. gracile (transverse fibers) might represent a separate population of glia‐like cells or sarcoplasmic projections of the peripheral muscles of the central nervous system. The lateral nerve cords in T. cf. candidum lack peripheral muscles but have muscles similar in their position and orientation to the transverse fibers. The musculature of the central nervous system is hypothesized to function as a support system for the lateral nerve cords and brain, preventing rupturing and herniation of the nervous tissue during locomotion. The occurrence of muscles of the central nervous system in nemerteans and other groups and their possible relevance in taxonomy are discussed. J. Morphol. 2012. © 2012 Wiley Periodicals, Inc.     

Developmental origin of the adult nervous system in a holothurian: an attempt to unravel the enigma of neurogenesis in echinoderms

In adult echinoderms, the nervous system includes the ectoneural and hyponeural subsystems. The former has been believed to develop from the ectoderm, whereas the latter is considered to be mesodermal in origin. However, this view has not been substantially supported by embryological examinations. Our study deals with the developmental origin of the nervous system in the direct-developing sea cucumber Eupentacta fraudatrix. The rudiment of the adult nervous system develops from ectodermally derived cells, which ingress into the primary body cavity from the floor of the vestibule. At the earliest stages, only the rudiment of the ectoneural nerve ring is laid down. The radial nerve cords and tentacular nerves grow out from this subcutaneous rudiment. The ectoneural cords do not develop simultaneously but make their appearance in the following order: unpaired mid-ventral cord, paired dorsal lateral cords, and ventral lateral cords. These transitional developmental stages probably recapitulate the evolution of the echinoderm body plan. The holothurian hyponeural subsystem, as other regions of the metazoan nervous system, has an ectodermal origin. It originally appears as a narrow band of tissue, which bulges out of the basal region of the ectoneural neuroepithelium. Our data combined with those of other workers strongly suggest that the adult nervous tissue in echinoderms develops separately from the superficial larval system of ciliary nerves. Therefore, our data are neither in strict accordance with Garstang's hypothesis nor do they allow to refuse it. Nevertheless, in addition to ciliary bands, other areas of neurogenetic epidermis must be taken into account.     

Ultrastructural and immunocytochemical observations of the nervous systems of three macrodasyidan gastrotrichs

The nervous systems of three macrodasyidan gastrotrichs, Dactylopodola baltica, Macrodasys caudatus and Dolichodasys elongatus, were investigated using immunocytochemistry and electron microscopy. Labelling of neural structures against serotonin revealed the presence of two pairs of cerebral cells, a dorsal cerebral connective, and paired ventral nerve cords in D. baltica. In M. caudatus and D. elongatus serotonin immunoreactivity was present in a single pair of dorsal cerebral cells and the ventral nerve cords; the dorsal connective of D. elongatus was also immunoreactive to serotonin and acetylated α‐tubulin. The presence of paired, serotonin‐like immunoreactive cells in D. baltica and other species may represent the plesiomorphic condition in Macrodasyida. The fine structure of the photoreceptors in D. baltica was also investigated to explore the potential ground pattern for eyes in the Macrodasyida. The pigmented photoreceptors of D. baltica contain a unicellular pigment cup, sheath cell and sensory receptor. The pigment cup contains numerous osmiophilic granules that presumably function to shield the eyes from downwelling light in the red part of the spectrum. Projecting into the pigment cup and sheath cell are numerous microvilli from a bipolar sensory cell. A single sensory cell may represent the plesiomorphic condition in Macrodasyida, with multiplication of sensory cells representative of more derived taxa.     

The nervous system of the tube-worm Chaetopterus variopedatus (Polychaeta)

Silver impregnation of serial histological sections of the tubeworm Chaetopterus variopedatus revealed the presence of a subepidermal nervous system. The anterior nervous system is delimited by the first 11 segments and comprises (1) two dorsolateral cerebral ganglia and lateral instead of ventral nerve cords which are widely separated and thus connected by unusually long commissures, (2) a pharyngeal ganglion in the fourth segment which is connected to the cerebral ganglia by pharyngeal nerves and constitutes along with the pharyngeal plexus a stomatogastric or enteric nervous system, and (3) small, presumably segmental ganglionic swellings along the lateral nerve cords from which emerge commissures and parapodial nerves. No subesophageal ganglion or periesophageal connective could be identified. The lateral nerve cords converge toward the midline in the 12th segment to form the posterior nervous system comprising a pair of ventromedian nerve cords with their repetitive segmental ganglia from which emerge numerous short commissures and three segmental nerves coursing toward the dorsal and ventral regions of parapods and toward the neuropod. Light and electron microscopic investigations of cerebral and segmental ganglia showed an arrangement of inner neuropile and of unipolar neuron somata at the periphery. The neuropile comprises numerous neurites ranging in diameter from 0.5 to 10 μm and making polarized or symmetrical synaptic junctions with each other. The pharyngeal ganglion consists of a similar neuropile and of a large mass of cell bodies which is traversed by an elaborate network of sinuses and harbors three types of neurosecretory cells in addition to the conventional neuron somata. These findings are interpreted in the framework of the highly specialized morphological features and habits of Chaetopterus, and the welldeveloped stomatogastric system is considered to be related to control of the feeding activities.     

Anatomy of the sensory nervous system in Craspedella pedum (Plathelminthes, Temnocephalida): DiO staining after fixation and in vivo

 The microanatomy of the nervous system of Craspedella pedum is described based on staining of the sensory nerves with DiO (3,3′-dioctadecyloxacarbocyanine perchlorate) in whole worms after fixation. The high resolution and reproducibility of this method revealed that the microanatomy of the nervous system is uniform within this species to rather minute details. Although the lateral and ventral cords have a similar diameter, the ventral nerve cords are very poor in sensory fibres. Such a high level of functional differentiation of nerve cords has not been known in representatives of the Plathelminthes. In vivo staining reveals numerous DiO-accumulating sensory neurons in the anterior portion of the body and a few pairs in its posterior half. The morphology of some neurons is described. Several neurons were identified and our data suggest that most, if not all, sensory neurons are identificable cells. Accepted: 16 December 1997     

Organization of the larval pre-ecdysis motor pattern in the tobacco hornworm,Manduca sexta

The tobacco hornworm, Manduca sexta, undergoes several larval molts before transforming into a pupa and then an adult moth. Each molt culminates in ecdysis, when the old cuticle is shed. Prior to each larval ecdysis, the old cuticle is loosened by pre-ecdysis behavior, which consists of rhythmic compressions that are synchronous along the abdomen and on both body sides, and rhythmic retractions of the abdominal prolegs. Both pre-ecdysis and ecdysis behaviors are triggered by a peptide, eclosion hormone. The aim of the present study was to investigate the neural circuitry underlying larval preecdysis behavior. The pre-ecdysis motor pattern was recorded in isolated nerve cords from eclosion hormone-treated larvae, and the effects of connective transections and ionic manipulations were tested. Our results suggest that the larval pre-ecdysis compression motor pattern is coordinated and maintained by interneurons in the terminal abdominal ganglion that ascend the nerve cord without chemical synaptic relays; these interneurons make bilateral, probably monosynaptic, excitatory connections with identified pre-ecdysis motor neurons throughout the abdominal nerve cord. This model of the organization of the larval pre-ecdysis motor pattern should facilitate identification of the relevant interneurons, allowing future investigation of the neural basis of the developmental weakening of the pre-ecdysis motor pattern that accompanies the larval-pupal transformation.Abbreviations A3, A4... abdominal ganglia 3, 4... - AT terminal abdominal ganglion - ALE anterior lateral external muscle - DN dorsal nerve - DN_A anterior branch of the dorsal nerve - DN_L lateral branch of the dorsal nerve - DN_P posterior branch of the dorsal nerve - EH eclosion hormone - TP tergopleural muscle - VN ventral nerve - VN_A anterior branch of the ventral nerve - VN_L lateral branch of the ventral nerve - VN_P posterior branch of the ventral nerve     

Localisation of GYIRFamide immunoreactivity inMacrostomum hystricinum marinum (Plathelminthes,Macrostomida)

The distribution of GYIRFamide immunoreactivity in the nervous system ofMacrostomum hystricinum marinum has been demonstrated by an indirect fluorescence technique in conjunction with confocal scanning laser microscopy (CSLM). Immunostaining was extensive in both the central (CNS) and peripheral (PNS) nervous systems, revealing detailed information on the microanatomy of the peptidergic nervous system of this free-living plathelminth. In the CNS, immunoreactive nerve cell bodies and nerve fibres occurred in the brain and along two pairs of longitudinal nerve cords: the main nerve cords and the ventral nerve cords. In the PNS, immunostaining was prevalent in nerve cells and fibres innervating the pharynx and the gut. The employed antibody is directed against a recently characterised FMRF-amide-related peptide (FaRP), GYIRFamide, isolated from two species of the Tricladida,Dugesia tigrina andBdelloura candida. Phylogenetically, GYIRFamide represents the most ancient neuropeptide thus far identified within the Bilateria     

Absence of sensory input does not affect persistent neurons in Tenebrio molitor metamorphosis (Insecta: Coleoptera)

Summary The ventral nerve cord of holometabolous insects is reorganized during metamorphosis. Certain elements, however, persist from the larval to the adult stage. A group of dorsal unpaired median neurons and a set of thoracic serotonin-immunoreactive interneurons are shown to be persistent elements in the ventral nerve cord of Tenebrio molitor. These persistent neurons retain their bauplan even after deprivation of the sensory neurons projecting onto them. These persistent neurons might provide a network into which newly developing sensory neurons fit during reorganization of the nervous tissue.     

Architecture of the nervous system in two Dactylopodola species (Gastrotricha, Macrodasyida)

Immunohistochemical stainings have become standard tools to describe the nervous system, but usually only singular or few markers are used and consequently show only subsets of neurons within the nervous system. We investigated two species of Dactylopodola (Gastrotricha, Macrodasyida) with a broad set and combination of markers, to represent the nervous system in a more holistic approach. We suggest that markers for both neurotubuli (tubulin) and neurotransmitters (e.g. serotonin, FMRF-amides, histamine) should be used. Combinations with markers for the musculature (phalloidin) and nuclei (propidiumiodide or other markers) help to reveal spatial patterns and when used with TEM can provide a more precise picture of the spatial relationships of particular nerves. Species of Dactylopodola have a brain consisting of a solid dorsal commissure and a fine ventral commissure. Cell somata of brain cells are arranged lateral to the dorsal commissure and form a dumbbell-like brain. Additionally, projections into the head region, head sensory organs, one pair of lateroventral nerve cords with three commissures and stomatogastric nerves are described. Obviously, some longitudinal transmitter-specific fibres run in parallel to the main longitudinal nerve and represent additional longitudinal fibres. In comparison with the nervous system architecture of other gastrotrich species and that of different bilaterian animals it is speculated that the gastrotrich nervous system retains several ancestral features, such as being commissural and not a compact brain.     

Morphogenesis defects are associated with abnormal nervous system regeneration following roboA RNAi in planarians

The process by which the proper pattern is restored to newly formed tissues during metazoan regeneration remains an open question. Here, we provide evidence that the nervous system plays a role in regulating morphogenesis during anterior regeneration in the planarian Schmidtea mediterranea. RNA interference (RNAi) knockdown of a planarian ortholog of the axon-guidance receptor roundabout (robo) leads to unexpected phenotypes during anterior regeneration, including the development of a supernumerary pharynx (the feeding organ of the animal) and the production of ectopic, dorsal outgrowths with cephalic identity. We show that Smed-roboA RNAi knockdown disrupts nervous system structure during cephalic regeneration: the newly regenerated brain and ventral nerve cords do not re-establish proper connections. These neural defects precede, and are correlated with, the development of ectopic structures. We propose that, in the absence of proper connectivity between the cephalic ganglia and the ventral nerve cords, neurally derived signals promote the differentiation of pharyngeal and cephalic structures. Together with previous studies on regeneration in annelids and amphibians, these results suggest a conserved role of the nervous system in pattern formation during blastema-based regeneration.     

The nervous system of Tricladida. I. Neuroanatomy ofProcerodes littoralis (Maricola,Procerodidae): An immunocytochemical study

The organization of the nervous system ofProcerodes littoralis (Tricladida, Maricola, Procerodidae) was studied by immunocytochemistry, using antibodies to authentic flatworm neuropeptide F (NPF) (Moniezia expansa). Compared to earlier investigations of the neuroanatomy of tricladid flatworms, the pattern of NPF immunoreactivity inProcerodes littoralis reveals differences in the following respects: 1. Shape and structure of the brain. 2. Number and composition of longitudinal nerve cords. 3. Shape of branches of, and transverse connections between, main ventral nerve cords. 4. Composition of the pharyngeal nervous system. The rich innervation by NPF immunoreactive (IR) fibres and cells of the subepithelial muscle layer, the pharynx musculature and the musculature of the male copulatory apparatus indicates a neurotransmitter or neuromodulatory influence on muscular activity.     

Immunohistochemical and ultrastructural analysis of the muscular and nervous systems in the interstitial polychaete Polygordius appendiculatus (Annelida)

The systematic position of Polygordiidae is still under debate. They have been assigned to various positions among the polychaetes. Recent molecular analyses indicate that they might well be part of a basal radiation in Annelida, suggesting that certain morphological characters could represent primitive character traits adopted from the annelid stem species. To test this hypothesis, an investigation of the muscular and nervous systems by means of immunological staining and confocal laser scanning microscopy and transmission electron microscopy was conducted. With the exception of the brain, the nervous system is entirely basiepidermal and consists of the brain, the esophageal connectives, the subesophageal region, the ventral nerve cord and several smaller longitudinal nerves. These are connected by a considerable number of ring nerves in each segment. The ventral nerve cord is made up of closely apposed longitudinal neurite bundles, a median and two larger lateral ones. Since distinct ganglia are lacking, it represents a medullary cord. The muscular system mainly consists of longitudinal fibers, regularly distributed oblique muscles and strong septa. The longitudinal fibers form a right and a left unit separated along the dorsal midline, each divided into a dorsal and ventral part by the oblique muscles. Anteriorly, the longitudinal musculature passes the brain and terminates in the prostomium. There is no musculature in the palps. In contrast to earlier observations, regularly arranged minute circular muscle fibers are present. Very likely, a basiepithelial and non-ganglionic organization of the ventral nerve cord as well as an orthogonal nervous system represent plesiomorphic characters. The same applies for the predominance of longitudinal muscle fibers.     

Nerve development, growth and differentiation during regeneration in Enchytraeus fragmentosus and Stylaria lacustris (Oligochaeta)

Enchytraeus fragmentosus (Enchytraeidae) and Stylaria lacustris (Naididae) are small terrestrial and limnetic oligochaetes that exclusively or seasonally reproduce by fragmentation and regeneration, respectively. We traced the neuronal development and differentiation during regeneration in order to gain information on the basic organization and evolution of the oligochaete nervous system. Subsequent to artificial amputation, the nervous systems have been stained with antibodies directed against acetylated alpha-tubulin. The staining was analyzed by indirect fluorescence in combination with confocal laser scanning microscopy. Both species show unique oligochaete neuronal regeneration patterns: (i) numerous fibers branch off from segmental nerves near the wound site and innervate the blastema; and (ii) the ventral cord is partly reestablished before the circumesophageal connectives develop. In the investigated 'Oligochaeta' the outgrowing fibers of the ventral nerve cord are soon bundled into at least two distinct connective pairs, which prolong into dorsal and ventral roots next to the mouth. Subsequent complete fusion of the doubled roots forms simple connectives. Thus, dorsal roots are not a unique feature for 'Polychaeta'. They occur as a transient structure in 'Oligochaeta' and might be part of the neuronal ground pattern of Annelida. The initially tetra or even pentaneuronal ventral nerve cord also differentiates into an unineuronal one by fusion.