Association of seasonal changes in circulating cortisol concentrations with the expression of cortisol biosynthetic enzymes and a glucocorticoid receptor in the blubber of common bottlenose dolphin.

Cortisol is secreted from the adrenal cortex in response to stress, and its circulating levels are used as robust physiological indicators of stress intensity in various animals. Cortisol is also produced locally in adipose tissue by the conversion of steroid hormones such as cortisone, which is related to fat accumulation. Circulating cortisol levels, probably induced by cold stress, increase in cetaceans under cold conditions. However, whether cortisol production in subcutaneous adipose tissue is enhanced when fat accumulation is renewed during the cold season remains unclear. Therefore, in this study, we examine the effect of environmental temperature on the expression of cortisol synthesis-related enzymes and a glucocorticoid receptor in the subcutaneous fat (blubber) and explore the association between these expressions and fluctuations in circulating cortisol levels in common bottlenose dolphins (Tursiops truncatus). Skin biopsies were obtained seasonally from eight female dolphins, and seasonal differences in the expression of target genes in the blubber were analyzed. Blood samples were collected throughout the year, and cortisol levels were measured. We found that the expressions of cytochrome P450 family 21 subfamily A member 2 (CYP21A2) and nuclear receptor subfamily 3 group C member 1 (NR3C1), a glucocorticoid receptor, were increased in the cold season, and 11 beta-hydroxysteroid dehydrogenase type 1 (HSD11B1) showed a similar trend. Blood cortisol levels increased when the water temperature decreased. These results suggest that the conversion of 17-hydroxyprogesterone to cortisol via 11-deoxycortisol and/or of cortisone to cortisol is enhanced under cold conditions, and the physiological effects of cortisol in subcutaneous adipose tissue may contribute to on-site lipid accumulation and increase the circulating cortisol concentrations. The results obtained in this study highlight the role of cortisol in the regulation of the blubber that has developed to adapt to aquatic life.

Blubber biopsy in common bottlenose dolphins using a novel biopsy puncher: Evaluation of the impact on living individuals and possibility of applications in cetacean research.

Biopsy has recently become a preferred protocol for sampling the skin and blubber of many cetacean species, although it is desirable to collect as minimally invasive as possible. Here, the effect of biopsy sampling on the captive common bottlenose dolphins was evaluated by analyzing the process of wound healing and changes in hematological and blood biochemical parameters after biopsy using a puncher developed to collect up to the inner layer of the dolphin blubber. Results showed that the wounds caused by biopsy were closed in as early as 1 day and completely covered with the epidermis within 5-11 days. Blood fibrinogen, which generally increases due to a wound-induced inflammatory response or activation of the coagulation system, was significantly elevated after the biopsy indicating ongoing tissue repair, while other parameters did not exhibit significant differences. Furthermore, histological observation and RNA extraction of samples were performed to investigate the versatility of this method to cetacean research. Histological examination revealed three distinct layers of the blubber in the biopsy samples. Moreover, total RNA extracted from biopsy samples exhibited sufficient quality and quantity for gene expression analyses. Overall, the puncher utilized in our study represents a valuable and minimally invasive tool for investigating various aspects of small cetacean studies.

Microbial decomposition of biodegradable plastics on the deep-sea floor.

Microbes can decompose biodegradable plastics on land, rivers and seashore. However, it is unclear whether deep-sea microbes can degrade biodegradable plastics in the extreme environmental conditions of the seafloor. Here, we report microbial decomposition of representative biodegradable plastics (polyhydroxyalkanoates, biodegradable polyesters, and polysaccharide esters) at diverse deep-sea floor locations ranging in depth from 757 to 5552 m. The degradation of samples was evaluated in terms of weight loss, reduction in material thickness, and surface morphological changes. Poly(L-lactic acid) did not degrade at either shore or deep-sea sites, while other biodegradable polyesters, polyhydroxyalkanoates, and polysaccharide esters were degraded. The rate of degradation slowed with water depth. We analysed the plastic-associated microbial communities by 16S rRNA gene amplicon sequencing and metagenomics. Several dominant microorganisms carried genes potentially encoding plastic-degrading enzymes such as polyhydroxyalkanoate depolymerases and cutinases/polyesterases. Analysis of available metagenomic datasets indicated that these microorganisms are present in other deep-sea locations. Our results confirm that biodegradable plastics can be degraded by the action of microorganisms on the deep-sea floor, although with much less efficiency than in coastal settings.

Translation and validation of the Japanese version of the Birth Satisfaction Scale-Revised.

AIM: This study aimed to develop a Japanese version of the Birth Satisfaction Scale-Revised and evaluate its reliability and validity. METHODS: After translating the Birth Satisfaction Scale-Revised into Japanese, we conducted an Internet-based cross-sectional study with 445 Japanese-speaking women within 2 months of childbirth. Of these, 98 participated in the retest 1 month later. Data were analyzed using the COSMIN study design checklist for patient-reported outcome measurement instruments. Content validity was evaluated through cognitive debriefing during the translation process into Japanese. Confirmatory factor analysis was conducted to verify structural and cross-cultural validities. For hypothesis testing, we tested correlations with existing measures for convergent and divergent validities, and for known-group discriminant validity, we made comparisons between types of childbirth. Internal consistency was calculated using Cronbach's α, and test-retest reliability was evaluated using the intraclass correlation coefficient. RESULTS: For the Japanese-Birth Satisfaction Scale-Revised, the established three-factor model fit poorly, whereas the four-factor model fit better. Full metric invariance was observed in both the nulliparous and multiparous groups. Good convergent, divergent, and known-group discriminant validities and test-retest reliability were established. Internal consistency observations were suboptimal; however for vaginal childbirth, the Cronbach's α of the total score was .71. CONCLUSIONS: The Japanese-Birth Satisfaction Scale-Revised is a valid and reliable scale, with the exception of internal consistency that requires further investigation. If limited to vaginal childbirth, research, clinical applications, and international comparisons can be drawn.

Establishment and characterization of a novel lung cell line derived from the common bottlenose dolphin.

Cetaceans are specialized marine mammals with a unique respiratory system adapted for diving behavior. Furthermore, respiratory diseases are commonly observed in these mammals. Nevertheless, much of their respiratory physiology remains unknown due to the limited supply and poor quality of their biological samples for research. In this study, we established a novel lung cell line, dLu, derived from the common bottlenose dolphin (Tursiops truncatus), which can prove useful in cetacean research, including for understanding the pathogenesis of respiratory diseases in cetaceans. The cells were cultured in a simple medium consisting of Dulbecco's modified Eagle's medium supplemented with 10% fetal bovine serum. The morphology of the cells was fibroblast-like. dLu was produced by transfecting the simian virus 40 large T antigen into primary cultured cells. Although dLu exhibited approximately 80 cell divisions, it was unable to achieve complete immortalization, as the cells stopped proliferating beyond this number. dLu cells expressed toll-like receptor 3 but not toll-like receptor 4. Immunostimulation with poly(I:C) altered the gene expressions of interferon beta 1 and tumor necrosis factor alpha in dLu cells. In summary, dLu established in this study is a novel cetacean cell resource that can be easily cultured and is a useful in vitro tool in cetacean research, particularly for studying host immune responses in the lungs.

Difference in sulfur regulation mechanism between tube-dwelling and free-moving polychaetes sympatrically inhabiting deep-sea hydrothermal chimneys.

The environment around deep sea hydrothermal vents is characterized by an abundance of sulfur compounds, including toxic hydrogen sulfide. However, numerous communities of various invertebrates are found in it. It is suggested that invertebrates in the vicinity of hydrothermal vents detoxify sulfur compounds by biosynthesis of taurine-related compounds in the body. On the other hand, the vent endemic polychaete Alvinella pompejana has spherocrystals composed of sulfur and other metals in its digestive tract. It was considered that the spherocrystals contribute to the regulation of sulfur in body fluids. Paralvinella spp. and Polynoidae. gen. sp. live sympatrically and in areas most affected by vent fluid. In this study, we focused on the digestive tract of Paralvinella spp. and Polynoidae. gen. sp. to examine whether they have spherocrystals. We also investigated the possible involvement of bacteria in the digestive tract in spherulization. Examination with a scanning electron microscope (SEM) equipped with Energy Disperse X-ray Spectroscopy (EDS) detected spherocrystals containing sulfur and iron in the digestive tract of Paralvinella spp. In contrast, such spherocrystals were not observed in that of Polynoidae. gen. sp. although sulfur is detected there by inductively coupled plasma-optical emission spectrometry (ICP-OES). Meta-16S rRNA analysis indicated that the floras of the digestive tracts of the two species were very similar, suggesting that enteric bacteria are not responsible for spherocrystal formation. Analysis of taurine-related compounds indicated that the digestive tissues of Polynoidae. gen. sp. contain a higher amount of hypotaurine and thiotaurine than those of Paralvinella spp. Therefore, the two sympatric polychaetes use different strategies for controlling sulfur, i.e., Paralvinella spp. forms spherocrystals containing elemental sulfur and iron in the digestive tract, but Polynoidae. gen. sp. accumulates taurine-related compounds instead of spherocrystals. Such differences may be related to differences in their lifestyles, i.e., burrow-dweller or free-moving, or may have been acquired phylogenetically in the evolutionary process.

Establishment and characterization of a novel kidney cell line derived from the common bottlenose dolphin.

Common bottlenose dolphin (Tursiops truncatus) is a well-known cetacean species that inhabits temperate and tropical seas worldwide. Limited supply and poor quality of samples hinder the investigation of the effects of various pathogens and environmental pollutants on this cetacean species. Cultured cells are useful for experimental studies; however, no cell lines derived from cetaceans are generally available. Therefore, in this study, we established a novel kidney cell line, TK-ST, derived from T. truncatus. Primary cells exhibited the morphological characteristics of epithelial and fibroblast cells, but their immortalization and passaging resulted in a predominantly epithelial cell morphology. TK-ST was immortalized using the large T SV40 antigen and human telomerase reverse transcriptase and exhibited long-term stable cell growth. TK-ST cells are generally cultured in Dulbecco's modified Eagle's medium with 10% fetal bovine serum at 37°C and 5% CO2 but can also be cultured in 5-20% fetal bovine serum and several other classical media commonly used for common animal cell culture. TK-ST cells were found to be susceptible to several viruses, including the dolphin morbillivirus (most important virus in cetaceans), and exhibited cytopathic effects, facilitating the replication of the dolphin morbillivirus. Furthermore, mRNA expression levels of cytokine genes were increased in TK-ST cells after stimulation with lipopolysaccharides and poly(I:C). Therefore, the novel TK-ST cell line derived in this study can potentially be used for further in vitro studies on cetaceans.

Possible contribution of phosphate to the pathogenesis of chronic kidney disease in dolphins.

This study aimed to investigate whether phosphate contributes to the pathogenesis of chronic kidney disease (CKD) in dolphins. Renal necropsy tissue of an aged captive dolphin was analyzed and in vitro experiments using cultured immortalized dolphin proximal tubular (DolKT-1) cells were performed. An older dolphin in captivity died of myocarditis, but its renal function was within the normal range until shortly before death. In renal necropsy tissue, obvious glomerular and tubulointerstitial changes were not observed except for renal infarction resulting from myocarditis. However, a computed tomography scan showed medullary calcification in reniculi. Micro area X-ray diffractometry and infrared absorption spectrometry showed that the calcified areas were primarily composed of hydroxyapatite. In vitro experiments showed that treatment with both phosphate and calciprotein particles (CPPs) resulted in cell viability loss and lactate dehydrogenase release in DolKT-1 cells. However, treatment with magnesium markedly attenuated this cellular injury induced by phosphate, but not by CPPs. Magnesium dose-dependently decreased CPP formation. These data support the hypothesis that continuous exposure to high phosphate contributes to the progression of CKD in captive-aged dolphins. Our data also suggest that phosphate-induced renal injury is mediated by CPP formation in dolphins, and it is attenuated by magnesium administration.

Genomics reveals a genetically isolated population of the Pacific white-sided dolphin (Lagenorhynchus obliquidens) distributed in the Sea of Japan.

The processes by which animals become genetically isolated in an open environment such as the ocean have not yet been fully elucidated. Morphologically different populations of Pacific white-sided dolphin Lagenorhynchus obliquidens are observed sympatrically in the Sea of Japan. However, genetic studies that exclusively used limited mitochondrial loci or microsatellite DNA have failed to demonstrate the existence of genetically distinct populations. Here, to reveal the population structure, we analysed genome-wide population genetic data using single nucleotide polymorphisms (SNPs) gathered in 2018-2021 from all domestic captive individuals in aquaria, the majority of which originated from the wild, as well as from some stranded individuals, together covering a wide area of coastal water around Japan (n = 123). Multiplexed intersimple sequence repeat genotyping-by-sequencing analysis was performed to obtain the SNP data. Principal coordinate analysis and the clustering method structure indicated that two genetically-distinct populations exist, with little interpopulation gene flow revealed. In addition, the genotypic segregation was reflected in differences in external morphotype. Furthermore, a population demographic analysis based on the whole-genome sequences of an individual from each population indicated that sea-level changes during the Last Glacial Period probably led to allopatric divergence of this species in a limited area of the Sea of Japan, with that group subsequently sharing a distribution area with the other population. These findings yield insights into the formation of genetically isolated sympatric populations in the ocean.

Vitamin B12 biosynthesis of Cetobacterium ceti isolated from the intestinal content of captive common bottlenose dolphins (Tursiops truncatus).

In comparison with terrestrial mammals, dolphins require a large amount of haemoglobin in blood and myoglobin in muscle to prolong their diving time underwater and increase the depth they can dive. The genus Cetobacterium is a common gastrointestinal bacterium in dolphins and includes two species: C. somerae and C. ceti. Whilst the former produces vitamin B12, which is essential for the biosynthesis of haem, a component of haemoglobin and myoglobin, but not produced by mammals, the production ability of the latter remains unknown. The present study aimed to isolate C. ceti from dolphins and reveal its ability to biosynthesize vitamin B12. Three strains of C. ceti, identified by phylogenetic analyses with 16S rRNA gene and genome-based taxonomy assignment and biochemical features, were isolated from faecal samples collected from two captive common bottlenose dolphins (Tursiops truncatus). A microbioassay using Lactobacillus leichmannii ATCC 7830 showed that the average concentration of vitamin B12 produced by the three strains was 11 (standard deviation: 2) pg ml-1. The biosynthesis pathway of vitamin B12, in particular, adenosylcobalamin, was detected in the draft genome of the three strains using blastKOALA. This is the first study to isolate C. ceti from common bottlenose dolphins and reveal its ability of vitamin B12 biosynthesis, and our findings emphasize the importance of C. ceti in supplying haemoglobin and myoglobin to dolphins.

Blood pressure adaptation in vertebrates: comparative biology.

With evolution from water to land, the osmotic regulation of body fluids and cardiovascular systems of vertebrates evolved to cope with dryness and gravity. While aquatic vertebrates can use buoyancy to compensate for the effects of gravity, terrestrial vertebrates cannot and must circulate blood throughout their body-a necessity that likely led to the development of strong hearts and high blood pressure. These changes may be supported by anatomic evolution of the cardiovascular system and by functional evolution, with alterations in hormonal systems. Thus, during the evolution of terrestrial animals, increased performance of body functions to endure harsher environments was required, necessitating increased blood pressure. In an age of overeating and insufficient exercise, modern man does not fully use the high levels of physical functions acquired through evolution. Drastic changes in our living environment cause hypertension, the pathogenesis of which remains unknown. To survive in new environments, as might be expected in outer space or underwater, an understanding is required of how changes in blood pressure have occurred that enabled adaptation through evolution in vertebrates.

Cytologic features of oral squamous cell carcinoma in an Indo-Pacific bottlenose dolphin (Tursiops aduncus): Papanicolaou stain and immunocytochemistry using liquid-based cytology.

Although oral cytology using Papanicolaou stain is useful for the early detection of oral premalignant lesions and squamous cell carcinoma (SCC) in people, little work has been conducted on this topic in veterinary medicine. This paper describes the features of oral cytology using Papanicolaou stain and immunocytochemistry on liquid-based cytology slides in a case of oral SCC in an Indo-Pacific bottlenose dolphin (Tursiops aduncus). In this case, dysplastic cells with koilocyte-like changes and SCC cells were identified using the Papanicolaou stain. These cells were positive for p53 using an immunocytochemistry analysis. A cytologic diagnosis of SCC was made. We believe that the early detection of premalignant oral lesions and SCC in dolphins can be significantly improved with cytology using liquid-based cytology, Papanicolaou staining, and immunocytochemistry.

The role of toxic planocerid flatworm larvae on tetrodotoxin accumulation in marine bivalves.

Tetrodotoxin (TTX), also known as pufferfish toxin, has been detected in marine edible bivalves worldwide. In this study, several bivalve species, Azumapecten farreri subsp. akazara, Patinopecten yessoensis and Mytilus galloprovincialis, collected from the Pacific side of the northern Japanese Islands, were studied for the accumulation of TTX in the presence of toxic planocerid larvae. LC-MS/MS analysis demonstrated that TTX was detected only in the midgut gland of A. farreri subsp. akazara. Toxic flatworm-specific PCR and direct sequencing of the amplicons showed that the DNA fragments of the Planocera multitentaculata COI gene were detected in the gut contents of the toxified bivalves. The planocerid larvae were also detected in the environmental seawaters. Toxification experiments in the aquarium demonstrated that the mussel M. galloprovincialis was also toxified by feeding on the toxic flatworm larvae. These results suggest that the source of TTX accumulation in edible bivalves is toxic flatworm larvae.

Identification of the causative gene of a transparent phenotype of juvenile red sea bream Pagrus major.

Deformities in cultured fish species may be genetic, and identifying causative genes is essential to expand production and maintain farmed animal welfare. We previously reported a genetic deformity in juvenile red sea bream, designated a transparent phenotype. To identify its causative gene, we conducted genome-wide linkage analysis and identified two single nucleotide polymorphisms (SNP) located on LG23 directly linked to the transparent phenotype. The scaffold on which the two SNPs were located contained two candidate genes, duox and duoxa, which are related to thyroid hormone synthesis. Four missense mutations were found in duox and one in duoxa, with that in duoxa showing perfect association with the transparent phenotype. The mutation of duoxa was suggested to affect the transmembrane structure and thyroid-related traits, including an enlarged thyroid gland and immature erythrocytes, and lower thyroxine (T4) concentrations were observed in the transparent phenotype. The transparent phenotype was rescued by T4 immersion. Loss-of-function of duoxa by CRISPR-Cas9 induced the transparent phenotype in zebrafish. Evidence suggests that the transparent phenotype of juvenile red sea bream is caused by the missense mutation of duoxa and that this mutation disrupts thyroid hormone synthesis. The newly identified missense mutation will contribute to effective selective breeding of red sea bream to purge the causative gene of the undesirable phenotype and improve seed production of red sea bream as well as provide basic information of the mechanisms of thyroid hormones and its related diseases in fish and humans.

Composition of uterine milk and its changes with gestational period in red stingrays (Hemitrygon akajei).

Uterine milk is secreted in the uterus for embryo nutrition in several elasmobranch species and may contribute to rapid embryonic growth, but the details of its composition and its functions are poorly understood. In this study, to explore the roles of uterine milk for embryos, its components throughout the gestational period were analysed in detail. Uterine milk was collected from pregnant red stingrays (Hemitrygon akajei) in the early, middle and late gestational periods, respectively (n= 3 for each period). The crude composition, constituent proteins and fatty acids in the milk were analysed. The uterine milk was rich in proteins throughout the gestational period, whereas lipids dramatically increased in the middle period and reduced slightly towards the late period. Some proteins potentially associated with nutrition, cartilage growth and embryonic immunity were found. Several enzymes related to central metabolism were also detected. The constituent fatty acids in the middle and late periods were similar to those in the egg yolks of elasmobranchs, except for C18:2, which was rich only in the uterine milk. The most abundant fatty acid in the milk was C16:1, which could function as a lipokine to promote lipid metabolism in the embryo. This study's data suggest that uterine milk may be secreted in addition to the egg yolk in elasmobranchs to support rapid and healthy embryonic growth.

Antimicrobial Activity of Lactococcus lactis subsp. lactis Isolated from a Stranded Cuvier's Beaked Whale (Ziphius cavirostris) against Gram-Positive and -Negative Bacteria.

In the present study, we isolated and characterized Lactococcus lactis (L. lactis) subsp. lactis from a female Cuvier's beaked whale (Ziphius cavirostris) stranded in Shizuoka, Japan. Only five isolates (CBW1-5), grown on Lactobacilli de Man Rogosa Sharpe (MRS) agar plates prepared using 50% artificial seawater, were positive in L. lactis species-specific primer PCR. Their 16S rRNA sequences were highly similar to those of L. lactis subsp. lactis JCM 5805T. The Gram reaction, motility, gas production from glucose, catalase production, and growth conditions were consistent with those of the type strain. Additionally, carbohydrate utilization of the strains was consistent with previously reported marine organism-derived strains. The pH-neutralized cell-free culture supernatant of strain CBW2 inhibited the growth of Bacillus subtilis subsp. subtilis ATCC 6051 and Vibrio alginolyticus ATCC 17749, whereas protease treatment eliminated or diminished its inhibitory activity. The strain possesses a precursor of the nisin structural gene (nisA), which showed 100% homology with nisin Z, and nisin biosynthesis-related genes (nisB, nisC, nisT, nisP, nisF, nisI, and nisRK), suggesting that the strain produces a nisin-like substance. This study provides fundamental information on whale-derived L. lactis subsp. lactis which may be useful for reducing the carriage of B. subtilis subsp. subtilis and V. alginolyticus.

Antioxidative Potency of Dolphin Serum Albumin Is Stronger Than That of Human Serum Albumin Irrespective of Substitution of 34Cysteine With Serine.

Serum albumin (SA), the most abundant protein in circulation, functions as a carrier protein, osmoregulator, and antioxidant. Generally, SA exerts its antioxidative effects by scavenging reactive oxygen species. Because marine mammals are superior divers, they are intermittently exposed to oxidative stress induced by rapid reperfusion of oxygen to ischemic tissues after the dive. Although several antioxidants in marine mammals have been described, SA activity remains largely uncharacterized. In this study, we investigated the antioxidative activity of SA in marine mammals by comparing features of the primary and steric structures, biochemical properties, and antioxidative activities of common bottlenose dolphin SA (DSA) and human SA (HSA). Our results revealed that DSA lacked free cysteine at position 34 that is important for the antioxidative activity of HSA; however, the antioxidative capacity and thiol activity of DSA were stronger than those of HSA. Circular dichroism spectra showed different patterns in DSA and HSA. Ultraviolet fluorescence intensities of DSA were higher than those of HSA, suggesting lower surface hydrophobicity of DSA. Additionally, DSA showed higher excess heat capacity than HSA. We then compared a homology model of DSA with a 3D model of HSA. Our results indicate that DSA was more unstable than HSA at least in the body-temperature range, probably due to the mode of molecules involved in the disulfide bonds and/or the lower surface hydrophobicity, and it may be related to the equivalent or stronger antioxidant potency of DSA. These data show that DSA is an effective antioxidant in the circulation of the dolphin.

Fecal microbiota of captive Antillean manatee Trichechus manatus manatus.

Herbivorous animals have unique intestinal microbiota that greatly helps with plant digestion in the host; however, knowledge on the microbiota of marine herbivores is limited. To better understand the taxonomy of intestinal microbiota in manatees, and the possible effects of captive conditions on that, we characterized the fecal microbiota of captive Antillean manatee Trichechus manatus manatus and compared the bacterial community with that of wild Florida manatees Trichechus manatus latirostris. Fecal samples were collected from four captive Antillean manatees in Ocean Expo Park, Okinawa, Japan. The high-quality sequences of the V3-V4 region of bacterial 16S rRNA obtained using an Illumina MiSeq platform were assigned to 16 bacterial phyla, and the most dominant was Firmicutes (84.05 ± 3.50%), followed by Bacteroidetes (8.60 ± 1.71%). Seven of the top 20 bacterial genera were responsible for hydrolyzing cellulose and metabolizing bile acid. The microbiota composition was remarkably different from that found in wild Florida manatees and more diverse than the composition in wild Florida manatees; hence, this result may be dependent on a captive environment. Our results highlight the unique intestinal microbiota in captive manatees, reflecting their diet and possibly an impact of the captive environment.

Functional characterization of the GABA transporter GAT-1 from the deep-sea mussel Bathymodiolus septemdierum.

Mammalian γ-aminobutyric acid (GABA) transporter subtype 1 (GAT-1) is a specific transporter for GABA, an inhibitory neurotransmitter in GABA-ergic neurons. GAT-1 belongs to the GAT group, in which five related transporters, GAT-2, GAT-3, GAT-4, CT1, and TAUT are known in mammals. By contrast, the deep-sea mussel, Bathymodiolus septemdierum has only two GAT group members, BsGAT-1 and BsTAUT, and their function in environmental adaptation is of interest to better understand the physiology of deep-sea organisms. Compared with BsTAUT, the function of BsGAT-1 is unknown. Here, we report the functional characterization of BsGAT-1. Analyses of BsGAT-1 expressed in Xenopus oocytes showed that it could transport GABA in a Na+- and Cl--dependent manner, with Km and Vmax values of 0.58 µM and 1.97 pmol/oocyte/h, respectively. BsGAT-1 activity was blocked by the GAT-1 selective inhibitors SKF89976A and ACHC. Competition assays indicated that BsGAT-1 has no affinity for taurine and thiotaurine. These characteristics were common with those of mammalian GAT-1, suggesting its conserved function in the nervous system. However, BsGAT-1 showed a certain affinity for hypotaurine, which is involved in sulfide detoxification in hydrothermal vent-specific animals. This result suggests an additional role for BsGAT-1 in sulfide detoxification, which may be specific to the deep-sea mussel. In a tissue distribution analysis, BsGAT-1 mRNA expression was observed in various tissues. The expression in the adductor and byssus retractor muscles, labial palp, and foot, which possibly contain ganglia, suggested a function in the neural system, while BsGAT-1 expression in other tissues might be related to sulfide detoxification.

Plasma metabolomic analysis in mature female common bottlenose dolphins: profiling the characteristics of metabolites after overnight fasting by comparison with data in beagle dogs.

The present study was aimed at determining the characteristics of plasma metabolites in bottlenose dolphins to provide a greater understanding of their metabolism and to obtain information for the health management of cetaceans. Capillary electrophoresis-time-of-flight mass spectrometry (CE-TOFMS) and liquid chromatograph-time-of-flight mass spectrometry (LC-TOFMS) were conducted on plasma samples after overnight fasting from three common bottlenose dolphins as well as three beagle dogs (representative terrestrial carnivores) for comparison. In total, 257 and 227 plasma metabolites were identified in the dolphins and the dogs, respectively. Although a small number of animals were used for each species, the heatmap patterns, a principal component analysis and a cluster analysis confirmed that the composition of metabolites could be segregated from each other. Of 257 compounds detected in dolphin plasma, 24 compounds including branched amino acids, creatinine, urea, and methylhistidine were more abundant than in dogs; 26 compounds including long-chained acyl-carnitines and fatty acids, astaxanthin, and pantothenic acid were detected only in dolphins. In contrast, 25 compounds containing lactic acid and glycerol 3-phosphate were lower in dolphins compared to dogs. These data imply active protein metabolism, differences in usage of lipids, a unique urea cycle, and a low activity of the glycolytic pathway in dolphins.