Elsevier BV Acta Medica Okayama 2589-0042 28 9 2025 Extensive urine production in euryhaline red stingray for adaptation to hypoosmotic environments 113274 EN Naotaka Aburatani Atmosphere and Ocean Research Institute, The University of Tokyo Wataru Takagi Atmosphere and Ocean Research Institute, The University of Tokyo Marty Kwok-Shing Wong Atmosphere and Ocean Research Institute, The University of Tokyo Nobuhiro Ogawa Atmosphere and Ocean Research Institute, The University of Tokyo Shigehiro Kuraku Department of Genomics and Evolutionary Biology, National Institute of Genetics Mana Sato Department of Genomics and Evolutionary Biology, National Institute of Genetics Kazuhiro Saito Ushimado Marine Institute, Faculty of Science, Okayama University Waichiro Godo Ushimado Marine Institute, Faculty of Science, Okayama University Tatsuya Sakamoto Ushimado Marine Institute, Faculty of Science, Okayama University Susumu Hyodo Atmosphere and Ocean Research Institute, The University of Tokyo Maintaining water balance is a prerequisite for all organisms. Euryhaline elasmobranchs face the severest water-influx potential in fresh water (FW), as they retain high concentrations of urea even in hypotonic environments. To elucidate how they overcome this osmotic challenge, we assessed urine output in conscious euryhaline red stingrays (Hemitrygon akajei). Following acclimation to 5% diluted seawater, the stingrays increased urinary output by 87-fold—the greatest change observed in vertebrates—partly due to 6.8-fold increase in glomerular filtration rate (GFR). In the nephron, expressions of Aquaporin-1 (Aqp1), Aqp3, and Aqp15 were strongly downregulated in FW, indicating that tubular diuresis bridges the gap between GFR and final urine volume. Meanwhile, FW-acclimation upregulated Aqp1 and Aqp4 in the distinct bundle structure, which promotes urea reabsorption. Euryhaline elasmobranchs resolve the huge osmotic challenge of FW by excreting massive amounts of water and retaining osmolytes including urea through coordinated regulation of GFR and Aqp expressions. No potential conflict of interest relevant to this article was reported.

Zoological Society of Japan Acta Medica Okayama 0289-0003 41 3 2024 Volume X-Ray Micro-Computed Tomography Analysis of the Early Cephalized Central Nervous System in a Marine Flatworm, Stylochoplana pusilla 281 289 EN Takanori Ikenaga Graduate School of Science and Engineering, Kagoshima University Aoshi Kobayashi Ushimado Marine Institute (UMI), Faculty of Environmental, Life, Natural Science and Technology, Okayama University Akihisa Takeuchi Japan Synchrotron Radiation Research Institute/SPring-8 Kentaro Uesugi Japan Synchrotron Radiation Research Institute/SPring-8 Takanobu Maezawa Department of Integrated Science and Technology, National Institute of Technology, Tsuyama College Norito Shibata Department of Integrated Science and Technology, National Institute of Technology, Tsuyama College Tatsuya Sakamoto Ushimado Marine Institute (UMI), Faculty of Environmental, Life, Natural Science and Technology, Okayama University Hirotaka Sakamoto Ushimado Marine Institute (UMI), Faculty of Environmental, Life, Natural Science and Technology, Okayama University Platyhelminthes are a phylum of simple bilaterian invertebrates with prototypic body systems. Compared with non-bilaterians such as cnidarians, the bilaterians are likely to exhibit integrated free-moving behaviors, which require a concentrated nervous system “brain” rather than the distributed nervous system of radiatans. Marine flatworms have an early cephalized ‘central’ nervous system compared not only with non-bilaterians but also with parasitic flatworms or freshwater planarians. In this study, we used the marine flatworm Stylochoplana pusilla as an excellent model organism in Platyhelminthes because of the early cephalized central nervous system. Here, we investigated the three-dimensional structures of the flatworm central nervous system by the use of X-ray micro-computed tomography (micro-CT) in a synchrotron radiation facility. We found that the obtained tomographic images were sufficient to discriminate some characteristic structures of the nervous system, including nerve cords around the cephalic ganglion, mushroom body-like structures, and putative optic nerves forming an optic commissure-like structure. Through the micro-CT imaging, we could obtain undistorted serial section images, permitting us to visualize precise spatial relationships of neuronal subpopulations and nerve tracts. 3-D micro-CT is very effective in the volume analysis of the nervous system at the cellular level; the methodology is straightforward and could be applied to many other non-model organisms. No potential conflict of interest relevant to this article was reported.

Public Library Science Acta Medica Okayama 1932-6203 17 12 2022 Behavioural osmoregulation during land invasion in fish: Prandial drinking and wetting of the dry skin e0277968 EN Yukitoshi Katayama Ushimado Marine Institute, Faculty of Science, Okayama University Takehiro Tsukada Department of Biomolecular Science, Toho University Susumu Hyodo Laboratory of Physiology, Atmosphere and Ocean Research Institute, University of Tokyo Hirotaka Sakamoto Ushimado Marine Institute, Faculty of Science, Okayama University Tatsuya Sakamoto Ushimado Marine Institute, Faculty of Science, Okayama University Osmoregulatory behaviours should have evolutionarily modified for terrestrialisation of vertebrates. In mammals, sensations of buccal food and drying have immediate effects on postprandial thirst to prevent future systemic dehydration, and is thereby considered to be 'anticipatory thirst'. However, it remains unclear whether such an anticipatory response has been acquired in the non-tetrapod lineage. Using the mudskipper goby (Periophthalmus modestus) as a semi-terrestrial ray-finned fish, we herein investigated postprandial drinking and other unique features like full-body 'rolling' over on the back although these behaviours had not been considered to have osmoregulatory functions. In our observations on tidal flats, mudskippers migrated into water areas within a minute after terrestrial eating, and exhibited rolling behaviour with accompanying pectoral-fin movements. In aquarium experiments, frequency of migration into a water area for drinking increased within a few minutes after eating onset, without systemic dehydration. During their low humidity exposure, frequency of the rolling behaviour and pectoral-fin movements increased by more than five times to moisten the skin before systemic dehydration. These findings suggest anticipatory responses which arise from oral/gastrointestinal and cutaneous sensation in the goby. These sensation and motivation seem to have evolved in distantly related species in order to solve osmoregulatory challenges during terrestrialisation. No potential conflict of interest relevant to this article was reported.

The Royal Society Acta Medica Okayama 0962-8452 289 1985 2022 Footedness for scratching itchy eyes in rodents 20221126 EN Yukitoshi Katayama Ushimado Marine Institute (UMI), Graduate School of Natural Science and Technology, Okayama University, Ushimado, Setouchi Ayane Miura Ushimado Marine Institute (UMI), Graduate School of Natural Science and Technology, Okayama University, Ushimado, Setouchi Tatsuya Sakamoto Ushimado Marine Institute (UMI), Graduate School of Natural Science and Technology, Okayama University, Ushimado, Setouchi Keiko Takanami Mouse Genomics Resources Laboratory, National Institute of Genetics, Yata, Mishima Hirotaka Sakamoto Ushimado Marine Institute (UMI), Graduate School of Natural Science and Technology, Okayama University, Ushimado, Setouchi The neural bases of itchy eye transmission remain unclear compared with those involved in body itch. Here, we show in rodents that the gastrin-releasing peptide receptor (GRPR) of the trigeminal sensory system is involved in the transmission of itchy eyes. Interestingly, we further demonstrate a difference in scratching behaviour between the left and right hindfeet in rodents; histamine instillation into the conjunctival sac of both eyes revealed right-foot biased laterality in the scratching movements. Unilateral histamine instillation specifically induced neural activation in the ipsilateral sensory pathway, with no significant difference between the activations following left- and right-eye instillations. Thus, the behavioural laterality is presumably due to right-foot preference in rodents. Genetically modified rats with specific depletion of Grpr-expressing neurons in the trigeminal sensory nucleus caudalis of the medulla oblongata exhibited fewer and shorter histamine-induced scratching movements than controls and eliminated the footedness. These results taken together indicate that the Grp-expressing neurons are required for the transmission of itch sensation from the eyes, but that foot preference is generated centrally. These findings could open up a new field of research on the mechanisms of the laterality in vertebrates and also offer new potential therapeutic approaches to refractory pruritic eye disorders. No potential conflict of interest relevant to this article was reported.

CELL PRESS Acta Medica Okayama 2589-0042 25 7 2022 Lattice-patterned collagen fibers and their dynamics in axolotl skin regeneration 104524 EN Rena Kashimoto Division of Earth, Life, and Molecular Sciences, Graduate School of Natural Science and Technology, Okayama University Saya Furukawa Department of Biological Sciences, Faculty of Science, Okayama University Sakiya Yamamoto Department of Biological Sciences, Faculty of Science, Okayama University Yasuhiro Kamei National Institute for Basic Biology (NIBB), National Institutes for Natural Sciences Joe Sakamoto National Institute for Basic Biology (NIBB), National Institutes for Natural Sciences Shigenori Nonaka National Institute for Basic Biology (NIBB), National Institutes for Natural Sciences Tomonobu M. Watanabe Laboratory for Comprehensive Bioimaging, RIKEN Center for Biosystems Dynamics Research (BDR) Tatsuya Sakamoto Division of Earth, Life, and Molecular Sciences, Graduate School of Natural Science and Technology, Okayama University Hirotaka Sakamoto Division of Earth, Life, and Molecular Sciences, Graduate School of Natural Science and Technology, Okayama University Akira Satoh Research Core for Interdisciplinary Sciences (RCIS), Okayama University The morphology of collagen-producing cells and the structure of produced collagen in the dermis have not been well-described. This lack of insights has been a serious obstacle in the evaluation of skin regeneration. We succeeded in visualizing collagen-producing cells and produced collagen using the axolotl skin, which is highly transparent. The visualized dermal collagen had a lattice-like structure. The collagen-producing fibroblasts consistently possessed the lattice-patterned filopodia along with the lattice-patterned collagen network. The dynamics of this lattice-like structure were also verified in the skin regeneration process of axolotls, and it was found that the correct lattice-like structure was not reorganized after simple skin wounding but was reorganized in the presence of nerves. These findings are not only fundamental insights in dermatology but also valuable insights into the mechanism of skin regeneration. No potential conflict of interest relevant to this article was reported.

American Association for the Advancement of Science (AAAS) Acta Medica Okayama 2375-2548 8 9 2022 Vasopressin-oxytocin–type signaling is ancient and has a conserved water homeostasis role in euryhaline marine planarians eabk0331 EN Aoshi Kobayashi Ushimado Marine Institute (UMI), Graduate School of Natural Science and Technology, Okayama University Mayuko Hamada Ushimado Marine Institute (UMI), Graduate School of Natural Science and Technology, Okayama University Masa-aki Yoshida Oki Marine Biological Station, Shimane University Yasuhisa Kobayashi Ushimado Marine Institute (UMI), Graduate School of Natural Science and Technology, Okayama University Naoaki Tsutsui Ushimado Marine Institute (UMI), Graduate School of Natural Science and Technology, Okayama University Toshio Sekiguchi Noto Marine Laboratory, Institute of Nature and Environmental Technology, Division of Marine Environmental Studies, Kanazawa University Yuta Matsukawa Ushimado Marine Institute (UMI), Graduate School of Natural Science and Technology, Okayama University Sho Maejima Ushimado Marine Institute (UMI), Graduate School of Natural Science and Technology, Okayama University Joseph J. Gingell Vertex Pharmaceuticals (Europe) Ltd. Shoko Sekiguchi Ushimado Marine Institute (UMI), Graduate School of Natural Science and Technology, Okayama University Ayumu Hamamoto Ushimado Marine Institute (UMI), Graduate School of Natural Science and Technology, Okayama University Debbie L. Hay School of Biological Sciences and Maurice Wilkins Centre for Molecular Biodiscovery, University of Auckland John F. Morris Department of Physiology, Anatomy, and Genetic, Le Gros Clark Building, University of Oxford Tatsuya Sakamoto Ushimado Marine Institute (UMI), Graduate School of Natural Science and Technology, Okayama University Hirotaka Sakamoto Ushimado Marine Institute (UMI), Graduate School of Natural Science and Technology, Okayama University, Ushimado Vasopressin/oxytocin (VP/OT)–related peptides are essential for mammalian antidiuresis, sociosexual behavior, and reproduction. However, the evolutionary origin of this peptide system is still uncertain. Here, we identify orthologous genes to those for VP/OT in Platyhelminthes, intertidal planarians that have a simple bilaterian body structure but lack a coelom and body-fluid circulatory system. We report a comprehensive characterization of the neuropeptide derived from this VP/OT-type gene, identifying its functional receptor, and name it the “platytocin” system. Our experiments with these euryhaline planarians, living where environmental salinities fluctuate due to evaporation and rainfall, suggest that platytocin functions as an “antidiuretic hormone” and also organizes diverse actions including reproduction and chemosensory-associated behavior. We propose that bilaterians acquired physiological adaptations to amphibious lives by such regulation of the body fluids. This neuropeptide-secreting system clearly became indispensable for life even without the development of a vascular circulatory system or relevant synapses. No potential conflict of interest relevant to this article was reported.

MDPI Acta Medica Okayama 1422-0067 22 19 2021 Sexual Experience Induces the Expression of Gastrin-Releasing Peptide and Oxytocin Receptors in the Spinal Ejaculation Generator in Rats 10362 EN Takumi Oti Ushimado Marine Institute (UMI), Graduate School of Natural Science and Technology, Okayama University Ryota Ueda Ushimado Marine Institute (UMI), Graduate School of Natural Science and Technology, Okayama University Ryoko Kumagai Department of Animal Sciences, Teikyo University of Science Junta Nagafuchi Ushimado Marine Institute (UMI), Graduate School of Natural Science and Technology, Okayama University Takashi Ito Ushimado Marine Institute (UMI), Graduate School of Natural Science and Technology, Okayama University Tatsuya Sakamoto Ushimado Marine Institute (UMI), Graduate School of Natural Science and Technology, Okayama University Yasuhiko Kondo Department of Animal Sciences, Teikyo University of Science Hirotaka Sakamoto Ushimado Marine Institute (UMI), Graduate School of Natural Science and Technology, Okayama University Male sexual function in mammals is controlled by the brain neural circuits and the spinal cord centers located in the lamina X of the lumbar spinal cord (L3-L4). Recently, we reported that hypothalamic oxytocin neurons project to the lumbar spinal cord to activate the neurons located in the dorsal lamina X of the lumbar spinal cord (dXL) via oxytocin receptors, thereby facilitating male sexual activity. Sexual experiences can influence male sexual activity in rats. However, how this experience affects the brain-spinal cord neural circuits underlying male sexual activity remains unknown. Focusing on dXL neurons that are innervated by hypothalamic oxytocinergic neurons controlling male sexual function, we examined whether sexual experience affects such neural circuits. We found that >50% of dXL neurons were activated in the first ejaculation group and similar to 30% in the control and intromission groups in sexually naive males. In contrast, in sexually experienced males, similar to 50% of dXL neurons were activated in both the intromission and ejaculation groups, compared to similar to 30% in the control group. Furthermore, sexual experience induced expressions of gastrin-releasing peptide and oxytocin receptors in the lumbar spinal cord. This is the first demonstration of the effects of sexual experience on molecular expressions in the neural circuits controlling male sexual activity in the spinal cord. No potential conflict of interest relevant to this article was reported.

Zoological Society of Japan Acta Medica Okayama 0289-0003 39 1 2022 Observing Phylum-Level Metazoan Diversity by Environmental DNA Analysis at the Ushimado Area in the Seto Inland Sea 157 165 EN Takeshi Kawashima National Institute of Genetics Masa-aki Yoshida Marine Biological Science Section, Education and Research Center Biological Resources, Faculty of Life and Environmental Science, Shimane University Hideyuki Miyazawa National Institute of Genetics Hiroaki Nakano Shimoda Marine Research Center, University of Tsukuba Natumi Nakano Department of Biology, Nara Medical University Tatsuya Sakamoto Ushimado Marine Institute, Okayama University Mayuko Hamada Ushimado Marine Institute, Okayama University The dynamics of microscopic marine plankton in coastal areas is a fundamental theme in marine biodiversity research, but studies have been limited because the only available methodology was collection of plankton using plankton-nets and microscopic observation. In recent years, environmental DNA (eDNA) analysis has exhibited potential for conducting comprehensive surveys of marine plankton diversity in water at fixed points and depths in the ocean. However, few studies have examined how eDNA analysis reflects the actual distribution and dynamics of organisms in the field, and further investigation is needed to determine whether it can detect distinct differences in plankton density in the field. To address this, we analyzed eDNA in seawater samples collected at 1 km intervals at three depths over a linear distance of approximately 3.0 km in the Seto Inland Sea. The survey area included a location with a high density of Acoela (Praesagittifera naikaiensis). However, the eDNA signal for this was little to none, and its presence would not have been noticed if we did not have this information beforehand. Meanwhile, eDNA analysis enabled us to confirm the presence of a species of Placozoa that was previously undiscovered in the area. In summary, our results suggest that the number of sequence reads generated from eDNA samples in our project was not sufficient to predict the density of a particular species. However, eDNA can be useful for detecting organisms that have been overlooked using other methods. No potential conflict of interest relevant to this article was reported.

MDPI Acta Medica Okayama 1422-0067 22 17 2021 Immunoelectron Microscopic Characterization of Vasopressin-Producing Neurons in the Hypothalamo-Pituitary Axis of Non-Human Primates by Use of Formaldehyde-Fixed Tissues Stored at-25 degrees C for Several Years 9180 EN Akito Otubo Ushimado Marine Institute (UMI), Graduate School of Natural Science and Technology, Okayama University Sho Maejima Ushimado Marine Institute (UMI), Graduate School of Natural Science and Technology, Okayama University Takumi Oti Ushimado Marine Institute (UMI), Graduate School of Natural Science and Technology, Okayama University Keita Satoh Ushimado Marine Institute (UMI), Graduate School of Natural Science and Technology, Okayama University Yasumasa Ueda Department of Physiology, Kyoto Prefectural University of Medicine John F. Morris Department of Physiology, Anatomy & Genetics, University of Oxford, South Parks Road Tatsuya Sakamoto Ushimado Marine Institute (UMI), Graduate School of Natural Science and Technology, Okayama University Hirotaka Sakamoto Ushimado Marine Institute (UMI), Graduate School of Natural Science and Technology, Okayama University Translational research often requires the testing of experimental therapies in primates, but research in non-human primates is now stringently controlled by law around the world. Tissues fixed in formaldehyde without glutaraldehyde have been thought to be inappropriate for use in electron microscopic analysis, particularly those of the brain. Here we report the immunoelectron microscopic characterization of arginine vasopressin (AVP)-producing neurons in macaque hypothalamo-pituitary axis tissues fixed by perfusion with 4% formaldehyde and stored at -25 degrees C for several years (4-6 years). The size difference of dense-cored vesicles between magnocellular and parvocellular AVP neurons was detectable in their cell bodies and perivascular nerve endings located, respectively, in the posterior pituitary and median eminence. Furthermore, glutamate and the vesicular glutamate transporter 2 could be colocalized with AVP in perivascular nerve endings of both the posterior pituitary and the external layer of the median eminence, suggesting that both magnocellular and parvocellular AVP neurons are glutamatergic in primates. Both ultrastructure and immunoreactivity can therefore be sufficiently preserved in macaque brain tissues stored long-term, initially for light microscopy. Taken together, these results suggest that this methodology could be applied to the human post-mortem brain and be very useful in translational research. No potential conflict of interest relevant to this article was reported.

Nature Research Acta Medica Okayama 2045-2322 11 1 2021 The gastrin-releasing peptide/bombesin system revisited by a reverse-evolutionary study considering Xenopus 13315 EN Asuka Hirooka Ushimado Marine Institute (UMI), Graduate School of Natural Science and Technology, Okayama University Mayuko Hamada Ushimado Marine Institute (UMI), Graduate School of Natural Science and Technology, Okayama University Daiki Fujiyama Ushimado Marine Institute (UMI), Graduate School of Natural Science and Technology, Okayama University Keiko Takanami Ushimado Marine Institute (UMI), Graduate School of Natural Science and Technology, Okayama University Yasuhisa Kobayashi Ushimado Marine Institute (UMI), Graduate School of Natural Science and Technology, Okayama University Takumi Oti Ushimado Marine Institute (UMI), Graduate School of Natural Science and Technology, Okayama University Yukitoshi Katayama Ushimado Marine Institute (UMI), Graduate School of Natural Science and Technology, Okayama University Tatsuya Sakamoto Ushimado Marine Institute (UMI), Graduate School of Natural Science and Technology, Okayama University Hirotaka Sakamoto Ushimado Marine Institute (UMI), Graduate School of Natural Science and Technology, Okayama University Bombesin is a putative antibacterial peptide isolated from the skin of the frog, Bombina bombina. Two related (bombesin-like) peptides, gastrin-releasing peptide (GRP) and neuromedin B (NMB) have been found in mammals. The history of GRP/bombesin discovery has caused little attention to be paid to the evolutionary relationship of GRP/bombesin and their receptors in vertebrates. We have classified the peptides and their receptors from the phylogenetic viewpoint using a newly established genetic database and bioinformatics. Here we show, by using a clawed frog (Xenopus tropicalis), that GRP is not a mammalian counterpart of bombesin and also that, whereas the GRP system is widely conserved among vertebrates, the NMB/bombesin system has diversified in certain lineages, in particular in frog species. To understand the derivation of GRP system in the ancestor of mammals, we have focused on the GRP system in Xenopus. Gene expression analyses combined with immunohistochemistry and Western blotting experiments demonstrated that GRP peptides and their receptors are distributed in the brain and stomach of Xenopus. We conclude that GRP peptides and their receptors have evolved from ancestral (GRP-like peptide) homologues to play multiple roles in both the gut and the brain as one of the 'gut-brain peptide' systems. No potential conflict of interest relevant to this article was reported.

MDPI Acta Medica Okayama 1422-0067 22 7 2021 In Vivo Electrophysiology of Peptidergic Neurons in Deep Layers of the Lumbar Spinal Cord after Optogenetic Stimulation of Hypothalamic Paraventricular Oxytocin Neurons in Rats 3400 EN Daisuke Uta Department of Applied Pharmacology, Faculty of Pharmaceutical Sciences, University of Toyama Takumi Oti Ushimado Marine Institute (UMI), Graduate School of Natural Science and Technology, Okayama University Tatsuya Sakamoto Ushimado Marine Institute (UMI), Graduate School of Natural Science and Technology, Okayama University Hirotaka Sakamoto Ushimado Marine Institute (UMI), Graduate School of Natural Science and Technology, Okayama University The spinal ejaculation generator (SEG) is located in the central gray (lamina X) of the rat lumbar spinal cord and plays a pivotal role in the ejaculatory reflex. We recently reported that SEG neurons express the oxytocin receptor and are activated by oxytocin projections from the paraventricular nucleus of hypothalamus (PVH). However, it is unknown whether the SEG responds to oxytocin in vivo. In this study, we analyzed the characteristics of the brain-spinal cord neural circuit that controls male sexual function using a newly developed in vivo electrophysiological technique. Optogenetic stimulation of the PVH of rats expressing channel rhodopsin under the oxytocin receptor promoter increased the spontaneous firing of most lamina X SEG neurons. This is the first demonstration of the in vivo electrical response from the deeper (lamina X) neurons in the spinal cord. Furthermore, we succeeded in the in vivo whole-cell recordings of lamina X neurons. In vivo whole-cell recordings may reveal the features of lamina X SEG neurons, including differences in neurotransmitters and response to stimulation. Taken together, these results suggest that in vivo electrophysiological stimulation can elucidate the neurophysiological response of a variety of spinal neurons during male sexual behavior. No potential conflict of interest relevant to this article was reported.

Wiley Acta Medica Okayama 0953-8194 32 8 2020 Vasopressin gene products are colocalised with corticotrophin‐releasing factor within neurosecretory vesicles in the external zone of the median eminence of the Japanese macaque monkey (Macaca fuscata) e12875 EN Akito Otubo Ushimado Marine Institute (UMI), Graduate School of Natural Science and Technology, Okayama University Natsuko Kawakami Ushimado Marine Institute (UMI), Graduate School of Natural Science and Technology, Okayama University Sho Maejima Ushimado Marine Institute (UMI), Graduate School of Natural Science and Technology, Okayama University Yasumasa Ueda Department of Physiology, Kyoto Prefectural University of Medicine John F. Morris Department of Physiology, Anatomy & Genetics, University of Oxford Tatsuya Sakamoto Ushimado Marine Institute (UMI), Graduate School of Natural Science and Technology, Okayama University Hirotaka Sakamoto Ushimado Marine Institute (UMI), Graduate School of Natural Science and Technology, Okayama University Arginine vasopressin (AVP), when released into portal capillaries with corticotrophin‐releasing factor (CRF) from terminals of parvocellular neurones of the hypothalamic paraventricular nucleus (PVH), facilitates the secretion of adrenocorticotrophic hormone (ACTH) in stressed rodents. The AVP gene encodes a propeptide precursor containing AVP, AVP‐associated neurophysin II (NPII), and a glycopeptide copeptin, although it is currently unclear whether copeptin is always cleaved from the neurophysin and whether the NPII and/or copeptin have any functional role in the pituitary. Furthermore, for primates, it is unknown whether CRF, AVP, NPII and copeptin are all colocalised in neurosecretory vesicles in the terminal region of the paraventricular CRF neurone axons. Therefore, we investigated, by fluorescence and immunogold immunocytochemistry, the cellular and subcellular relationships of these peptides in the CRF‐ and AVP‐producing cells in unstressed Japanese macaque monkeys (Macaca fuscata). Reverse transcription‐polymerase chain reaction analysis showed the expression of both CRF and AVP mRNAs in the monkey PVH. As expected, in the magnocellular neurones of the PVH and supraoptic nucleus, essentially no CRF immunoreactivity could be detected in NPII‐immunoreactive (AVP‐producing) neurones. Immunofluorescence showed that, in the parvocellular part of the PVH, NPII was detectable in a subpopulation (approximately 39%) of the numerous CRF‐immunoreactive neuronal perikarya, whereas, in the outer median eminence, NPII was more prominent (approximately 52%) in the CRF varicosities. Triple immunoelectron microscopy in the median eminence demonstrated the presence of both NPII and copeptin immunoreactivity in dense‐cored vesicles of CRF‐containing axons. The results are consistent with an idea that the AVP propeptide is processed and NPII and copeptin are colocalised in hypothalamic‐pituitary CRF axons in the median eminence of a primate. The CRF, AVP and copeptin are all co‐packaged in neurosecretory vesicles in monkeys and are thus likely to be co‐released into the portal capillary blood to amplify ACTH release from the primate anterior pituitary. No potential conflict of interest relevant to this article was reported.

MDPI Acta Medica Okayama 1422-0067 21 18 2020 Introducing the Amphibious Mudskipper Goby as a Unique Model to Evaluate Neuro/Endocrine Regulation of Behaviors Mediated by Buccal Sensation and Corticosteroids 6748 EN Yukitoshi Katayama Ushimado Marine Institute, Faculty of Science, Okayama University Kazuhiro Saito Ushimado Marine Institute, Faculty of Science, Okayama University Tatsuya Sakamoto Ushimado Marine Institute, Faculty of Science, Okayama University Some fish have acquired the ability to breathe air, but these fish can no longer flush their gills effectively when out of water. Hence, they have developed characteristic means for defense against external stressors, including thirst (osmolarity/ions) and toxicity. Amphibious fish, extant air-breathing fish emerged from water, may serve as models to examine physiological responses to these stressors. Some of these fish, including mudskipper gobies such asPeriophthalmodon schlosseri,Boleophthalmus boddartiand ourPeriophthalmus modestus, display distinct adaptational behaviors to these factors compared with fully aquatic fish. In this review, we introduce the mudskipper goby as a unique model to study the behaviors and the neuro/endocrine mechanisms of behavioral responses to the stressors. Our studies have shown that a local sensation of thirst in the buccal cavity-this being induced by dipsogenic hormones-motivates these fish to move to water through a forebrain response. The corticosteroid system, which is responsive to various stressors, also stimulates migration, possibly via the receptors in the brain. We suggest that such fish are an important model to deepen insights into the stress-related neuro/endocrine-behavioral effects. No potential conflict of interest relevant to this article was reported.

Wiley Acta Medica Okayama 0021-9967 529 7 2020 Variation of pro‐vasopressin processing in parvocellular and magnocellular neurons in the paraventricular nucleus of the hypothalamus: Evidence from the vasopressin‐related glycopeptide copeptin 1372 1390 EN Natsuko Kawakami Ushimado Marine Institute (UMI), Graduate School of Natural Science and Technology, Okayama University Akito Otubo Ushimado Marine Institute (UMI), Graduate School of Natural Science and Technology, Okayama University Sho Maejima Ushimado Marine Institute (UMI), Graduate School of Natural Science and Technology, Okayama University Ashraf H. Talukder Laboratory of Information Biology, Graduate School of Information Sciences, Tohoku University Keita Satoh Ushimado Marine Institute (UMI), Graduate School of Natural Science and Technology, Okayama University Takumi Oti Ushimado Marine Institute (UMI), Graduate School of Natural Science and Technology, Okayama University Keiko Takanami Ushimado Marine Institute (UMI), Graduate School of Natural Science and Technology, Okayama University Yasumasa Ueda Department of Physiology, Kyoto Prefectural University of Medicine Keiichi Itoi Laboratory of Information Biology, Graduate School of Information Sciences, Tohoku University John F. Morris Department of Physiology, Anatomy and Genetics, University of Oxford Tatsuya Sakamoto Ushimado Marine Institute (UMI), Graduate School of Natural Science and Technology, Okayama University Hirotaka Sakamoto Ushimado Marine Institute (UMI), Graduate School of Natural Science and Technology, Okayama University Arginine vasopressin (AVP) is synthesized in parvocellular‐ and magnocellular neuroendocrine neurons in the paraventricular nucleus (PVN) of the hypothalamus. Whereas magnocellular AVP neurons project primarily to the posterior pituitary, parvocellular AVP neurons project to the median eminence (ME) and to extrahypothalamic areas. The AVP gene encodes pre‐pro‐AVP that comprises the signal peptide, AVP, neurophysin (NPII), and a copeptin glycopeptide. In the present study, we used an N‐terminal copeptin antiserum to examine copeptin expression in magnocellular and parvocellular neurons in the hypothalamus in the mouse, rat, and macaque monkey. Although magnocellular NPII‐expressing neurons exhibited strong N‐terminal copeptin immunoreactivity in all three species, a great majority (~90%) of parvocellular neurons that expressed NPII was devoid of copeptin immunoreactivity in the mouse, and in approximately half (~53%) of them in the rat, whereas in monkey hypothalamus, virtually all NPII‐immunoreactive parvocellular neurons contained strong copeptin immunoreactivity. Immunoelectron microscopy in the mouse clearly showed copeptin‐immunoreactivity co‐localized with NPII‐immunoreactivity in neurosecretory vesicles in the internal layer of the ME and posterior pituitary, but not in the external layer of the ME. Intracerebroventricular administration of a prohormone convertase inhibitor, hexa‐d‐arginine amide resulted in a marked reduction of copeptin‐immunoreactivity in the NPII‐immunoreactive magnocellular PVN neurons in the mouse, suggesting that low protease activity and incomplete processing of pro‐AVP could explain the disproportionally low levels of N‐terminal copeptin expression in rodent AVP (NPII)‐expressing parvocellular neurons. Physiologic and phylogenetic aspects of copeptin expression among neuroendocrine neurons require further exploration. No potential conflict of interest relevant to this article was reported.

Frontiers Media Acta Medica Okayama 1664-2392 11 2020 Transcriptomic Analysis of the Kuruma PrawnMarsupenaeus japonicusReveals Possible Peripheral Regulation of the Ovary 541 EN Naoaki Tsutsui Faculty of Science, Ushimado Marine Institute, Okayama University Yasuhisa Kobayashi Faculty of Science, Ushimado Marine Institute, Okayama University Kouichi Izumikawa Research Institute for Fisheries Science, Okayama Prefectural Technology Center for Agriculture, Forestry, and Fisheries Tatsuya Sakamoto Faculty of Science, Ushimado Marine Institute, Okayama University Crustacean reproduction has been hypothesized to be under complex endocrinological regulation by peptide hormones. To further improve our understanding of the mechanisms underlying this complex regulation, knowledge is needed regarding the hormones not only of the central nervous system (CNS) such as the X-organ/sinus gland (XOSG), brain, and thoracic ganglia, but also the peripheral gonadal tissues. For example, in vertebrates, some gonadal peptide hormones including activin, inhibin, follistatin, and relaxin are known to be involved in the reproductive physiology. Therefore, it is highly likely that some peptide factors from the ovary are serving as the signals among peripheral tissues and central nervous tissues in crustaceans. In this work, we sought to find gonadal peptide hormones and peptide hormone receptors by analyzing the transcriptome of the ovary of the kuruma prawnMarsupenaeus japonicus. The generated ovarian transcriptome data led to the identification of five possible peptide hormones, including bursicon-alpha and -beta, the crustacean hyperglycemic hormone (CHH)-like peptide, insulin-like peptide (ILP), and neuroparsin-like peptide (NPLP). Dominant gene expressions for the bursicons were observed in the thoracic ganglia and the ovary, in the CNS for the CHH-like peptide, in the heart for NPLP, and in the ovary for ILP. Since the gene expressions of CHH-like peptide and NPLP were affected by a CHH (Penaeus japonicussinus gland peptide-I) from XOSG, we produced recombinant peptides for CHH-like peptide and NPLP usingEscherichia coliexpression system to examine their possible peripheral regulation. As a result, we found that the recombinant NPLP increased vitellogenin gene expression in incubated ovarian tissue fragments. Moreover, contigs encoding putative receptors for insulin-like androgenic gland factor, insulin, neuroparsin, and neuropeptide Y/F, as well as several contigs encoding orphan G-protein coupled receptors and receptor-type guanylyl cyclases were also identified in the ovarian transcriptome. These results suggest that reproductive physiology in crustaceans is regulated by various gonadal peptide hormones, akin to vertebrates. No potential conflict of interest relevant to this article was reported.

岡山実験動物研究会 Acta Medica Okayama 34 2018 無腸動物Praesagittifera naikaiensis における細胞骨格要素 21 27 EN Risa Ikeda Graduate School of Education, Okayama University Chiho Fujiwara Graduate School of Education, Okayama University Mayuko Hamada Ushimado Marine Institute, Faculty of Science, Okayama University Tatsuya Sakamoto Ushimado Marine Institute, Faculty of Science, Okayama University Noboru Saito Graduate School of Environmental and Life Science, Okayama University Motonori Ando Graduate School of Education, Okayama University Acoel flatworms can move in a variety of ways such as muscular and ciliary movements via cytoskeletal elements and their neural regulations. However, those locomotive mechanisms have not yet been fully elucidated. In this study, we examined the distribution of cytoskeletal elements including filamentous actin (F-actin) and tubulin, and the neuroanatomical organization in an acoelomorph worm, Praesagittifera naikaiensis (P. naikaiensis). Video microscopy revealed the elongation/contraction and the bending/rotation processes, and the ciliary gliding movement of P. naikaiensis. Histochemical and morphological analysis demonstrated that F-actin networks of inner longitudinal and outer circular muscle fibers were positioned along the entire surface of the body, and that the average distance between the circular muscle fibers in the contracted organism was decreased in the anterior region compared with that in the elongated organism. Electron microscopy showed dense bodies on the muscle cells of P. naikaiensis, which indicates that those muscle cells have the appearance of vertebrate smooth muscle cells. Immunohistochemical analysis revealed that -tubulin-positive signals on the ciliary microtubules had close contact with the F-actin network, and that neurite bundles labelled with anti dSap47 antibody as a neuronal marker run along the anterior-posterior body axis. These results indicate that the well-organized cytoskeletal elements and their neural control systems are preserved in P. naikaiensis, and that their mechanisms involved in those regulation systems are similar to those vertebrate systems. Further studies are needed to clarify the physiological mechanisms underlying the muscular and ciliary movements in P. naikaiensis. No potential conflict of interest relevant to this article was reported.

Wistar Institute of Anatomy and Biology Acta Medica Okayama 0021-9967 525 7 2017 Identification of the sexually dimorphic gastrin-releasing peptide system in the lumbosacral spinal cord that controls male reproductive function in the mouse and Asian house musk shrew (Suncus murinus) 1586 1598 EN Kei Tamura Ushimado Marine Institute (UMI), Graduate School of Natural Science and Technology, Okayama University Yasuhisa Kobayashi Ushimado Marine Institute (UMI), Graduate School of Natural Science and Technology, Okayama University Asuka Hirooka Ushimado Marine Institute (UMI), Graduate School of Natural Science and Technology, Okayama University Keiko Takanami Ushimado Marine Institute (UMI), Graduate School of Natural Science and Technology, Okayama University Takumi Oti Ushimado Marine Institute (UMI), Graduate School of Natural Science and Technology, Okayama University Takamichi Jogahara Laboratory of Animal Management and Resources, Department of Zoology, Okayama University of Science Sen-ichi Oda Laboratory of Animal Management and Resources, Department of Zoology, Okayama University of Science Tatsuya Sakamoto Ushimado Marine Institute (UMI), Graduate School of Natural Science and Technology, Okayama University Hirotaka Sakamoto Ushimado Marine Institute (UMI), Graduate School of Natural Science and Technology, Okayama University Several regions of the brain and spinal cord control male reproductive function. We previously demonstrated that the gastrin-releasing peptide (GRP) system, located in the lumbosacral spinal cord of rats, controls spinal centers to promote penile reflexes during male copulatory behavior. However, little information exists on the male-specific spinal GRP system in animals other than rats. The objective of this study was to examine the functional generality of the spinal GRP system in mammals using the Asian house musk shrew (Suncus murinus; suncus named as the laboratory strain), a specialized placental mammal model. Mice are also used for a representative model of small laboratory animals. We first isolated complementary DNA encoding GRP in suncus. Phylogenetic analysis revealed that suncus preproGRP was clustered to an independent branch. Reverse transcription-PCR showed that GRP and its receptor mRNAs were both expressed in the lumbar spinal cord of suncus and mice. Immunohistochemistry for GRP demonstrated that the sexually dimorphic GRP system and male-specific expression/distribution patterns of GRP in the lumbosacral spinal cord in suncus are similar to those of mice. In suncus, we further found that most GRP-expressing neurons in males also express androgen receptors, suggesting that this male-dominant system in suncus is also androgen-dependent. Taken together, these results indicate that the sexually dimorphic spinal GRP system exists not only in mice but also in suncus, suggesting that this system is a conserved property in mammals. No potential conflict of interest relevant to this article was reported.