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Background: Regeneration plays a key role in energy recycling and homeostasis maintenance. Planarians, as ideal model animals for studying regeneration, stem cell proliferation, and apoptosis, have the strong regenerative abilities. Considerable evidence suggests that ubiquitin plays an important role in maintaining homeostasis and regulating regeneration, but the function of Ubiquitin specific proteases 7 (Usp7) on regeneration in planarians remains elusive., Methods: We identified an evolutionarily conserved gene, Usp7, and utilized RNA interference (RNAi), Quantitative real-time PCR (qRT-PCR), Whole-mount immunofluorescence, Tunnel, Whole-mount in situ hybridization (WISH), and western blotting to detect the function of Usp7 during the planarian regeneration., Results: In this study, we found that the regenerative trunk fragments in the Usp7 RNAi worms could not regenerate missing tails; meanwhile, the level of cell proliferation was decreased, while cell apoptosis was increased. Furthermore, the expression of Islet was inhibited in the Usp7 RNAi worms during planarian regeneration. The hybridization signal of wnt1/P-1 exhibited the dot-like pattern at the posterior of the regenerating planarians after Usp7 RNAi at regenerative 1 day (R 1 d). However, the concentrated expression pattern wnt1/P-1 dramatically declined at regenerative 3 days (R 3 d) and disappeared at regenerative 7 days (R 7 d). In addition, activating the Wnt pathway partially rescued regenerative defects induced by inhibition of Usp7., Conclusions: Collectively, Usp7 is necessary for tissue regeneration and tail blastema formation partially by regulating the cell proliferation and apoptosis during planarian regeneration. It could also promote the posterior polarity reconstruction of the regenerative planarians via the Islet/Wnt1 axis., Competing Interests: Declarations. Ethics approval and consent to participate: Not applicable. Consent for publication: All authors consent to publication. Competing interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2025. The Author(s).)

Using immunocytochemistry, serotonergic nerve elements were documented in the nervous system of the planarian Girardia tigrina. Serotonin-immunopositive components were observed in the brain, ventral, dorsal and longitudinal nerve cords, transverse nerve commissures connecting the nerve cords, and in the nerve plexus. Whole-mount preparations of G. tigrina were analyzed by fluorescent and confocal laser scanning microscopy. An essential quantitative morphometric measurement of serotonin-immunopositive structures was conducted in three body regions (anterior, middle, and posterior) of the planarian. The number of serotonin neurons was maximal in the head region. The ventral nerve cords gradually decreased in thickness from anterior to posterior body ends. Physiological action of exogenously applied serotonin was studied in G. tigrina for the first time. It was found that serotonin (0.1 and 1 µmol L -1 ) accelerated eye regeneration. The transcriptome sequencing performed for the first time for the planarian G. tigrina revealed the transcripts of the tryptophan hydroxylase (trph), amino acid decarboxylase (aadc) and serotonin transporter (sert) genes. The data obtained indicate the presence of the components of serotonin pathway in G. tigrina. The identified transcripts can take part in serotonin turnover and participate in the realization of biological effects of serotonin in planarians, associated with eyes regeneration and differentiation., (© 2024 Wiley Periodicals LLC.)

A new species of the genus Dugesia (Platyhelminthes, Tricladida, Dugesiidae) from Xiangxi River, Shennongjia Forestry District, Hubei Province, China, is described on the basis of an integrative approach, involving morphology, and molecular systematics. The new species Dugesia saccaria A-T. Wang & Sluys, sp. nov. is characterized by the following features: a dumb-bell-shaped, muscularized hump located just anterior to the knee-shaped bend in the bursal canal; a ventrally displaced ejaculatory duct, which, however, opens terminally through the dorsal portion of the blunt tip of the penis papilla; a ventrally located seminal vesicle, giving rise to a vertically running duct that eventually curves downwards to communicate with the ejaculatory duct via a small diaphragm; oviducts opening asymmetrically into the dorsal portion of the common atrium and at the knee-shaped part of the bursal canal. The phylogenetic position of the new species was determined using four molecular markers (18S rDNA; ITS-1; 28S rDNA; COI), which suggested that it groups with other species of Dugesia from the Australasian and Oriental biogeographical regions.

Background: Despite large morphological differences between the nervous systems of lower animals and humans, striking functional similarities have been reported. However, little is known about how these functional similarities translate to cognitive similarities. As a first step towards studying the cognitive abilities of simple nervous systems, we here characterize the ongoing electrophysiological activity of the planarian Schmidtea mediterranea. One previous report using invasive microelectrodes describes that the ongoing neural activity is characterized by a 1/f x power spectrum with the exponent 'x' of the power spectrum close to 1. To extend these findings, we aimed to establish a recording protocol to measure ongoing neural activity safely and securely from alive and healthy planarians under different lighting conditions using non-invasive surface electrodes., Results: As a replication and extension of the previous results, we show that the ongoing neural activity is characterized by a 1/f x power spectrum, that the exponent 'x' in living planarians is close to 1, and that changes in lighting induce changes in neural activity likely due to the planarian photophobia., Conclusions: We confirm the existence of continuous EEG activity in planarians and show that it is possible to noninvasively record this activity with surface wire electrodes. This opens up broad possibilities for continuous recordings across longer intervals, and repeated recordings from the same animals to study cognitive processes., (© 2023. The Author(s).)

Simultaneous longitudinal imaging across multiple conditions and replicates has been crucial for scientific studies aiming to understand biological processes and disease. Yet, imaging systems capable of accomplishing these tasks are economically unattainable for most academic and teaching laboratories around the world. Here, we propose the Picroscope, which is the first low-cost system for simultaneous longitudinal biological imaging made primarily using off-the-shelf and 3D-printed materials. The Picroscope is compatible with standard 24-well cell culture plates and captures 3D z-stack image data. The Picroscope can be controlled remotely, allowing for automatic imaging with minimal intervention from the investigator. Here, we use this system in a range of applications. We gathered longitudinal whole organism image data for frogs, zebrafish, and planaria worms. We also gathered image data inside an incubator to observe 2D monolayers and 3D mammalian tissue culture models. Using this tool, we can measure the behavior of entire organisms or individual cells over long-time periods., (© 2021. The Author(s).)

Many biological organisms regenerate structure and function after damage. Despite the long history of research on molecular mechanisms, many questions remain about algorithms by which cells can cooperate towards the same invariant morphogenetic outcomes. Therefore, conceptual frameworks are needed not only for motivating hypotheses for advancing the understanding of regeneration processes in living organisms, but also for regenerative medicine and synthetic biology. Inspired by planarian regeneration, this study offers a novel generic conceptual framework that hypothesizes mechanisms and algorithms by which cell collectives may internally represent an anatomical target morphology towards which they build after damage. Further, the framework contributes a novel nature-inspired computing method for self-repair in engineering and robotics. Our framework, based on past in vivo and in silico studies on planaria, hypothesizes efficient novel mechanisms and algorithms to achieve complete and accurate regeneration of a simple in silico flatwormlike organism from any damage, much like the body-wide immortality of planaria, with minimal information and algorithmic complexity. This framework that extends our previous circular tissue repair model integrates two levels of organization: tissue and organism. In Level 1, three individual in silico tissues (head, body, and tail-each with a large number of tissue cells and a single stem cell at the centre) repair themselves through efficient local communications. Here, the contribution extends our circular tissue model to other shapes and invests them with tissue-wide immortality through an information field holding the minimum body plan. In Level 2, individual tissues combine to form a simple organism. Specifically, the three stem cells form a network that coordinates organism-wide regeneration with the help of Level 1. Here we contribute novel concepts for collective decision-making by stem cells for stem cell regeneration and large-scale recovery. Both levels (tissue cells and stem cells) represent networks that perform simple neural computations and form a feedback control system. With simple and limited cellular computations, our framework minimises computation and algorithmic complexity to achieve complete recovery. We report results from computer simulations of the framework to demonstrate its robustness in recovering the organism after any injury. This comprehensive hypothetical framework that significantly extends the existing biological regeneration models offers a new way to conceptualise the information-processing aspects of regeneration, which may also help design living and non-living self-repairing agents., (© 2021 Massachusetts Institute of Technology.)

Complex anatomical form is regulated in part by endogenous physiological communication between cells; however, the dynamics by which gap junctional (GJ) states across tissues regulate morphology are still poorly understood. We employed a biophysical modeling approach combining different signaling molecules (morphogens) to qualitatively describe the anteroposterior and lateral morphology changes in model multicellular systems due to intercellular GJ blockade. The model is based on two assumptions for blocking-induced patterning: (i) the local concentrations of two small antagonistic morphogens diffusing through the GJs along the axial direction, together with that of an independent, uncoupled morphogen concentration along an orthogonal direction, constitute the instructive patterns that modulate the morphological outcomes, and (ii) the addition of an external agent partially blocks the intercellular GJs between neighboring cells and modifies thus the establishment of these patterns. As an illustrative example, we study how the different connectivity and morphogen patterns obtained in presence of a GJ blocker can give rise to novel head morphologies in regenerating planaria. We note that the ability of GJs to regulate the permeability of morphogens post-translationally suggests a mechanism by which different anatomies can be produced from the same genome without the modification of gene-regulatory networks. Conceptually, our model biosystem constitutes a reaction-diffusion information processing mechanism that allows reprogramming of biological morphologies through the external manipulation of the intercellular GJs and the resulting changes in instructive biochemical signals., (Copyright © 2021 The Authors. Published by Elsevier B.V. All rights reserved.)

As the planarian research community expands, the need for an interoperable data organization framework for tool building has become increasingly apparent. Such software would streamline data annotation and enhance cross-platform and cross-species searchability. We created the Planarian Anatomy Ontology (PLANA), an extendable relational framework of defined Schmidtea mediterranea (Smed) anatomical terms used in the field. At publication, PLANA contains over 850 terms describing Smed anatomy from subcellular to system levels across all life cycle stages, in intact animals and regenerating body fragments. Terms from other anatomy ontologies were imported into PLANA to promote interoperability and comparative anatomy studies. To demonstrate the utility of PLANA as a tool for data curation, we created resources for planarian embryogenesis, including a staging series and molecular fate-mapping atlas, and the Planarian Anatomy Gene Expression database, which allows retrieval of a variety of published transcript/gene expression data associated with PLANA terms. As an open-source tool built using FAIR (findable, accessible, interoperable, reproducible) principles, our strategy for continued curation and versioning of PLANA also provides a platform for community-led growth and evolution of this resource., Competing Interests: Competing interests The authors declare no competing or financial interests., (© 2021. Published by The Company of Biologists Ltd.)

Two mostly white terrestrial planarians, about 7 mm long, found in a garden in Yorkshire, UK, are described. They have a single pair of eyes and both specimens are fully mature, with a single pair of ovaries, several testes, a conical penis papilla and a genito-intestinal duct, characters of the genus Microplana, but differ in color and size from other species of the genus and are described as Microplana edwardsi sp. nov.

The genus Phagocata (Platyhelminthes, Tricladida) is represented in Israel by two pigmented species, namely, Phagocata armeniaca (Komárek, 1916), reported previously from Armenia, the Caucasus and eastern Turkey, and more recently known from the constantly cold headwaters of the River Jordan in northern Israel, and Phagocata punctata sp. nov., which was found inhabiting the littoral of the south-eastern shore of Lake Kinneret where there are large seasonal temperature fluctuations. Several samples of both species were collected in different seasons and raised in the laboratory at various temperatures and their habitats, morphology, karyology (2n = 34) and breeding behaviour were compared. The results of cross-breeding experiments are given, with most of the F1 offspring showing a range of patterns of colouration, gut branching and fecundity which were intermediate to those of the parent species. However, crossbreeding also resulted in a small proportion of abnormalities among the F1, and especially the F2, generations, indicating an incomplete reproductive compatibility, but supporting the hypothesis of their common origin. An emended description of Dugesia salina (Whitehouse, 1914), is also given together with karyological data (2n = 16) from material collected from a saline spring, En Sheva (Tabgha), situated on the north-western shore of Lake Kinneret.

Background Information: Dense multicilia in protozoa and metazoa generate a strong force important for locomotion and extracellular fluid flow. During ciliogenesis, multiciliated cells produce hundreds of centrioles to serve as basal bodies through various pathways including deuterosome-dependent (DD), hyper-activated mother centriole-dependent (MCD) and basal bodydependent (BBD) pathways. The centrosome-free planarian Schmidtea mediterranea is widely used for regeneration studies because its neoblasts are capable of regenerating any body part after injury. However, it is currently unclear how the flatworms generate massive centrioles for multiciliated cells in the pharynx and body epidermis when their cells are initially centriole-free., Results: In this study, we investigate the progress of centriole amplification during the pharynx regeneration. We observe that the planarian pharyngeal epithelial cells generate their centrioles asynchronously through a de novo pathway. Most of the de novo centrioles are formed individually, whereas the remaining ones are assembled in pairs, possibly by sharing a cartwheel, or in small clusters lacking a nucleation center. Further RNAi experiments show that the known key factors of centriole duplication, including Cep152, Plk4 and Sas6, are crucial for the centriole amplification., Conclusions and Significance: Our study demonstrates the distinct process of massive centriole biogenesis in S. mediterranea and helps to understand the diversity of centriole biogenesis during evolution., (© 2020 Société Française des Microscopies and Société de Biologie Cellulaire de France. Published by John Wiley & Sons Ltd.)

An extraordinary micro-structural similarity between squid sucker teeth and the egg shell of a terrestrial planarian worm has been reported, but to date only the amino acid content of the squid sucker tooth has been available. This prompted us to analyse the amino acid content of the planarian egg shell. Although both share an absence of detectable chitin and metal ions and both possess relatively high amounts of the amino acids GLY and HIS, the planarian egg shell is considerably richer in GLU, LYS and ASP. Most dramatic was the difference in TYR, which was the second most abundant amino acid in the squid, but hardly featured at all in the planarian egg shell. In the light of these new findings the different functional roles that the structures in question play in the lives of the two taxa are discussed. An EDS-analysis revealed clear C, N, and O peaks and additional very small peaks less than 0.1w% suggesting the presence of one or both S and Cl., (Copyright © 2020 Elsevier GmbH. All rights reserved.)

The keystone of planarian taxonomy traditionally has been the anatomy of the copulatory apparatus. However, many planarian species comprise asexual fissiparous populations, with the fissiparous animals not developing a copulatory apparatus, thus precluding their morphological identification. Incorporation of molecular data into planarian systematics has been of great value, not only in the identification of fissiparous individuals but also as an additional source of information for determining species boundaries. Nevertheless, the discrepancy between morphological and molecular data has highlighted the need for extra sources of taxonomic information. Moreover, a recent study has pointed out that fissiparous reproduction may lead to high levels of intraindividual genetic diversity in planarians, which may mislead molecular analyses. In the present study we aim to test a new up-to-date integrative taxonomic procedure for planarians, including intraindividual genetic data and additional sources of taxonomic information, besides morphology and DNA, using Dugesia subtentaculata sensu lato as a model organism, a species with an intricate taxonomic history. First, we used three different methods for molecular species delimitation on single locus datasets, both with and without intraindividual information, for formulating Primary Species Hypotheses (PSHs). Subsequently, Secondary Species Hypotheses (SSHs) were formulated on the basis of three types of information: (1) a coalescent-based species delimitation method applied to multilocus data, (2) morphology of the copulatory apparatus, and (3) karyological metrics. This resulted in the delimitation of four morphologically cryptic species within the nominal species D. subtentaculata. Our results provide evidence that the analysis of intraindividual genetic data is essential for properly developing PSHs in planarians. Our study reveals also that karyological differentiation, rather than morphological differentiation, may play an important role in speciation processes in planarians, thus suggesting that the currently known diversity of the group could be highly underestimated., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Planarians are a group of flatworms. Some planarian species have remarkable regenerative abilities, which involve abundant pluripotent adult stem cells. This makes these worms a powerful model system for understanding the molecular and evolutionary underpinnings of regeneration. By providing a succinct overview of planarian taxonomy, anatomy, available tools and the molecular orchestration of regeneration, this Primer aims to showcase both the unique assets and the questions that can be addressed with this model system., Competing Interests: Competing interestsThe authors declare no competing or financial interests., (© 2019. Published by The Company of Biologists Ltd.)

Differential coordination of growth and patterning across metazoans gives rise to a diversity of sizes and shapes at tissue, organ and organismal levels. Although tissue size and tissue function can be interdependent 1-5 , mechanisms that coordinate size and function remain poorly understood. Planarians are regenerative flatworms that bidirectionally scale their adult body size 6,7 and reproduce asexually, via transverse fission, in a size-dependent manner 8-10 . This model offers a robust context to address the gap in knowledge that underlies the link between size and function. Here, by generating an optimized planarian fission protocol in Schmidtea mediterranea, we show that progeny number and the frequency of fission initiation are correlated with parent size. Fission progeny size is fixed by previously unidentified mechanically vulnerable planes spaced at an absolute distance along the anterior-posterior axis. An RNA interference screen of genes for anterior-posterior patterning uncovered components of the TGFβ and Wnt signalling pathways as regulators of the frequency of fission initiation rather than the position of fission planes. Finally, inhibition of Wnt and TGFβ signalling during growth altered the patterning of mechanosensory neurons-a neural subpopulation that is distributed in accordance with worm size and modulates fission behaviour. Our study identifies a role for TGFβ and Wnt in regulating size-dependent behaviour, and uncovers an interdependence between patterning, growth and neurological function.

Planarian flatworms are known as the masters of regeneration, re-growing an entire organism from as little as 1/279th part of their body. While the proteomics of these processes has been studied extensively, the planarian lipodome remains relatively unknown. In this study we investigate the lipid profile of planarian tissue sections with imaging Time-of-Flight - Secondary-Ion-Mass-Spectrometry (ToF-SIMS). ToF-SIMS is a label-free technique capable of gathering intact, location specific lipid information on a cellular scale. Lipid identities are confirmed using LC-MS/MS. Our data shows that different organ structures within planarians have unique lipid profiles. The 22-carbon atom poly unsaturated fatty acids (PUFAs) which occur in unusually high amounts in planarians are found to be mainly located in the testes. Additionally, we observe that planarians contain various odd numbered fatty acid species, that are usually found in bacteria, localized in the reproductive and ectodermal structures of the planarian. An abundance of poorly understood ether fatty acids and ether lipids were found in unique areas in planarians as well as a new, yet unidentified class of potential lipids in planarian intestines. Identifying the location of these lipids in the planarian body provides insights into their bodily functions and, in combination with knowledge about their diet and their genome, enables drawing conclusions about planarian fatty acid processing., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Control of axial polarity during regeneration is a crucial open question. We developed a quantitative model of regenerating planaria, which elucidates self-assembly mechanisms of morphogen gradients required for robust body-plan control. The computational model has been developed to predict the fraction of heteromorphoses expected in a population of regenerating planaria fragments subjected to different treatments, and for fragments originating from different regions along the anterior-posterior and medio-lateral axis. This allows for a direct comparison between computational and experimental regeneration outcomes. Vector transport of morphogens was identified as a fundamental requirement to account for virtually scale-free self-assembly of the morphogen gradients observed in planarian homeostasis and regeneration. The model correctly describes altered body-plans following many known experimental manipulations, and accurately predicts outcomes of novel cutting scenarios, which we tested. We show that the vector transport field coincides with the alignment of nerve axons distributed throughout the planarian tissue, and demonstrate that the head-tail axis is controlled by the net polarity of neurons in a regenerating fragment. This model provides a comprehensive framework for mechanistically understanding fundamental aspects of body-plan regulation, and sheds new light on the role of the nervous system in directing growth and form., Competing Interests: The authors have declared that no competing interests exist.

Planarian behavior, physiology, and pattern control offer profound lessons for regenerative medicine, evolutionary biology, morphogenetic engineering, robotics, and unconventional computation. Despite recent advances in the molecular genetics of stem cell differentiation, this model organism's remarkable anatomical homeostasis provokes us with truly fundamental puzzles about the origin of large-scale shape and its relationship to the genome. In this review article, we first highlight several deep mysteries about planarian regeneration in the context of the current paradigm in this field. We then review recent progress in understanding of the physiological control of an endogenous, bioelectric pattern memory that guides regeneration, and how modulating this memory can permanently alter the flatworm's target morphology. Finally, we focus on computational approaches that complement reductive pathway analysis with synthetic, systems-level understanding of morphological decision-making. We analyze existing models of planarian pattern control and highlight recent successes and remaining knowledge gaps in this interdisciplinary frontier field., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

A key requirement of tissue/organ regeneration is the ability to induce appropriate shape in situ. Regenerated structures need to be integrated with pre-existing ones, through the combined regulation of new tissue growth and the scaling of surrounding tissues. This requires a tightly coordinated control of individual cell functions such as proliferation and stem cell differentiation. While great strides have been made in elucidating cell growth and differentiation mechanisms, how overall shape is generated during regeneration remains unknown. This is because a significant gap remains in our understanding of how cell behaviors are coordinated at the level of tissues and organs. The highly regenerative planarian flatworm has emerged as an important model for defining and understanding regenerative shape mechanisms. This review provides an overview of the main processes known to regulate tissue and animal shape during planarian regeneration: adult stem cell regulation, the reestablishment of body axes, tissue remodeling in pre-existing structures, organ scaling and the maintenance of body proportion, and the bioelectrical regulation of animal morphology. In order for the field to move forward, it will be necessary to identify shape mutants as a means to uncover the molecular mechanisms that synchronize all these separate processes to produce the worm's final regenerative shape. This knowledge will also aid efforts to define the mechanisms that control the termination of regenerative processes., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Large gradient high magnetic field (LG-HMF) is a powerful tool to study the effects of altered gravity on organisms. In our study, a platform for the long-term culture of aquatic organisms was designed based on a special superconducting magnet with an LG-HMF, which can provide three apparent gravity levels (µ g, 1 g, and 2 g), along with a control condition on the ground. Planarians, Dugesia japonica, were head-amputated and cultured for 5 days in a platform for head reconstruction. After planarian head regeneration, all samples were taken out from the superconducting magnet for a behavioral test under geomagnetic field and normal gravity conditions. To analyze differences among the four groups, four aspects of the planarians were considered, including head regeneration rate, phototaxis response, locomotor velocity, and righting behavior. Data showed that there was no significant difference in the planarian head regeneration rate under simulated altered gravity. According to statistical analysis of the behavioral test, all of the groups had normal functioning of the phototaxis response, while the planarians that underwent head reconstruction under the microgravity environment had significantly slower locomotor velocity and spent more time in righting behavior. Furthermore, histological staining and immunohistochemistry results helped us reveal that the locomotor system of planarians was affected by the simulated microgravity environment. We further demonstrated that the circular muscle of the planarians was weakened (hematoxylin and eosin staining), and the epithelial cilia of the planarians were reduced (anti-acetylated tubulin staining) under the simulated microgravity environment. Bioelectromagnetics. 2018;39:428-440. © 2018 Wiley Periodicals, Inc., (© 2018 Wiley Periodicals, Inc.)

Land planarians have a simple anatomy and simple behavioral repertoire in relation to most bilaterian animals, which makes them adequate for the study of biological processes. In this study, we investigate the behavior of land planarians during interaction events with other invertebrates found in the same environment. We observed 16 different behavioral units, including seven different capture behaviors and three different prey ingestion behaviors. The capture behavior varied from very simple, such as simply covering the prey with the body, to more complex ones, including two forms of tube formation that are described for the first time. In general, the capture behaviors were similar among different predators but different for different prey. Similarly, prey ingestion type was more related to prey type than to predator species, with small soft prey being swallowed without fragmentation, large prey being crushed, and prey with a hard skeleton being perforated. Considering that land planarians face limitations due to their lack of efficient ways to retain water, thus being highly dependent on a moist environment, the set of behaviors shown by them in this study was considerably rich, especially concerning strategies to capture prey., (Copyright © 2017 Elsevier GmbH. All rights reserved.)

Hundreds of planarian species exist worldwide, representing a rich phenotypic diversity. This chapter presents an overview of the morphology and anatomy of various taxonomic groups of planarian flatworms, focusing on features enabling recognition and identification of the animals. The most recent view on the phylogenetic relationships of the planarians is presented, together with geographic distribution patterns of major groups of triclads. The chapter concludes with a brief methodological section outlining species identification on basis of anatomical features. In conjunction with the established laboratory model species, the phenotypic diversity of planarians provides rich opportunities for comparative studies, which this chapter aims to inspire.

The ability to regenerate missing body parts exists throughout the animal kingdom. Positional information is crucial for regeneration, but how it is harboured and used by differentiated tissues is poorly understood. In planarians, positional information has been identified from study of phenotypes caused by RNA interference in which the wrong tissues are regenerated. For example, inhibition of the Wnt signalling pathway leads to regeneration of heads in place of tails. Characterization of these phenotypes has led to the identification of position control genes (PCGs)-genes that are expressed in a constitutive and regional manner and are associated with patterning. Most PCGs are expressed within planarian muscle; however, how muscle is specified and how different muscle subsets affect regeneration is unknown. Here we show that different muscle fibres have distinct regulatory roles during regeneration in the planarian Schmidtea mediterranea. myoD is required for formation of a specific muscle cell subset: the longitudinal fibres, oriented along the anterior-posterior axis. Loss of longitudinal fibres led to complete regeneration failure because of defects in regeneration initiation. A different transcription factor-encoding gene, nkx1-1, is required for the formation of circular fibres, oriented along the medial-lateral axis. Loss of circular fibres led to a bifurcated anterior-posterior axis with fused heads forming in single anterior blastemas. Whereas muscle is often viewed as a strictly contractile tissue, these findings reveal that different muscle types have distinct and specific regulatory roles in wound signalling and patterning to enable regeneration.

Cytoplasmic polyadenylation is a mechanism of mRNA regulation prevalent in metazoan germ cells; it is largely dependent on Cytoplasmic Polyadenylation Element Binding proteins (CPEBs). Two CPEB homologs were identified in the planarian Schmidtea mediterranea. Smed-CPEB1 is expressed in ovaries and yolk glands of sexually mature planarians, and required for oocyte and yolk gland development. In contrast, Smed-CPEB2 is expressed in the testes and the central nervous system; its function is required for spermatogenesis as well as non-autonomously for development of ovaries and accessory reproductive organs. Transcriptome analysis of CPEB knockdown animals uncovered a comprehensive collection of molecular markers for reproductive structures in S. mediterranea, including ovaries, testes, yolk glands, and the copulatory apparatus. Analysis by RNA interference revealed contributions for a dozen of these genes during oogenesis, spermatogenesis, or capsule formation. We also present evidence suggesting that Smed-CPEB2 promotes translation of Neuropeptide Y-8, a prohormone required for planarian sexual maturation. These findings provide mechanistic insight into potentially conserved processes of germ cell development, as well as events involved in capsule deposition by flatworms., (Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.)

How positional information instructs adult tissue maintenance is poorly understood. Planarians undergo whole-body regeneration and tissue turnover, providing a model for adult positional information studies. Genes encoding secreted and transmembrane components of multiple developmental pathways are predominantly expressed in planarian muscle cells. Several of these genes regulate regional identity, consistent with muscle harboring positional information. Here, single-cell RNA-sequencing of 115 muscle cells from distinct anterior-posterior regions identified 44 regionally expressed genes, including multiple Wnt and ndk/FGF receptor-like (ndl/FGFRL) genes. Two distinct FGFRL-Wnt circuits, involving juxtaposed anterior FGFRL and posterior Wnt expression domains, controlled planarian head and trunk patterning. ndl-3 and wntP-2 inhibition expanded the trunk, forming ectopic mouths and secondary pharynges, which independently extended and ingested food. fz5/8-4 inhibition, like that of ndk and wntA, caused posterior brain expansion and ectopic eye formation. Our results suggest that FGFRL-Wnt circuits operate within a body-wide coordinate system to control adult axial positioning.

Background: We investigated whether images obtained through X-ray micro-computed tomography (μCT) can be used in conjunction with traditional methods for morphological studies of soft-bodied land planarians. μCT is non-invasive and provides true-to-scale three-dimensional imagery at high resolution. We compared μCT-based images of a recently described land planarian species of Obama otavioi (Platyhelminthes) with those obtained from light microphotography of histological sections, most of which were also digitized at high magnification., Findings: The specimens studied were collected in 2012. Subsequent μCT-based images of the stained body of a paratype show nearly all morphological features provided by traditional histology, with the exception of particularly minute structures, smaller than 5 μm, such as the sensory pits and single muscle fibers, which are best visible on traditional histological sections. Because the technique is non-destructive, the scanned specimen is preserved without damage. The raw and derivative μCT data and virtual histological sections are freely available in GigaDB., Conclusions: The μCT datasets of these stained soft-bodied organisms reveal images of external and internal structures that support previous taxonomic studies. This technique can be particularly important for non-destructively revealing internal details of whole museum specimens at a faster rate than histology alone. High-resolution virtual histological slides also allow further searches for new, previously unstudied morphological features. The use of X-ray equipment with higher resolution can enable smaller sensory organ and muscle fiber details to be seen. The image sets, μCT-based images and digitized histological slides can be disseminated without the constraints of specimen loans.

The shape of an animal body plan is constructed from protein components encoded by the genome. However, bioelectric networks composed of many cell types have their own intrinsic dynamics, and can drive distinct morphological outcomes during embryogenesis and regeneration. Planarian flatworms are a popular system for exploring body plan patterning due to their regenerative capacity, but despite considerable molecular information regarding stem cell differentiation and basic axial patterning, very little is known about how distinct head shapes are produced. Here, we show that after decapitation in G. dorotocephala, a transient perturbation of physiological connectivity among cells (using the gap junction blocker octanol) can result in regenerated heads with quite different shapes, stochastically matching other known species of planaria (S. mediterranea, D. japonica, and P. felina). We use morphometric analysis to quantify the ability of physiological network perturbations to induce different species-specific head shapes from the same genome. Moreover, we present a computational agent-based model of cell and physical dynamics during regeneration that quantitatively reproduces the observed shape changes. Morphological alterations induced in a genomically wild-type G. dorotocephala during regeneration include not only the shape of the head but also the morphology of the brain, the characteristic distribution of adult stem cells (neoblasts), and the bioelectric gradients of resting potential within the anterior tissues. Interestingly, the shape change is not permanent; after regeneration is complete, intact animals remodel back to G. dorotocephala-appropriate head shape within several weeks in a secondary phase of remodeling following initial complete regeneration. We present a conceptual model to guide future work to delineate the molecular mechanisms by which bioelectric networks stochastically select among a small set of discrete head morphologies. Taken together, these data and analyses shed light on important physiological modifiers of morphological information in dictating species-specific shape, and reveal them to be a novel instructive input into head patterning in regenerating planaria.

Two new land planarian species, collected in the State of Rio de Janeiro, Brazil, are described. Their external aspect is similar to that of Imbira marcusi Carbayo et al., 2013 and Pseudogeoplana theresopolitana (Schirch, 1929), respectively. The analysis of the internal organs, however, revealed they belong to the genus Cratera. The male copulatory organs of one species is very different from any other geoplaninid, for the penis papilla holds a large, distal cavity receiving the ejaculatory duct and, furthermore, the papilla projects vertically downwards from the roof of the male atrium. Thus we consider it as a new species, Cratera cuarassu sp. nov. The second species differs from its congeners in that the dorsal insertion of the penis papilla is anterior to the ventral one, and in that the female atrium is narrowed in the anterior portion. The species was found in the type locality of Pseudogeoplana theresopolitana (Schirch, 1929) and compares well with it in the external features. However, since its internal organs are unknown and the type material of the species is seemingly lost, we describe it as Cratera anamariae Carbayo, sp. nov.

Land planarians are an interesting group of free-living flatworms that can be useful as bioindicators because of their high sensitivity to environmental changes and low dispersal capacity. In this study, we describe and compare assemblages of land planarians from areas with different conservation degrees of the Interior Atlantic Forest (Misiones, Argentina), and assess factors that could be related to their abundance and richness. Eight sites were tracked in search of land planarians in Reserva de Vida Silvestre Urugua-í (RVSU) and Campo Anexo Manuel Belgrano (CAMB). Diurnal and nocturnal surveys were performed in each site along nine sampling campaigns. We collected 237 individuals belonging to 18 species of the subfamily Geoplaninae. All sites were dominated by Geoplana sp. 1 and Pasipha hauseri. The richness estimators showed that there would be more species in RVSU than in CAMB. The abundance and richness of land planarians was high during the night and after rainfalls, suggesting an increased activity of flatworms under such conditions. The abundance and richness of land planarians were also related to the conservation condition of the sites. Disturbed sites showed less abundance and richness, and were segregated from non-disturbed ones by nmMDS analysis. Beta diversity between sites was higher than expected, indicating that the species turnover between sites contributed more to the total richness (gamma diversity) than the alpha diversity.

Species capable of regenerating lost body parts occur throughout the animal kingdom, yet close relatives are often regeneration incompetent. Why in the face of 'survival of the fittest' some animals regenerate but others do not remains a fascinating question. Planarian flatworms are well known and studied for their ability to regenerate from minute tissue pieces, yet species with limited regeneration abilities have been described even amongst planarians. Here we report the characterization of the regeneration defect in the planarian Dendrocoelum lacteum and its successful rescue. Tissue fragments cut from the posterior half of the body of this species are unable to regenerate a head and ultimately die. We find that this defect originates during the early stages of head specification, which require inhibition of canonical Wnt signalling in other planarian species. Notably, RNA interference (RNAi)-mediated knockdown of Dlac-β-catenin-1, the Wnt signal transducer, restored the regeneration of fully functional heads on tail pieces, rescuing D. lacteum's regeneration defect. Our results demonstrate the utility of comparative studies towards the reactivation of regenerative abilities in regeneration-deficient animals. Furthermore, the availability of D. lacteum as a regeneration-impaired planarian model species provides a first step towards elucidating the evolutionary mechanisms that ultimately determine why some animals regenerate and others do not.

ImagePlane is a modular pipeline for automated, high-throughput image analysis and information extraction. Designed to support planarian research, ImagePlane offers a self-parameterizing adaptive thresholding algorithm; an algorithm that can automatically segment animals into anterior-posterior/left-right quadrants for automated identification of region-specific differences in gene and protein expression; and a novel algorithm for quantification of morphology of animals, independent of their orientations and sizes. ImagePlane also provides methods for automatic report generation, and its outputs can be easily imported into third-party tools such as R and Excel. Here we demonstrate the pipeline's utility for identification of genes involved in stem cell proliferation in the planarian Schmidtea mediterranea. Although designed to support planarian studies, ImagePlane will prove useful for cell-based studies as well.

The planarian Dugesia japonica can regenerate a complete individual from a head, trunk or tail fragment via activation of somatic pluripotent stem cells. About a century ago, Thomas Hunt Morgan attempted to explain the extraordinary regenerative ability of planarians by positing two opposing morphogenetic gradients of formative "head stuff" and "tail stuff" along the anterior-posterior axis. However, Morgan's hypothesis remains open to debate. Here we show that extracellular signal-related kinase (ERK) and Wnt/β-catenin signalling pathways establish a solid framework for planarian regeneration. Our data suggest that ERK signalling forms a spatial gradient in the anterior region during regeneration. The fibroblast growth factor receptor-like gene nou-darake (which serves as an output of ERK signalling in the differentiating head) and posteriorly biased β-catenin activity negatively regulate ERK signalling along the anterior-posterior axis in distinct manners, and thereby posteriorize regenerating tissues outside the head region to reconstruct a complete head-to-tail axis. On the basis of this knowledge about D. japonica, we proposed that β-catenin signalling is responsible for the lack of head-regenerative ability of tail fragments in the planarian Phagocata kawakatsui, and our confirmation thereof supports the notion that posterior β-catenin signalling negatively modulates the ERK signalling involved in anteriorization across planarian species. These findings suggest that ERK signalling has a pivotal role in triggering globally dynamic differentiation of stem cells in a head-to-tail sequence through a default program that promotes head tissue specification in the absence of posteriorizing signals. Thus, we have confirmed the broad outline of Morgan's hypothesis, and refined it on the basis of our proposed default property of planarian stem cells.

Variability of regenerative potential among animals has long perplexed biologists. On the basis of their exceptional regenerative abilities, planarians have become important models for understanding the molecular basis of regeneration. However, planarian species with limited regenerative abilities are also found. Despite the importance of understanding the differences between closely related, regenerating and non-regenerating organisms, few studies have focused on the evolutionary loss of regeneration, and the molecular mechanisms leading to such regenerative loss remain obscure. Here we examine Procotyla fluviatilis, a planarian with restricted ability to replace missing tissues, using next-generation sequencing to define the gene expression programs active in regeneration-permissive and regeneration-deficient tissues. We found that Wnt signalling is aberrantly activated in regeneration-deficient tissues. Notably, downregulation of canonical Wnt signalling in regeneration-deficient regions restores regenerative abilities: blastemas form and new heads regenerate in tissues that normally never regenerate. This work reveals that manipulating a single signalling pathway can reverse the evolutionary loss of regenerative potential.

Having an almost unlimited capacity to regenerate tissues lost to age and injury, planarians have long fascinated naturalists. In the Western hemisphere alone, their documented history spans more than 200 years. Planarians were described in the early 19th century as being 'immortal under the edge of the knife', and initial investigation of these remarkable animals was significantly influenced by studies of regeneration in other organisms and from the flourishing field of experimental embryology in the late 19th and early 20th centuries. This review strives to place the study of planarian regeneration into a broader historical context by focusing on the significance and evolution of knowledge in this field. It also synthesizes our current molecular understanding of the mechanisms of planarian regeneration uncovered since this animal's relatively recent entrance into the molecular-genetic age., (Copyright © 2012 Wiley Periodicals, Inc.)

Summary: Understanding the mechanisms governing the regeneration capabilities of many organisms is a fundamental interest in biology and medicine. An ever-increasing number of manipulation and molecular experiments are attempting to discover a comprehensive model for regeneration, with the planarian flatworm being one of the most important model species. Despite much effort, no comprehensive, constructive, mechanistic models exist yet, and it is now clear that computational tools are needed to mine this huge dataset. However, until now, there is no database of regenerative experiments, and the current genotype-phenotype ontologies and databases are based on textual descriptions, which are not understandable by computers. To overcome these difficulties, we present here Planform (Planarian formalization), a manually curated database and software tool for planarian regenerative experiments, based on a mathematical graph formalism. The database contains more than a thousand experiments from the main publications in the planarian literature. The software tool provides the user with a graphical interface to easily interact with and mine the database. The presented system is a valuable resource for the regeneration community and, more importantly, will pave the way for the application of novel artificial intelligence tools to extract knowledge from this dataset., Availability: The database and software tool are freely available at http://planform.daniel-lobo.com.

In this study we analyzed the phylogenetic relationships of eastern Mediterranean freshwater planarians of the genus Dugesia, estimated divergence times for the various clades, and correlated their phylogeographic patterns with geological and paleoclimatic events, in order to discover which evolutionary processes have shaped the present-day distribution of these animals. Specimens were collected from freshwater courses and lakes in continental and insular Greece. Genetic divergences and phylogenetic relationships were inferred by using the mitochondrial gene subunit I of cytochrome oxidase (COI) and the nuclear ribosomal internal transcribed spacer-1 (ITS-1) from 74 newly collected individuals from Greece. Divergence time estimates were obtained under a Bayesian framework, using the COI sequences. Two alternative geological dates for the isolation of Crete from the mainland were tested as calibration points. A clear phylogeographic pattern was present for Dugesia lineages in the Eastern Mediterranean. Morphological data, combined with information on genetic divergences, revealed that eight out of the nine known species were represented in the samples, while additional new, and still undescribed species were detected. Divergence time analyses suggested that Dugesia species became isolated in Crete after the first geological isolation of the island, and that their present distribution in the Eastern Mediterranean has been shaped mainly by vicariant events but also by dispersal. During the Messinian salinity crisis these freshwater planarians apparently were not able to cross the sea barrier between Crete and the mainland, while they probably did disperse between islands in the Aegean Sea. Their dependence on freshwater to survive suggests the presence of contiguous freshwater bodies in those regions. Our results also suggest a major extinction of freshwater planarians on the Peloponnese at the end of the Pliocene, while about 2Mya ago, when the current Mediterranean climate was established, these Peloponnese populations probably began to disperse again. At the end of the Pliocene or during the Pleistocene, mainland populations of Dugesia colonized the western coast, including the Ionian Islands, which were then part of the continent., (Copyright © 2012 Elsevier Inc. All rights reserved.)

Self-resolving inflammatory exudates and lipid mediator metabolomics recently uncovered a new family of potent anti-inflammatory and proresolving mediators biosynthesized by macrophages (MΦs), denoted maresins. Here we determined that maresin 1 (MaR1) produced by human MΦs from endogenous docosahexaenoic acid (DHA) matched synthetic 7R,14S-dihydroxydocosa-4Z,8E,10E,12Z,16Z,19Z-hexaenoic acid. The MaR1 alcohol groups and Z/E geometry of conjugated double bonds were matched using isomers prepared by total organic synthesis. MaR1's potent defining actions were confirmed with synthetic MaR1, i.e., limiting polymorphonuclear neutrophil (PMN) infiltration in murine peritonitis (ng/mouse range) as well as enhancing human macrophage uptake of apoptotic PMNs. At 1 nM, MaR1 was slightly more potent than resolvin D1 in stimulating human MΦ efferocytosis, an action not shared by leukotriene B(4). MaR1 also accelerated surgical regeneration in planaria, increasing the rate of head reappearance. On injury of planaria, MaR1 was biosynthesized from deuterium-labeled (d(5))-DHA that was blocked with lipoxygenase (LOX) inhibitor. MaR1 dose-dependently inhibited TRPV1 currents in neurons, blocked capsaicin (100 nM)-induced inward currents (IC(50) 0.49±0.02 nM), and reduced both inflammation- and chemotherapy-induced neuropathic pain in mice. These results demonstrate the potent actions of MaR1 in regulating inflammation resolution, tissue regeneration, and pain resolution. These findings suggest that chemical signals are shared in resolution cellular trafficking, a key process in tissue regeneration. Moreover, immunoresolvents of the innate immune response, such as MaR1, offer new opportunities for assessing MΦs and their local DHA metabolome in the return to tissue homeostasis.

There have been multiple claims that exposing water to a static magnetic field affects its properties which influence living systems. To test this hypothesis, planarian subsequent to dissection were maintained in spring water that had been previously exposed for only one day to one of three (16, 160, or 1,600 G) intensity static magnetic fields or to a reference condition. Although there was no significant difference in regeneration rates over the subsequent seven-day period, there was a statistically significant nonlinear effect for planarian mobility and diffusion rates. Both mobility rates and diffusion velocity of a liquid within the water that had been exposed to the 16 G field was about twice that for water exposed to the other intensities. These results imply that nonlinear biophysical effects may emerge under specific conditions of intensity ranges for particular volumes of water.

Free-living freshwater planarians are considered to have the potential for development as an experimental model for toxicological studies on xenobiotics, including metals. However, little was known about the distribution patterns of metals in the body of treated planarians. This study was conducted to determine the tissue distribution patterns of cadmium (Cd) in different body portions of the treated planarian, Dugesia japonica. Results showed that Cd accumulated in the head of planarians at a significantly higher concentration than in the tail. After examining the level of metallothionein (MT), we suggested that the tissue distribution pattern of Cd might be related to MT induction patterns. In contrast, in planarians treated with copper (Cu), neither the tissue accumulation of Cu nor the multiples of induction of MTs significantly differed between different portions. Furthermore, a higher Cd accumulation rate in the head of planarians caused more-severe oxidative stress to appear in this portion and also a higher susceptibility to a lethal concentration of Cd. Finally, both in vitro and in vivo acetylcholinesterase activities in both body portions of planarians were inhibited by Cd. The present study provides the first report that different metals are distributed in various body portions with different patterns in the planarian.

Bioactive peptides (i.e., neuropeptides or peptide hormones) represent the largest class of cell-cell signaling molecules in metazoans and are potent regulators of neural and physiological function. In vertebrates, peptide hormones play an integral role in endocrine signaling between the brain and the gonads that controls reproductive development, yet few of these molecules have been shown to influence reproductive development in invertebrates. Here, we define a role for peptide hormones in controlling reproductive physiology of the model flatworm, the planarian Schmidtea mediterranea. Based on our observation that defective neuropeptide processing results in defects in reproductive system development, we employed peptidomic and functional genomic approaches to characterize the planarian peptide hormone complement, identifying 51 prohormone genes and validating 142 peptides biochemically. Comprehensive in situ hybridization analyses of prohormone gene expression revealed the unanticipated complexity of the flatworm nervous system and identified a prohormone specifically expressed in the nervous system of sexually reproducing planarians. We show that this member of the neuropeptide Y superfamily is required for the maintenance of mature reproductive organs and differentiated germ cells in the testes. Additionally, comparative analyses of our biochemically validated prohormones with the genomes of the parasitic flatworms Schistosoma mansoni and Schistosoma japonicum identified new schistosome prohormones and validated half of all predicted peptide-encoding genes in these parasites. These studies describe the peptide hormone complement of a flatworm on a genome-wide scale and reveal a previously uncharacterized role for peptide hormones in flatworm reproduction. Furthermore, they suggest new opportunities for using planarians as free-living models for understanding the reproductive biology of flatworm parasites., Competing Interests: The authors have declared that no competing interests exist.

Wnt signal transduction plays a crucial role in stem cell proliferation and regeneration. When canonical Wnt signaling is low, heads develop, and when it is high, tails are formed. In planarians, Wnt transcription is activated by wounding in a beta-catenin-independent way. Hedgehog is one of the signals involved, because it induces regeneration of tails (instead of heads) through the activation of Wnt transcription. Depletion of Smad4 blocks regeneration entirely, which suggests that the bone morphogenetic protein signaling pathway and the Wnt pathway are required for regeneration and body patterning.

Planarians are an ideal model system to study in vivo the dynamics of adult pluripotent stem cells. However, our knowledge of the factors necessary for regulating the 'stemness' of the neoblasts, the adult stem cells of planarians, is sparse. Here, we report on the characterization of the first planarian member of the LSm protein superfamily, Smed-SmB, which is expressed in stem cells and neurons in Schmidtea mediterranea. LSm proteins are highly conserved key players of the splicing machinery. Our study shows that Smed-SmB protein, which is localized in the nucleus and the chromatoid body of stem cells, is required to safeguard the proliferative ability of the neoblasts. The chromatoid body, a cytoplasmatic ribonucleoprotein complex, is an essential regulator of the RNA metabolism required for the maintenance of metazoan germ cells. However, planarian neoblasts and neurons also rely on its functions. Remarkably, Smed-SmB dsRNA-mediated knockdown results in a rapid loss of organization of the chromatoid body, an impairment of the ability to post-transcriptionally process the transcripts of Smed-CycB, and a severe proliferative failure of the neoblasts. This chain of events leads to a quick depletion of the neoblast pool, resulting in a lethal phenotype for both regenerating and intact animals. In summary, our results suggest that Smed-SmB is an essential component of the chromatoid body, crucial to ensure a proper RNA metabolism and essential for stem cell proliferation.

More than 100 years ago, early workers realized that planarians offer an excellent system for regeneration studies. Another unique aspect of planarians is that they occupy an interesting phylogenetic position with respect to the nervous system in that they possess an evolutionarily primitive brain structure and can regenerate a functional brain from almost any tiny body fragment. Recent molecular studies have revisited planarian regeneration and revealed key information about the cellular and molecular mechanisms underlying brain regeneration in planarians. One of our great advances was identification of a gene, nou-darake, which directs the formation of a proper extrinsic environment for pluripotent stem cells to differentiate into brain cells in the planarian Dugesia japonica. Our recent findings have provided mechanistic insights into stem cell biology and also evolutionary biology.

This review aims to demonstrate the importance of freshwater planarians as model organisms, particularly emphasizing those characteristics of the animal that make them a good model to study autophagy. The aim of this review is to provide a better understanding of autophagy in this model for the nonplanarian reader, and elucidate the relevance of autophagy research in this peculiar model organism. Furthermore, I will try to synthesize the evidence showing the importance of autophagy in planarian body remodeling, and I will discuss some ideas about the role of autophagy in stem cell biology. In light of these new developments, it is likely that the planarian field will make an important contribution to the study of the molecular mechanisms involved in autophagy in the future.

Many animals produce continuous brainwaves, known as the electroencephalogram (EEG), but it is not known at what point in evolution the EEG developed. Planarians possess the most primitive form of brain, but still exhibit learning and memory behaviors. Here, we observed and characterized the EEG waveform of the planarian. We inserted a monopole electrode into the head of a planarian on a cold stage, and were able to observe the EEG at sub-microvolt amplitudes. The EEG had a continuous waveform, similar to that of evolutionarily advanced animals with more developed brains. Occasional myogenic potential spikes were observed in the EEG due to sticking of the electrode, but this was markedly diminished by cooling the sample, which enabled us to investigate the intrinsic character of the continuous EEG waveform. The frequency spectrum of the EEG was observed in the range of 0.1-5 Hz, showing a broad rise below 0.5 Hz and a monotonic decrease above 1 Hz, apparently following the 1/f law. The intensity of the total EEG diminished during anesthesia by cooling to 2-3 degrees C, and recovered when the sample was warmed to about 10 degrees C. The EEG signal was sustained for 30-40 min, and gradually weakened as the animal died. Stimulation of the planarian with water vibration at 0.5-2 Hz induced chaotic resonance with a broad peak spectrum of around the stimulation frequency. Strong illumination suppressed the EEG signals for several minutes, with the degree of suppression positively correlating with the intensity of the light. This provides evidence that the EEG responds to optical signals, although there were no synchronous reactions to light flashes. The continuous EEG waveform suggests the existence of feedback loop circuits in the neural network of the planarian, which was supposed in electric shock memory experiments [McConnell JV, Cornwell P, Clay M (1960) An apparatus for conditioning planaria. Am J Psychol 73:618-622]. However, because of the broad band character of chaotic resonance observed, these loops appear to be loose couplings between ganglia.

The obtained and published data on pharynx regeneration in planarians have been reviewed. Planarians can regenerate from a small body fragment and restore all missing organs including the pharynx. The pharynx is a relatively autonomous organ with a differentiated structure and specialized function. Pharynx regeneration has specific features, and its studies are of considerable theoretical interest. Pharynx regeneration can also be a convenient model to study the molecular mechanisms of regeneration that remain undisclosed. In addition, this model can be used to test biologically active compounds in order to elucidate their effect on morphogenesis. This subject of investigation benefits by a simpler and more adequate analysis as well as a possibility to use large numbers of animals and small quantities of analyzed substances.

Freshwater planarians are an emerging model in which to study regeneration at the molecular level. These animals can regenerate a complete central nervous system (CNS) in only a few days. In recent years, hundreds of genes expressed in the nervous system have been identified in two popular planarian species used by several laboratories: Dugesia japonica and Schmidtea mediterranea. Functional analyses of some of those neural genes have allowed the process of CNS regeneration to begin to be elucidated in those animals. However, additional work is required to characterize the different neuronal populations. Thus, the identification or generation of antibodies that act as markers for specific neuronal cell types would be extremely useful not only in obtaining a more detailed characterization of the planarian nervous system but also for the analysis of phenotypes obtained by RNA interference. Here, I have used five different antibodies to describe different neuronal populations in the freshwater planarian S. mediterranea. This study represents a first step in characterizing the organization of the nervous system of this species at the cellular level.

Understanding stem cells is a major goal of current research because of its potential medical applications. Although great advances have been made, such as the culturing and differentiation of embryonic stem cells and reprogramming of cell fates, many basic questions remain unanswered. Describing the mechanisms underlying regeneration will help to understand the biology of stem cells and therefore to control their behavior. While regeneration is being studied in a variety of models, the planarian is particularly noteworthy. In this model system a fragment as small as 1/279 of the animal can regenerate completely within a few weeks. These animals can also grow and degrow--specifically degenerating certain tissues--according to environmental conditions, thus demonstrating a complete control of their stem cell dynamics. However, one of the most interesting aspects of the planarian model system is the presence of a unique type of stem cell that can differentiate into all cell types found in the organism, including the germ line. This represents a simple, extremely powerful, and accessible stem cell system in which to address a variety of important questions. In the last ten years, molecular, cellular, and bioinformatics tools have been established for use in this model, making it ideally placed for in vivo analysis of stem cells in their natural environment without ethical complications.