Metabolic Water As a Route for Water Acquisition in Vertebrates Inhabiting Dehydrating Environments.

Vertebrates have expanded their habitats during evolution, which accompanies diversified routes for water acquisition. Water is acquired by oral intake and subsequent absorption by the intestine in terrestrial and marine animals which are subjected to constant dehydration, whereas most water is gained osmotically across body surfaces in freshwater animals. In addition, a significant amount of water, called metabolic water, is produced within the body by the oxidation of hydrogen in organic substrates. The importance of metabolic water production as a strategy for water acquisition has been well documented in desert animals, but its role has attracted little attention in marine animals which also live in a dehydrating environment. In this article, the author has attempted to reevaluate the role of metabolic water production in body fluid regulation in animals inhabiting desiccating environments. Because of the exceptional ability of their kidney, marine mammals are thought to typically gain water by drinking environmental seawater and excreting excess NaCl in the urine. On the other hand, it is established that marine teleosts drink seawater to enable intestinal water and ion absorption, and the excess NaCl is excreted by branchial ionocytes. In addition to the oral route, we suggest through experiments using eels that water production by lipid metabolism is an additional route for water acquisition when they encounter seawater. It seems that metabolic water production contributes to counteract dehydration before mechanisms for water regulation are reversed from excretion in freshwater to acquisition in seawater.

Spatial and ontogenetic modulation of fatty acid composition in juvenile European sea bass (Dicentrarchus labrax) from two French estuaries.

This study evaluated how estuary of origin and ontogenetic stage influence the fatty acid (FA) composition in the tissues of wild European sea bass juvenile. We evidenced tissue-specific patterns, with the brain exhibiting a distinct FA composition from the liver and muscle. Ontogenetic stage and estuary influenced the general FA profile, and particularly the essential FA (EFA) like docosahexaenoic acid (DHA), eicosapentaenoic acid (EPA), and arachidonic acid (ARA) in all tissues. The data also revealed the ability of wild sea bass to modulate, at the molecular level, FA biosynthesis pathways and suggest a potential dietary DHA limitation in the natural environment. The distribution of FA within tissues might reflect shifts in diet, metabolic demands, or adaptations to environmental conditions. This study provides insights about FA dynamics in euryhaline fish during juvenile life stage, improving our understanding of the metabolism need and EFA trophic availability in a changing environment.

NADK-mediated proline synthesis enhances high-salinity tolerance in the razor clam.

Euryhaline organisms can accumulate organic osmolytes to maintain osmotic balance between their internal and external environments. Proline is a pivotal organic small molecule and plays an important role in osmoregulation that enables marine shellfish to tolerate high-salinity conditions. During high-salinity challenge, NAD kinase (NADK) is involved in de novo synthesis of NADP(H) in living organisms, which serves as a reducing agent for the biosynthetic reactions. However, the role of shellfish NADK in proline biosynthesis remains elusive. In this study, we show the modulation of NADK on proline synthesis in the razor clam (Sinonovacula constricta) in response to osmotic stress. Under acute hypersaline conditions, gill tissues exhibited a significant increase in the expression of ScNADK. To elucidate the role of ScNADK in proline biosynthesis, we performed dsRNA interference in the expression of ScNADK in gill tissues to assess proline content and the expression levels of key enzyme genes involved in proline biosynthesis. The results indicate that the knock-down of ScNADK led to a significant decrease in proline content (P<0.01), as well as the expression levels of two proline synthetase genes P5CS and P5CR involved in the glutamate pathway. Razor clams preferred to use ornithine as substrate for proline synthesis when the glutamate pathway is blocked. Exogenous administration of proline greatly improved cell viability and mitigated cell apoptosis in gills. In conclusion, our results demonstrate the important role of ScNADK in augmenting proline production under high-salinity stress, by which the razor clam is able to accommodate salinity variations in the ecological niche.

Molecular evidence for the adaptive evolution in euryhaline bivalves.

Marine bivalves inhabiting intertidal and estuarine areas are frequently exposed to salinity stress due to persistent rainfall and drought. Through prolonged adaptive evolution, numerous bivalves have developed eurysalinity, which are capable of tolerating a wide range of salinity fluctuations through the sophisticated regulation of physiological metabolism. Current research has predominantly focused on investigating the physiological responses of bivalves to salinity stress, leaving a significant gap in our understanding of the adaptive evolutionary characteristics in euryhaline bivalves. Here, comparative genomics analyses were performed in two groups of bivalve species, including 7 euryhaline species and 5 stenohaline species. We identified 24 significantly expanded gene families and 659 positively selected genes in euryhaline bivalves. A significant co-expansion of solute carrier family 23 (SLC23) facilitates the transmembrane transport of ascorbic acids in euryhaline bivalves. Positive selection of antioxidant genes, such as GST and TXNRD, augments the capacity of active oxygen species (ROS) scavenging under salinity stress. Additionally, we found that the positively selected genes were significantly enriched in KEGG pathways associated with carbohydrates, lipids and amino acids metabolism (ALDH, ADH, and GLS), as well as GO terms related to transmembrane transport and inorganic anion transport (SLC22, CLCND, and VDCC). Positive selection of MCT might contribute to prevent excessive accumulation of intracellular lactic acids during anaerobic metabolism. Positive selection of PLA2 potentially promote the removal of damaged membranes lipids under salinity stress. Our findings suggest that adaptive evolution has occurred in osmoregulation, ROS scavenging, energy metabolism, and membrane lipids adjustments in euryhaline bivalves. This study enhances our understanding of the molecular mechanisms underlying the remarkable salinity adaption of euryhaline bivalves.

When ecological transitions are not so infrequent: independent colonizations of athalassohaline water bodies by Arcellidae (Arcellinida; Amoebozoa), with descriptions of four new species.

The salinity and humidity barriers divide biodiversity and strongly influence the distribution of organisms. Crossing them opens the possibility for organisms to colonize new niches and diversify, but requires profound physiological adaptations and is supposed to happen rarely in evolutionary history. We tested the relative importance of each ecological barrier by building the phylogeny, based on mitochondrial cytochrome oxidase gene (COI) sequences, of a group of microorganisms common in freshwater and soils, the Arcellidae (Arcellinida; Amoebozoa). We explored the biodiversity of this family in the sediments of athalassohaline water bodies (i.e. of fluctuating salinity that have non-marine origins). We found three new aquatic species, which represent, to the best of our knowledge, the first reports of Arcellinida in these salt-impacted ecosystems, plus a fourth terrestrial one in bryophytes. Culturing experiments performed on Arcella euryhalina sp. nov. showed similar growth curves in pure freshwater and under 20 g/L salinity, and long-term survival at 50 g/L, displaying a halotolerant biology. Phylogenetic analyses showed that all three new athalassohaline species represent independent transition events through the salinity barrier by freshwater ancestor, in contrast to the terrestrial species, which are monophyletic and represent a unique ecological transition from freshwater to soil environments.

Genomic and Transcriptomic Insights into Salinity Tolerance-Based Niche Differentiation of Synechococcus Clades in Estuarine and Coastal Waters.

Cluster 5 Synechococcus is one of the most important primary producers on earth. However, ecotypes of this genus exhibit complex geographical distributions, and the genetic basis of niche partitioning is still not fully understood. Here, we report distinct distributions of subcluster 5.1 (SC5.1) and subcluster 5.2 (SC5.2) Synechococcus in estuarine waters, and we reveal that salinity is the main factor determining their distribution. Clade III (belonging to SC5.1) and CB4 (belonging to SC5.2) are dominant clades in the study region, with different ecological distributions. We further conducted physiological, genomic, and transcriptomic studies of Synechococcus strains YX04-3 and HK05, which are affiliated with clade III and CB4, respectively. Laboratory tests showed that HK05 could grow at low salinity (13 ppt), whereas the growth of YX04-3 was suppressed when salinity decreased to 13 ppt. Genomic and transcriptomic analysis suggested that euryhaline clade CB4 is capable of dealing with a sudden drop of salinity by releasing compatible solutes through mechanosensitive channels that are coded by the mscL gene, decreasing biosynthesis of organic osmolytes, and increasing expression of heat shock proteins and high light-inducible proteins to protect photosystem. Furthermore, CB4 strain HK05 exhibited a higher growth rate when growing at low salinity than at high salinity. This is likely achieved by reducing its biosynthesis of organic osmolyte activity and increasing its photosynthetic activity at low salinity, which allowed it to enhance the assimilation of inorganic carbon and nitrogen. Together, these results provide new insights regarding the ecological distribution of SC5.2 and SC5.1 ecotypes and their underlying molecular mechanisms. IMPORTANCE Synechococcus is a group of unicellular Cyanobacteria that are widely distributed in global aquatic ecosystems. Salinity is a factor that affects the distribution of microorganisms in estuarine and coastal environments. In this study, we studied the distribution pattern of Synechococcus community along the salinity gradient in a subtropical estuary. By using omic methods, we unveiled genetic traits that determine the niche partitioning of euryhaline and strictly marine Synechococcus. We also explored the strategies employed by euryhaline Synechococcus to cope with a sudden drop of salinity, and revealed possible mechanisms for the higher growth rate of euryhaline Synechococcus in low salinity conditions. This study provides new insight into the genetic basis of niche partitioning of Synechococcus clades.

Factors affecting 137Cs radioactivity and water-to-body concentration ratios of fish in river and pond environments near the Fukushima Dai-ichi Nuclear Power Plant.

To elucidate 137Cs contamination levels and mechanisms of fish inhabiting river and pond environments near the Fukushima Dai-ichi Nuclear Power Plant, 137Cs activity concentrations in fish (15 species, n = 164) and water collected from Maeda River (3.3-8.9 km from the plant) and Shimofukazawa Pond (2.9 km) in 2017 were analyzed. Also, an 8-week rearing experiment using Japanese dace Pseudaspius hakonensis fed on non-contaminated pellets and the pond water (mean 137Cs concentration of 2.0 Bq/L) was conducted to evaluate 137Cs accumulation from water to fish. The 137Cs concentrations in Japanese dace, the only species collected throughout five sampling sites from estuarine to upstream areas in Maeda River, were found to be correlated with ambient air dose rates and fish size, exhibiting large variations (16.5-2.6×103 Bq/kg-wet). By contrast, dissolved 137Cs in river waters increased from the upper to lower course (0.025-0.28 Bq/L), which caused large variations of the water-to-body concentration ratio (CR) in Japanese dace (60.0-35700 L/kg-wet). These CRs (geometric mean of 3670 L/kg-wet) were much higher than the steady-state CR of reared fish (9.7 L/kg-wet), indicating that river fish uptake 137Cs mainly from prey items from aquatic and riparian zones, rather than from water. Statistically significant negative correlations between K+ concentrations in water and river fish CRs were detected, resulting in the decreasing trend of CRs from upstream to estuarine areas. These results suggest that the large heterogeneity of air dose rates, K+ concentration, and estuarine processes in brackish water habitats, in association with the feeding habit and size effect in fish, can engender wide variation of 137Cs concentrations and CRs of river fish along a river course. In contrast, 137Cs concentrations in pond fish (4.3-14.6 kBq/kg-wet) were higher than in river fish. The CRs of pond fish were constantly high but the range was smaller (1010-3440 L/kg-wet) with larger values in fish of higher trophic levels. These findings suggest that biomagnification within a pond was inferred as the main cause of 137Cs contamination of pond fish.

Transcriptome Analysis Revealed Osmoregulation Related Regulatory Networks and Hub Genes in the Gills of Hilsa shad, Tenualosa ilisha, during the Migratory Osmotic Stress.

Tenualosa ilisha (Hilsa shad), an anadromous fish, usually inhabits coastal and estuarine waters, and migrates to freshwater for spawning. In this study, large-scale gill transcriptome analyses from three salinity regions, i.e., fresh, brackish and marine water, revealed 3277 differentially expressed genes (DEGs), out of which 232 were found to be common between marine vs freshwater and brackish vs freshwater. These genes were mapped into 54 KEGG Pathways, and the most significant of these were focal adhesion, adherens junction, tight junction, and PI3K-Akt signaling pathways. A total of 24 osmoregulatory genes were found to be differentially expressed in different habitats. The gene members of slc16 and slc2 families showed a dissimilar pattern of expressions, while two claudin genes (cldn11 & cldn10), transmembrane tm56b, and voltage-gated potassium channel gene kcna10 were downregulated in freshwater samples, as compared to that of brackish and marine environment. Protein-protein interaction (PPI) network analysis of 232 DEGs showed 101 genes to be involved in PPI, while fn1 gene was found to be interacting with the highest number of genes (36). Twenty-five hub genes belonged to 12 functional groups, with muscle structure development with seven genes, forming the major group. These results provided valuable information about the genes, potentially involved in the molecular mechanisms regulating water homeostasis in gills, during migration for spawning and low-salinity adaptation in Hilsa shad. These genes may form the basis for the bio-marker development for adaptation to the stress levied by major environmental changes, due to hatchery/culture conditions.

Effects of seawater and freshwater challenges on the Gh/Igf system in the saline-tolerant blackchin tilapia (Sarotherodon melanotheron).

Prolactin (Prl) and growth hormone (Gh) as well as insulin-like growth factor 1 (Igf1) are involved in the physiological adaptation of fish to varying salinities. The Igfs have been also ascribed other physiological roles during development, growth, reproduction and immune regulation. However, the main emphasis in the investigation of osmoregulatory responses has been the endocrine, liver-derived Igf1 route and local regulation within the liver and osmoregulatory organs. Few studies have focused on the impact of salinity alterations on the Gh/Igf-system within the neuroendocrine and immune systems and particularly in a salinity-tolerant species, such as the blackchin tilapia Sarotherodon melanotheron. This species is tolerant to hypersalinity and saline variations, but it is confronted by severe climate changes in the Saloum inverse estuary. Here we investigated bidirectional effects of increased salinity followed by its decrease on the gene regulation of prl, gh, igf1, igf2, Gh receptor and the tumor-necrosis factor a. A mixed population of sexually mature 14-month old blackchin tilapia adapted to freshwater were first exposed to seawater for one week and then to fresh water for another week. Brain, pituitary, head kidney and spleen were excised at 4 h, 1, 2, 3 and 7 days after both exposures and revealed differential expression patterns. This investigation should give us a better understanding of the role of the Gh/Igf system within the neuroendocrine and immune organs and the impact of bidirectional saline challenges on fish osmoregulation in non-osmoregulatory organs, notably the complex orchestration of growth factors and cytokines.

Compensation mechanism for membrane potential against hypoosmotic stress in the Onchidium neuron.

The effect of acute hypoosmotic stress on the neural response was investigated using the neurons identified in the abdominal ganglion of the amphibious mollusk Onchidium. The membrane potential of an identified neuron (Ip-1/2) was not significantly altered in 50% hypoosmotic artificial sea water. In isotonic 50% artificial seawater (ASW) with osmolarity that was compensated for using glycerol or urea, the membrane potentials of Ip-1/2 were also not altered compared to those in 50% hypoosmotic ASW. However, hyperpolarization was induced in isotonic 50% ASW when osmolarity was compensated for using sucrose or mannose. In the presence of volume-regulated anion channel (VRAC) inhibitors (niflumic acid and glibenclamide), the Ip-1/2 membrane potentials were hyperpolarized in 50% hypoosmotic ASW. These results suggest that there is a compensatory mechanism involving aquaglyceroporin and VRAC-like channels that maintains membrane potential under hypoosmotic conditions. Here, we detected the expression of aquaglyceroporin mRNA in neural tissues of Onchidium.

Quantifying euryhaline histories in red drum Sciaenops ocellatus: otolith chemistry and muscle isotope ratios.

The combined use of otolith chemistry and tissue isotopes has the potential to reveal movements, habitat associations and food web interactions at a variety of spatial and temporal scales. Here, we used a combination of otolith Ba:Ca life history transects with muscle tissue δ13 C and δ15 N values to assess habitat use and oligohaline residence in red drum Sciaenops ocellatus in subtropical estuaries in the northwestern Gulf of Mexico. Tissue isotopes were distinct among capture locations, particularly between bays with differing proximity to freshwater inflow sources. Otolith edge Ba:Ca values and tissue δ13 C values were not correlated. These results indicated that fish were not residing in nor feeding in oligohaline waters for significant periods of time within the tissue turnover window of several months prior to capture. However, spatial differences in tissue isotope values indicated limited mixing among bays and relatively high site fidelity during estuarine occupancy. Lifetime otolith Ba:Ca transects revealed individual variability in the magnitude of residence in oligohaline waters. Using a medium oligohaline occupancy threshold, an estimated 82% of individuals used oligohaline waters at some point in their life. However, 66% of individuals spent less than 20% of their life histories in oligohaline waters, suggesting intermittent and infrequent excursions into low salinity waters. Finally, a literature survey identified 56 peer-reviewed publications using combinations of otolith chemistry and tissue stable isotope ratios with a wide range of marker pairings and study aims. The diversity of ecological questions that can be asked with the combined use of these two approaches will provide valuable insight into fish ecology. This article is protected by copyright. All rights reserved.

Endocrine and osmoregulatory responses to tidally-changing salinities in fishes.

Salinity is one of the main physical properties that govern the distribution of fishes across aquatic habitats. In order to maintain their body fluids near osmotic set points in the face of salinity changes, euryhaline fishes rely upon tissue-level osmotically-induced responses and systemic endocrine signaling to direct adaptive ion-transport processes in the gill and other critical osmoregulatory organs. Some euryhaline teleosts inhabit tidally influenced waters such as estuaries where salinity can vary between fresh water (FW) and seawater (SW). The physiological adaptations that underlie euryhalinity in teleosts have been traditionally identified in fish held under steady-state conditions or following unidirectional transfers between FW and SW. Far fewer studies have employed salinity regimes that simulate the tidal cycles that some euryhaline fishes may experience in their native habitats. With an emphasis on prolactin (Prl) signaling and branchial ionocytes, this mini-review contrasts the physiological responses between euryhaline fish responding to tidal versus unidirectional changes in salinity. Three patterns that emerged from studying Mozambique tilapia (Oreochromis mossambicus) subjected to tidally-changing salinities include, 1) fish can compensate for continuous and marked changes in external salinity to maintain osmoregulatory parameters within narrow ranges, 2) tilapia maintain branchial ionocyte populations in a fashion similar to SW-acclimated fish, and 3) there is a shift from systemic to local modulation of Prl signaling.

Chromosome-level Genome Assembly of Acanthopagrus latus Provides Insights into Salinity Stress Adaptation of Sparidae.

The yellowfin seabream, Acanthopagrus latus, is widely distributed throughout the Indo-West Pacific. This species, as a euryhaline Sparidae fish, inhabits in coastal environments with large and frequent salinity fluctuation. So the A. latus can be considered as an ideal species for elucidating the evolutionary mechanism of salinity stress adaption on teleost fish species. Here, a chromosome-scale assembly of A. latus was obtained with PacBio and Hi-C hybrid sequencing strategy. The final assembly genome of A. latus is 685.14 Mbp. The values of contig N50 and scaffold N50 are 14.88 Mbp and 30.72 Mbp, respectively. 29,227 genes were successfully predicted for A. latus in total. Then, the comparative genomics and phylogenetic analysis were employed for investigating the different osmoregulation strategies of salinity stress adaption on multiple whole genome scale of Sparidae species. The highly accurate chromosomal information provides the important genome resources for understanding the osmoregulation evolutionary pattern of the euryhaline Sparidae species.

Optimal composition of chloride cells for osmoregulation in a randomly fluctuating environment.

Fish live in water with a different osmotic pressure from that in the body. Their gills have chloride cells that transport ions to maintain an appropriate level of osmotic pressure in the body. The direction of ion transport is different between seawater and freshwater. There are two types of chloride cells that specialize in unidirectional transport and generalist cells that can switch their function quickly in response to environmental salinity. In species that experience salinity changes throughout life (euryhaline species), individuals may replace some chloride cells with cells of different types upon a sudden change in environmental salinity. In this paper, we develop a dynamic optimization model for the chloride cell composition of an individual living in an environment with randomly fluctuating salinity. The optimal solution is to minimize the sum of the workload of chloride cells in coping with the difference in osmotic pressure, the maintenance cost, and the temporal cost due to environmental change. The optimal fraction of generalist chloride cells increases with the frequency of salinity changes and the time needed for new cells to be fully functional but decreases with excess maintenance cost.

Transcriptomic analysis of brine shrimp Artemia franciscana across a wide range of salinities.

Brine shrimp Artemia franciscana, a commercially important species, can thrive in a wide range of salinities and is commonly found in hypersaline lakes and solar salterns. Transcriptome analysis can enhance the understanding of the adaptative mechanisms of brine shrimp in aquaculture. RNA sequencing (RNAseq) data was generated from A. franciscana adults that were salt-adapted for 2-4 weeks at five salinities: 35, 50, 100, 150, and 230 psu. Long-read isoform sequencing (IsoSeq) data was used to construct a high-quality transcriptome assembly. Also, the gene expression patterns in A. franciscana adults were examined. Notably, the transcriptional response of A. franciscana's acclimation to intermediate salinities (50-150 psu) displayed frequently and differentially U-shaped or inverted U-shaped expression patterns. In addition, the types of genes showing two nonmonotonic expression patterns were distinct from each other. The coordinated shifts in gene expression suggest different homeostatic strategies of A. franciscana at specific salinities; such strategies may enhance population fitness at extreme salinities. Our study should promote a scientific concept for the gene expression patterns of A. franciscana along a broad salinity gradient, and a variety of salinity and prey should be monitored for testing the gene expression pattern of this important aquaculture species.

A genetically based ecological trade-off contributes to setting a geographic range limit.

Understanding the ecological factors that shape geographic range limits and the evolutionary constraints that prevent populations from adaptively evolving beyond these limits is an unresolved question. Here, we investigated why the euryhaline fish, Poecila reticulata, is confined to freshwater within its native range, despite being tolerant of brackish water. We hypothesised that competitive interactions with a close relative, Poecilia picta, in brackish water prevents P. reticulata from colonising brackish water. Using a combination of field transplant, common garden breeding, and laboratory behaviour experiments, we find support for this hypothesis, as P. reticulata are behaviourally subordinate and have lower survival in brackish water with P. picta. We also found a negative genetic correlation between P. reticulata growth in brackish water versus freshwater in the presence of P. picta, suggesting a genetically based trade-off between salinity tolerance and competitive ability could constrain adaptive evolution at the range limit.

Estrogen-related receptor γ2 controls NaCl uptake to maintain ionic homeostasis.

Estrogen-related receptors (ERRs) are known to function in mammalian kidney as key regulators of ion transport-related genes; however, a comprehensive understanding of the physiological functions of ERRs in vertebrate body fluid ionic homeostasis is still elusive. Here, we used medaka (Oryzias melastigma), a euryhaline teleost, to investigate how ERRs are involved in ion regulation. After transferring medaka from hypertonic seawater to hypotonic freshwater (FW), the mRNA expression levels of errγ2 were highly upregulated, suggesting that Errγ2 may play a crucial role in ion uptake. In situ hybridization showed that errγ2 was specifically expressed in ionocytes, the cells responsible for Na+/Cl- transport. In normal FW, ERRγ2 morpholino knockdown caused reductions in the mRNA expression of Na+/Cl- cotransporter (Ncc), the number of Ncc ionocytes, Na+/Cl- influxes of ionocytes, and whole-body Na+/Cl- contents. In FW with low Na+ and low Cl-, the expression levels of mRNA for Na+/H+ exchanger 3 (Nhe3) and Ncc were both decreased in Errγ2 morphants. Treating embryos with DY131, an agonist of Errγ, increased the whole-body Na+/Cl- contents and ncc mRNA expression in Errγ2 morphants. As such, medaka Errγ2 may control Na+/Cl- uptake by regulating ncc and/or nhe3 mRNA expression and ionocyte number, and these regulatory actions may be subtly adjusted depending on internal and external ion concentrations. These findings not only provide new insights into the underpinning mechanism of actions of ERRs, but also enhance our understanding of their roles in body fluid ionic homeostasis for adaptation to changing environments during vertebrate evolution.

Effects of temperature and salinity on antioxidant responses in livers of temperate (Dicentrarchus labrax) and tropical (Chanos Chanos) marine euryhaline fish.

Temperature and salinity are abiotic factors that affect physiological responses in aquaculture species. The European sea bass (Dicentrarchus labrax) is a temperate species that is generally farmed at 18 °C in seawater (SW). In the wild, its incursions in shallow habitats such as lagoons may result in hyperthermal damage despite its high thermal tolerance. Meanwhile, the milkfish (Chanos chanos), a tropical species, is generally reared at 28 °C, and in winter, high mortality usually occurs under hypothermal stress such as cold snaps. This study compared changes in hepatic antioxidant enzymes (superoxide dismutase, SOD; and catalase, CAT) in these two important marine euryhaline aquaculture species in Europe and Southeast Asia, respectively, under temperature challenge combined with hypo-osmotic (fresh water, FW) stress. After a four-week hyper- or hypo-thermal treatment, hepatic SOD activity was upregulated in both species reared in SW and FW, indicating enhanced oxidative stress in European sea bass and milkfish. The expression profiles of sod isoforms suggested that in milkfish, the increase in reactive oxygen species (ROS) was mainly at the cytosol level, leading to increased sod1 expression. In European sea bass, however, no obvious difference was found between the expression of sod isoforms at different temperatures. A lower expression of sod2 was observed in FW compared to SW in the latter species. Moreover, no significant change was observed in the mRNA expression and activity of CAT in the livers of these two species under the different temperature treatments, with the exception of the lower CAT activity in milkfish challenged with SW at 18 °C. Taken together, our results indicated that the antioxidant responses were not changed under long-term hypoosmotic challenge but were enhanced during the four-week temperature treatments in livers of both the temperate and tropical euryhaline species.

Enhanced expression of ncc1 and clc2c in the kidney and urinary bladder accompanies freshwater acclimation in Mozambique tilapia.

Euryhaline fishes maintain hydromineral balance in a broad range of environmental salinities via the activities of multiple osmoregulatory organs, namely the gill, gastrointestinal tract, skin, kidney, and urinary bladder. Teleosts residing in freshwater (FW) environments are faced with the diffusive loss of ions and the osmotic gain of water, and, therefore, the kidney and urinary bladder reabsorb Na+ and Cl- to support the production of dilute urine. Nonetheless, the regulated pathways for Na+ and Cl- transport by euryhaline fishes, especially in the urinary bladder, have not been fully resolved. Here, we first investigated the ultrastructure of epithelial cells within the urinary bladder of FW-acclimated Mozambique tilapia (Oreochromis mossambicus) by electron microscopy. We then investigated whether tilapia employ Na+/Cl- cotransporter 1 (Ncc1) and Clc family Cl- channel 2c (Clc2c) for the reabsorption of Na+ and Cl- by the kidney and urinary bladder. We hypothesized that levels of their associated gene transcripts vary inversely with environmental salinity. In whole kidney and urinary bladder homogenates, ncc1 and clc2c mRNA levels were markedly higher in steady-state FW- versus SW (seawater)-acclimated tilapia. Following transfer from SW to FW, ncc1 and clc2c in both the kidney and urinary bladder were elevated within 48 h. A concomitant increase in branchial ncc2, and decreases in Na+/K+/2Cl-cotransporter 1a (nkcc1a) and cystic fibrosis transmembrane regulator 1 (cftr1) levels indicated a transition from Na+ and Cl- secretion to absorption by the gills in parallel with the identified renal and urinary bladder responses to FW transfer. Our findings suggest that Ncc1 and Clc2c contribute to the functional plasticity of the kidney and urinary bladder in tilapia.

Ion Transporters and Osmoregulation in the Kidney of Teleost Fishes as a Function of Salinity.

Euryhaline teleosts exhibit major changes in renal function as they move between freshwater (FW) and seawater (SW) environments, thus tolerating large fluctuations in salinity. In FW, the kidney excretes large volumes of water through high glomerular filtration rates (GFR) and low tubular reabsorption rates, while actively reabsorbing most ions at high rates. The excreted product has a high urine flow rate (UFR) with a dilute composition. In SW, GFR is greatly reduced, and the tubules reabsorb as much water as possible, while actively secreting divalent ions. The excreted product has a low UFR, and is almost isosmotic to the blood plasma, with Mg2+, SO4 2-, and Cl- as the major ionic components. Early studies at the organismal level have described these basic patterns, while in the last two decades, studies of regulation at the cell and molecular level have been implemented, though only in a few euryhaline groups (salmonids, eels, tilapias, and fugus). There have been few studies combining the two approaches. The aim of the review is to integrate known aspects of renal physiology (reabsorption and secretion) with more recent advances in molecular water and solute physiology (gene and protein function of transporters). The renal transporters addressed include the subunits of the Na+, K+- ATPase (NKA) enzyme, monovalent ion transporters for Na+, Cl-, and K+ (NKCC1, NKCC2, CLC-K, NCC, ROMK2), water transport pathways [aquaporins (AQP), claudins (CLDN)], and divalent ion transporters for SO4 2-, Mg2+, and Ca2+ (SLC26A6, SLC26A1, SLC13A1, SLC41A1, CNNM2, CNNM3, NCX1, NCX2, PMCA). For each transport category, we address the current understanding at the molecular level, try to synthesize it with classical knowledge of overall renal function, and highlight knowledge gaps. Future research on the kidney of euryhaline fishes should focus on integrating changes in kidney reabsorption and secretion of ions with changes in transporter function at the cellular and molecular level (gene and protein verification) in different regions of the nephrons. An increased focus on the kidney individually and its functional integration with the other osmoregulatory organs (gills, skin and intestine) in maintaining overall homeostasis will have applied relevance for aquaculture.