Wound repair during arm regeneration in the red starfish Echinaster sepositus.

Starfish can regenerate entire arms following their loss by both autotomic and traumatic amputation. Although the overall regenerative process has been studied several times in different asteroid species, there is still a considerable gap of knowledge as far as the detailed aspects of the repair phase at tissue and cellular level are concerned, particularly in post-traumatic regeneration. The present work is focused on the arm regeneration model in the Mediterranean red starfish Echinaster sepositus; to describe the early cellular mechanisms of arm regeneration following traumatic amputation, different microscopy techniques were employed. In E. sepositus, the repair phase was characterized by prompt wound healing by a syncytial network of phagocytes and re-epithelialisation followed by a localized subepidermal oedematous area formation. Scattered and apparently undifferentiated cells, intermixed with numerous phagocytes, were frequently found in the wound area during these first stages of regeneration and extensive dedifferentiation phenomena were seen at the level of the stump, particularly in the muscle bundles. A true localized blastema did not form. Our results confirm that regeneration in asteroids mainly relies on morphallactic processes, consisting in extensive rearrangement of the existing tissues which contribute to the new tissues through cell dedifferentiation, redifferentiation, and/or migration.

Re-growth, morphogenesis, and differentiation during starfish arm regeneration.

The red starfish Echinaster sepositus is an excellent model for studying arm regeneration processes following traumatic amputation. The initial repair phase was described in a previous paper in terms of the early cicatrisation phenomena, and tissue and cell involvement. In this work, we attempt to provide a further comprehensive description of the later regenerative stages in this species. Here, we present the results of a detailed microscopic and submicroscopic investigation of the long regenerative phase, which can be subdivided into two subphases: early and advanced regenerative phases. The early regenerative phase (1-6 weeks p.a.) is characterized by tissue rearrangement, morphogenetic processes and initial differentiation events (mainly neurogenesis and skeletogenesis). The advanced regenerative phase (after 6 weeks p.a.) is characterized by further differentiation processes (early myogenesis), and obvious morphogenesis and re-growth of the regenerate. As in other starfish, the regenerative process in E. sepositus is relatively slow in comparison with that of crinoids and many ophiuroids, which is usually interpreted as resulting mainly from size-related aspects and of the more conspicuous involvement of morphallactic processes. Light and electron microscopy analyses suggest that some of the amputated structures, such as muscles, are not able to replace their missing parts by directly re-growing them from the remaining tissues, whereas others tissues, such as the skeleton and the radial nerve cord, appear to undergo direct re-growth. The overall process is in agreement with the distalization-intercalation model proposed by Agata and co-workers. Further experiments are needed to confirm this hypothesis.

Lessons learned from implementing education on dual-use in Austria, Italy, Pakistan and Sweden.

This paper provides insights into the achievements and challenges of implementing education on dual-use in four countries: Austria, Italy, Pakistan and Sweden. It draws attention to the different institutional mechanisms through which dual-use education may be introduced into academic curricula and some of the difficulties encountered in this process. It concludes that there is no 'one size fits all' approach to the implementation of dual-use education. Rather, initiatives must be tailored to suit the teaching traditions, geographical and historical context in which they are being delivered. However, a number of common principles and themes can be derived from all four cases. All these courses bring together a number of different topics that place 'dual-use' in the broader context of biosafety, biosecurity, ethics, law and the environment. The case studies suggest that success in this area depends largely on the leadership and commitment of individuals directly involved in teaching, who are active within the scientific community.

Chemical fate and biological effects of several endocrine disrupters compounds in two echinoderm species.

Two echinoderm species, the sea urchin Paracentrotus lividus and the feather star Antedon mediterranea, were exposed for 28 days to several EDCs: three putative androgenic compounds, triphenyltin (TPT), fenarimol (FEN), methyltestosterone (MET), and two putative antiandrogenic compounds, p,p'-DDE (DDE) and cyproterone acetate (CPA). The exposure nominal concentrations were from 10 to 3000 ng L(-1), depending on the compound. This paper is an attempt to join three different aspects coming from our ecotoxicological tests: (1) the chemical behaviour inside the experimental system; (2) the measured toxicological endpoints; (3) the biochemical responses, to which the measured endpoints may depend. The chemical fate of the different compounds was enquired by a modelling approach throughout the application of the 'Aquarium model'. An estimation of the day-to-day concentration levels in water and biota were obtained together with the amount assumed each day by each animal (uptake in microg animal(-1) d(-1) or ng g-wet weight(-1) d(-1)). The toxicological endpoints investigated deal with the reproductive potential (gonad maturation stage, gonad index and oocyte diameter) and with the regenerative potential (growth and histology). Almost all the compounds exerted some kind of effect at the tested concentrations, however TPT was the most effective in altering both reproductive and regenerative parameters (also at the concentration of few ng L(-1)). The biochemical analyses of testosterone (T) and 17beta-estradiol (E(2)) also showed the ability of the selected compounds to significantly alter endogenous steroid concentrations.

Steroid levels in crinoid echinoderms are altered by exposure to model endocrine disruptors.

Sexual steroids (testosterone and estradiol) were measured in the whole body of wild specimens of the crinoid Antedon mediterranea collected from the Tyrrhenian Sea (Italy). Testosterone levels (274-1,488 pg/g wet weight (w.w.)) were higher than those of estradiol (60-442 pg/g w.w.) and no significant differences between males and females were observed. No clear seasonal trend was either detected - individuals from February, June and October 2004 analyzed - apart from a peak of estradiol in males in autumn. Nonetheless, dramatic changes on tissue steroid levels were observed when individuals were exposed to model androgenic and anti-androgenic compounds for 2 and 4 weeks. The selected compounds were 17 alpha-methyltestosterone (17 alpha-MT), triphenyltin (TPT), fenarimol (FEN), cyproterone acetate (CPA), and p,p'-DDE. Endogenous testosterone levels were significantly increased after exposure to 17 alpha-MT, TPT and FEN, while different responses were observed for estradiol; 17 alpha-MT and FEN increased endogenous estradiol (up to seven-fold), and TPT lead to a significant decrease. Concerning the anti-androgenic compounds, CPA significantly reduced testosterone in a dose-dependent manner without altering estradiol levels, whereas specimens exposed to p,p'-DDE at a low dose (24 ng/L) for 4 weeks showed a four-fold increase in T levels. Overall, the data show the ability of the selected compounds to alter endogenous steroid concentrations in A. mediterranea, and suggest the existence in this echinoderm species of vertebrate-like mechanisms that can be affected by exposure to androgenic and anti-androgenic chemicals.

The elusive role of L-glutamate as an echinoderm neurotransmitter: evidence for its involvement in the control of crinoid arm muscles.

Although l-glutamate is the most widespread excitatory neurotransmitter in vertebrate and invertebrate nervous systems, there is only sparse evidence that it has this role in echinoderms. Following our previous finding that l-glutamate is widely distributed in the arms of the featherstar (crinoid echinoderm) Antedon mediterranea and initiates arm autotomy (defensive detachment), we now provide evidence of glutamatergic involvement in the control of the arm muscles of the same species using immunocytochemical and physiological methods. Immunofluorescence and immunoenzymatic techniques, which employed the same polyclonal antibody against l-glutamate conjugated to glutaraldehyde, revealed a high level of glutamate-like reactivity in the brachial muscles. By recording the mechanical responses of isolated arm pieces, we found that l-glutamate, l-aspartate and elevated [K(+)](o) induced rhythmic muscle contractions, while glycine, γ-aminobutyric acid, adrenaline and acetylcholine had either no, or no consistent, effect. The frequency and duration of the dominant component of the rhythmic contractions indicated that these may be responsible for the rhythmic activity of the arms that occurs during swimming and after autotomy. We conclude that it is highly likely that l-glutamate has at least a neuromodulatory role in the neural pathways controlling the brachial muscles of A. mediterranea.

Muscle system organization in the echinoderms: III. Fine structure of the contractile apparatus of the arm flexor muscles of the comatulids (Antedon mediterranea).

The crinoid arm muscles consist of obliquely striated fibers with striking differences in function and in ultrastructural features. These fibers can be distinguished mainly on the basis of different myofilament arrangements (A- and B-type patterns) and are variously combined at different levels (proximal, intermediate, and distal portions) of the arm. Some rare smooth fibers (C-type) are irregularly distributed in the periphery and in the core of the bundle. The characteristic features of the A- and B-type obliquely striated fibers are (1) a continuous and homogeneous structure of the Z line and (2) a very heterogeneous arrangement of myosin filaments which vary widely in size, number, and distribution from section to section. The significance of such an atypical, obliquely striated muscle may be related to the double skeletal system combination (endoskeleton and hydroskeleton) of the crinoid arms.

Muscle system organization in the echinoderms: II. Microscopic anatomy and functional significance of the muscle-ligament-skeleton system in the arm of the comatulids (Antedon mediterranea).

Comatulids are able to perform quick and complex movements of the arms which are used to swim, creep, walk, and also form a rigid, feeding-filtration fan. To perform such versatile movements, the arms of these animals are equipped with a classical endoskeletal system, with joints, muscles, ligaments, and a hydroskeleton of three different coelomic channels. Light microscopic study of the detailed anatomical organization of the arm clarifies both the complex relations between the parts involved in the movements and their functional responsibilities. In particular, (1) the ventral muscle bundles show a heterogeneous structure that consists of different and variously arranged populations of fibers, which allows the different flexing movements of the arms (i.e., flexion and maintaining the flexed state); (2) the ligaments (both dorsal and interarticular) consist only of collagen fibrils and, therefore, have a passive function in binding the skeletal pieces together: their possible active engagement in the extending movements of the arms is thus excluded; (3) owing to the absence of other suitable antagonists to the flexor muscles, the only efficient antagonist system seems to be the coelomic cavities, which are well separated from each other and are also provided with muscular valves. They thus function as typical hydraulic systems, which allows the arm to perform both simple extensions and very complex combined movements and to maintain some rigid straight or twisted positions.