Epitheliomuscular Cells Research Articles

The Infection Route of Tomato Zonate Spot Virus in the Digestive System of Its Insect Vector Frankliniella occidentalis.

Tomato zonate spot virus (TZSV) is a phytopathogen of the genus Orthotospovirus (Bunyaviridae) that is widespread in many areas of Southwest China. TZSV is mainly transmitted by Frankliniella occidentalis, but its exact infection route remains unclear. To explore this issue, we detected the nucleocapsid protein of TZSV in the digestive systems of first-instar F. occidentalis nymphs fed with TZSV-infected pepper leaves. TZSV infection in the F. occidentalis digestive system begins within 4 h post-first access to diseased plants: The foregut is likely the primary site of infection, and primary salivary glands (PSGs) are the destination. There are three potential routes for TZSV transmission from the alimentary canal to the PSGs: (1) virus dissemination from the midgut to hemocoel followed by movement to the PSGs; (2) accumulation in midgut epithelial cells and arrival at PSGs via tubular salivary glands and efferent ducts; and (3) arrival at epitheliomuscular cells of the forepart of the midgut and movement along the ligament to the PSGs. We tested the transmission efficiency of F. occidentalis in second-instar nymphs and female and male adults. TZSV was transmitted in a persistent-propagative mode by both nymphs and adults, with adults appearing to show slightly higher transmission efficiency than nymphs. We confirmed the presence of all three routes for TZSV transmission in F. occidentalis and determined that like other Orthotospoviruses, TZSV is transmitted in a persistent-propagative manner. These results should facilitate the control of TZSV-related diseases and further our understanding of the transmission biology of Orthotospoviruses in general.

How to build single-celled cnidarians with worm-like motility: Lessons from Myxozoa.

Metazoans with worm-like morphologies across diverse and disparate groups typically demonstrate motility generated by hydrostatic skeletons involving tissue layers (muscles and epithelia). Here we present representative morphological, behavioural and molecular data for parasitic cnidarians (myxozoans) that demonstrate unprecedented variation in form and function, developing as cellular hydrostats. Motile elongate plasmodia characterise a remarkable radiation of species in the genus Ceratomyxa. The vermiform plasmodia inhabit gall bladders of a range of South American freshwater fish and exhibit undulatory motility reminiscent of nematodes but achieved at the cellular level. Collective insights from ultrastructure, confocal and light microscopy along with videos depicting movements highlight key features that we propose explain the unique motility of the plasmodia. These features include cytoskeletal elements (net forming microfilaments and microtubules), a large internal vacuole, a relatively rigid outer glycocalyx and peripherally arranged mitochondria. These constituents provide collective evidence for repurposing of the cnidarian epitheliomuscular cell to support worm-like motility at the cellular level. The apparent restriction of vermiform ceratomyxids to South American freshwaters suggests an origination via Cretaceous or Miocene marine transgressions and subsequent radiation.

Motor Evolution: Lit-Up Hydra Bare All

Whole-animal Hydra imaging shows that epitheliomuscular calcium influx dynamics and inter-cell progression speeds are very different for different behaviors. Hydra movements therefore likely arise from fast (ionotropic) and slow (metabotropic) neural mechanisms, and from interactions among the epitheliomuscular cells themselves.

Mapping the Whole-Body Muscle Activity of Hydra vulgaris

Hydra is a cnidarian polyp with an anatomically simple neuromuscular system that can offer evolutionary insights on the functional design of animal body plans. Using calcium imaging to map the activity of the entire epitheliomuscular system of behaving Hydra, we find seven basic spatiotemporal patterns of muscle activity. Patterns include global and local activation events with widely varying kinetics of initiation and wave-like propagation. The orthogonally oriented endodermal and ectodermal muscle fibers are jointly activated during longitudinal contractions. Individual epitheliomuscular cells can participate in multiple patterns, even with very different kinetics. This cellular multifunctionality could enable the structurally simple epitheliomuscular tissue of basal metazoans to implement a diverse behavioral output.

Mechanisms of cnidocyte development in the moon jellyfish Aurelia.

Stinging cells called cnidocytes are a defining trait of the cnidarians (sea anemones, corals, jellyfish, and their relatives). In hydrozoan cnidarians such as Hydra, cnidocytes develop from interstitial stem cells set aside in the ectoderm. It is less clear how cnidocytes develop outside the Hydrozoa, as other cnidarians appear to lack interstitial stem cells. We addressed this question by studying cnidogenesis in the moon jellyfish (Aurelia) through the visualization of minicollagen-a protein associated with cnidocyte development-as well as transmission electron microscopy. We discovered that developing cnidoblasts are rare or absent in feeding structures rich in mature cnidocytes, such as tentacles and lappets. Using transmission electron microscopy, we determined that the progenitors of cnidocytes have traits consistent with epitheliomuscular cells. Our data suggests a dynamic where cnidocytes develop at high concentrations in the epithelium of more proximal regions, and subsequently migrate to more distal regions where they exhibit high usage and turnover. Similar to some anthozoans, cnidocytes in Aurelia do not appear to be generated by interstitial stem cells; instead, epitheliomuscular cells appear to be the progenitor cell type. This observation polarizes the evolution of cnidogenesis, and raises the question of how interstitial stem cells came to regulate cnidogenesis in hydrozoans.

Neuronal photoactivation through second-harmonic near-infrared absorption by gold nanoparticles

Optical activation of neurons requires genetic manipulation or the use of chemical photoactivators with undesirable side effects. As a solution to these disadvantages, here, we demonstrate optically evoked neuronal activity in mouse cortical neurons in acute slices and in vivo by nonlinear excitation of gold nanoparticles. In addition, we use this approach to stimulate individual epitheliomuscular cells and evoke body contractions in Hydra vulgaris. To achieve this, we use a low-power pulsed near-infrared excitation at the double-wavelength of the plasmon resonance of gold nanoparticles, which enables optical sectioning and allows for high spatial precision and large penetration depth. The effect is explained by second-harmonic Mie scattering, demonstrating light absorption by a second-order nonlinear process, which enables photothermal stimulation of the cells. Our approach also minimizes photodamage, demonstrating a major advancement towards precise and harmless photoactivation for neuroscience and human therapeutics.

Hydra's feeding response: Effect of GABAB ligands on GSH-induced electrical activity in the hypostome of H. vulgaris

The feeding response in the Cnidarian, hydra, consists of mouth opening, tentacle writhing, and the cessation of pacemaker-controlled tentacle and body contractions. The behavior can be induced by reduced glutathione (GSH), contained in body fluids that leak from prey impaled by hydra's cnidocysts. Mouth, tentacle, and body-contraction behavior is carried out by hydra's ectodermal and endodermal epitheliomuscular cells. Here, we present the first evidence of GSH-induced electrical activity in the hypostome and its modification by GABA and GABAB ligands. The ‘heads’ of hydra were ablated and the tentacles removed. Suction electrodes, positioned on the mouth, recorded electrical activity produced by GSH, contained either within the electrode, or in the surrounding bath, the mouth being shielded. Recorded impulses were characterized, according to size and temporal pattern, as small, medium and (large) pacemaker impulses. GSH applied in the bath caused a frequency increase of small and medium impulses and a decrease in pacemaker bursts. The changes in frequencies of medium and pacemaker bursts, though not obviously affected by GABA, were counteracted by blocking GABAB inhibition with phaclofen. Only the highest concentration of GSH applied at the mouth potentially decreased pacemaker frequency and potentially increased medium impulses, without affecting small impulses. GABA caused a significant increase in small and medium impulses relative to GSH which was counteracted by baclofen and/or baclofen plus phaclofen. The results indicate that considerable GSH-receptor circuitry is located in hypostomal tissue proximal to hydra's mouth, and substantiate GABA and GABAB inhibition within the neuroeffector network of the feeding response.

Apical and basal epitheliomuscular F-actin dynamics duringHydrabud evagination

ABSTRACTBending of 2D cell sheets is a fundamental morphogenetic mechanism during animal development and reproduction. A critical player driving cell shape during tissue bending is the actin cytoskeleton. Much of our current knowledge about actin dynamics in whole organisms stems from studies of embryonic development in bilaterian model organisms. Here, we have analyzed actin-based processes during asexual bud evagination in the simple metazoan Hydra. We created transgenic Hydra strains stably expressing the actin marker Lifeact-GFP in either ectodermal or endodermal epitheliomuscular cells. We then combined live imaging with conventional phalloidin staining to directly follow actin reorganization. Bending of the Hydra epithelial double layer is initiated by a group of epitheliomuscular cells in the endodermal layer. These cells shorten their apical-basal axis and arrange their basal muscle processes in a circular configuration. We propose that this rearrangement generates the initial forces to bend the endoderm towards the ectoderm. Convergent tissue movement in both epithelial layers towards the centre of evagination then leads to elongation and extension of the bud along its new body axis. Tissue movement into the bud is associated with lateral intercalation of epithelial cells, remodelling of apical septate junctions, and rearrangement of basal muscle processes. The work presented here extends the analysis of morphogenetic mechanisms beyond embryonic tissues of model bilaterians.

Sequential development of apical-basal and planar polarities in aggregating epitheliomuscular cells of Hydra

Apical-basal and planar cell polarities are hallmarks of metazoan epithelia required to separate internal and external environments and to regulate trans- and intracellular transport, cytoskeletal organization, and morphogenesis. Mechanisms of cell polarization have been intensively studied in bilaterian model organisms, particularly in early embryos and cultured cells, while cell polarity in pre-bilaterian tissues is poorly understood. Here, we have studied apical-basal and planar polarization in regenerating (aggregating) clusters of epitheliomuscular cells of Hydra, a simple representative of the ancestral, pre-bilaterian phylum Cnidaria. Immediately after dissociation, single epitheliomuscular cells do not exhibit cellular polarity, but they polarize de novo during aggregation. Reestablishment of the Hydra-specific epithelial bilayer is a result of short-range cell sorting. In the early phase of aggregation, apical-basal polarization starts with an enlargement of the epithelial apical-basal diameter and by the development of belt-like apical septate junctions. Specification of the basal pole of epithelial cells occurs shortly later and is linked to synthesis of mesoglea, development of hemidesmosome-like junctions, and formation of desmosome-like junctions connecting the basal myonemes of neighbouring cells. Planar polarization starts, while apical-basal polarization is already ongoing. It is executed gradually starting with cell-autonomous formation, parallelization, and condensation of myonemes at the basal end of each epithelial cell and continuing with a final planar alignment of epitheliomuscular cells at the tissue level. Our findings reveal that epithelial polarization in Hydra aggregates occurs in defined steps well accessible by histological and ultrastructural techniques and they will provide a basis for future molecular studies.

Loss of neurogenesis in Hydra leads to compensatory regulation of neurogenic and neurotransmission genes in epithelial cells.

Hydra continuously differentiates a sophisticated nervous system made of mechanosensory cells (nematocytes) and sensory–motor and ganglionic neurons from interstitial stem cells. However, this dynamic adult neurogenesis is dispensable for morphogenesis. Indeed animals depleted of their interstitial stem cells and interstitial progenitors lose their active behaviours but maintain their developmental fitness, and regenerate and bud when force-fed. To characterize the impact of the loss of neurogenesis in Hydra, we first performed transcriptomic profiling at five positions along the body axis. We found neurogenic genes predominantly expressed along the central body column, which contains stem cells and progenitors, and neurotransmission genes predominantly expressed at the extremities, where the nervous system is dense. Next, we performed transcriptomics on animals depleted of their interstitial cells by hydroxyurea, colchicine or heat-shock treatment. By crossing these results with cell-type-specific transcriptomics, we identified epithelial genes up-regulated upon loss of neurogenesis: transcription factors (Dlx, Dlx1, DMBX1/Manacle, Ets1, Gli3, KLF11, LMX1A, ZNF436, Shox1), epitheliopeptides (Arminins, PW peptide), neurosignalling components (CAMK1D, DDCl2, Inx1), ligand-ion channel receptors (CHRNA1, NaC7), G-Protein Coupled Receptors and FMRFRL. Hence epitheliomuscular cells seemingly enhance their sensing ability when neurogenesis is compromised. This unsuspected plasticity might reflect the extended multifunctionality of epithelial-like cells in early eumetazoan evolution.

The comprehensive analysis of DEG/ENaC subunits in Hydra reveals a large variety of peptide-gated channels, potentially involved in neuromuscular transmission.

BackgroundIt is generally the case that fast transmission at neural synapses is mediated by small molecule neurotransmitters. The simple nervous system of the cnidarian Hydra, however, contains a large repertoire of neuropeptides and it has been suggested that neuropeptides are the principal transmitters of Hydra. An ion channel directly gated by Hydra-RFamide neuropeptides has indeed been identified in Hydra – the Hydra Na+ channel (HyNaC) 2/3/5, which is expressed at the oral side of the tentacle base. Hydra-RFamides are more widely expressed, however, being found in neurons of the head and peduncle region. Here, we explore whether further peptide-gated HyNaCs exist, where in the animal they are expressed, and whether they are all gated by Hydra-RFamides.ResultsWe report molecular cloning of seven new HyNaC subunits – HyNaC6 to HyNaC12, all of which are members of the DEG/ENaC gene family. In Xenopus oocytes, these subunits assemble together with the four already known subunits into thirteen different ion channels that are directly gated by Hydra-RFamide neuropeptides with high affinity (up to 40 nM). In situ hybridization suggests that HyNaCs are expressed in epitheliomuscular cells at the oral and the aboral side of the tentacle base and at the peduncle. Moreover, diminazene, an inhibitor of HyNaCs, delayed tentacle movement in live Hydra.ConclusionsOur results show that Hydra has a large variety of peptide-gated ion channels that are activated by a restricted number of related neuropeptides. The existence and expression pattern of these channels, and behavioral effects induced by channel blockers, suggests that Hydra co-opted neuropeptides for fast neuromuscular transmission.Electronic supplementary materialThe online version of this article (doi:10.1186/s12915-014-0084-2) contains supplementary material, which is available to authorized users.

Development and epithelial organisation of muscle cells in the sea anemone Nematostella vectensis.

IntroductionNematostella vectensis, a member of the cnidarian class Anthozoa, has been established as a promising model system in developmental biology, but while information about the genetic regulation of embryonic development is rapidly increasing, little is known about the cellular organization of the various cell types in the adult. Here, we studied the anatomy and development of the muscular system of N. vectensis to obtain further insights into the evolution of muscle cells.ResultsThe muscular system of N. vectensis is comprised of five distinct muscle groups, which are differentiated into a tentacle and a body column system. Both systems house longitudinal as well as circular portions. With the exception of the ectodermal tentacle longitudinal muscle, all muscle groups are of endodermal origin. The shape and epithelial organization of muscle cells vary considerably between different muscle groups. Ring muscle cells are formed as epitheliomuscular cells in which the myofilaments are housed in the basal part of the cell, while the apical part is connected to neighboring cells by apical cell-cell junctions. In the longitudinal muscles of the column, the muscular part at the basal side is connected to the apical part by a long and narrow cytoplasmic bridge. The organization of these cells, however, remains epitheliomuscular. A third type of muscle cell is represented in the longitudinal muscle of the tentacle. Using transgenic animals we show that the apical cell-cell junctions are lost during differentiation, resulting in a detachment of the muscle cells to a basiepithelial position. These muscle cells are still located within the epithelium and outside of the basal matrix, therefore constituting basiepithelial myocytes. We demonstrate that all muscle cells, including the longitudinal basiepithelial muscle cells of the tentacle, initially differentiate from regular epithelial cells before they alter their epithelial organisation.ConclusionsA wide range of different muscle cell morphologies can already be found in a single animal. This suggests how a transition from an epithelially organized muscle system to a mesenchymal could have occurred. Our study on N. vectensis provides new insights into the organisation of a muscle system in a non-bilaterian organism.

The comprehensive analysis of DEG/ENaC subunits in Hydra reveals a large variety of peptide-gated channels, potentially involved in neuromuscular transmission

It is generally the case that fast transmission at neural synapses is mediated by small molecule neurotransmitters. The simple nervous system of the cnidarian Hydra, however, contains a large repertoire of neuropeptides and it has been suggested that neuropeptides are the principal transmitters of Hydra. An ion channel directly gated by Hydra-RFamide neuropeptides has indeed been identified in Hydra - the Hydra Na+ channel (HyNaC) 2/3/5, which is expressed at the oral side of the tentacle base. Hydra-RFamides are more widely expressed, however, being found in neurons of the head and peduncle region. Here, we explore whether further peptide-gated HyNaCs exist, where in the animal they are expressed, and whether they are all gated by Hydra-RFamides. We report molecular cloning of seven new HyNaC subunits - HyNaC6 to HyNaC12, all of which are members of the DEG/ENaC gene family. In Xenopus oocytes, these subunits assemble together with the four already known subunits into thirteen different ion channels that are directly gated by Hydra-RFamide neuropeptides with high affinity (up to 40 nM). In situ hybridization suggests that HyNaCs are expressed in epitheliomuscular cells at the oral and the aboral side of the tentacle base and at the peduncle. Moreover, diminazene, an inhibitor of HyNaCs, delayed tentacle movement in live Hydra. Our results show that Hydra has a large variety of peptide-gated ion channels that are activated by a restricted number of related neuropeptides. The existence and expression pattern of these channels, and behavioral effects induced by channel blockers, suggests that Hydra co-opted neuropeptides for fast neuromuscular transmission.

Histological staining approaches for high-quality imaging of Actinia equina and Anemonia sulcata anatomy

Sea anemones species identification of can be difficult, especially in the field. The taxonomic keys currently utilised are based mainly on histological differences and therefore require collection of whole animals. Furthermore, histological analysis of sea anemones requires considerable expertise and worldwide sea anemones histological descriptions are still limited. Here it is reported a detailed histological description of the internal structure of the two most conspicuous species in tidal pool communities in European shores, Actinia equina and Anemonia sulcata. This species description makes them excellent models for studying reproductive strategies that are instrumental in establishing and maintaining new populations. Therefore they are potential indicator species important both for micro and macro scale monitoring programs. The anatomy and histology of main organs of the studied sea anemones were described. The different colour morphs of the species were studied. Fine histological details of their muscular and digestive systems and appendages, down to the cellular level, were studied. They are two-germinal layer animals and possess radial symmetry in body form. Samples of the main organs of sea anemones including epidermis, tentacles, mesentery, gonads and digestive tract (mouth, stomodaeum and gastrovascular cavity) were processed for histological examination according to procedures described elsewhere [1,2]. The epithelia were composed of epithelio-muscular cells, in which many gland cells were distributed. Three types of cnidoblasts (nematooysts) were observed in the epithelioma of the tentacles, mesentery filament and stomodaeum. Cnidoblasts in different places have different functions. The gonads were of the follicular type and were composed of germ cells and nutritional cells. The investigation of the internal organization of invertebrate communities provides important information on the organism’s biology with special interest for the study of ecological processes to evaluate their reproductive modes in order to understand how they may reflect their dispersal strategies [1, 2]. Traits that influence reproduction in animal and plant species have been a central focus of evolutionary biology. Therefore, considering their basal position in evolution, the Anthozoa (Cnidaria) provide important models for further understanding processes affecting reproductive strategies in the eumetazoan (i.e., cnidarian-bilaterian) ancestor and in modern vertebrates.This work was supported by the Portuguese Foundation for the Science and Technology – Portugal and FEDER funds, through the Projects: PTDC/MAR/464729/2006 and FCT/CNPq (Brazil), Project 6818, Programme 19/ 004.

FoxO is a critical regulator of stem cell maintenance in immortal Hydra

Immortality has always been an issue of curiosity among scientific community. Hydra's indefinite self renewal capacity of stem cells has attracted natural scientists to seek insights about longevity studies in humans. Three stem cell lineages ectodermal and endodermal epitheliomuscular stem cells and interstitial stem cells contribute to indefinite self renewal capacity of epithelial cells. Moreover, forkhead boxO (foxO) which plays a key role in increased life span in lower creatures like flies and worms has shown to have a strong association of FoxO3a gene in human longevity and maintainence of adult haematopoitic stem cells and neural stem cells. doi : 10.5214/ans.0972.7531.200107

Identification and characterization of VEGF and FGF from Hydra

Vascular endothelial growth factor (VEGF) and fibroblast growth factor (FGF) play important roles in the formation of the blood vascular system and in axon guidance, nervous system development and function. Here, we report isolation and characterization of VEGF and FGF homologues from Hydra vulgaris Ind-Pune, a Cnidarian which exhibits an organized nervous system and primitive epithelio-muscular cells. VEGF expression was prominent in the endoderm of the peduncle region and tentacles, as evident from in situ hybridization of whole polyps and its transverse sections. High levels of FGF were detected in the ectoderm of the budding region. The expression of VEGF in endodermal and FGF in interstitial cells was confirmed using sf-1 hydra, a temperature-sensitive mutant strain of Hydra magnipapillata. Tissue-specific expression of VEGF and FGF was confirmed by semi quantitative RT-PCR for ectodermal and endodermal tissues in H. vulgaris Ind-Pune. Treatment with SU5416, a specific inhibitor of the VEGF receptor, did not affect the whole polyp, but did delay both budding and head regeneration, suggesting a possible role of VEGF in nerve cell development, tube formation and/or in branching. FGF expression in the ectoderm of budding region, where the majority of interstitial stem cells reside suggests its role in interstitial stem cell maintenance. Further, activation of canonical Wnt signalling with the glycogen synthase kinase-3β (GSK-3β) inhibitor alsterpaullone caused down-regulation of VEGF and FGF, suggesting an antagonistic relationship between the Wnt and VEGF/FGF pathways. Our results indicate that VEGF and FGF evolved early in evolution, before the development of the blood vascular system, and open up the possibility of elucidating the evolutionarily ancient functions of VEGF and FGF.

Development and myogenesis of the vermiform Buddenbrockia (Myxozoa) and implications for cnidarian body plan evolution

BackgroundThe enigmatic wormlike parasite Buddenbrockia plumatellae has recently been shown to belong to the Myxozoa, which are now supported as a clade within Cnidaria. Most myxozoans are morphologically extremely simplified, lacking major metazoan features such as epithelial tissue layers, gut, nervous system, body axes and gonads. This hinders comparisons to free-living cnidarians and thus an understanding of myxozoan evolution and identification of their cnidarian sister group. However, B. plumatellae is less simplified than other myxozoans and therefore is of specific significance for such evolutionary considerations.MethodsWe analyse and describe the development of major body plan features in Buddenbrockia worms using a combination of histology, electron microscopy and confocal microscopy.ResultsEarly developmental stages develop a primary body axis that shows a polarity, which is manifested as a gradient of tissue development, enabling distinction between the two worm tips. This polarity is maintained in adult worms, which, in addition, often develop a pore at the distal tip. The musculature comprises tetraradially arranged longitudinal muscle blocks consisting of independent myocytes embedded in the extracellular matrix between inner and outer epithelial tissue layers. The muscle fibres are obliquely oriented and in fully grown worms consistently form an angle of 12° with respect to the longitudinal axis of the worm in each muscle block and hence confer chirality. Connecting cells form a link between each muscle block and constitute four rows of cells that run in single file along the length of the worm. These connecting cells are remnants of the inner epithelial tissue layer and are anchored to the extracellular matrix. They are likely to have a biomechanical function.ConclusionsThe polarised primary body axis represents an ancient feature present in the last common ancestor of Cnidaria and Bilateria. The tetraradial arrangement of musculature is consistent with a medusozoan affinity for Myxozoa. However, the chiral pattern of muscle fibre orientation is apparently novel within Cnidaria and could thus be a specific adaptation. The presence of independent myocytes instead of Cnidaria-like epitheliomuscular cells can be interpreted as further support for the presence of mesoderm in cnidarians, or it may represent convergent evolution to a bilaterian condition.

Stemness in Hydra - a current perspective

Hydra is a classic and simple model for pattern formation and regeneration research. More recently, it has also been promoted as a model to study ancestral stem cell biology. Three independent cell lineages form the body of the polyp and exhibit characteristics of stem cell systems. In order to define differences in stemness between the ectodermal and endodermal epitheliomuscular cell lineages and the interstitial cell lineage, we compare cellular properties and decision making. We argue that these three lineages are expected to show substantial variation in their stemness-related gene regulatory networks. Finally, we discuss Wnt signalling pathways and Myc oncoproteins, which are beginning to offer a perspective on how proliferation and differentiation might be regulated.

GABA and glutamate immunoreactivity in tentacles of the sea anemone Phymactis papillosa (LESSON 1830)

Sea anemones have a structurally simple nervous system that controls behaviors like feeding, locomotion, aggression, and defense. Specific chemical and tactile stimuli are transduced by ectodermal sensory cells and transmitted via a neural network to cnidocytes and epithelio-muscular cells, but the nature of the neurotransmitters operating in these processes is still under discussion. Previous studies demonstrated an important role of peptidergic transmission in cnidarians, but during the last decade the contribution of conventional neurotransmitters became increasingly evident. Here, we used immunohistochemistry on light and electron microscopical preparations to investigate the localization of glutamate and GABA in tentacle cross-sections of the sea anemone Phymactis papillosa. Our results demonstrate strong glutamate immunoreactivity in the nerve plexus, while GABA labeling was most prominent in the underlying epithelio-muscular layer. Immunoreactivity for both molecules was also found in glandular epithelial cells, and putative sensory cells were GABA positive. Under electron microscopy, both glutamate and GABA immunogold labeling was found in putative neural processes within the neural plexus. These data support a function of glutamate and GABA as signaling molecules in the nervous system of sea anemones.

Nucleolar binding of an anti‐NMDA receptor antibody in hydra: a non‐canonical role for an NMDA receptor protein?

Two emerging concepts in cell biology are the back-and-forth trafficking of receptor proteins and nuclear transcription factors between the nucleus and the cell membrane, and the alternative splicing of messenger RNA to produce similar proteins targeted to different cell sites. Recent evidence suggests that the nucleolus is a dynamic structure whose components may be involved in both types of trafficking. In the nervous system of higher animals, the N-methyl-D-aspartate (NMDA)-specific glutamate receptor has various roles in development and cell communication. It is involved in learning, memory, axonal guidance and nerve regeneration. We have reported earlier that the NR1 subunit of the NMDA receptor is present in the cell periphery and the nucleus of stem cells, neurons and epitheliomuscular cells of the early-evolved cnidarian, Hydra vulgaris (Scappaticci et al., 2004. Cell Tissue Res 316:263-270); it is involved in coordinating hydra's neuroeffector systems (Kass-Simon and Scappaticci, 2003. Hydrobiologia 530/531:67-71; Pierobon et al., 2004. Eur J Neurosci 20:2598-2604; Scappaticci and Kass-Simon, 2008. Comp Biochem Physiol A 150:415-422; Kay and Kass-Simon, 2009. Bio Bull 216:113-129). Here we report immunocytochemical experiments, using a mouse monoclonal antibody raised against the mammalian NR1 receptor subunit, and an in silico genomic and gene expression analysis identifying the homologues in hydra of mammalian NR1 and fibrillarin (FBL) genes, and their expressed proteins. The experiments reveal that the NR1 antibody specifically labels the nucleoli of large and small interstitial cells (stem cells), nematoblasts, neuroblasts, and epitheliomusclar cells; antibody labeling of the nucleolar marker, FBL, confirms the nucleolar localization of NR1 antibody labeling. Genomic analysis reveals that NR1 and FBL genes are conserved in hydra, and suggests that there are at least two NR1 splice variants, one of which contains both nuclear and nucleolar targeting signals. The finding that an NR1 receptor subunit (or a portion of it) appears in nucleoli of hydra cells is unique, and has not been reported for any other organism. Its presence in nucleoli of hydra may signal the existence of a yet-undescribed shuttle mechanism between the cell surface and the nucleous, or the alternative deployment of NR1 splice variants to different cell sites.