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Neuroendocrine systems in animals maintain organismal homeostasis and regulate stress response. Although a great deal of work has been done on the neuropeptides and hormones that are released and act on target organs in the periphery, the synaptic inputs onto these neuroendocrine outputs in the brain are less well understood. Here, we use the transmission electron microscopy reconstruction of a whole central nervous system in the Drosophila larva to elucidate the sensory pathways and the interneurons that provide synaptic input to the neurosecretory cells projecting to the endocrine organs. Predicted by network modeling, we also identify a new carbon dioxide-responsive network that acts on a specific set of neurosecretory cells and that includes those expressing corazonin (Crz) and diuretic hormone 44 (Dh44) neuropeptides. Our analysis reveals a neuronal network architecture for combinatorial action based on sensory and interneuronal pathways that converge onto distinct combinations of neuroendocrine outputs., Competing Interests: SH, PS, AM, AS, IZ, AH, CS, JT, RF, AC, MP No competing interests declared, (© 2021, Hückesfeld et al.)

Objective: The ependymal cells, considered today as an active participant in neuroendocrine functions, were investigated by electron microscopy in the central canal of the lowest spinal cord, the filum terminale (FT), in adult rats. In this area of the spinal cord, the central canal is covered by a heterogeneous population of ependymal cells. The aim of the present work was to compare the regional features of the ependymal cells in two different parts of the FT with a special regard to their ultrastructure., Methods: Two parts of the FT were selected for the ultrastructural observations: the rostral (rFT) and the caudal (cFT) ones. The rTF was removed at the level of the immediate continuation of the conus medullaris, while the cFT 30 mm further caudally. After formaldehyde fixation, the spinal cord was removed and cut into small blocks for electron microscopic processing. The material was embedded into durcupan, contrasted with uranyl acetate, lead citrate as well as osmium tetroxide, and investigated under JEOL 1200 EX electron microscope., Results: In the rFT, the ependymal lining is pseudostratified and one-layered in the cFT, whereas the shape of the ependymal cells may vary from cuboidal to flatten in the rostro-caudal direction. The basal membrane of many ependymal cells possesses deep invaginations, so called "filum terminale labyrinths". Many neuronal processes occur in the pericanalicular neuropil. In contrast to the rFT, the cFT is less rich in the neuropil particles. Some of the ependymal cells concurrently reach both the intracanalicular and extracanalicular cerebrospinal fluid (CSF), thus they may represent a new variant of the ependymal cells designated as "bridge cells of the FT"., Conclusions: The present data indicate that the FT ependymal cells exhibit clear differences in anatomy as well as ultrastructure that may reflect their distinct functional activity. Therefore, observations presented here may serve for the better understanding of the physiological role of the individual ependymal areas in this special portion of the rat spinal cord.

The aim of this investigation was to demonstrate the regulatory elements in the skin epithelium of Enteropneusta which are supposed to be related to the chordate ancestors. Using electron microscopy, it was found that in the skin epithelium of a representative of enteropneusts Saccoglossus mereschkowskii, the basal parts of some epitheliocytes took part in formation of a nerve layer. These cells were considered as receptor ciliated cells. The granular epithelial cells were shown to release secretion according to both exocrine and endocrine mechanism; these cells were characterized as endocrine-like regulatory cells. Fine granular cells possibly represent special receptor-endocrine-like cell type. The immunocytochemical detection of FMRFamid neuropeptide localization in histological sections confirmed the electron microscopic data on the presence of receptor and endocrine-like cells in the epithelium. It is suggested that the skin epithelium of Enteropneusta contains a peculiar neuro-endocrine regulatory system that is represented by receptor cells, receptor-endocrine-like cells of an open type and nerve elements of the nerve layer.

Nerve cells in a parasitic cnidarian Polypodium hydriforme at the parasitic and free-living stages of the life cycle have been localized immunocytochemically using antibodies to FMRF-amide, and their ultrastructure has been described. Ganglion cells form a net under epidermis consisting of bi- and tripolar neurons which cross the mesoglea and usually contact muscle cells and cnidocytes. Fusiform sensory and neurosecretory cells, especially characteristic to sensory tentacles, are interspersed among epidermal cells. All three types of nerve cells have dense cored vesicles about 80-120 nm in diameter. The sensory cells demonstrate a sensory flagellum-like immobile structure. Neurosecretory and sensory cells form septate junctions with epidermal cells. Ganglion cells show gap junctions between them. A centriole encircled by a fragment of nuclear envelope which is a marker of ectodermal lineage cells in Polypodium has been described in the cytoplasm of a sensory cell, thus proving the ectodermal nature of the nervous system.

The clinical, histologic, immunophenotypic, ultrastructural, and molecular features of a distinctive gastrointestinal tumor are described. Sixteen patients, 8 women and 8 men aged 17 to 77 years (mean age, 42 y; 63% less than 40 y) presented with abdominal pain, intestinal obstruction, and an abdominal mass. Mean tumor size was 5.2 cm (range, 2.4 to 15.0 cm). The tumors arose in the small bowel (10), stomach (4), and colon (2) and were histologically characterized by a sheet-like or nested population of epithelioid or oval-to-spindle cells with small nucleoli and scattered mitoses. Five cases showed focal clearing of the cytoplasm. Scattered osteoclast-type multinucleated giant cells were present in 8 cases. The tumor cells were positive for S-100 protein, SOX10, and vimentin in 100% of cases, for CD56 in 70%, for synaptophysin in 56%, for NB84 in 50%, for NSE in 45%, and for neurofilament protein in 14% of cases. All cases tested were negative for specific melanocytic, gastrointestinal stromal tumors, epithelial, and myoid markers. Ultrastructural examination of 5 cases showed features of primitive neuroectodermal cells with clear secretory vesicles, dense-core granules, occasional gap junctions, and no evidence of melanogenesis. EWSR1 gene rearrangement was assessed by fluorescence in situ hybridization in 14 cases. Twelve cases (86%) showed split EWSR1 signal consistent with a chromosomal translocation involving EWSR1. One case showed extra intact signals, indicating that the nuclei possessed either extra copies of the EWSR1 gene or chromosome 22 polysomy. Only 1 case showed no involvement of the EWSR1 gene. Six cases demonstrated rearrangement of the partner fusion gene ATF1 (46%), and 3 showed rearrangement of CREB1 (23%); 2 cases lacked rearrangement of either partner gene. Clinical follow-up was available in 12 patients and ranged from 1.5 to 106 months. Six patients died of their tumors (mean survival, 32 mo; 83% less than 24 mo). At last follow-up, 4 patients were alive with regional, lymph node, and liver metastases, and 2 patients were alive with no evidence of disease. The tumor described here is an aggressive form of neuroectodermal tumor that should be separated from other primitive epithelioid and spindle cell tumors of the gastrointestinal tract. The distinctive ultrastructural features and absence of melanocytic differentiation serve to separate them from soft tissue clear cell sarcomas involving the gastrointestinal tract. The designation "malignant gastrointestinal neuroectodermal tumor" is proposed for this tumor type.

4-Nonylphenol (4-NP), an estrogen mimicking compound is produced by biodegradation of alkylethoxylates. It is well established that 4-NP can affect the development of aquatic animals by disrupting the endocrine signals. Here we show for the first time in zebrafish that 4-NP does not only target the neuroendocrine system but also the notochord and the muscle. The notochord malformation was first evident as distortions at 24hourspostfertilization (hpf) which within 24h appeared as kinks and herniations. The notochord phenotype was accompanied by reduced motility and impaired swimming behavior. Whole-mount in situ hybridization using chordamesoderm markers and electron microscopic analysis showed failure in the notochord differentiation and disruption of the perinotochordal basement membrane. Late larval stages of 4-NP treated embryos displayed abnormal mineralization, vertebral curvature, fusion of vertebral bodies and abnormal extension of haemal arches. The muscle structure and the maximal active force in isolated muscle preparations were similar between 4-NP exposed and of control embryos, suggesting that 4-NP did not induce major changes in striated muscle function. However, repeated electrical stimulation (>40Hz) of the 4-NP exposed larvae revealed an impaired relaxation between stimuli, possibly reflecting an alteration in the relaxant mechanisms (e.g. in cellular Ca(2+) removal) which could explain the abnormal swimming pattern exhibited by 4-NP exposed larvae. Additionally, we demonstrate that the expression levels of the stress hormone, corticotropin releasing hormonewere elevated in the brain following 4-NP treatment. We also observed a significant decrease in the transcript levels of luteinizing hormone b at early larval stages. Collectively, our results show that 4-NP is able to disrupt the notochord morphogenesis, muscle function and the neuroendocrine system. These data suggest that 4-NP enduringly affects the embryonic development in zebrafish and that this compound might exert these deleterious effects through diverse signaling pathways., (Copyright © 2011 Elsevier Inc. All rights reserved.)

SNARE protein-driven secretion of neurotransmitters from synaptic vesicles is at the center of neuronal communication. In the absence of the cytosolic protein Munc18-1, synaptic secretion comes to a halt. Although it is believed that Munc18-1 orchestrates SNARE complexes, its mode of action is still a matter of debate. In particular, it has been challenging to clarify the role of a tight Munc18/syntaxin 1 complex, because this interaction interferes strongly with syntaxin's ability to form a SNARE complex. In this complex, two regions of syntaxin, the N-peptide and the remainder in closed conformation, bind to Munc18 simultaneously. Until now, this binary complex has been reported for neuronal tissues only, leading to the hypothesis that it might be a specialization of the neuronal secretion apparatus. Here we aimed, by comparing the core secretion machinery of the unicellular choanoflagellate Monosiga brevicollis with that of animals, to reconstruct the ancestral function of the Munc18/syntaxin1 complex. We found that the Munc18/syntaxin 1 complex from M. brevicollis is structurally and functionally highly similar to the vertebrate complex, suggesting that it constitutes a fundamental step in the reaction pathway toward SNARE assembly. We thus propose that the primordial secretion machinery of the common ancestor of choanoflagellates and animals has been co-opted for synaptic roles during the rise of animals.

Decapod crustaceans have long served as important models for the study of neuroendocrine signaling. For example, the process of neurosecretion was first formally demonstrated by using a member of this order. In this review, the major decapod neuroendocrine organs are described, as are their phylogenetic conservation and neurochemistry. In addition, recent advances in crustacean neurohormone discovery and tissue mapping are discussed, as are several recent advances in our understanding of hormonal control in this group of animals.

Small cell neuroendocrine carcinoma (SCNEC) of the urinary bladder is a rare but aggressive neoplasm that usually exhibits neuroendocrine differentiation. Here, the authors report a case of SCNEC in an 80-year-old man. The patient had gross hematuria and nodular mass involving the wall of the urinary bladder. Total cystectomy was done. The tumor consisted of small, uniform, round, and spindled-shaped cells with chromatin dark nuclei and numerous mitotic figures. The cells were reactive for chromogranin, neuron-specific enolase (diffuse), and keratin (focal). Ultrastructural studies revealed neurosecretory granules and intermediate filaments. The diagnosis of SCNEC with focal high-grade urothelial component was established. No metastasis was found at the time of diagnosis and the patient refused further chemotherapy or radiotherapy. The histogenesis, differential diagnosis, and prognosis of SCNEC of the urinary bladder were discussed.

In experiment the action of Thymalin, alpha-tocopherol and stress on the centers of hypothalamus related to the regulation of the gonadotropic function has been investigated and the age-specific features of such influence as well. The results of experiments indicate the presence of neuron's reaction of the rostra preoptical area and arcuate nucleus for experimental influences. As a stress exposure and alpha-tocopherol injection to the young animals a reduction of neuron's kernels of the arcuate nucleus was observed, a sexual distinction wasn't revealed in the process. The response age-specific feature for alpha-tocopherol injection of the nucleus neuron's rostra preoptical and arcuate area of the hypothalamus of white rats is a reduction in a degree of response. Thymalin reduces a threshold of sensitivity of the neuron's arcuate center to action of stress.

Neuropeptide Y (NPY) functions as a peptide neurotransmitter and as a neuroendocrine hormone. The active NPY peptide is generated in secretory vesicles by proteolytic processing of proNPY. Novel findings from this study show that cathepsin L participates as a key proteolytic enzyme for NPY production in secretory vesicles. Notably, NPY levels in cathepsin L knockout (KO) mice were substantially reduced in brain and adrenal medulla by 80% and 90%, respectively. Participation of cathepsin L in producing NPY predicts their colocalization in secretory vesicles, a primary site of NPY production. Indeed, cathepsin L was colocalized with NPY in brain cortical neurons and in chromaffin cells of adrenal medulla, demonstrated by immunofluorescence confocal microscopy. Immunoelectron microscopy confirmed the localization of cathepsin L with NPY in regulated secretory vesicles of chromaffin cells. Functional studies showed that coexpression of proNPY with cathepsin L in neuroendocrine PC12 cells resulted in increased production of NPY. Furthermore, in vitro processing indicated cathepsin L processing of proNPY at paired basic residues. These findings demonstrate a role for cathepsin L in the production of NPY from its proNPY precursor. These studies illustrate the novel biological role of cathepsin L in the production of NPY, a peptide neurotransmitter, and neuroendocrine hormone.

The ultrastructural features of neuroendocrine neurons in the dorsomedial nucleus (DMN) of the amygdaloid body of the brain - one of the major zones of sexual dimorphism - in 12 Wistar rats weighing 250-300 g were studied in three males and nine females at different stages of the estral cycle. On the basis of ultrastructural characteristics, analysis of the functional states of an average of 50 DMN neurons were studied in each animal. A morphofunctional classification reflecting hormone-dependent variations in neuron activity is proposed. DMN neurons were found to be in different structural-functional states, which could be classified as the states of rest, moderate activity, elevated activity, tension (maximal activity), decreased activity (types 1 and 2, depending on prior history), return to the initial state, and apoptosis. At the estrus stage, there was a predominance of neurons in the states of elevated activity (40% of all cells) and maximal activity (26%). At the metestrus stage, neurons in the state of decreased activity type 1 (with increased nuclear heterochromatin content) predominated (30% of cells), while 25% and 20% of cells were in the states of maximal activity and elevated activity respectively. In diestrus, neurons in the resting state, in moderate and elevated activity, in maximal activity, and in decreased activity type 1 were present in essentially identical proportions (18%, 21%, 18%, 20%, and 16% respectively). In males, 35% and 22% of neurons were in the states of elevated and maximal activity respectively. Neuron death was seen only in males.

We report two cases of a large-cell neuroendocrine carcinoma of the skin as a distinct histological category. The patients are a 63-year-old woman with a 7-mm reddish round nodule on her nose for 4 months, and a 95-year-old man with several hemorrhagic nodules on the external ear for 28 months. The first patient had two positive sentinel lymph nodes. The tumors from both patients consisted of islands of pleomorphic large cells within a lymphoid stroma.

The histopathology, ultrastructure, and immunohistochemistry of the alimentary canal of flounder Platichthys flesus (L.), naturally infected with the nematode Anisakis simplex s.l. (Rudolphi 1809) from the River Forth (Scotland), were investigated and described. Eight of the 16 flounders were infected with A. simplex s.l. larvae (L3); parasites were encapsulated by serosa on the external surface of the host's digestive tract (intensity of infection 1-8 parasites per host), although nematode larvae were found encysted under the peritoneal visceral serosa of the host spleen and liver and, occasionally, in the liver parenchyma (intensity of infection 3-10 parasites per host). In all sites, different structural elements were recognized within the capsule surrounding larvae. Among the epithelial cells of the intestine of 5 flounders with larvae encysted on external surface of the gut, the presence of several rodlet cells (RCs) was observed. Furthermore, often the occurrence of macrophage aggregates (MAs) was noticed in infected liver and spleen, mainly around the parasite larvae. Eight neuropeptide antisera were tested with immunohistochemistry methods on gut sections of 4 P. flesus infected with extraintestinal nematodes. Sections from the gut of 5 uninfected flounder were used for comparative purposes. In the tunica mucosa of parasitized P. flesus, several endocrine epithelial cells were immunoreactive to anti-CCK-39 (cholecystokinin 39) and -NPY (neuropeptide Y) sera; furthermore, in the myenteric plexus, a high number of neurons were immunoreactive to antibombesin, -galanin, and several to -NPY and -PHI (peptide histidine isoleucine) sera.

Several evidences suggest that the posterodorsal medial amygdala (MePD) can be a relevant part of the rat neural circuitry for the regulation of hypothalamic neuroendocrine secretion and for ontogenetically different behavioral displays. The dendritic spine density of Golgi-impregnated neurons from the MePD was evaluated in young rats following acute or chronic restraint stress and in aged animals (24 months old). Compared to the control group, a single 1 h restraint stress session promoted a decreased spine density (p<0.01) whereas a single 6 h restraint stress session or daily 6-h restraint sessions for 28 consecutive days did not lead to the same effect (p>0.05). Aged rats showed no difference in this dendritic spine parameter when compared to young adults (p>0.05). These results indicate that short-term stress (1 h) can affect MePD dendritic spines and that neural plasticity is involved with adaptive responses onwards in restrained rats. On the other hand, brain structural modifications related with ageing appear not to influence the number of certain postsynaptic sites in the MePD of rats.

Angiotensin IV and LVV-hemorphin 7 promote robust enhancing effects on learning and memory. These peptides are also competitive inhibitors of the insulin-regulated membrane aminopeptidase, suggesting that the biological actions of these peptides may result from inhibition of IRAP activity. However, the normal function of IRAP in the brain is yet to be determined. The present study investigated the sub-cellular distribution of IRAP in four neuronal cell lines and in the mouse brain. Using sub-cellular fractionation, IRAP was found to be enriched in low density microsomes, while lower levels of IRAP were also present in high density microsomes, plasma membrane and mitochondrial fractions. Dual-label immunohistochemistry confirmed the presence of IRAP in vesicles co-localized with the vesicular maker VAMP2, in the trans Golgi network co-localized with TGN 38 and in endosomes co-localized with EEA1. Finally using electron microscopy, IRAP specific immunoreactivity was predominantly associated with large 100-200 nm vesicles in hippocampal neurons. The location, appearance and size of these vesicles are consistent with neurosecretory vesicles. IRAP precipitate was also detected in intracellular structures including the rough endoplasmic reticulum, Golgi stack and mitochondrial membranes. The sub-cellular localization of IRAP in neurons demonstrated in the present study bears striking parallels with distribution of IRAP in insulin responsive cells, where the enzyme plays a role in insulin-regulated glucose uptake. Therefore, we propose that the function of IRAP in neurons may be similar to that in insulin responsive cells.

Neurosecretory control centers form part of the forebrain in many animal phyla, including vertebrates, insects, and annelids. The evolutionary origin of these centers is largely unknown. To identify conserved, and thus phylogenetically ancient, components of neurosecretory brain centers, we characterize and compare neurons that express the prohormone vasotocin (vasopressin/oxytocin)-neurophysin in the developing forebrain of the annelid Platynereis dumerilii and of the zebrafish. These neurons express the same tissue-restricted microRNA, miR-7, and conserved, cell-type-specific combinations of transcription factors (nk2.1, rx, and otp) that specify their identity, as evidenced by the specific requirement of zebrafish rx3 for vasotocin-neurophysin expression. MiR-7 also labels another shared population of neurons containing RFamides. Since the vasotocinergic and RFamidergic neurons appear to be directly sensory in annelid and fish, we propose that cell types with dual sensory-neurosecretory properties were the starting point for the evolution of neurosecretory brain centers in Bilateria.

We developed a classification of neuroendocrine neurons of the dorsomedial nucleus of the cerebral amigdaloid complex of male rats; characteristics of these cells are presented.

Heat shock proteins (Hsps) are evolutionary conserved peptides well known as molecular chaperones and stress proteins. Elevated levels of extracellular Hsps in blood plasma have been observed during the stress responses and some diseases. Information on the cellular sources of extracellular Hsps and mechanisms regulating their release is still scanty. Here we showed the presence and localization of Hsp70 in the neuroendocrine system in the atrium of the snail, Achatina fulica. The occurrence of the peptide in snail atrium lysate was detected by Western blot analysis. Immunoperoxidase and immunogold staining demonstrated that Hsp70-immunoreactivity is mainly confined to the peculiar atrial neuroendocrine units which are formed by nerve fibers tightly contacted with large granular cells. Immunolabelling intensity differed in morphologically distinct types of secretory granules in the granular cells. The pictures of exocytosis of Hsp70-immunolabeled granules from the granular cells were observed. In nerve bundles, axon profiles with Hsp70-immunoreactive and those with non-immunoreactive neurosecretory granules were found. In addition, Hsp70-like material was also revealed in the granules of glia-interstitial cells that accompanied nerve fibers. Our findings provide an immuno-morphological basis for a role of Hsp70 in the functioning of the neuroendocrine system in the snail heart, and show that the atrial granular cells are a probable source of extracellular Hsp70 in the snail hemolymph.

Recent studies have localized the glutamatergic cell marker type-2 vesicular glutamate transporter (VGLUT2) to distinct peptidergic neurosecretory systems that regulate hypophysial functions in rats. The present studies were aimed to map the neuronal sources of VGLUT2 in the median eminence and the posterior pituitary, the main terminal fields of hypothalamic neurosecretory neurons. Neurons innervating these regions were identified by the uptake of the retrograde tract-tracer Fluoro-Gold (FG) from the systemic circulation, whereas glutamatergic perikarya of the hypothalamus were visualized via the radioisotopic in situ hybridization detection of VGLUT2 mRNA. The results of dual-labeling studies established that the majority of neurons accumulating FG and also expressing VGLUT2 mRNA were located within the paraventricular, periventricular and supraoptic nuclei and around the organum vasculosum of the lamina terminalis and the preoptic area. In contrast, only few FG-accumulating cells exhibited VGLUT2 mRNA signal in the arcuate nucleus. Dual-label immunofluorescent studies of the median eminence and posterior pituitary to determine the subcellular location of VGLUT2, revealed the association of VGLUT2 immunoreactivity with SV2 protein, a marker for small clear vesicles in neurosecretory endings. Electron microscopic studies using pre-embedding colloidal gold labeling confirmed the localization of VGLUT2 in small clear synaptic vesicles. These data suggest that neurosecretory neurons located mainly within the paraventricular, anterior periventricular and supraoptic nuclei and around the organum vasculosum of the lamina terminalis and the preoptic area secrete glutamate into the fenestrated vessels of the median eminence and posterior pituitary. The functional aspects of the putative neuropeptide/glutamate co-release from neuroendocrine terminals remain to be elucidated.

Background: Extraventricular neurocytoma is a well-established entity, and non-classical sites like the cerebellum and spinal cord have occasionally been reported., Case: We report a rare case of intramedullary neurocytoma in an 8-year-old boy who presented with paraparesis and urinary incontinence. This tumour was isointense on T1-weighted image and hyperintense on T2-weighted image, and showed heterogenous contrast enhancement., Findings: Pathological examination showed atypical histology in the form of vascular endothelial proliferation, increased proliferation index, mitosis and necrosis., Conclusions: Atypical location, younger age and histopathological features are some of the interesting findings in this case, which need to be documented. To the best of our knowledge, this is the youngest patient of spinal cord neurocytoma.

FXPRLamides are insect neuropeptides that mediate such diverse functions as pheromone biosynthesis, visceral muscle contraction, and induction of diapause. Although multiple forms occur in every insect studied so far, little is known about a possible functional differentiation and/or differences in the cellular expression pattern of these messenger molecules. In this study, we performed a mass spectrometric survey of all FXPRLamide-expressing neurosecretory neurons in the CNS of Periplaneta americana. That species combines a very well characterized peptidergic system with relatively easy accessible neurosecretory cells suitable for dissection. In addition to the extensive mass spectrometric analyses of single cells, the projection of the FXPRLamide-expressing neurons was studied with three antisera specifically recognizing different FXPRLamides. The following conclusions can be drawn from this first comprehensive peptidomic approach on insect neurons. 1) A high degree of differentiation in the expression of FXPRLamides exists; not fewer then four cell types containing different sets of FXPRLamides were observed. 2) A low level of colocalization with other neuropeptides was found in these neurons. 3) A comparison with FXPRLamide-expressing neurons of other insects shows a high degree of conservation in the localization and projection of these neurons, which is not corroborated by a similar conservation of the corresponding peptide sequences. 4) Although the methods for cell identification, dissection, and sample preparation for mass spectrometry were kept as simple as possible, it was unambiguously shown that this approach is generally suitable for routine analysis of single identified neurons of insects., (Copyright 2006 Wiley-Liss, Inc.)

The saccus vasculosus (SV) is a circumventricular organ of the hypothalamus of many jawed fishes whose functions have not yet been clarified. It is a vascularized neuroepithelium that consists of coronet cells, cerebrospinal fluid-contacting (CSF-c) neurons and supporting cells. To assess the organization, development and evolution of the SV, the expression of glial fibrillary acidic protein (GFAP) and the neuronal markers gamma-aminobutyric acid (GABA), glutamic acid decarboxylase (GAD; the GABA synthesizing enzyme), neuropeptide Y (NPY), neurophysin II (NPH), tyrosine hydroxylase (TH; the rate-limiting catecholamine-synthesizing enzyme) and serotonin (5-HT), were investigated by immunohistochemistry in developing and adult sharks. Coronet cells showed GFAP immunoreactivity from embryos at stage 31 to adults, indicating a glial nature. GABAergic CSF-c neurons were evidenced just when the primordium of the SV becomes detectable (at stage 29). Double immunolabeling revealed colocalization of NPY and GAD in these cells. Some CSF-c cells showed TH immunoreactivity in postembryonic stages. Saccofugal GABAergic fibers formed a defined SV tract from the stage 30 and scattered neurosecretory (NPH-immunoreactive) and monoaminergic (5-HT- and TH-immunoreactive) saccopetal fibers were first detected at stages 31 and 32, respectively. The early differentiation of GABAergic neurons and the presence of a conspicuous GABAergic saccofugal system are shared by elasmobranch and teleosts (trout), suggesting that GABA plays a key function in the SV circuitry. Monoaminergic structures have not been reported in the SV of bony fishes, and were probably acquired secondarily in sharks. The existence of saccopetal monoaminergic and neurosecretory fibers reveals reciprocal connections between the SV and hypothalamic structures which have not been previously detected in teleosts., (Copyright (c) 2007 S. Karger AG, Basel.)

The caudal neurosecretory system is described here for the first time in the zebrafish, one of the most important models used to study biological processes. Light- and electron-microscopical approaches have been employed to describe the structural organization of Dahlgren cells and the urophysis, together with the immunohistochemical localization of urotensin I and II (UI and UII) peptides. Two latero-ventral bands of neuronal perikarya in the caudal spinal cord project axons to the urophysis. The largest secretory neurons (approximately 20 microm) are located rostrally. UII-immunoreactive perikarya are much more numerous than those immunoreactive for UI. A few neurons are immunopositive for both peptides. Axons contain 75-nm to 180-nm dense-core vesicles comprising two populations distributed in two axonal types (A and B). Large dense vesicles predominate in type A axons and smaller ones in type B. Immunogold double-labelling has revealed that some fibres contain both UI and UII, sometimes even within the same neurosecretory granule. UII is apparently the major peptide present and predominates in type A axons, with UI predominating in type B. A surprising finding, not previously reported in other fish, is the presence of dense-core vesicles, similar to those in neurons, in astrocytes including their end-feet around capillaries. Secretory type vesicles are also evident in ependymocytes and cerebrospinal-fluid-contacting neurons in the terminal spinal cord. Thus, in addition to the urophysis, this region may possess further secretory systems whose products and associated targets remain to be established. These results provide the basis for further experimental, genetic and developmental studies of the urophysial system in the zebrafish.

Light and electron microscopic methods were used to analyze changes in secretory neurons in the supraoptic (SON) and paraventricular (PVN) nuclei in the hypothalamus in 100 adult male rats at time points from the first minutes to 180 days after 48 hours of full-time exposure to bright light. At the early time points after exposure, the cellular formulae of the SON and PVN shifted towards functionally active neurons with minimal quantities of secretory granules, large nuclei and nucleoli, low RNA contents, small numbers of rough endoplasmic reticulum cisterns, vacuoles, and lysosomes in the perikarya. The number of cells depositing secretion was greater than in controls at 24 h in the SON and PVN and at 10 days in the SON. Normalization of the cellular formula and the structural organization of the protein-synthesizing apparatus of PVN neurons occurred at 10-30 days, with normalization in the SON at 30-180 days. These data provide evidence that the range of plasticity of neurons in the PVN on exposure to full-time bright light was more significant than that in the SON.

Ultrastructural characteristics of neuroendocrine neurons were studied in amygdala dorsomedial nucleus (DMN), which is one of main zones of sexual dimorphism, in 12 Wistar rats with the body mass of 250-300 g, 3 males and 9 females at different stages of the estrous cycle. In each animal, an average of 50 DMN neurons were examined and their functional state was analyzed on the basis of their ultrastructural characteristics. Morpho-functional classification is proposed, that reflects hormone-dependent variations in neuronal activity. It was established that DMN neurons could be found in different structural and functional states, that may be classified as the state of rest, moderate activity, increased activity, stress (peak activity), decrease in activity (two types), return to an initial state and apoptosis. At the stage of estrus, the neurons in the state of increased activity (40% of the total population) and peak activity (26%) were found to prevail. In metestrus, most of the neurons were in the state of type I decrease in activity (with an increase in nuclear heterochromatin content--30% of the cells); peak activity and increased activity were found in 25% and 20% of neurons, respectively. In diestrus, the neurons in the state of rest, moderate and increased activity, peak activity and type I decrease in activity were represented in approximately equal proportions (18%, 21%, 18%, 20% and 16%, respectively). In males, 35% and 22% of neurons, respectively, were found in the state of increased activity and peak activity. Neuronal death was detected only in males.

In crustaceans, circulating hormones influence many physiological processes. Two neuroendocrine organs, the sinus gland (SG) and the pericardial organ (PO), are the sources of many of these compounds. As a first step in determining the roles played by hemolymph-borne agents in the crab Cancer productus, we characterized the hormone complement of its SG and PO. We show via transmission electron microscopy that the nerve terminals making up each site possess dense-core and/or electron-lucent vesicles, suggesting diverse complements of bioactive molecules for both structures. By using immunohistochemistry, we show that small molecule transmitters, amines and peptides, are among the hormones present in these tissues, with many differentially distributed between the two sites (e.g., serotonin in the PO but not the SG). With several mass spectrometric (MS) methods, we identified many of the peptides responsible for the immunolabeling and surveyed the SG and PO for peptides for which no antibodies exist. By using MS, we characterized 39 known peptides [e.g., beta-pigment-dispersing hormone (beta-PDH), crustacean cardioactive peptide, and red pigment-concentrating hormone] and de novo sequenced 23 novel ones (e.g., a new beta-PDH isoform and the first B-type allatostatins identified from a non-insect species). Collectively, our results show that diverse and unique complements of hormones, including many previously unknown peptides, are present in the SG and PO of C. productus. Moreover, our study sets the stage for future biochemical and physiological studies of these molecules and ultimately the elucidation of the role(s) they play in hormonal control in C. productus., (Copyright (c) 2005 Wiley-Liss, Inc.)

In females of the blow fly Protophormia terraenovae, neurons with cell bodies in the pars lateralis (PL) projecting to the retrocerebral complex (designated as PL neurons) are necessary for the induction of reproductive diapause under short-day and low-temperature conditions. In the present study, neural connections between PL neurons and pigment-dispersing factor (PDF)-immunoreactive neurons were examined via immunolight microscopy and immunoelectron microscopy combined with backfills through the cardiac-recurrent nerve. Immunolight microscopy showed that fibers of PL neurons overlapped with PDF-immunoreactive fibers in the dorsolateral region of the superior protocerebral neuropil. Immunoelectron microscopy showed that PDF-immunoreactive fibers formed output synapses with fibers of PL neurons and unlabeled neurons in a region dorsoanteriorly located with respect to the calyx of the mushroom body. The distribution of synaptic connections between PDF-immunoreactive fibers and the fibers of PL neurons was sparse. According to the projection patterns, PDF-immunoreactive fibers with synaptic connections with PL neurons appeared to originate from PDF-immunoreactive neurons with cell bodies at the base of the medulla of the optic lobe (medulla PDF neurons), which are putative circadian clock neurons in P. terraenovae. PDF immunoreactivity was restrictively detected in dense-core vesicles but not in clear synaptic vesicles. The present results suggest that medulla PDF neurons convey time or photoperiodic information to PL neurons for diapause induction through direct synaptic connections., (2005 Wiley-Liss, Inc.)

The structural changes of neurons of the rat hypothalamic supraoptic (SON) and of paraventricular (PVN) nucleus after 48 h of bright light exposure, of 5 Gy whole-body X-irradiation and of their combination subjected to the analysis by means of light-optic and of electron microscopy for the estimation of radimodificated effect of light exposure lasted 24 h a day and plasticity of neuroendocrine transducers interacted with the optic sensory system. The structural changes of neurons of the SON after combined action are less considerable and more prolonged in comparison with the PVN that loas defermined by their direct connection with the optic sensory system via the retinohypothalamic tract.

Neuroendocrine (NE) carcinoma was diagnosed in 10 dogs. In six cases examined by cephalometric radiography and computerized tomography, a large mass was seen to fill the nasal cavity. Histopathologically, sheets, nests or ribbons of neoplastic cells were separated by delicate or thick fibrovascular stroma. The neoplastic cells were round, oval, or spindle-shaped; cytoplasmic granules and hyperchromatic nuclei with prominent nucleoli were present. Neoplastic cells were invariably immunohistochemically positive for cytokeratin (CK) AE1/AE3, neuron-specific enolase, chromogranin A and vasoactive intestinal polypeptide. Eight dogs were positive for S100 protein, seven for synaptophysin, five for protein gene product 9.5, two for somatostatin, and one for Leu-7. Immunolabelling gave negative results for CK 8, CK 19, calcitonin, calcitonin gene-related polypeptide, neurofilaments, serotonin, gastrin and glial fibrillary acidic protein. Ultrastructurally, the neoplastic cells contained a large number of round, membrane-bounded, densely-cored granules corresponding to neurosecretory granules. These observations were consistent with the neuroendocrine nature of the carcinomas.

Cryo field emission scanning electron microscopy (cryo-FE-SEM) is a versatile technique that allows the investigation of the three-dimensional organization of cells at the ultrastructural level over a wide range of magnifications. Unfortunately, cryopreparation of the specimens for this technique remains cumbersome, in particular because ice crystal formation must be prevented during freezing. Here we report that a light prefixation with glutaraldehyde and incubation in glycerol as cryoprotectant or a high-pressure freezing approach are both excellent procedures for cryopreparation of animal cells to be used in combination with cryo-FE-SEM. Using the proopiomelanocortin-producing intermediate pituitary melanotrope cells of Xenopus laevis as a physiologically inducible neuroendocrine system, we compared the ultrastructural characteristics of inactive and hyperactive neuroendocrine cells. The overall quality of the ultrastructural images was comparable for the two cryopreparation procedures, although some fine structures were better conserved using high-pressure freezing. Melanotrope cells in a secretory inactive state contained numerous storage granules and a poorly developed endoplasmic reticulum (ER), while large amounts of rough ER were present in hyperactive cells. Thus, the cryo-FE-SEM approach described here allows a fast ultrastructural study on the secretory activity of neuroendocrine cells.

A 17-year-old horse developed severe proptosis of the left eye over a period of 1.5 years. At post-mortem examination a neoplasm was found involving the left ethmoid bone, left maxillary sinus, left orbit, left superior turbinate, and the left eye. Tumour cells were arranged in nests separated by a fine fibrovascular stroma. Immunohistochemically, the tumour cells were labelled by antibodies against neurofilament protein, synaptophysin, glial fibrillary acidic protein and S-100 protein antigen, but were negative for chromogranin A, cytokeratin and desmin. Electronmicroscopically, the cells showed neurosecretory granules with an electron-dense centre and a light halo, and microfilaments. On the basis of macroscopical, light microscopical and ultrastructural findings a diagnosis of a metastatic esthesioneuroblastoma was made.

A unique costorage of neuropeptides was recently found in the abdominal perisympathetic organs (PSOs) of the American cockroach, Periplaneta americana. Having specific antisera directed against all peptides belonging to this neurosecretory system, we examined the sorting of PSO-peptides in the soma of the median neurosecretory cells of abdominal ganglia by using immunoelectron microscopic double stainings. The data indicate that all six abundant neuropeptides of this neurohormonal system, which includes three capa-gene related products, are primarily incorporated into separate vesicles. These vesicles fuse with each other in the cytoplasm and become translucent on their way to the axon hillock. By means of light microscopy and MALDI-TOF mass spectrometry, an identical population of neuropeptides was found in interneurons of the brain. As revealed by subsequent immunoelectron microscopic analysis, the peptides of these cells are separately packed into dense core vesicles but do not fuse with each other. Thus, hitherto unknown cell-type-specific sorting mechanisms occur in neurosecretory cells and interneurons, respectively.

Glial and endothelial cells interact throughout the brain to define specific functional domains. Whether endothelial cells convey signals to glia in the mature brain is unknown but is amenable to examination in circumventricular organs. Here we report that purified endothelial cells of one of these organs, the median eminence of the hypothalamus, induce acute actin cytoskeleton remodeling in isolated ependymoglial cells and show that this plasticity is mediated by nitric oxide (NO), a diffusible factor. We found that both soluble guanylyl cyclase and cyclooxygenase products are involved in this endothelial-mediated control of ependymoglia cytoarchitecture. We also demonstrate by electron microscopy that activation of endogenous NO release in the median eminence induces rapid structural changes, allowing a direct access of neurosecretory axons containing gonadotropin-releasing hormone (GnRH) (the neuropeptide controlling reproductive function) to the portal vasculature. Local in vivo inhibition of NO synthesis disrupts reproductive cyclicity, a process that requires a pulsatile, coordinated delivery of GnRH into the hypothalamic-adenohypophyseal portal system. Our results identify a previously unknown function for endothelial cells in inducing neuroglial plasticity and raise the intriguing possibility that endothelial cells throughout the brain may use a similar signaling mechanism to regulate glial-neuronal interactions.

Trypsin and chymotrypsin-like enzymes were detected in the gut of Aedes aegypti in the four larval instar and pupal developmental stages. Although overall the amount of trypsin synthesized in the larval gut was 2-fold higher than chymotrypsin, both enzymes are important in food digestion. Feeding Aea-Trypsin Modulating Oostatic Factor (TMOF) to Ae. aegypti and Culex quinquefasciatus larvae inhibited trypsin biosynthesis in the larval gut, stunted larval growth and development, and caused mortality. Aea-TMOF induced mortality in Ae. aegypti, Cx. quinquefasciatus, Culex nigripalpus, Anopheles quadrimaculatus, and Aedes taeniorhynchus larvae, indicating that many mosquito species have a TMOF-like hormone. The differences in potency of TMOF on different mosquito species suggest that analogues in other species are similar but may differ in amino acid sequence or are transported differently through the gut. Feeding of 29 different Aea-TMOF analogues to mosquito larvae indicated that full biological activity of the hormone is achieved with the tetrapeptide YDPA. Using cytoimmunochemical analysis, intrinsic TMOF was localized to ganglia of the central nervous system in larvae and male and female Ae. aegypti adults. The subesophageal, thoracic, and abdominal ganglia of both larval and adult mosquitoes contained immunoreactive cells. Immunoreactive cells were absent in the corpus cardiacum of newly molted 4th instar larvae but were found in late 4th instar larvae. In both males and females, the intrinsic neurosecretory cells of the corpus cardiacum were filled with densely stained immunoreactive material. These results indicate that TMOF-immunoreactive material is synthesized in sugar-fed male and female adults and larvae by the central nervous system cells., (Copyright 2004 Wiley-Liss, Inc.)

To study the morpho-physiological changes of the nervous and endocrine regulatory systems following long-term influence of morphine hydrochloride, morphometric and correlation analysis of the structural components of the cerebral cortex, hypothalamus, adenohypophysis and thyroid gland was performed. The interrelation between the values of some parameters and the dose of morphine hydrochloride, that caused the tolerance to the drug, was shown. Dose-dependent effect was found in the degree of morphological remodeling of organ components that was demonstrated by the changes in cell and nuclear dimensions, staining properties of cytoplasm, ultrastructural components of thyrocytes, neurons and their processes, hormonal background, the number of the functioning blood capillaries. Interrelations of the morphological parameters are demonstrated by the correlations of different strength and direction.

Electron microscopy of the lungs of Melanophryniscus stelzneri stelzneri (Anura) revealed the presence of a complex pattern of corpuscular cells (CCs). The respiratory surface over the septa presents small areas where the CCs are grouped forming neuroepithelial bodies (NEBs). These corpuscular structures can also be localized in the inner layer of the lung wall. Although in both cases NEBs protrude slightly into the airway lumen, they are separated from the airway lumen and the basal connective tissue by thin apical and basal cytoplasmic processes of neighbouring pneumocytes. Ultrastructurally, the CCs possess a large nucleus, clear cytoplasm and vesicles of variable morphology and size, containing an electron dense material surrounded by a lucent space in some cases. The size of these dense-core vesicles (DCVs) ranged from 40 to 100 nm. The NEBs are associated with afferent and efferent terminal nerves. These types of nerve endings are located between the CCs and in the basal part of the NEBs. The location of the NEBs in strategic positions on the septa and in the wall of the lung, the presence of the DCVs in the basolateral region of the CCs, the occurrence of synaptic contacts between nerve endings and the CCs and the occurrence of capillaries close to the NEBs, suggest a receptosecretory function for NEBs in the lung of M.s. stelzneri.

Neuropeptides were used in experiments to assess their effects on planarian cells. Intact and decapitated planarians exposed to 10(-6) M neuropeptides for two days were examined electron microscopically and quantitative changes in the nuclear pores and chromatoid bodies in various types of cells were ascertained. The data clearly indicated the diversity of effects produced in planarian cells by neuropeptide treatments. The number of nuclear pores increased considerably in each cell type treated with neuropeptides. In particular, the effects of neuropeptides were strongest in differentiating cells which were forming the regeneration blastema. Neuropeptide-treated cells also experienced a dramatic increase in the number of chromatoid bodies. The results obtained in this study suggest that synthesis of RNAs leading to increases in the numbers of nuclear pores and chromatoid bodies is facilitated in neuropeptide-treated cells which are undergoing cell differentiation. The different mechanisms of the effects induced in undifferentiated cells by neuropeptides are discussed.

Isomerization of the third amino acid residue (a phenylalanine) of crustacean hyperglycemic hormone (CHH) has been previously reported to occur as a late step of hormone precursor maturation in a few neurosecretory cells in the X-organ-sinus gland complex of the crayfish Orconectes limosus. In the present report, using conformation-specific antisera combined with immunogold labeling, we have studied, at the ultrastructural level, the distribution of L- and D-CHH immunoreactivity in CHH-secreting cells of the crayfish Astacus leptodactylus. Two CHH-secreting cell populations were observed, the first one (L-cells), the most numerous, exhibited only labeling for L-CHH. In the second one (D-cells), four secretory granule populations were distinguished according to their labeling: unlabeled, either L- or D- exclusively or both L- and D-granules. Labeling quantification by image analysis in D-cells showed a marked increase in D-labeling from the cell body to the axon terminal. However some L- and mixed granules remain in axon terminals. Our results demonstrate that Phe3 isomerization of CHH occurs within the secretory granules of specialized neurosecretory cells and progresses as the granules migrate along the axonal tract. The observation that not all the CHH synthesized is isomerized, and the great variability in the proportion of L- and D-immunoreactivity in granules in every cell region may suggest an heterogeneous distribution of the putative enzyme involved in Phe3 isomerization, a peptide isomerase, within the secretory pathway.

The SNARE complex, consisting of synaptobrevin, syntaxin, and SNAP-25, is essential for calcium-triggered exocytosis in neurosecretory cells. Little is known, however, about how developmentally regulated isoforms and other cognate SNARE components regulate vesicular fusion. To address this question, we examined neuroexocytosis from chromaffin cells of Snap25 null mice rescued by the two splice variants SNAP-25a and SNAP-25b and the ubiquitously expressed homolog SNAP-23. In the absence of SNAP-25, vesicle docking persisted, but primed vesicle pools were empty and fast calcium-triggered release abolished. Single vesicular fusion events showed normal characteristics, except for a shorter duration of the fusion pore. Overexpression of SNAP-25a, SNAP-25b, and SNAP-23 resulted in three distinct phenotypes; SNAP-25b induced larger primed vesicle pools than SNAP-25a, whereas SNAP-23 did not support a standing pool of primed vesicles. We conclude that three alternative SNARE components support exocytosis, but they differ in their ability to stabilize vesicles in the primed state.

Objective: The urethra is the main port of entry of sexually transmitted pathogens. However, papers on the morphology of the urethra are scarce. The Mongolian gerbil is a rodent native of the Mongolia and China and has been utilized as a laboratory animal since the 1960s. This work describes the ultrastructure of the urethra of the Mongolian gerbil to provide data for future experimental studies., Methods: The urethra of ten adult male gerbils was studied by transmission electron microscopy., Results: The epithelium of the pelvic urethra possesses two cell types: I and II, without the formation of cellular layers, while the penile urethra possesses cellular layers: basal, intermediate and superficial. The urethra presents neurosecretory cells belonging to the amine precursor uptake and decarboxylation system., Conclusions: The urethral epithelium of the gerbil is a neurosecretory epithelium, part of the amine precursor uptake and decarboxylation system.

A case of a 58-year-old woman with ovarian carcinoid exhibiting double function is reported. She had suffered from constipation and hirsutism before surgery. Pathological examination revealed that many carcinoid tumor cells were immunohistochemically positive for peptide YY, which inhibits intestinal motility and many peripheral steroid cells. After surgery the patient recovered from constipation immediately. Although the serum level of testosterone also immediately decreased, hirsutism remained for about 2 years. These clinical manifestations are considered to be due to peptide hormone-producing tumor parenchymal cells and testosterone-producing functioning stromal cells. This is the first report of clinically manifested double-functioning ovarian carcinoid; one function is due to tumor cells themselves and another function is due to stromal cells.

A 56-year-old Japanese man underwent total gastrectomy with lymph-node dissection for a gastric tumor. In the oral rim of the excised large ulcerated tumor, a small nodular mass demonstrated a unique histology of a carcinosarcoma composed of more than three distinctive components. A major part of the large ulcerated tumor tissue consisted of conventional tubular adenocarcinoma, with the coincidental focal distribution of solid cell nests of poorly differentiated or neuroendocrine cell (small cell) carcinoma. Sarcoma cells were another of the constituents of the small nodular tumor with atypical spindle-shaped cells. All the excised lymph-node metastasis demonstrated the histology of tubular adenocarcinoma. In the sarcoma cells, many of the lineage-specific immunohistochemical markers were negative. Only vimentin was positive. Electron microscopy revealed that the neuroendocrine cell carcinoma contained cells with abundant small aggregations of neurosecretory-type granules in their cellular cytoplasm. Primitive cellular junctions and incomplete basal lamina were seen in the sarcoma cells. No transitional or hybrid-type cells were seen between carcinoma and sarcoma cells. The interfaces between the cancer cell nests and the proliferation of sarcoma cells are clearly discerned. It was speculated that the unique histology of carcinosarcoma might have progressed from the conventional type of pre-existing advanced tubular adenocarcinoma.

The median neurosecretory cells in abdominal ganglia of insects synthesize a number of putative hormones, which are abundant in the abdominal perisympathetic organs (PSOs). The peptide inventory of these prominent neurohemal release sites is best investigated in the American cockroach and strongly differs from that of head/thoracic neurohemal organs. In this study, we found a complete colocalization of all abundant neuropeptides in this hormonal system, including periviscerokinin-1 and -2, pyrokinin-5, YLSamide, VEAacid, and SKNacid. The first immunoreactive cells were detected on day 18 of embryonic development and already contained the complete set of peptides. By using antisera against the above-mentioned peptides, the development of this neurohormonal system could be studied and is described in detail. Subsequent electron microscopic immunogold stainings in PSO preparations revealed the costorage of PSO peptides in a single vesicle species. Surprisingly, all these peptides were found in axons containing clear vesicles, whereas all axons with dense core vesicles were totally devoid of immunoreactivity. Unlike the axons with dense core vesicles, immunostained axons ramify in the center of the PSO but exhibit only rare morphological signs of exocytosis. Instead, putative release sites of the clear vesicle-containing axons were detected peripherally to the PSOs, namely, on the hyperneural muscle., (Copyright 2002 Wiley-Liss, Inc.)

18 adrenocortical tumours and 5 pheochromocytomas were studied immunohistochemically. Expression of synaptophysin and chromogranin A was found in cells of cortical adenomas, "frontier" neoplasms and in 20-75% of carcinoma cells, this ultramicroscopically was confirmed by observation of typical neuroendocrine granules. Some groups of cells of cortico-medullary tumours also expressed proteins of neural differentiation (protein S-100).

Eighty-two mesenchymal tumors of the gastrointestinal tract were examined by electron microscopy for the purposes of subtyping for diagnostic precision and of understanding cellular differentiation. Tumors were subclassified into leiomyoma/leiomyosarcoma, tumors of the interstitial cell of Cajal (equivalent to traditionally defined GISTs [Miettinen et al. Hum Pathol. 1999; 30:1213-1220; Mod Pathol. 2000; 13:1134-1142]), gastrointestinal autonomic nerve tumors (GANTs), and fibroblastic and myofibroblastic tumors, using criteria from the literature. Leiomyoma/leiomyosarcoma were diagnosed by myofilaments, attachment plaques, plasmalemmal caveolae, and lamina; GIST by processes or cell bodies full of intermediate filaments, solitary focal densities amid intermediate filaments, attachment plaques with incomplete lamina, scarce myofilaments, and smooth endoplasmic reticulum; GANTs by neuroendocrine granules, cell bodies/processes full of intermediate filaments (more rarely microtubules), and smooth endoplasmic reticulum; fibroblastic/myofibroblastic tumors by abundant rough endoplasmic reticulum, myofilaments, and fibronexuses. Seventy-three tumors (89%) were successfully subclassified, as 5 leiomyoma/leiomyosarcoma (6%), 36 GISTs (44%), 22 GANTs (27%), 10 fibroblastic and myofibroblastic tumors (12%). Results indicated overlap between poorly differentiated leiomyosarcoma and GIST, and between GIST and GANT. GANT is emphasized as a neuronal tumor identifiable by electron microscopy, and thereby distinguishable from GIST.

Neuronal calcium sensor-1 (NCS-1) and its non-mammalian homologue, frequenin, have been implicated in a spectrum of cellular processes, including regulation of stimulated exocytosis of synaptic vesicles and secretory granules (SGs) in neurons and neuroendocrine cells and regulation of phosphatidylinositol 4-kinase beta activity in yeast. However, apart from these intriguing putative functions, NCS-1 and frequenin are relatively poorly understood. Here, the distribution, dynamics and function of NCS-1 were studied using PC12 cells that stably express NCS-1-EYFP (NCS-1 fused to enhanced yellow fluorescent protein) or that stably overexpress NCS-1. Fluorescence and electron microscopies show that NCS-1-EYFP is absent from SGs but is present on small clear organelles, some of which are just below the plasma membrane. Total internal reflection fluorescence microscopy shows that NCS-1-EYFP is associated with synaptic-like microvesicles (SLMVs) in growth cones. Overexpression studies show that NCS-1 enhances exocytosis of synaptotagmin-labeled regulated secretory organelles (RSOs) under basal conditions and during stimulation by UTP. Significantly, these studies implicate NCS-1 in the enhancement of both basal and stimulated phosphoinositide-dependent exocytosis of RSOs in PC12 cells, and they show that NCS-1 is distributed strategically to interact with putative targets on the plasma membrane and on SLMVs. These studies also reveal that SLMVs undergo both fast directed motion and highly hindered diffusive motion in growth cones, suggesting that cytoskeletal constituents can both facilitate and hinder SLMV motion. These results also reveal interesting similarities and differences between transport organelles in differentiated neuroendocrine cells and neurons.

Approximately 5% to 10% of paragangliomas occur in extra-adrenal sites, which can extend from the upper cervical region to the pelvis, parallel to the autonomic nervous system. This distribution corresponds to the embryologic development of the paraganglia from neural crest cells. Rarely, extra-adrenal paragangliomas can also occur aberrantly outside this distribution. We report such a case of extra-adrenal paraganglioma occurring in the anterior mesentery in a 76-year-old man. Two case reports exist in the literature describing extra-adrenal paragangliomas in the posterior mesentery. Normal paraganglionic tissue has been described at the roots of the superior and inferior mesenteric arteries, theoretically explaining the origin of the posterior mesenteric paragangliomas. Our case can best be attributed to the ventral migration of paraganglionic tissue through these vessels to reach the anterior mesentery, where they could potentially give rise to paragangliomas in this site.

Routing of membrane proteins to large dense core vesicles in neuroendocrine cells can depend on information in both the lumenal and cytoplasmic domains. This study in PC12 cells focuses on the routing, cleavage, and secretion of an integral membrane protein, peptidylglycine alpha-amidating monooxygenase (PAM), examining both endogenous and virally derived membrane PAM. The role of the lumenal catalytic domains in membrane PAM trafficking was examined by replacement with an epitope tag. Virally derived membrane PAM is localized to the perinuclear area and to slender processes where the large dense core vesicles are located. Expression of PAM along with a neuroendocrine-specific endoprotease liberates a soluble monooxygenase domain, yielding regulated secretion of both the monooxygenase and the prohormone convertase from large dense core vesicles. The subcellular distribution of the epitope-substituted version of PAM within the cells is similar to that of membrane PAM, and both proteins are internalized from the plasma membrane. When secretion is stimulated, Serine937 in the cytoplasmic domain of PAM is phosphorylated to a similar extent in endogenous membrane PAM, virally encoded membrane PAM, and epitope-substituted PAM. Thus, the lumenal PAM catalytic domains are not required for routing or phosphorylation of PAM in PC12 cells.

Two cases of the diffuse type of giant-cell tumor of the tendon sheath (GCTTS) are described. Both tumors arose in the vicinity of large joints of the lower extremity, showing similar clinical and radiological features. Histologically, a proliferation of polygonal mononuclear cells was seen, together with osteoclastlike giant cells, foam cells, and siderophages. The tumors were poorly delineated, displaying an infiltrative pattern into the neighboring soft tissues. Immunohistochemically, strong expression of vimentin, neuron-specific enolase, A1-antitrypsin, and CD68 was found in both mono- and multinucleated tumor cells. At the ultrastructural level, mononuclear cells revealed a diverse morphology, displaying features of histiocytelike and fibroblastlike cells, with the former being more numerous. Scarce neurosecretorylike granules, made up of electrondense membrane-bound material, were found in the cytoplasm of the mononuclear cells. Cytogenetic analysis of one case shows the presence of a clonal population with 47 chromosomes and two different translocations, t(2;3) and der(8) t(8;12). Present findings provide further support regarding the neoplasic nature of this tumoral entity.