Membrane molecule bouncer regulates sperm binding activity in immature oocytes in the viviparous teleost species Poecilia reticulata (guppy).

Generally, in vertebrates, the first step toward fertilization is the ovulation of mature oocytes, followed by their binding to sperm cells outside of the ovary. Exceptionally, the oocytes of poeciliid fish are fertilized by sperm cells within the follicle, and the developmental embryo is subsequently released into the ovarian lumen before delivery. In the present study, we aimed to identify the factor(s) responsible for intrafollicular fertilization in a viviparous teleost species, Poecilia reticulata (guppy). Sperm tracking analysis in this regard indicated that in this species, sperm cells reached immature oocytes including the germinal vesicle, and the insemination assay indicated that the immature oocytes robustly adhered to the sperm cells; similar binding was not observed in Danio rerio (zebrafish) and Oryzias latipes (medaka). We also identified the Ly6/uPAR protein bouncer as the factor responsible for the observed sperm binding activity of the immature oocytes in this species. The recombinant bouncer peptide acted as an inhibitory decoy for the sperm-oocyte binding in guppy. On the other hand, ectopic expression of guppy bouncer in zebrafish oocytes resulted in interspecific sperm-oocyte binding. These results argue that bouncer is responsible for sperm-immature oocyte binding. Our findings highlight the unique reproductive strategies of guppy fish and enhance our understanding of the diverse reproductive mechanisms in vertebrates.

Expression and antimicrobial activity of liver-expressed antimicrobial peptides in the ovaries of the viviparous teleost Xenotoca eiseni.

The mechanism via which the mothers of viviparous animals regulate the internal environment of pregnancy-associated organs for maintaining offspring growth is poorly understood. Environmental niches in organs contain fluid components for supporting embryonic growth; however, they may serve as nutrients for microbes. Therefore, microbial control is essential in viviparous animals to reduce the risk of infection in the ovarian lumen. Its importance may be higher than that in the case of oviparous animals. In this study, we investigated the antimicrobial factors in a viviparous teleost, Xenotoca eiseni. Four transcripts of the liver-expressed antimicrobial peptide (LEAP) were identified via RNA-Seq analysis. Some of the genes were expressed in the ovaries or intraovarian embryos of the fish. In particular, high expression of leap1a was detected in the ovaries of both pregnant and non-pregnant fish. Moreover, the ovary extracts from X. eiseni and transformed leap genes exhibited antimicrobial activity against Escherichia coli. Our results suggest that viviparous teleosts utilize antimicrobial peptides to reduce the risk of infection in the ovarian lumen.

Single and chronic L-serine treatments exert antidepressant-like effects in rats possibly by different means.

In the present study, the effects of both single (6 mmol L-serine/10 ml/kg orally administrated) and chronic (2% L-serine solution freely given for 28 days) treatments on depression-like behavior were evaluated in Wistar rats, representing the control, and Wistar Kyoto rats, representing an animal model of depression. Both single and chronic L-serine treatments decreased the duration of immobility, which is an index of a depressive-like state, in the forced swimming test in both strains. However, the decreases in the duration of immobility appear to be regulated differently by the different mechanisms involved in single and chronic L-serine treatments. In the prefrontal cortex and hippocampus, single L-serine treatment increased the concentrations of L-serine, but not D-serine, while chronic L-serine treatment increased those of D-serine, but not L-serine. These data suggest that the antidepressant-like effects of single and chronic L-serine treatments may have been induced by the increased L-serine and D-serine concentrations, respectively, in the brain. In addition, chronic L-serine treatment increased cystathionine concentrations in the hippocampus and prefrontal cortex in Wistar rats, but not in Wistar Kyoto rats, suggesting that Wistar Kyoto rats have an abnormality in the serine-cystathionine metabolic pathway. In conclusion, single and chronic L-serine treatments may induce antidepressant-like effects via the different mechanisms related to serine metabolism in the brain.

Orally administered whole egg demonstrates antidepressant-like effects in the forced swimming test on rats.

OBJECTIVE: Several studies have reported that vegetarian diets are associated with a higher prevalence of major depression. Therefore, we hypothesised that the consumption of animal products, especially eggs, may have positive effects on mental health, especially on major depression, because a previous study reported that egg consumption produces numerous beneficial effects in humans. The purpose of the present study was to evaluate the effects of chronic whole-egg treatment on depression-like behaviours in Wistar rats, a control strain, and Wistar Kyoto rats, an animal model of depression. METHODS: In both the rats, either whole-egg solution (5 ml/kg) or distilled water (5 ml/kg) was orally administrated for 35 days. During these periods, the open-field test (OFT) was conducted on the 21st day, and a forced swimming test (FST) was enforced on the 27th and 28th days. On the 36th day, the plasma and brain were collected. RESULTS: Chronic whole-egg treatment did not affect line crossing in the OFT, whereas it reduced the total duration of immobility in the FST on both strains. Furthermore, interestingly, the results indicated the possibility that whole-egg treatment elevated the incorporation of tryptophan into the brain, and the tryptophan concentration in the prefrontal cortex was actually increased by the treatment. CONCLUSION: This study demonstrated that whole-egg treatment exerts an antidepressant-like effect in the FST. It is suggested that whole egg may be an excellent food for preventing and alleviating the conditions of major depression.

Evolutionary insights from an anatomical network analysis of the hyolaryngeal apparatus in extant archosaurs (birds and crocodilians).

Adaptive radiation of archosaurs, represented by crocodilians, non-avian dinosaurs, and birds, since the Mesozoic has been studied mainly based on their major skeletal elements (skull, vertebrae, and limbs). However, little is known about the evolution of their hyolaryngeal apparatus, which is involved with feeding, respiration, and vocalization, because of poor fossil preservation and the difficulty in determining the musculoskeletal homology of the apparatus. Network analysis is a framework to quantitatively characterize the topological organization of anatomical structures for comparing structural integration and modularity regardless of ambiguous homology. Herein, we modeled the musculoskeletal system of hyolarynx in six species of extant archosaurs and its sister-taxon turtle, and conducted a network analysis using network parameters, modular partition, and bone centrality in a phylogenetic framework. The network parameters reveal that ancestral archosaurs have reduced the numbers of elements and links and acquired complex networks as a whole domain with strong modularity in the hyolarynx. Furthermore, the modular partition and centrality reveal that the hyoids are highly evolvable, while the larynx is constrained and less evolvable. The archosaur hyolarynx exhibits different evolutionary trends: crocodilians with the system integration, basihyal simplification, and ceratobranchial centralization; and birds with the simplicity, weak integration, and modularity of the hyolarynx, laryngeal integration with cricoid centrality, and tongue-module expansion with the acquisition of paraglossal. Four hyolaryngeal bones (ceratobranchial, basihyal, paraglossal, and cricoid) have played important roles in archosaur evolution, and their fossil records are keys to understanding the two major archosaur lineages toward crocodilians and birds.

An ankylosaur larynx provides insights for bird-like vocalization in non-avian dinosaurs.

A voice box (larynx) is unique for tetrapods and plays functional roles in respiration, airway protection, and vocalization. However, in birds and other reptiles, the larynx fossil is extremely rare, and the evolution of this structure remains largely unknown. Here we report the fossil larynx found in non-avian dinosaurs from ankylosaur Pinacosaurus grangeri. The larynx of Pinacosaurus is composed of the cricoid and arytenoid like non-avian reptiles, but specialized with the firm and kinetic cricoid-arytenoid joint, prominent arytenoid process, long arytenoid, and enlarged cricoid, as a possible vocal modifier like birds rather than vocal source like non-avian reptiles. Although bird-unique vocal source (syrinx) have never been reported in non-avian dinosaurs, Pinacosaurus could have employed bird-like vocalization with the bird-like large, kinetic larynx. This oldest laryngeal fossil from the Cretaceous dinosaur provides the first step for understanding the vocal evolution in non-avian dinosaurs toward birds.

Glucose absorption activity and gene expression of sugar transporters in the trophotaenia of the viviparous teleost Xenotoca eiseni.

In viviparous reproductive systems, nutrient transfer from mother to embryo plays a critical role in the generation of offspring. Herein, we investigated the mother-to-embryo nutrient transfer machinery in the viviparous teleost Xenotoca eiseni, which belongs to the family Goodeidae. The intraovarian embryo absorbs maternal supplements via the hindgut-derived placental structure termed the trophotaenia. Tracer analysis indicated that the trophotaenia can take up glucose analogs in ex vivo cultured embryos. The candidate genes for absorption, sglt1, glut2, atp1a, and atp1b, were determined from published transcriptomes. These genes were expressed in the trophotaenia of X. eiseni embryos. Fluorescent immunohistochemistry of Na+/K+ ATPase indicated the polarity of epithelial cells in the trophotaenia. The presented evidence suggests that the epithelial cell layer transports monosaccharides from the apical membrane of epithelial cells in a basolateral direction. Taken together, this study provides insight into how maternal fish maintain their offspring during gestation and will aid in the development of strategies to improve offspring generation in these fish.

Central administration of L- and D-aspartate attenuates stress behaviors by social isolation and CRF in neonatal chicks.

Intracerebroventricular (i.c.v.) administration of L-aspartate (L-Asp) attenuates stress responses in neonatal chicks, but the mechanism has not been clarified. In the present study, three behavioral experiments were carried out under socially isolated stressful conditions exacerbated by the use of corticotrophin-releasing factor (CRF). In Experiment 1, i.c.v. injection of L-Asp attenuated behavioral stress responses (distress vocalization and active wakefulness) in a dose-dependent manner. Furthermore, L-Asp increased time spent standing/sitting motionless with eyes open and sitting motionless with head dropped (sleeping posture) in comparison with the group receiving CRF alone. In Experiment 2, i.c.v. injection of D-Asp dose-dependently decreased the number of distress vocalizations and the amount of time spent in active wakefulness. D-Asp increased the time spent standing/sitting motionless with eyes open compared with the group receiving CRF alone. In Experiment 3, we directly compared the effect of L-Asp with that of D-Asp. Both L- and D-Asp induced sedative effects under an acutely stressful condition. However, L-Asp, but not D-Asp, increased the time spent in a sleeping posture. These results indicate that both L- and D-Asp, when present in the brain, could induce a sedative effect, while the mechanism for hypnosis in neonatal chicks may be different for L-Asp in comparison with D-Asp.

Endocytosis-mediated vitellogenin absorption and lipid metabolism in the hindgut-derived placenta of the viviparous teleost Xenotoca eiseni.

Certain viviparous animals possess mechanisms for mother-to-embryo nutrient transport during gestation. Xenotoca eiseni is one such viviparous teleost species in which the mother supplies proteins and other components to the offspring developing in the ovary. The embryo possesses trophotaenia, hindgut-derived placental structure, to receive the maternal supplement. However, research on the molecular mechanisms underlying viviparous species is scarce in non-mammalian vertebrates, including teleosts. Thus, we conducted this study to investigate the mechanism for nutrient absorption and degradation in trophotaeniae of X. eiseni. A tracer assay indicated that a lipid transfer protein, vitellogenin (Vtg), was absorbed into the epithelial layer cells of the trophotaeniae. Vtg uptake was significantly suppressed by Pitstop-2, an inhibitor of clathrin-mediated endocytosis. Gene expression analysis indicated that the genes involved in endocytosis-mediated lipolysis and lysosomal cholesterol transport were expressed in the trophotaeniae. In contrast, plasma membrane transporters expressed in the intestinal tract were not functional in the trophotaeniae. Our results suggested that endocytosis-mediated lysosomal lipolysis is one of the mechanisms underlying maternal component metabolism. Thus, our study demonstrated how viviparous teleost species have acquired a unique developmental system that is based on the hindgut-derived placenta.

A new goniopholidid from the Upper Jurassic Morrison Formation, USA: novel insight into aquatic adaptation toward modern crocodylians.

Goniopholididae is a group of basal neosuchian crocodyliforms closely related to Paralligatoridae and Eusuchia that lived during the Jurassic and Early Cretaceous. Goniopholidids have the long, flat snout and secondary palate of modern crocodylians, the acquisition of which is regarded as a key feature in the early evolution of crocodylian body plan and their aquatic adaptation. Here, we report a new species, Amphicotylus milesi, with the description from the best-preserved specimen to date of Goniopholididae from Wyoming, USA. Its posterior extension of the nasopharyngeal passage (pterygoid secondary palate) and the shortening and dorsal deflection of the ceratobranchial suggest that basal neosuchians could raise their gular valve to separate oral and pharyngeal cavities as in modern crocodylians. The anatomy of Amphicotylus milesi sheds light on the acquisition of this new respiratory system in the crocodyliform evolution and their early aquatic adaptation, leading to modern crocodylians.

Effects of S-adenosylmethionine decarboxylase, polyamines, amino acids, and weak bases (amines and ammonia) on development and ribosomal RNA synthesis in Xenopus embryos.

We have been studying control mechanisms of gene expression in early embryogenesis in a South African clawed toad Xenopus laevis, especially during the period of midblastula transition (MBT), or the transition from the phase of active cell division (cleavage stage) to the phase of extensive morphogenesis (post-blastular stages). We first found that ribosomal RNA synthesis is initiated shortly after MBT in Xenopus embryos and those weak bases, such as amines and ammonium ion, selectively inhibit the initiation and subsequent activation of rRNA synthesis. We then found that rapidly labeled heterogeneous mRNA-like RNA is synthesized in embryos at pre-MBT stage. We then performed cloning and expression studies of several genes, such as those for activin receptors, follistatin and aldolases, and then reached the studies of S-adenosylmethionine decarboxylase (SAMDC), a key enzyme in polyamine metabolism. Here, we cloned a Xenopus SAMDC cDNA and performed experiments to overexpress the in vitro-synthesized SAMDC mRNA in Xenopus early embryos, and found that the maternally preset program of apoptosis occurs in cleavage stage embryos, which is executed when embryos reach the stage of MBT. In the present article, we first summarize results on SAMDC and the maternal program of apoptosis, and then describe our studies on small-molecular-weight substances like polyamines, amino acids, and amines in Xenopus embryos. Finally, we summarize our studies on weak bases, especially on ammonium ion, as the specific inhibitor of ribosomal RNA synthesis in Xenopus embryonic cells.

Dinosaur ichnology and sedimentology of the Chignik Formation (Upper Cretaceous), Aniakchak National Monument, southwestern Alaska; Further insights on habitat preferences of high-latitude hadrosaurs.

While there are now numerous records of dinosaurs from Cretaceous rocks around the state of Alaska, very few fossil records of terrestrial vertebrates are known from the Mesozoic rocks of the southwestern part of the state. Here we report the new discovery of extensive occurrences of dinosaur tracks from Aniakchak National Monument of the Alaska Peninsula. These tracks are in the Late Cretaceous (Maastrichtian) Chignik Formation, a cyclic sequence of rocks, approximately 500-600 m thick, representing shallow marine to nearshore marine environments in the lower part and continental alluvial coastal plain environments in the upper part of the section. These rocks are part of the Peninsular Terrane and paleomagnetic reconstructions based on the volcanic rocks of this terrane suggest that the Chignik Formation was deposited at approximately its current latitude which is almost 57° N. Recent field work in Aniakchak National Monument has revealed over 75 new track sites, dramatically increasing the dinosaur record from the Alaska Peninsula. Most of the combined record of tracks can be attributed to hadrosaurs, the plant-eating duck-billed dinosaurs. Tracks range in size from those made by full-grown adults to juveniles. Other tracks can be attributed to armored dinosaurs, meat-eating dinosaurs, and two kinds of fossil birds. The track size of the predatory dinosaur suggests a body approximately 6-7 m long, about the estimated size of the North Slope tyrannosaurid Nanuqsaurus. The larger bird tracks resemble Magnoavipes denaliensis previously described from Denali National Park, while the smaller bird tracks were made by a bird about the size of a modern Willet. Previous interdisciplinary sedimentologic and paleontologic work in the correlative and well-known dinosaur bonebeds of the Prince Creek Formation 1400km-1500km further north in Alaska suggested that high-latitude hadrosaurs preferred distal coastal plain or lower delta plain habitats. The ichnological record being uncovered in the Chignik Formation of southwestern Alaska is showing that the hadrosaur tracks here were also made in distal coastal and delta plain conditions. This similarity may corroborate the habitat preference model for Cretaceous high-latitude dinosaurs proposed for the data gathered from the Prince Creek Formation, and may indicate that at least Beringian hadrosaurids had similar habitat preferences regardless of latitude.

Central Injection of Glucose Modifies Behavior, Amino Acid and Monoamine Metabolism in Neonatal Chicks under Acute Stressful Conditions.

The effect of intracerebroventricular (i.c.v.) injection of a wide range of glucose concentrations on the behavioral response, central amino acid and monoamine contents was investigated in chicks exposed to a social isolation stressful condition. The chicks were given an i.c.v. injection of 0.21, 0.42, 0.84, and 1.68 µmol of D-glucose, and then behavioral changes were observed over 10 min. The behavioral stress response was dose-dependently decreased and calm behavior was increased by i.c.v. administration of glucose. In the diencephalon, glutamine was positively correlated, whereas glycine was negatively correlated with the dose of glucose. In the telencephalon, the dopamine metabolite and dopamine turnover rates were positively correlated, whereas dopamine was negatively correlated with doses of glucose. In the plasma, isoleucine and hydroxyproline were positively correlated with the dose of glucose, and several amino acids were also influenced by glucose levels. These results suggest that the possible pathways of the sedative effect of glucose include: (1) amino acids synthesized from injected glucose, which can induce the sedative and/or hypnotic effects; (2) amino acids modified by injected glucose transported in the brain from the peripheral tissues; and (3) injected glucose-induced decreases in brain dopamine levels. In conclusion, these changes induced by central glucose interact and induce the sedative effect in neonatal chicks.

Comparison of centrally injected tryptophan-related substances inducing sedation in acute isolation stress-induced neonatal chicks.

In the present study, we first focused on the function of l-tryptophan (TRP) metabolites which are synthesized in different metabolic pathways, namely, the kynurenine (KYN) pathway and serotonin (5-HT) pathway during an acute isolation stress. When l-TRP metabolites were intracerebroventricularly injected on an equimolar basis (100 nmol), 5-HT induced a sedative effect in neonatal chicks. Additionally, plasma corticosterone, dopamine, 5-HT, and its metabolite 5-hydroxyindoleacetic acid concentrations were increased in the diencephalon of the 5-HT treated group compared with other groups. Second, the two doses (400 or 800 nmol) of l- and d-TRP were compared under a corticotrophin-releasing hormone-augmented social isolation stress. When comparing the efficacy between l- and d-TRP against stress behavior, both amino acids had a similar effect and quickly suppressed distress vocalizations. Finally, d-amino acid levels in the diencephalon and telencephalon were measured but d-TRP was not found. These results indicate that l- and d-TRP induce the same effect in attenuating stress but the mode of action of TRP derivatives, namely 5-HT differs during an acute isolation stress in neonatal chick. The absence of d-TRP in the diencephalon further suggests that instead of being an endogenous factor it may play role as a pharmacological factor.