Molecular profiling of sponge deflation reveals an ancient relaxant-inflammatory response.

A hallmark of animals is the coordination of whole-body movement. Neurons and muscles are central to this, yet coordinated movements also exist in sponges that lack these cell types. Sponges are sessile animals with a complex canal system for filter-feeding. They undergo whole-body movements resembling "contractions" that lead to canal closure and water expulsion. Here, we combine live 3D optical coherence microscopy, pharmacology, and functional proteomics to elucidate the sequence and detail of shape changes, the tissues and molecular physiology involved, and the control of these movements. Morphometric analysis and targeted perturbation suggest that the movement is driven by the relaxation of actomyosin stress fibers in epithelial canal cells, which leads to whole-body deflation via collapse of the incurrent and expansion of the excurrent canal system. Thermal proteome profiling and quantitative phosphoproteomics confirm the control of cellular relaxation by an Akt/NO/PKG/PKA pathway. Agitation-induced deflation leads to differential phosphorylation of proteins forming epithelial cell junctions, implying their mechanosensitive role. Unexpectedly, untargeted metabolomics detect a concomitant decrease in antioxidant molecules during deflation, reflecting an increase in reactive oxygen species. Together with the secretion of proteinases, cytokines, and granulin, this indicates an inflammation-like state of the deflating sponge reminiscent of vascular endothelial cells experiencing oscillatory shear stress. These results suggest the conservation of an ancient relaxant-inflammatory response of perturbed fluid-carrying systems in animals and offer a possible mechanism for whole-body coordination through diffusible paracrine signals and mechanotransduction.

Molecular profiling of sponge deflation reveals an ancient relaxant-inflammatory response.

A hallmark of animals is the coordination of whole-body movement. Neurons and muscles are central to this, yet coordinated movements also exist in sponges that lack these cell types. Sponges are sessile animals with a complex canal system for filter-feeding. They undergo whole-body movements resembling "contractions" that lead to canal closure and water expulsion. Here, we combine 3D optical coherence microscopy, pharmacology, and functional proteomics to elucidate anatomy, molecular physiology, and control of these movements. We find them driven by the relaxation of actomyosin stress fibers in epithelial canal cells, which leads to whole-body deflation via collapse of the incurrent and expansion of the excurrent system, controlled by an Akt/NO/PKG/A pathway. A concomitant increase in reactive oxygen species and secretion of proteinases and cytokines indicate an inflammation-like state reminiscent of vascular endothelial cells experiencing oscillatory shear stress. This suggests an ancient relaxant-inflammatory response of perturbed fluid-carrying systems in animals.

Body-Plan Reorganization in a Sponge Correlates with Microbiome Change.

Mounting evidence suggests that animals and their associated bacteria interact via intricate molecular mechanisms, and it is hypothesized that disturbances to the microbiome influence animal development. Here, we show that the loss of a key photosymbiont (i.e., bleaching) upon shading correlates with a stark body-plan reorganization in the common aquarium cyanosponge Lendenfeldia chondrodes. The morphological changes observed in shaded sponges include the development of a thread-like morphology that contrasts with the flattened, foliose morphology of control specimens. The microanatomy of shaded sponges markedly differed from that of control sponges, with shaded specimens lacking a well-developed cortex and choanosome. Also, the palisade of polyvacuolar gland-like cells typical in control specimens was absent in shaded sponges. The morphological changes observed in shaded specimens are coupled with broad transcriptomic changes and include the modulation of signaling pathways involved in animal morphogenesis and immune response, such as the Wnt, transforming growth factor ß (TGF-ß), and TLR-ILR pathways. This study provides a genetic, physiological, and morphological assessment of the effect of microbiome changes on sponge postembryonic development and homeostasis. The correlated response of the sponge host to the collapse of the population of symbiotic cyanobacteria provides evidence for a coupling between the sponge transcriptomic state and the state of its microbiome. This coupling suggests that the ability of animals to interact with their microbiomes and respond to microbiome perturbations has deep evolutionary origins in this group.

Profiling cellular diversity in sponges informs animal cell type and nervous system evolution.

The evolutionary origin of metazoan cell types such as neurons and muscles is not known. Using whole-body single-cell RNA sequencing in a sponge, an animal without nervous system and musculature, we identified 18 distinct cell types. These include nitric oxide­sensitive contractile pinacocytes, amoeboid phagocytes, and secretory neuroid cells that reside in close contact with digestive choanocytes that express scaffolding and receptor proteins. Visualizing neuroid cells by correlative x-ray and electron microscopy revealed secretory vesicles and cellular projections enwrapping choanocyte microvilli and cilia. Our data show a communication system that is organized around sponge digestive chambers, using conserved modules that became incorporated into the pre- and postsynapse in the nervous systems of other animals.

A new flow-regulating cell type in the Demosponge Tethya wilhelma - functional cellular anatomy of a leuconoid canal system.

Demosponges possess a leucon-type canal system which is characterized by a highly complex network of canal segments and choanocyte chambers. As sponges are sessile filter feeders, their aquiferous system plays an essential role in various fundamental physiological processes. Due to the morphological and architectural complexity of the canal system and the strong interdependence between flow conditions and anatomy, our understanding of fluid dynamics throughout leuconoid systems is patchy. This paper provides comprehensive morphometric data on the general architecture of the canal system, flow measurements and detailed cellular anatomical information to help fill in the gaps. We focus on the functional cellular anatomy of the aquiferous system and discuss all relevant cell types in the context of hydrodynamic and evolutionary constraints. Our analysis is based on the canal system of the tropical demosponge Tethya wilhelma, which we studied using scanning electron microscopy. We found a hitherto undescribed cell type, the reticuloapopylocyte, which is involved in flow regulation in the choanocyte chambers. It has a highly fenestrated, grid-like morphology and covers the apopylar opening. The minute opening of the reticuloapopylocyte occurs in an opened, intermediate and closed state. These states permit a gradual regulation of the total apopylar opening area. In this paper the three states are included in a theoretical study into flow conditions which aims to draw a link between functional cellular anatomy, the hydrodynamic situation and the regular body contractions seen in T. wilhelma. This provides a basis for new hypotheses regarding the function of bypass elements and the role of hydrostatic pressure in body contractions. Our study provides insights into the local and global flow conditions in the sponge canal system and thus enhances current understanding of related physiological processes.

The need for data standards in zoomorphology.

eScience is a new approach to research that focuses on data mining and exploration rather than data generation or simulation. This new approach is arguably a driving force for scientific progress and requires data to be openly available, easily accessible via the Internet, and compatible with each other. eScience relies on modern standards for the reporting and documentation of data and metadata. Here, we suggest necessary components (i.e., content, concept, nomenclature, format) of such standards in the context of zoomorphology. We document the need for using data repositories to prevent data loss and how publication practice is currently changing, with the emergence of dynamic publications and the publication of digital datasets. Subsequently, we demonstrate that in zoomorphology the scientific record is still limited to published literature and that zoomorphological data are usually not accessible through data repositories. The underlying problem is that zoomorphology lacks the standards for data and metadata. As a consequence, zoomorphology cannot participate in eScience. We argue that the standardization of morphological data requires i) a standardized framework for terminologies for anatomy and ii) a formalized method of description that allows computer-parsable morphological data to be communicable, compatible, and comparable. The role of controlled vocabularies (e.g., ontologies) for developing respective terminologies and methods of description is discussed, especially in the context of data annotation and semantic enhancement of publications. Finally, we introduce the International Consortium for Zoomorphology Standards, a working group that is open to everyone and whose aim is to stimulate and synthesize dialog about standards. It is the Consortium's ultimate goal to assist the zoomorphology community in developing modern data and metadata standards, including anatomy ontologies, thereby facilitating the participation of zoomorphology in eScience.

Deep metazoan phylogeny: when different genes tell different stories.

Molecular phylogenetic analyses have produced a plethora of controversial hypotheses regarding the patterns of diversification of non-bilaterian animals. To unravel the causes for the patterns of extreme inconsistencies at the base of the metazoan tree of life, we constructed a novel supermatrix containing 122 genes, enriched with non-bilaterian taxa. Comparative analyses of this supermatrix and its two non-overlapping multi-gene partitions (including ribosomal and non-ribosomal genes) revealed conflicting phylogenetic signals. We show that the levels of saturation and long branch attraction artifacts in the two partitions correlate with gene sampling. The ribosomal gene partition exhibits significantly lower saturation levels than the non-ribosomal one. Additional systematic errors derive from significant variations in amino acid substitution patterns among the metazoan lineages that violate the stationarity assumption of evolutionary models frequently used to reconstruct phylogenies. By modifying gene sampling and the taxonomic composition of the outgroup, we were able to construct three different yet well-supported phylogenies. These results show that the accuracy of phylogenetic inference may be substantially improved by selecting genes that evolve slowly across the Metazoa and applying more realistic substitution models. Additional sequence-independent genomic markers are also necessary to assess the validity of the phylogenetic hypotheses.

Cell death and renewal during prey capture and digestion in the carnivorous sponge Asbestopluma hypogea (Porifera: Poecilosclerida).

The sponge Asbestopluma hypogea is unusual among sponges due to its peculiar carnivorous feeding habit. During various stages of its nutrition cycle, the sponge is subjected to spectacular morphological modifications. Starved animals are characterized by many elongated filaments, which are crucial for the capture of prey. After capture, and during the digestion process, these filaments actively regress before being regenerated during a subsequent period of starvation. Here, we demonstrate that these morphological events rely on a highly dynamic cellular turnover, implying a coordinated sequence of programmed cell death (apoptosis and autophagy), cell proliferation and cell migration. A candidate niche for cell renewal by stem cell proliferation and differentiation was identified at the base of the sponge peduncle, characterized by higher levels of BrdU/EdU incorporation. Therefore, BrdU/EdU-positive cells of the peduncle base are candidate motile cells responsible for the regeneration of the prey-capturing main sponge body, i.e. the dynamic filaments. Altogether, our results demonstrate that dynamics of cell renewal in sponge appear to be regulated by cellular mechanisms as multiple and complex as those already identified in bilaterian metazoans.

Independent evolution of striated muscles in cnidarians and bilaterians.

Striated muscles are present in bilaterian animals (for example, vertebrates, insects and annelids) and some non-bilaterian eumetazoans (that is, cnidarians and ctenophores). The considerable ultrastructural similarity of striated muscles between these animal groups is thought to reflect a common evolutionary origin. Here we show that a muscle protein core set, including a type II myosin heavy chain (MyHC) motor protein characteristic of striated muscles in vertebrates, was already present in unicellular organisms before the origin of multicellular animals. Furthermore, 'striated muscle' and 'non-muscle' myhc orthologues are expressed differentially in two sponges, compatible with a functional diversification before the origin of true muscles and the subsequent use of striated muscle MyHC in fast-contracting smooth and striated muscle. Cnidarians and ctenophores possess striated muscle myhc orthologues but lack crucial components of bilaterian striated muscles, such as genes that code for titin and the troponin complex, suggesting the convergent evolution of striated muscles. Consistently, jellyfish orthologues of a shared set of bilaterian Z-disc proteins are not associated with striated muscles, but are instead expressed elsewhere or ubiquitously. The independent evolution of eumetazoan striated muscles through the addition of new proteins to a pre-existing, ancestral contractile apparatus may serve as a model for the evolution of complex animal cell types.

RNA interference in marine and freshwater sponges: actin knockdown in Tethya wilhelma and Ephydatia muelleri by ingested dsRNA expressing bacteria.

BACKGROUND: The marine sponge Tethya wilhelma and the freshwater sponge Ephydatia muelleri are emerging model organisms to study evolution, gene regulation, development, and physiology in non-bilaterian animal systems. Thus far, functional methods (i.e., loss or gain of function) for these organisms have not been available. RESULTS: We show that soaking developing freshwater sponges in double-stranded RNA and/or feeding marine and freshwater sponges bacteria expressing double-stranded RNA can lead to RNA interference and reduction of targeted transcript levels. These methods, first utilized in C. elegans, have been adapted for the development and feeding style of easily cultured marine and freshwater poriferans. We demonstrate phenotypic changes result from 'knocking down' expression of the actin gene. CONCLUSION: This technique provides an easy, efficient loss-of-function manipulation for developmental and gene regulatory studies in these important non-bilaterian animals.

The contractile sponge epithelium sensu lato--body contraction of the demosponge Tethya wilhelma is mediated by the pinacoderm.

Sponges constitute one of the two metazoan phyla that are able to contract their bodies despite a complete lack of muscle cells. Two competing hypotheses on the mechanisms behind this have been postulated to date: (1) mesohyl-mediated contraction originating from fusiform smooth muscle-like actinocytes ('myocytes') and (2) epidermal contraction originating in pinacocytes. No direct support exists for either hypothesis. The question of agonist-antagonist interaction in sponge contraction seems to have been completely neglected so far. In the present study we addressed this by studying sponge contraction kinetics. We also tested both hypotheses by carrying out volumetric studies of 3D synchrotron radiation-based x-ray microtomography data obtained from contracted and expanded specimens of Tethya wilhelma. Our results support the pinacoderm contraction hypothesis. Should mesohyl contraction be present, it is likely to be part of the antagonist system. We conclude that epithelial contraction plays a major role in sponges. Contractile epithelia sensu lato may be regarded as part of the ground pattern of the Metazoa.

Community-based partnered research: new directions in mental health services research.

OBJECTIVE: Community-based participatory research has the potential to improve implementation of best practices to reduce disparities but has seldom been applied in mental health services research. This article presents the content and lessons learned from a national conference designed to stimulate such an application. DESIGN: Mental health program developers collaborated in hosting a two-day conference that included plenary and break-out sessions, sharing approaches to community-academic partnership development, and preliminary findings from partnered research studies. Sessions were audiotaped, transcribed and analyzed by teams of academic and community conference participants to identify themes about best practices, challenges faced in partnered research, and recommendations for development of the field. Themes were illustrated with selections from project descriptions at the conference. SETTING AND PARTICIPANTS: Participants, representing 9 academic institutions and 12 community-based agencies from four US census regions, were academic and community partners from five research centers funded by the National Institute of Mental Health, and also included staff from federal and non-profit funding agencies. RESULTS: Five themes emerged: 1) Partnership Building; 2) Implementing and Supporting Partnered Research; 3) Developing Creative Dissemination Strategies; 4) Evaluating Impact; and 5) Training. CONCLUSIONS: Emerging knowledge of the factors in the partnership process can enhance uptake of new interventions in mental health services. Conference proceedings suggested that further development of this field may hold promise for improved approaches to address the mental health services quality chasm and service disparities.

Is veteran status and suicide risk assessed in community long-term care? A review of the states' assessment instruments.

Given recent policy initiatives to address suicide risk among older persons and veterans, community-based elder serving agencies may serve an important role in identifying and referring individuals at risk for suicide. A review of state-level long-term assessment instruments was conducted to determine whether veteran status and suicide are assessed. Data from forty-three state's Units on Aging instruments were content analyzed. Results indicate that over two thirds of the states in this review included questions about suicide and veterans in their assessments, 69.8% and 67.4% respectively. Suicide risk among elders and veterans must be addressed at local, state, and federal levels so that concerted attention and oversight can be provided for matching elders to the services they need.

Sponge budding is a spatiotemporal morphological patterning process: Insights from synchrotron radiation-based x-ray microtomography into the asexual reproduction of Tethya wilhelma.

BACKGROUND: Primary agametic-asexual reproduction mechanisms such as budding and fission are present in all non-bilaterian and many bilaterian animal taxa and are likely to be metazoan ground pattern characters. Cnidarians display highly organized and regulated budding processes. In contrast, budding in poriferans was thought to be less specific and related to the general ability of this group to reorganize their tissues. Here we test the hypothesis of morphological pattern formation during sponge budding. RESULTS: We investigated the budding process in Tethya wilhelma (Demospongiae) by applying 3D morphometrics to high resolution synchrotron radiation-based x-ray microtomography (SR-muCT) image data. We followed the morphogenesis of characteristic body structures and identified distinct morphological states which indeed reveal characteristic spatiotemporal morphological patterns in sponge bud development. We discovered the distribution of skeletal elements, canal system and sponge tissue to be based on a sequential series of distinct morphological states. Based on morphometric data we defined four typical bud stages. Once they have reached the final stage buds are released as fully functional juvenile sponges which are morphologically and functionally equivalent to adult specimens. CONCLUSION: Our results demonstrate that budding in demosponges is considerably more highly organized and regulated than previously assumed. Morphological pattern formation in asexual reproduction with underlying genetic regulation seems to have evolved early in metazoans and was likely part of the developmental program of the last common ancestor of all Metazoa (LCAM).

GABA and glutamate specifically induce contractions in the sponge Tethya wilhelma.

Sponges (Porifera) are nerve- and muscleless. Nevertheless, they react to external stimuli in a coordinated way, by body contraction, oscule closure or stopping pumping activity. The underlying mechanisms are still unknown, but evidence has been found for chemical messenger-based systems. We used the sponge Tethya wilhelma to test the effect of gamma-aminobutyric acid (GABA) and glutamate (L: -Glu) on its contraction behaviour. Minimal activating concentrations were found to be 0.5 microM (GABA) and 50 microM (L: -Glu), respectively. Taking maximum relative contraction speed and minimal relative projected body area as a measure of the sponge's response, a comparison of the dose-response curves indicated a higher sensitivity of the contractile tissue for GABA than for L: -Glu. The concentrations eliciting the same contractile response differ by about 100-fold more than the entire concentration range tested. In addition, desensitising effects and spasm-like reactions were observed. Presumably, a GABA/L: -Glu metabotropic receptor-based system is involved in the regulation of contraction in T. wilhelma. We discuss a coordination system for sponges based on hypothetical chemical messenger pathways.

Like a 'rolling stone': quantitative analysis of the body movement and skeletal dynamics of the sponge Tethya wilhelma.

Although sponges (Porifera) are basal Metazoa without muscles and a central nervous system, they are able to locomote, which is generally correlated to drastic morphological changes. This behaviour has been known for more almost 150 years, but it is only partly understood. The sponge T. wilhelma displays extraordinary movement and rhythmic body contractions, and is thus a valuable model for the investigation of sponge movement. The aims of the present study were to track T. wilhelma quantitatively on natural and artificial substrates, to test for a peristaltic movement mechanism and to check for the influence of morphological changes. T. wilhelma displays a unique mode of locomotion among sponges, without reorganizing the whole sponge body. The overall morphology was stable, and skeletal rotation during movement was shown; this is the first time that such movement has been demonstrated in a sponge. The stability of the skeletal superstructure arrangement during movement suggests that only the cortical tissue is involved in movement, with only local tissue rearrangements. The movement track followed a straight direction for long periods, but directions could be altered instantly. It is most likely that environmental conditions play an important roll in induction of movement. In summary, T. wilhelma resembles the proverbial ;rolling stone' that stays at a given location if the conditions are favourable and starts moving when conditions change for the worse.

Neuroactive substances specifically modulate rhythmic body contractions in the nerveless metazoon Tethya wilhelma (Demospongiae, Porifera).

BACKGROUND: Sponges (Porifera) are nerve- and muscleless metazoa, but display coordinated motor reactions. Therefore, they represent a valuable phylum to investigate coordination systems, which evolved in a hypothetical Urmetazoon prior to the central nervous system (CNS) of later metazoa. We have chosen the contractile and locomotive species Tethya wilhelma (Demospongiae, Hadromerida) as a model system for our research, using quantitative analysis based on digital time lapse imaging. In order to evaluate candidate coordination pathways, we extracorporeally tested a number of chemical messengers, agonists and antagonists known from chemical signalling pathways in animals with CNS. RESULTS: Sponge body contraction of T. wilhelma was induced by caffeine, glycine, serotonine, nitric oxide (NO) and extracellular cyclic adenosine monophosphate (cAMP). The induction by glycine and cAMP followed patterns varying from other substances. Induction by cAMP was delayed, while glycine lead to a bi-phasic contraction response. The frequency of the endogenous contraction rhythm of T. wilhelma was significantly decreased by adrenaline and NO, with the same tendency for cAMP and acetylcholine. In contrast, caffeine and glycine increased the contraction frequency. The endogenous rhythm appeared irregular during application of caffeine, adrenaline, NO and cAMP. Caffeine, glycine and NO attenuated the contraction amplitude. All effects on the endogenous rhythm were neutralised by the washout of the substances from the experimental reactor system. CONCLUSION: Our study demonstrates that a number of chemical messengers, agonists and antagonists induce contraction and/or modulate the endogenous contraction rhythm and amplitude of our nerveless model metazoon T. wilhelma. We conclude that a relatively complex system of chemical messengers regulates the contraction behaviour through auto- and paracrine signalling, which is presented in a hypothetical model. We assume that adrenergic, adenosynergic and glycinergic pathways, as well as pathways based on NO and extracellular cAMP are candidates for the regulation and timing of the endogenous contraction rhythm within pacemaker cells, while GABA, glutamate and serotonine are candidates for the direct coordination of the contractile cells.

Kinetics and rhythm of body contractions in the sponge Tethya wilhelma (Porifera: Demospongiae).

Sponges of the species Tethya wilhelma display rhythmic body contractions, which were analyzed by digital timelapse imaging and semi-automated image analysis. For the first time, differential, quantitative data on sponge behaviour could be obtained. The sponges are able to reduce their body volume by up to 73.3% during regular contractions. Each contraction cycle follows a characteristic pattern of four phases, permitting analysis of the kinetics of contraction and expansion. Long-term observations (for >7 days) reveal that the sponge contractions display a day-night periodicity in which contraction cycles are significantly longer during the dark hours. The contractions seem to be mediated by the pinacoderm; they are triggered locally and spread over the sponge surface at 12.5 microm s(-1). If two individuals of a clone are fused, the individual contraction rhythm of both sponges persists for several days, until a single new individual sponge is formed with a synchronized rhythm. The reported results and techniques establish T. wilhelma as a model organism for research on the development of aneural signal transduction and integration during early Metazoan evolution.

In vitro sponge fragment culture of Chondrosia reniformis (Nardo, 1847).

In vitro cultivation systems for sponges (Porifera) have to be developed to produce compounds of value in biotechnological processes. Organotypic culture attempts, which maintain or mimic the natural tissue structure, are promising ways towards a biotechnology of sponges. We used the Mediterranean species Chondrosia reniformis for sponge fragment in vitro cultivation. The species is common throughout the Mediterranean, easy to keep in aquariums and shows good recovery and regeneration after fragmentation. The regeneration process of the 50-80 mm(3) fragments lasted for several days and resulted in a rounded or ovoid body shape. The aquiferous system was reduced. Cells performed proliferation during the first weeks as we could demonstrate by 5-bromo-2'-deoxy-uridine (BrdU) incorporation. No proliferation could be demonstrated after a culture period of 3 months, but silicate uptake. Cellular density decreased with cultivation length, but collagen production increased. Fragments have been kept in culture up to 19 months. C. reniformis can be used as a model system to develop feeding strategies and evaluate the biotechnological potential of sponge fragment in vitro cultivation.