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.     

The morphology and anatomy of the vestimentiferan worm <Emphasis Type="Italic">Oasisia alvinae</Emphasis> Jones, 1985 (Annelida: Siboglinidae). II. Integument,nervous system and musculature

The histological structure of the epidermis, nervous system, and musculature of the vestimentiferan worm Oasisia alvinae is examined. The differentiation of the integument of different regions of the body and the histological structure of the pyriform glands and papillae with cuticular plaques are described. All the main elements of the nervous system lie in the thickness of the epidermis. The brain structure is examined and an attempt is made to homologize parts of the vestimentiferan brain with the polychaete brain. Part of the vestimentiferan brain above the intrabrain channel corresponds to the supraesophageal ganglion of polychaetes, while part of the vestimentiferan brain under the channel corresponds to the subesophageal ganglion. Two commissures are located in the brain region that corresponds to the supraesophageal ganglion. These commissures could be related to the ventral and dorsal roots of the circumesophageal connectives of sedentary polychaetes. Bundles of fiber occur, which presumably correspond to the circumesophageal connectives and the transverse commissure inside the subesophageal ganglion. The musculature differentiation in the trunk region is a characteristic feature of O. alvinae, i.e., feather-like muscles that occur only in the anterior part of the trunk.     

Polychaete nervous systems: Ground pattern and variations--cLS microscopy and the importance of novel characteristics in phylogenetic analysis

In Annelida, as well as in other invertebrate taxa, the nervoussystem is considered to be a very conservative organ system.Immunohistochemical investigations [use of anti-5-HT (serotonin),FMRFamide, and acetylated -tubulin antibodies] in combinationwith laser scanning microscopy enable more detailed reanalysesof known structures and detection of new characteristics thatare useful for phylogenetic analyses. One hypothesis enabledby such studies is outlined for the evolution of arrangementsof the dorsal circumesophageal roots in polychaetes and oligochaetes.These roots are not a unique feature of polychaetes; they alsooccur in oligochaetes. According to the Articulata hypothesisof metazoan relationships, the specific structure of the rope-ladder-likenervous system is, among others, an autapomorphic characteristicthat unifies Annelida and Arthropoda. Recent studies applyingthe techniques mentioned here, however, demonstrate that theannelidan bauchmark (central nervous system of the trunk), incontrast to the arthropod pattern, is highly variable in termsof the number and position of connectives and the number ofcommissures per segment. The variability of the neuronal architectureas well as a hypothesis on how it evolved will be introducedwith the aid of regeneration and developmental studies. Furthermore,it is shown that hitherto unknown nerves are present in theperipheral nervous system.     

The formation of the nervous system during larval development in Triops cancriformis (Bosc) (crustacea,Branchiopoda): An immunohistochemical survey

We provide data of the development of thenervous system during the first five larval stages of Triops cancriformis. We use immunohistochemical labeling (against acetylated α‐tubulin, serotonin, histamine, and FMRFamide), confocal laser scanning microscopy analysis, and 3D‐reconstruction. The development of the nervous system corresponds with the general anamorphic development in T. cancriformis. In larval stage I (L I), all brain parts (proto‐, deuto‐, and tritocerebrum), the circumoral connectives, and the mandibular neuromere are already present. Also, the frontal filaments and the developing nauplius eye are already present. However, until stage L III, the nauplius eye only consists of three cups. Throughout larval development, the protocerebral network differentiates into distinct subdivisions. In the postnaupliar region, additional neuromeres and their commissures emerge in an anteroposterior gradient. The larval nervous system in L V consists of a differentiated protocerebrum including a central body, a nauplius eye comprising four cups, a circumoral nerve ring, mandibular‐ and postnaupliar neuromeres up to the seventh thoracic segment, each featuring an anterior and a posterior commissure, and two parallel connectives. The presence of a protocerebral bridge is questionable. The distribution of neurotransmitters in L I is restricted to the naupliar nervous system. Over the course of the five stages of development, neurotransmitter distribution also follows an anteroposterior gradient. Each neuromere is equipped with two ganglia innervating the locomotional appendages and possesses a specific neurotransmitter distribution pattern. We suggest a correlation between neurotransmitter expression and locomotion. J. Morphol., 2010. © 2010 Wiley‐Liss, Inc.     

Regeneration of Identified Neurons and Their Synaptic Connections in the Central Nervous System of Aplysia

SYNOPSIS. In the CNS of Aplysia the number and sequence of lesionsinfluence the regeneration of identified neurons and their synapticconnections The left pleural ganglion giant neuron (LGC) failsto regenerate its severed axon following crushes of the cerebropleural (C-PL) connectses A conditioning lesion paradigm inwhich first a foot nerve and then 10 days later, the left C-PLconnective is crushed, results in the LGC regenerating its axonSynaptic inputs from identified neurons in the cerebral ganglionrarely regenerate when only the C-PL connectives are crushedConditioning lesions also result in the regeneration of synapticconnections from the cerebral A neurons Multiple and sequentiallesions appear to be more effective than single lesions in promotingboth axonal regeneration and synapse formation The possiblemechanisms by which axonal regeneration and synaptogenesis arecontrolled are discussed.     

Axogenesis in the stomatopod crustacean Gonodactylaceus falcatus (Malacostraca)

Abstract. The formation of the central nervous system of the stomatopod crustacean Gonodactylaceus falcatus is described by means of antibody stainings against synapsin and α-tubulin. It is shown that the longitudinal fiber tracts of the ventral nervous system are formed by two centers of origin comprising a number of pioneer neurons, one at the posterior part of the forming brain, the other in the area of the telson anlage at the posteriormost region of the embryo. In addition to the lateral anlagen of the connectives, a median longitudinal nerve is formed beginning in the mandibular segment neuromere. In contrast to those of other segments, the mandibular ganglia are connected by a single commissure. The brain forms a circumoral ring. There is evidence that the deutocerebrum possesses praestomodeal and poststomodeal commissural fibers. The anlage of the nauplius eye reveals a specific pattern of pigment and sensory cells with the two pigment cells expressing synapsin. Clear differences between the expression patterns of synapsin and α-tubulin recommend the combination of a variety of antibodies to gain a complete picture of embryonic neuroanatomy. Our results show overall similarities to other malacostracan and non-malacostracan crustaceans. The comparisons with other crustaceans and arthropods indicate homology of crustacean nauplius eyes, a circumoral deutocerebrum, and a more widespread occurrence of posterior pioneer neurons forming the axon scaffold of the ventral central nervous system than previously thought.     

日本血吸虫胆碱酯酶的组织化学定位

血吸虫的胆碱酯酶(ChE)与血吸虫神经介质的传递、肌肉活动以及与其它物质代谢均有密切关系,也是抗血吸虫药物作用的靶子之一。1959年沈美玲等进行了药物对日本血吸虫乙酰胆碱酯酶(AChE)活力的研究。最近,姚民一等(1981)采用等电聚焦电泳分离出日本血吸虫胆碱酯酶的同功酶。然而有关血吸虫胆碱酯酶的组织化学定位研究主要限于曼氏血吸虫(Pepler,1958;Lewert等,1965;Fripp,1967;Bueding等,1967;Bruckner等,1974;Diconza等,1975)除Bueding简单述及日本血吸虫成虫的AChE外,迄今国内外尚无日本血吸虫CbE的组化资料,而且日本血吸虫的神经系统也未被专门观察。由于ChE的组化定位能细致地显示出血吸虫的神经系统构造,并能进一步提     

Immunocytochemical studies on the naupliar nervous system of Balanus improvisus (Crustacea, Cirripedia, Thecostraca)

The nervous system of nauplii of the crustacean taxon Cirripedia was analysed in the species Balanus improvisus Darwin, 1854 using for the first time immunocytochemical staining against serotonin, RFamide and α-tubulin in combination with confocal laser scanning microscopy. This approach revealed a circumoesophageal neuropil ring with nerves extending to the first and second antennae and to the mandibles, all features typical for Crustacea. In addition, RFamidergic structures are present in the region of the thoraco-abdomen. A pair of posterior nerves and a pair of lateral nerves run in anterior-posterior direction and are connected by a thoracic nerve ring and a more posteriorly situated commissure. A median nerve is situated along the ventral side of the thoraco-abdomen. The innervation of frontolateral horns and the frontal filaments are α-tubulin-positive. Several pairs of large neurons in the protocerebrum, along the circumoesophageal connectives and in the mandibular ganglion stain only for serotonin. Due to the almost complete absence of comparable data on the neuroanatomy of early (naupliar) stages in other Crustacea, we include immunocytochemical data on the larvae of the branchiopod, Artemia franciscana Kellogg, 1906 in our analysis. We describe several characteristic neurons in the brains of the nauplius larvae of both species which are also found in decapod larvae and in adult brains of other crustaceans. Furthermore, our data reveal that the naupliar brain of cirripedes is more complex than the adult brain. It is concluded that this ontogenetic brain reduction is related to the sessile life style of adult Cirripedia.     

Structure of the nervous system of Myzostoma cirriferum (Annelida) as revealed by immunohistochemistry and cLSM analyses

The nervous systems of juvenile and adult Myzostoma cirriferum Leuckart, 1836, were stained with antisera against 5-HT (5-hydroxytryptamine, serotonin), FMRFamide, and acetylated alpha-tubulin in combination with the indirect fluorescence technique and analyzed by confocal laser scanning microscopy. The central nervous system consists of two small cerebral ganglia, connected by a dorsal commissure, a ventral nerve mass, and a pair of long circumesophageal connectives joining the former to the latter. The two neuropil cords within the ventral nerve mass curve outward and are joined to one another anteriorly and posteriorly. They are connected by 12 commissures, forming a ladder-like system. A single median nerve runs along the midventral axis. In addition to the circumesophageal connectives, 11 peripheral nerves arise from each main cord. The first innervates the anterior body region. The others form five groups of two nerves each, the first and thicker nerve of which is the parapodial nerve, innervating the parapodium and two corresponding cirri. Except for those in the most posterior group, the second nerves innervate the lateral organs and the body periphery. Serotonergic perikarya are arranged in six more or less distinct clusters, the first lying in front of and the other five between the main nerve cords. The distribution pattern of the FMRFamidergic perikarya is less clear and the somata lie between and outside the cords. One pair of dorsolateral longitudinal nerves was visualized by tubulin staining. Peripheral nerves and the commissures, in particular, demonstrate a segmental organization of the nervous system of M. cirriferum. Furthermore, their arrangement indicates that the body consists of six segments, the first of which is identifiable only by the first pair of peripheral nerves, the first two commissures, and the anteriormost ventral ganglion. The nervous system M. cirriferum thus exhibits several structures also found in the basic plan of the polychaete nervous system.     

Organization of the nervous system in the pygmy cuttlefish,Idiosepius paradoxus ortmann (Idiosepiidae,Cephalopoda)

The idiosepiid cuttlefish is a suitable organism for behavioral, genetic, and developmental studies. As morphological bases for these studies, organization of the nervous system was examined in Idiosepius paradoxus Ortmann, 1881, using Cajal's silver technique and immunohistochemical staining with anti-acetylated alpha-tubulin antibody. The nervous architecture is generally identical to that described in Sepia and Loligo, but some features characterize the idiosepiid nervous system. The olfactory system is highly developed in the optic tract region. The dorsolateral lobes show large neuropils, connected with each other by a novel well-fasciculated commissure. Each olfactory lobe is subdivided into two lobules. The neuropils of the anterior and the posterior chromatophore lobes are very poorly developed. Neuronal gigantism is not extensive in the brain; enlarged neuronal cells are visible only in the perikaryal layer of the posterior subesophageal mass. The giant nerve fiber system is of the Sepia type; the axons are not markedly thick and the first-order giant fibers do not fuse with each other at the chiasma. Three-dimensional images by whole-mount immunostaining clarified the innervation pattern in the peripheral nervous system in detail. Two commissural fibers link the left and right posterior funnel nerves ventrally and dorsally. The stellate commissure, which is absent in Sepia and Sepiola, connects the stellate ganglia with each other. A branch of the visceral nerve innervating the median pallial adductor muscle is characteristically thick. Tubulinergic reactivity of the cilia and axons reveals the presence of many ciliated cells giving off an axon toward brain nerves in the surface of the funnel, head integument, arm tips, and epidermal lines. Some of these features seem to reflect the inactive nekto-benthic life of the idiosepiid cuttlefish in the eelgrass bed.     

The ganglion-connective junction in the central nervous system of the leech,Hirudo medicinalis

Classical studies of the nervous system of the leech revealed that there were specific types of very large glial cells associated with various parts of the neuron. Recent microelectrode studies demonstrated that there was a low resistance to the flow charge from any one of these large glial cells to another. The present study describes a previously unreported type of glial cell, the glial cell of the fascicles. These cells, which resemble the glial cells of the connectives but are smaller, are found in the fascicles of axons that unite the connectives to the neuropil. Thus, these cells are located between the glial cells of the connectives on the one hand and the glial cells of the neuropil and packets on the other and must be taken into account in considerations of the low resistance to the transfer of charge from one glial cell to another.     

Neural control of somatic muscle function in the earthworm, Allobophora longa, and in the leech, Hirudo medicinalis

Studies have been made on the electrical activity of the segmentary nerves and connectives of the abdominal nervous chain in the earthworm and leech. It was shown that the electrical activity of the isolated piece of the abdominal chain of the leech is manifested of periodic outbursts of impulsation. Presumably this central periodicity accounts for the discharge-like pattern of muscle rhythmic activity which was revealed in our earlier investigations. The electrical activity in the central nervous system of the earthworm depends on afferent influences which pass to the ganglia from the peripheral sensory nervous cells. Stimulation of the abdominal nervous chain did not result in extra discharges of muscle activity, but only affected some of the parameters of the latter.     

The midline glial cells are required for regionalization of commissural axons in the embryonic CNS of Drosophila

 The ventral nerve cord of arthropods is characterised by the organisation of major axon tracts in a ladder-like pattern. The individual neuromeres are connected by longitudinal connectives whereas the contra-lateral connections are brought about through segmental commissures. In each neuromere of the embryonic central nervous system (CNS) of Drosophila an anterior and a posterior commissure is found. The development of these commissures requires a set of neurone-glia interactions at the midline. Here we show that both the anterior as well as the posterior commissures are subdivided into three axon-containing regions. Electron microscopy of the ventral nerve cord of mutations affecting CNS midline cells indicates that the midline glial cells are required for this subdivision. In addition the midline glial cells appear required for a crossing of commissural growth cones perpendicular to the longitudinal tracts, since in mutants with defective midline glial cells commissural axons frequently cross the midline at aberrant angles. Received: 6 July 1997 / Accepted: 27 August 1997     

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.     

Structure and anterior regeneration of musculature and nervous system in Cirratulus cf. cirratus (Cirratulidae,Annelida)

Annelids provide suitable models for studying regeneration. By now, comprehensive information is restricted to only a few taxa. For many other annelids, comparative data are scarce or even missing. Here, we describe the regeneration of a member of the Cirratulus cirratus species complex. Using phalloidin‐labeling and antibody‐stainings combined with subsequent confocal laser scanning microscopy, we provide data about the organization of body wall musculature and nervous system of intact specimens, as well as about anteriorly regenerating specimens. Our analyses show that C. cf. cirratus exhibits a prominent longitudinal muscle layer forming a dorsal muscle plate, two ventral muscle strands and a ventral‐median muscle fiber. The circular musculature forms closed rings which are interrupted in the area of parapodia. The nervous system of C. cf. cirratus shows a typical rope‐ladder like arrangement and the circumesophageal connectives exhibit two separate roots leading to the brain. During regeneration, the nervous system redevelops remarkably earlier than the musculature, first constituting a tripartite loop‐like structure which later become the circumesophageal connectives. Regeneration of longitudinal musculature starts with diffuse ingrowth and subsequent structuring into the blastema. In contrast, circular musculature develops independently inside the blastema. Our findings constitute the first analysis of regeneration for a member of the Cirratuliformia on a structural level. Summarizing the regeneration process in C. cf. cirratus, five main phases can be subdivided: 1) wound closure, 2) blastema formation, 3) blastema differentiation, 4) resegmentation, and 5) growth, respectively elongation. Additionally, the described tripartite loop‐like structure of the regenerating nervous system has not been reported for any other annelid taxon. In contrast, the regeneration of circular and longitudinal musculature originating from different groups of cells seems to be a general pattern in annelid regeneration. J. Morphol. 275:1418–1430, 2014. © 2014 Wiley Periodicals, Inc.     

SALMFamide2 and serotonin immunoreactivity in the nervous system of some acoels (Xenacoelomorpha)

Acoel worms are simple, often microscopic animals with direct development, a multiciliated epidermis, a statocyst, and a digestive parenchyma instead of a gut epithelium. Morphological characters of acoels have been notoriously difficult to interpret due to their relative scarcity. The nervous system is one of the most accessible and widely used comparative features in acoels, which have a so‐called commissural brain without capsule and several major longitudinal neurite bundles. Here, we use the selective binding properties of a neuropeptide antibody raised in echinoderms (SALMFamide2, or S2), and a commercial antibody against serotonin (5‐HT) to provide additional characters of the acoel nervous system. We have prepared whole‐mount immunofluorescent stainings of three acoel species: Symsagittifera psammophila (Convolutidae), Aphanostoma pisae, and the model acoel Isodiametra pulchra (both Isodiametridae). The commissural brain of all three acoels is delimited anteriorly by the ventral anterior commissure, and posteriorly by the dorsal posterior commissure. The dorsal anterior commissure is situated between the ventral anterior commissure and the dorsal posterior commissure, while the statocyst lies between dorsal anterior and dorsal posterior commissure. S2 and serotonin do not co‐localise, and they follow similar patterns to each other within an animal. In particular, S2, but not 5‐HT, stains a prominent commissure posterior to the main (dorsal) posterior commissure. We have for the first time observed a closed posterior loop of the main neurite bundles in S. psammophila for both the amidergic and the serotonergic nervous system. In I. pulchra, the lateral neurite bundles also form a posterior loop in our serotonergic nervous system stainings.     

The development of a simple scaffold of axon tracts in the brain of the embryonic zebrafish, Brachydanio rerio

We have examined neuronal differentiation and the formation of axon tracts in the embryonic forebrain and midbrain of the zebrafish, between 1 and 2 days postfertilisation. Axons were visualised with three techniques; immunocytochemistry (using HNK-1 and antiacetylated tubulin antibodies) and horseradish peroxidase (HRP) labelling in whole-mounted brains, and transmission electron microscopy. Differentiation was monitored by histochemical staining for acetylcholinesterase (AChE). These independent methods demonstrated that a simple grid of tracts and commissures forms the initial axon scaffold of the brain. At 1 day, the olfactory nerve, four commissures, their associated tracts and three other non-commissural tracts are present. By 2 days, these tracts and commissures have all greatly enlarged and, in addition, the optic nerve and tract, and three new commissures and their associated tracts have been added. Small applications of HRP at various sites revealed the origins and projections of some of these earliest axons. Retrogradely labelled cell bodies originated from regions that were also positive for AChE activity. At 1 day, HRP-labelled axons were traced: (1) from the olfactory placode through the olfactory nerve to the dorsal telencephalon; (2) from the telencephalon into the tract of the anterior commissure and also to the postoptic region of the diencephalon; (3) from the hindbrain through the ventral midbrain and diencephalon to the postoptic commissure; (4) from the dorsal diencephalon (in or near the epiphysis) to the tract of the postoptic commissure; (5) from ventral and rostral midbrain through the posterior commissure. Three new projections were demonstrated at 2 days: (1) from the retina through the tract of the postoptic commissure to the tectum; (2) from the telencephalon to the contralateral diencephalon; and (3) from the telencephalon to the ventral flexure. These results show that at 1 day, the zebrafish brain is impressively simple, with a few small, well-separated tracts but by 2 days the brain is already considerably more complex. Most of the additional axons added onto pre-existent tracts rather than pioneered new ones supporting the notion that other axons play a crucial role in the guidance of early central nervous system (CNS) axons.