Selective concentration of iron, titanium, and zirconium substrate minerals within Gregory's diverticulum, an organ unique to derived sand dollars (Echinoidea: Scutelliformes).

Gregory's diverticulum, a digestive tract structure unique to a derived group of sand dollars (Echinoidea: Scutelliformes), is filled with sand grains obtained from the substrate the animals inhabit. The simple methods of shining a bright light through a specimen or testing response to a magnet can reveal the presence of a mineral-filled diverticulum. Heavy minerals with a specific gravity of >2.9 g/cm3 are selectively concentrated inside the organ, usually at concentrations one order of magnitude, or more, greater than found in the substrate. Analyses of diverticulum content for thirteen species from nine genera, using optical mineralogy, powder X-ray diffraction, scanning electron microscopy and energy dispersive X-ray spectroscopy, as well as micro-computed tomography shows the preference for selection of five major heavy minerals: magnetite (Fe3O4), hematite (Fe2O3), ilmenite (FeTiO3), rutile (TiO2), and zircon (ZrSiO4). Minor amounts of heavy or marginally heavy amphibole, pyroxene and garnet mineral grains may also be incorporated. In general, the animals exhibit a preference for mineral grains with a specific gravity of >4.0 g/cm3, although the choice is opportunistic and the actual mix of mineral species depends on the mineral composition of the substrate. The animals also select for grain size, with mineral grains generally in the range of 50 to 150 µm, and do not appear to alter this preference during ontogeny. A comparison of analytical methods demonstrates that X-ray attenuation measured using micro-computed tomography is a reliable non-destructive method for heavy mineral quantification when supported by associated analyses of mineral grains extracted destructively from specimens or from substrate collected together with the specimens. Commonalities in the electro-chemical surface properties of the ingested minerals suggest that such characteristics play an important role in the selection process.

Ion Mobility-Based Enrichment-Free N-Terminomics Analysis Reveals Novel Legumain Substrates in Murine Spleen.

Aberrant levels of the asparaginyl endopeptidase legumain have been linked to inflammation, neurodegeneration, and cancer, yet our understanding of this protease is incomplete. Systematic attempts to identify legumain substrates have been previously confined to in vitro studies, which fail to mirror physiological conditions and obscure biologically relevant cleavage events. Using high-field asymmetric waveform ion mobility spectrometry (FAIMS), we developed a streamlined approach for proteome and N-terminome analyses without the need for N-termini enrichment. Compared to unfractionated proteomic analysis, we demonstrate FAIMS fractionation improves N-termini identification by >2.5 fold, resulting in the identification of >2882 unique N-termini from limited sample amounts. In murine spleens, this approach identifies 6366 proteins and 2528 unique N-termini, with 235 cleavage events enriched in WT compared to legumain-deficient spleens. Among these, 119 neo-N-termini arose from asparaginyl endopeptidase activities, representing novel putative physiological legumain substrates. The direct cleavage of selected substrates by legumain was confirmed using in vitro assays, providing support for the existence of physiologically relevant extra-lysosomal legumain activity. Combined, these data shed critical light on the functions of legumain and demonstrate the utility of FAIMS as an accessible method to improve depth and quality of N-terminomics studies.

Deep-sea benthic crustacean and annelid data from the Bering Sea.

Samples of Crustacea and Annelida (Polychaeta, Sipuncula, and Hirudinea) were collected in the Bering Sea and the northwestern Pacific Ocean during scientific cruise SO-249 BERING in 2016. Biological samples were collected from 32 locations by the team on-board RV Sonne using a chain bag dredge at depths ranging between 330-5,070 m, and preserved in 96% ethanol. Specimens were morphologically identified to the lowest taxonomic level possible using a Leica M60 stereomicroscope. The generated data here comprise taxonomic information as well as annotated bathymetric and biogeographic information from a total of 78 samples (26 Crustacea, 47 Polychaeta, 4 Sipuncula, and 1 Hirudinea). The dataset was prepared following Darwin Core Biodiversity standards for FAIR data sharing based on Ocean Biodiversity Information System (OBIS) and Global Biodiversity Facility (GBIF) guidelines. The standardised digitised data were then mobilised to both OBIS and GBIF under CC BY 4.0 licence to publicly share and adopt the data. As records of these important marine taxa from bathyal and abyssal depths are sparse, especially from the deep Bering Sea, the herein generated and digitised data aid in filling existing knowledge gaps on their diversity and distribution in that region. As part of the "Biogeography of the NW Pacific deep-sea fauna and their possible future invasions into the Arctic Ocean" (BENEFICIAL) project, this dataset thus not only increases our knowledge in re-assessing and uncovering the deep-sea diversity of these taxa, but also serves policy and management sectors by providing first-hand data for global report assessments.

Holistic description of new deep sea megafauna (Cephalopoda: Cirrata) using a minimally invasive approach.

BACKGROUND: In zoology, species descriptions conventionally rely on invasive morphological techniques, frequently leading to damage of the specimens and thus only a partial understanding of their structural complexity. More recently, non-destructive imaging techniques have successfully been used to describe smaller fauna, but this approach has so far not been applied to identify or describe larger animal species. Here, we present a combination of entirely non-invasive as well as minimally invasive methods that permit taxonomic descriptions of large zoological specimens in a more comprehensive manner. RESULTS: Using the single available representative of an allegedly novel species of deep-sea cephalopod (Mollusca: Cephalopoda), digital photography, standardized external measurements, high-field magnetic resonance imaging, micro-computed tomography, and DNA barcoding were combined to gather all morphological and molecular characters relevant for a full species description. The results show that this specimen belongs to the cirrate octopod (Octopoda: Cirrata) genus Grimpoteuthis Robson, 1932. Based on the number of suckers, position of web nodules, cirrus length, presence of a radula, and various shell characters, the specimen is designated as the holotype of a new species of dumbo octopus, G. imperator sp. nov. The digital nature of the acquired data permits a seamless online deposition of raw as well as derived morphological and molecular datasets in publicly accessible repositories. CONCLUSIONS: Using high-resolution, non-invasive imaging systems intended for the analysis of larger biological objects, all external as well as internal morphological character states relevant for the identification of a new megafaunal species were obtained. Potentially harmful effects on this unique deep-sea cephalopod specimen were avoided by scanning the fixed animal without admixture of a contrast agent. Additional support for the taxonomic placement of the new dumbo octopus species was obtained through DNA barcoding, further underlining the importance of combining morphological and molecular datasets for a holistic description of zoological specimens.

Deep regional hyperthermia with preoperative radiochemotherapy in locally advanced rectal cancer, a prospective phase II trial.

PURPOSE: The goal of the present study was to investigate the effect of deep regional hyperthermia on early and long-term oncological outcomes in the context of preoperative radiochemotherapy in rectal cancer. METHODS: In this prospective phase II trial, patients with locally advanced rectal cancer were treated with 5-fluorouracil based preoperative radiochemotherapy with 50.4 Gy in 28 fractions. Deep regional hyperthermia was scheduled twice weekly. Pathological tumor regression was scored according to the Dworak regression system. The primary endpoint was pathological complete response (pCR). Further endpoints were local control (LC), distant control (DC), disease-free survival (DFS) and overall survival (OS). Hyperthermia was defined as feasible if 70% of patients received at least eight treatments. Quality of life was assessed at follow-up by the EORTC-QLQ-C30 and QLQ-CR29 questionnaires. Time to event data was analyzed according to Kaplan-Meier based on first-events. The study was registered on clinicaltrials.gov (NCT02353858). RESULTS: From 2012 until 2017, 78 patients were recruited. Median follow-up was 54 months. Based on magnetic resonance imaging, the mesorectal fascia was involved or threatened in 60% of the patients. Compliance with radiotherapy was 99%, 91% received both cycles of chemotherapy and 77% had eight or more hyperthermia treatments. Median time from the end of radiotherapy to surgery was 6.7 weeks. A pathological complete response was reported in 14% of the patients, 50% had either Dworak 4 (complete regression) or Dworak 3 regression (scattered tumor cells only). Three year estimates for OS, DFS, LC and DC were 94%, 81%, 96% and 87%. Patients with higher hyperthermia related cumulative temperatures showed stronger tumor regression. Global health status based on EORTC-QLQ-C30 was comparable with data from the general population. CONCLUSION: Deep regional hyperthermia was feasible, did not compromise standard treatments and resulted in promising long-term oncological outcomes and QoL.

Effect of Pacifier Design on Nonnutritive Suck Maturation and Weight Gain in Preterm Infants: A Pilot Study.

BACKGROUND: Pacifiers are effective in promoting oral feeding by increasing the maturation of nonnutritive sucking to nutritive suck in preterm neonates. It is unclear whether pacifier design can influence suck dynamics and weight loss during the first week of life. OBJECTIVES: This pilot study examined the feasibility of studying the effect of pacifier design on suck maturation and weight loss in preterm neonates. METHODS: Twenty-five preterm neonates (mean [SD] birth weight 1791 [344.9] grams, mean [SD] gestational age 33.1 [1.2] weeks) were studied in a single newborn intensive care unit. Neonates were assigned to either an orthodontic pacifier (n = 13) or a bulb-shaped pacifier (n = 12) immediately after birth. Suck dynamics (cycles per minute, total compressions per minute, cycle bursts, and amplitude) were assessed with an NTrainer (Innara Health, Olathe, Kansas). Weight was recorded during the first week of life on day 1.2 (±2.5 days) and day 6.0 (±2.1 days). Descriptive statistics were applied to analyze data. RESULTS: No significant differences were seen between groups with respect to birth weight and gestational age. Reproducible nonnutritive sucking measurements could be obtained with the NTrainer, with both types of pacifiers. No differences were detected in nonnutritive sucking dynamics or weight loss over time within each group or between groups. CONCLUSIONS: Data indicate that it is feasible to measure nonnutritive sucking dynamics and associated weight loss in relation to pacifier design in preterm neonates. Larger trials over longer time periods are needed to determine whether pacifier design influences suck dynamics and maturation, oromotor function, feeding/weight loss, and dental formation in preterm neonates. (Curr Ther Res Clin Exp. 2020; 81:XXX-XXX).

Schizasterid Heart Urchins Host Microorganisms in a Digestive Symbiosis of Mesozoic Origin.

Because of their lifestyles, abundance, and feeding habits, infaunal marine deposit feeders have a significant impact on the ocean floor. As these animals also ingest microorganisms associated with their sediment and seawater diet, their digestive tract usually contains a diverse array of bacteria. However, while most of these microorganisms are transients, some may become part of a resident gut microbiome, in particular when sheltered from the main flow of digesta in specialized gut compartments. Here, we provide an in-depth analysis of the structure and contents of the intestinal caecum (IC), a hindgut diverticulum found exclusively in schizasterid heart urchins (Echinoidea: Spatangoida: Schizasteridae). Based on specimens of Brisaster townsendi, in addition to various other schizasterid taxa, our structural characterization of the IC shows that the organ is a highly specialized gut compartment with unique structural properties. Next generation sequencing shows that the IC contains a microbial population composed predominantly of Bacteroidales, Desulfobacterales, and Spirochaetales. The microbiome of this gut compartment is significantly different in composition and lower in diversity than the microbial population in the sediment-filled main digestive tract. Inferences on the function and evolution of the IC and its microbiome suggest that this symbiosis plays a distinct role in host nutrition and that it evolved at least 66 million years ago during the final phase of the Mesozoic.

Purification of a Crenarchaeal ATP Synthase in the Light of the Unique Bioenergetics of Ignicoccus Species.

In this study, the ATP synthase of Ignicoccus hospitalis was purified, characterized, and structurally compared to the respective enzymes of the other Ignicoccus species, to shed light on energy conservation in this unique group of archaea. The crenarchaeal genus Ignicoccus comprises three described species, i.e., I. hospitalis and Ignicoccus islandicus from hot marine sediments near Iceland and Ignicoccus pacificus from a hydrothermal vent system in the Pacific Ocean. This genus is unique among all archaea due to the unusual cell envelope, consisting of two membranes that enclose a large intermembrane compartment (IMC). I. hospitalis is the best studied member of this genus, mainly because it is the only known host for the potentially parasitic archaeon Nanoarchaeum equitansI. hospitalis grows chemolithoautotrophically, and its sole energy-yielding reaction is the reduction of elemental sulfur with molecular hydrogen, forming large amounts of hydrogen sulfide. This reaction generates an electrochemical gradient, which is used by the ATP synthase, located in the outer cellular membrane, to generate ATP inside the IMC. The genome of I. hospitalis encodes nine subunits of an A-type ATP synthase, which we could identify in the purified complex. Although the maximal in vitro activity of the I. hospitalis enzyme was measured around pH 6, the optimal stability of the A1AO complex seemed to be at pH 9. Interestingly, the soluble A1 subcomplexes of the different Ignicoccus species exhibited significant differences in their apparent molecular masses in native electrophoresis, although their behaviors in gel filtration and chromatography-mass spectrometry were very similar.IMPORTANCE The Crenarchaeota represent one of the major phyla within the Archaea domain. This study describes the successful purification of a crenarchaeal ATP synthase. To date, all information about A-type ATP synthases is from euryarchaeal enzymes. The fact that it has not been possible to purify this enzyme complex from a member of the Crenarchaeota until now points to significant differences in stability, possibly caused by structural alterations. Furthermore, the study subject I. hospitalis has a particular importance among crenarchaeotes, since it is the only known host of N. equitans The energy metabolism in this system is still poorly understood, and our results can help elucidate the unique relationship between these two microbes.

Dumbo octopod hatchling provides insight into early cirrate life cycle.

Cirrate octopods (Cephalopoda: Cirrata) are among the largest invertebrates of the deep sea. These organisms have long been known to lay single, large egg capsules on hard substrates on the ocean bottom [1], including cold-water octocorals (Anthozoa: Octocorallia). The egg capsule is comprised of an external egg case as well as the chorion and developing embryo. Development in cirrates proceeds for an extended time without parental care [2]. Although juveniles have previously been collected in the midwater [3], cirrate hatchlings have so far never been observed. Here, we provide the first video of a living hatchling and use magnetic resonance imaging (MRI) to analyze its anatomy and assign the specimen to the genus Grimpoteuthis, the so-called dumbo octopods. The specimen's behavior and advanced state of organ development show that cirrate hatchlings possess all morphological features required for movement via fin-swimming, for visually and chemically sensing their environment, and for prey capture. In addition, the presence of a large internal yolk sac reduces the risk of failure at first feeding. These data provide evidence that dumbo octopods hatch as competent juveniles.

Oxidative Stress in Neurodegenerative Diseases: From Molecular Mechanisms to Clinical Applications.

Increasing numbers of individuals, particularly the elderly, suffer from neurodegenerative disorders. These diseases are normally characterized by progressive loss of neuron cells and compromised motor or cognitive function. Previous studies have proposed that the overproduction of reactive oxygen species (ROS) may have complex roles in promoting the disease development. Research has shown that neuron cells are particularly vulnerable to oxidative damage due to their high polyunsaturated fatty acid content in membranes, high oxygen consumption, and weak antioxidant defense. However, the exact molecular pathogenesis of neurodegeneration related to the disturbance of redox balance remains unclear. Novel antioxidants have shown great potential in mediating disease phenotypes and could be an area of interest for further research. In this review, we provide an updated discussion on the roles of ROS in the pathological mechanisms of Alzheimer's disease, Huntington's disease, Parkinson's disease, amyotrophic lateral sclerosis, and spinocerebellar ataxia, as well as a highlight on the antioxidant-based therapies for alleviating disease severity.

Comprehensive analysis and reinterpretation of Cenozoic mesofossils reveals ancient origin of the snapping claw of alpheid shrimps.

Alpheid snapping shrimps (Decapoda: Caridea: Alpheidae) constitute one of the model groups for inferences aimed at understanding the evolution of complex structural, behavioural, and ecological traits among benthic marine invertebrates. Despite being a super-diverse taxon with a broad geographical distribution, the alpheid fossil record is still poorly known. However, data presented herein show that the strongly calcified fingertips of alpheid snapping claws are not uncommon in the fossil record and should be considered a novel type of mesofossil. The Cenozoic remains analysed here represent a compelling structural match with extant species of Alpheus. Based on the presence of several distinct snapping claw-fingertip morphotypes, the major radiation of Alpheus lineages is estimated to have occurred as early as 18 mya. In addition, the oldest fossil record of alpheids in general can now be confirmed for the Late Oligocene (27-28 mya), thus providing a novel minimum age for the entire group as well as the first reliable calibration point for deep phylogenetic inferences.

Form and function of the teleost lateral line revealed using three-dimensional imaging and computational fluid dynamics.

Fishes sense weak water motion using the lateral line. Among the thousands of described fish species, this organ may differ in size, shape and distribution of individual mechanoreceptors or lateral line canals. The reasons for this diversity remain unclear, but are very likely related to habitat preferences. To better understand the performance of the organ in natural hydrodynamic surroundings, various three-dimensional imaging datasets of the cephalic lateral line were gathered using Leuciscus idus as representative freshwater teleost. These data are employed to simulate hydrodynamic phenomena around the head and within lateral line canals. The results show that changes in canal dimensions alter the absolute stimulation amplitudes, but have little effect on the relation between bulk water flow and higher frequency signals. By contrast, depressions in the skin known as epidermal pits reduce bulk flow stimulation and increase the ratio between higher-frequency signals and the background flow stimulus.

Increased Conformational Flexibility of HLA-B*27 Subtypes Associated With Ankylosing Spondylitis.

OBJECTIVE: Dissimilarities in antigen processing and presentation are known to contribute to the differential association of HLA-B*27 subtypes with the inflammatory rheumatic disease ankylosing spondylitis (AS). In support of this notion, previous x-ray crystallographic data showed that peptides can be displayed by almost identical HLA-B*27 molecules in a subtype-dependent manner, allowing cytotoxic T lymphocytes to distinguish between these subtypes. For example, a human self-peptide derived from vasoactive intestinal peptide receptor type 1 (pVIPR; sequence RRKWRRWHL) is displayed in a single conformation by B*27:09 (which is not associated with AS), while B*27:05 (which is associated with AS) presents the peptide in a dual binding mode. In addition, differences in conformational flexibility between these subtypes might affect their stability or antigen presentation capability. This study was undertaken to investigate B*27:04 and B*27:06, another pair of minimally distinct HLA-B*27 subtypes, to assess whether dual peptide conformations or structural dynamics play a role in the initiation of AS. METHODS: Using x-ray crystallography, we determined the structures of the pVIPR-B*27:04 and pVIPR-B*27:06 complexes and used isotope-edited infrared (IR) spectroscopy to probe the dynamics of these HLA-B*27 subtypes. RESULTS: As opposed to B*27:05 and B*27:09, B*27:04 (which is associated with AS) displays pVIPR conventionally and B*27:06 (which is not associated with AS) presents the peptide in a dual conformation. Comparison of the 4 HLA-B*27 subtypes using IR spectroscopy revealed that B*27:04 and B*27:05 possess elevated molecular dynamics compared to the nonassociated subtypes B*27:06 and B*27:09. CONCLUSION: Our results demonstrate that an increase in conformational flexibility characterizes the disease-associated subtypes B*27:04 and B*27:05.

Huge Mediastinal Lipoblastoma in a Nine Year-old Boy Successfully Removed Surgically.

A nine year-old boy presented with symptoms of dyspnoea. The chest radiograph and computed tomography scan revealed a large mediastinal tumour. A decision to operate on the patient was made. A huge 2.45 kg mediastinal lipoblastoma was successfully removed from the boy's chest. At three years no evidence of recurrence was found.

A dataset comprising 141 magnetic resonance imaging scans of 98 extant sea urchin species.

BACKGROUND: Apart from its application in human diagnostics, magnetic resonance imaging (MRI) can also be used to study the internal anatomy of zoological specimens. As a non-invasive imaging technique, MRI has several advantages, such as rapid data acquisition, output of true three-dimensional imagery, and provision of digital data right from the onset of a study. Of particular importance for comparative zoological studies is the capacity of MRI to conduct high-throughput analyses of multiple specimens. In this study, MRI was applied to systematically document the internal anatomy of 98 representative species of sea urchins (Echinodermata: Echinoidea). FINDINGS: The dataset includes raw and derived image data from 141 MRI scans. Most of the whole sea urchin specimens analyzed were obtained from museum collections. The attained scan resolutions permit differentiation of various internal organs, including the digestive tract, reproductive system, coelomic compartments, and lantern musculature. All data deposited in the GigaDB repository can be accessed using open source software. Potential uses of the dataset include interactive exploration of sea urchin anatomy, morphometric and volumetric analyses of internal organs observed in their natural context, as well as correlation of hard and soft tissue structures. CONCLUSIONS: The dataset covers a broad taxonomical and morphological spectrum of the Echinoidea, focusing on 'regular' sea urchin taxa. The deposited files significantly expand the amount of morphological data on echinoids that are electronically available. The approach chosen here can be extended to various other vertebrate and invertebrate taxa. We argue that publicly available digital anatomical and morphological data gathered during experiments involving non-invasive imaging techniques constitute one of the prerequisites for future large-scale genotype-phenotype correlations.

A dataset comprising four micro-computed tomography scans of freshly fixed and museum earthworm specimens.

BACKGROUND: ALTHOUGH MOLECULAR TOOLS ARE INCREASINGLY EMPLOYED TO DECIPHER INVERTEBRATE SYSTEMATICS, EARTHWORM (ANNELIDA: Clitellata: 'Oligochaeta') taxonomy is still largely based on conventional dissection, resulting in data that are mostly unsuitable for dissemination through online databases. In order to evaluate if micro-computed tomography (µCT) in combination with soft tissue staining techniques could be used to expand the existing set of tools available for studying internal and external structures of earthworms, µCT scans of freshly fixed and museum specimens were gathered. FINDINGS: Scout images revealed full penetration of tissues by the staining agent. The attained isotropic voxel resolutions permit identification of internal and external structures conventionally used in earthworm taxonomy. The µCT projection and reconstruction images have been deposited in the online data repository GigaDB and are publicly available for download. CONCLUSIONS: The dataset presented here shows that earthworms constitute suitable candidates for µCT scanning in combination with soft tissue staining. Not only are the data comparable to results derived from traditional dissection techniques, but due to their digital nature the data also permit computer-based interactive exploration of earthworm morphology and anatomy. The approach pursued here can be applied to freshly fixed as well as museum specimens, which is of particular importance when considering the use of rare or valuable material. Finally, a number of aspects related to the deposition of digital morphological data are briefly discussed.

Sine systemate chaos? A versatile tool for earthworm taxonomy: non-destructive imaging of freshly fixed and museum specimens using micro-computed tomography.

In spite of the high relevance of lumbricid earthworms ('Oligochaeta': Lumbricidae) for soil structure and functioning, the taxonomy of this group of terrestrial invertebrates remains in a quasi-chaotic state. Earthworm taxonomy traditionally relies on the interpretation of external and internal morphological characters, but the acquisition of these data is often hampered by tedious dissections or restricted access to valuable and rare museum specimens. The present state of affairs, in conjunction with the difficulty of establishing primary homologies for multiple morphological features, has led to an almost unrivaled instability in the taxonomy and systematics of certain earthworm groups, including Lumbricidae. As a potential remedy, we apply for the first time a non-destructive imaging technique to lumbricids and explore the future application of this approach to earthworm taxonomy. High-resolution micro-computed tomography (µCT) scanning of freshly fixed and museum specimens was carried out using two cosmopolitan species, Aporrectodea caliginosa and A. trapezoides. By combining two-dimensional and three-dimensional dataset visualization techniques, we demonstrate that the morphological features commonly used in earthworm taxonomy can now be analyzed without the need for dissection, whether freshly fixed or museum specimens collected more than 60 years ago are studied. Our analyses show that µCT in combination with soft tissue staining can be successfully applied to lumbricid earthworms. An extension of the approach to other families is poised to strengthen earthworm taxonomy by providing a versatile tool to resolve the taxonomic chaos currently present in this ecologically important, but taxonomically neglected group of terrestrial invertebrates.

The songbird syrinx morphome: a three-dimensional, high-resolution, interactive morphological map of the zebra finch vocal organ.

BACKGROUND: Like human infants, songbirds learn their species-specific vocalizations through imitation learning. The birdsong system has emerged as a widely used experimental animal model for understanding the underlying neural mechanisms responsible for vocal production learning. However, how neural impulses are translated into the precise motor behavior of the complex vocal organ (syrinx) to create song is poorly understood. First and foremost, we lack a detailed understanding of syringeal morphology. RESULTS: To fill this gap we combined non-invasive (high-field magnetic resonance imaging and micro-computed tomography) and invasive techniques (histology and micro-dissection) to construct the annotated high-resolution three-dimensional dataset, or morphome, of the zebra finch (Taeniopygia guttata) syrinx. We identified and annotated syringeal cartilage, bone and musculature in situ in unprecedented detail. We provide interactive three-dimensional models that greatly improve the communication of complex morphological data and our understanding of syringeal function in general. CONCLUSIONS: Our results show that the syringeal skeleton is optimized for low weight driven by physiological constraints on song production. The present refinement of muscle organization and identity elucidates how apposed muscles actuate different syringeal elements. Our dataset allows for more precise predictions about muscle co-activation and synergies and has important implications for muscle activity and stimulation experiments. We also demonstrate how the syrinx can be stabilized during song to reduce mechanical noise and, as such, enhance repetitive execution of stereotypic motor patterns. In addition, we identify a cartilaginous structure suited to play a crucial role in the uncoupling of sound frequency and amplitude control, which permits a novel explanation of the evolutionary success of songbirds.

Evolution of a novel muscle design in sea urchins (Echinodermata: Echinoidea).

The sea urchin (Echinodermata: Echinoidea) masticatory apparatus, or Aristotle's lantern, is a complex structure composed of numerous hard and soft components. The lantern is powered by various paired and unpaired muscle groups. We describe how one set of these muscles, the lantern protractor muscles, has evolved a specialized morphology. This morphology is characterized by the formation of adaxially-facing lobes perpendicular to the main orientation of the muscle, giving the protractor a frilled aspect in horizontal section. Histological and ultrastructural analyses show that the microstructure of frilled muscles is largely identical to that of conventional, flat muscles. Measurements of muscle dimensions in equally-sized specimens demonstrate that the frilled muscle design, in comparison to that of the flat muscle type, considerably increases muscle volume as well as the muscle's surface directed towards the interradial cavity, a compartment of the peripharyngeal coelom. Scanning electron microscopical observations reveal that the insertions of frilled and flat protractor muscles result in characteristic muscle scars on the stereom, reflecting the shapes of individual muscles. Our comparative study of 49 derived "regular" echinoid species using magnetic resonance imaging (MRI) shows that frilled protractor muscles are found only in taxa belonging to the families Toxopneustidae, Echinometridae, and Strongylocentrotidae. The onset of lobe formation during ontogenesis varies between species of these three families. Because frilled protractor muscles are best observed in situ, the application of a non-invasive imaging technique was crucial for the unequivocal identification of this morphological character on a large scale. Although it is currently possible only to speculate on the functional advantages which the frilled muscle morphology might confer, our study forms the anatomical and evolutionary framework for future analyses of this unusual muscle design among sea urchins.

Analysis of freshly fixed and museum invertebrate specimens using high-resolution, high-throughput MRI.

Magnetic resonance imaging (MRI) is now considered a routine tool for comparative morphological imaging in small vertebrate model organisms. However, the application of high-resolution imaging protocols to visualize the anatomy of invertebrate organisms has not yet become a generally accepted tool among zoologists. Here, we describe MRI protocols that permit visualization of both the internal and the external anatomy of freshly fixed invertebrates and specimens from museum collections. The choice of protocols has been optimized to allow the assembly of the large numbers of datasets that are necessary for comparative morphological analyses. Although the primary focus of our work is on sea urchin internal anatomy, we also present results from a variety of other invertebrate taxa to demonstrate the principal feasibility of MRI studies to obtain anatomical information at high resolutions. Furthermore, we briefly describe procedures suitable for 3D modelling.