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Abstract Background Cyprinidae, the largest fish family, encompasses approximately 367 genera and 3006 species. While they exhibit remarkable adaptability to diverse aquatic environments, it is exceptionally rare to find them in seawater, with the Far Eastern daces being of few exceptions. Therefore, the Far Eastern daces serve as a valuable model for studying the genetic mechanisms underlying seawater adaptation in Cyprinidae. Results Here, we sequenced the chromosome-level genomes of two Far Eastern daces (Pseudaspius brandtii and P. hakonensis), the two known cyprinid fishes found in seawater, and performed comparative genomic analyses to investigate their genetic mechanism of seawater adaptation. Demographic history reconstruction of the two species reveals that their population dynamics are correlated with the glacial-interglacial cycles and sea level changes. Genomic analyses identified Pseudaspius-specific genetic innovations related to seawater adaptation, including positively selected genes, rapidly evolving genes, and conserved non-coding elements (CNEs). Functional assays of Pseudaspius-specific variants of the prolactin (prl) gene showed enhanced cell adaptation to greater osmolarity. Functional assays of Pseudaspius specific CNEs near atg7 and usp45 genes suggest that they exhibit higher promoter activity and significantly induced at high osmolarity. Conclusions Our results reveal the genome-wide evidence for the evolutionary adaptation of cyprinid fishes to seawater, offering valuable insights into the molecular mechanisms supporting the survival of migratory fish in marine environments. These findings are significant as they contribute to our understanding of how cyprinid fishes navigate and thrive in diverse aquatic habitats, providing useful implications for the conservation and management of marine ecosystems.

Background: Cyprinidae, the largest fish family, encompasses approximately 367 genera and 3006 species. While they exhibit remarkable adaptability to diverse aquatic environments, it is exceptionally rare to find them in seawater, with the Far Eastern daces being of few exceptions. Therefore, the Far Eastern daces serve as a valuable model for studying the genetic mechanisms underlying seawater adaptation in Cyprinidae. Results: Here, we sequenced the chromosome-level genomes of two Far Eastern daces (Pseudaspius brandtii and P. hakonensis), the two known cyprinid fishes found in seawater, and performed comparative genomic analyses to investigate their genetic mechanism of seawater adaptation. Demographic history reconstruction of the two species reveals that their population dynamics are correlated with the glacial-interglacial cycles and sea level changes. Genomic analyses identified Pseudaspius-specific genetic innovations related to seawater adaptation, including positively selected genes, rapidly evolving genes, and conserved non-coding elements (CNEs). Functional assays of Pseudaspius-specific variants of the prolactin (prl) gene showed enhanced cell adaptation to greater osmolarity. Functional assays of Pseudaspius specific CNEs near atg7 and usp45 genes suggest that they exhibit higher promoter activity and significantly induced at high osmolarity. Conclusions: Our results reveal the genome-wide evidence for the evolutionary adaptation of cyprinid fishes to seawater, offering valuable insights into the molecular mechanisms supporting the survival of migratory fish in marine environments. These findings are significant as they contribute to our understanding of how cyprinid fishes navigate and thrive in diverse aquatic habitats, providing useful implications for the conservation and management of marine ecosystems. [ABSTRACT FROM AUTHOR]

Smoltification (parr-smolt transformation) is a complex developmental process consisting of developmental changes that lead to remodeling of the Atlantic salmon gill. Here, the expression changes of miRNAs and mRNAs were studied by small-RNA sequencing and microarray analysis, respectively, to identify miRNAs and their predicted targets associated with smoltification and subsequent sea water adaptation (SWA). In total, 18 guide miRNAs were identified as differentially expressed (gDE miRNAs). Hierarchical clustering analysis of expression changes divided these into one cluster of 13 gDE miRNAs with decreasing expression during smoltification and SWA that included the miRNA-146, miRNA-30 and miRNA-7132 families. Another smaller cluster that showed increasing expression consisted of miR-101a-3p, miR-193b-5p, miR-499a-5p, miR-727a-3p and miR-8159-5p. The gDE miRNAs were predicted to target 747 of the genes (DE mRNAs), showing expression changes in the microarray analysis. The predicted targets included genes encoding NKA-subunits, aquaporin-subunits, cystic fibrosis transmembrane conductance regulator and the solute carrier family. Furthermore, the predicted target genes were enriched in biological processes associated with smoltification and SWA (e.g., immune system, reactive oxygen species, stress response and extracellular matrix organization). Collectively, the results indicate that remodeling of the gill involves the post-transcriptional regulation of gene expression by the characterized gDE miRNAs. [ABSTRACT FROM AUTHOR]

Simple Summary: Smoltification is a developmental process that preadapts Atlantic salmon for a life in seawater. Suboptimal smoltification and poor timing of transfer to seawater is associated with increased mortality. MicroRNAs (miRNAs) are small non-coding genes. They regulate gene expression post-transcriptionally as part of the miRNA induce silencing complex (miRISC) where they guide miRISC to particular mRNAs (target genes). The aim of this study was to identify Atlantic salmon miRNAs expressed in liver that are associated with smoltification and adaptation to seawater as well as to predict their target genes. In total, 62 guide miRNAs were identified, and by their expression patterns they were clustered into three groups. Target gene predictions followed by gene enrichment analysis of the predicted targets indicated that the guide miRNAs were involved in post-transcriptional regulation of important smoltification associated biological processes. Some of these were energy metabolism, protein metabolism and transport, circadian rhythm, stress and immune response. Together, the results indicate that certain miRNAs are involved in the regulation of many of the important changes occurring in the liver during this developmental transition. Optimal smoltification is crucial for normal development, growth, and health of farmed Atlantic salmon in seawater. Here, we characterize miRNA expression in liver to reveal whether miRNAs regulate gene expression during this developmental transition. Expression changes of miRNAs and mRNAs was studied by small-RNA sequencing and microarray analysis, respectively. This revealed 62 differentially expressed guide miRNAs (gDE-miRNAs) that could be divided into three groups with characteristic dynamic expression patterns. Three of miRNA families are known as highly expressed in liver. A rare arm shift was observed during smoltification in the Atlantic salmon-specific novel-ssa-miR-16. The gDE-miRNAs were predicted to target 2804 of the genes revealing expression changes in the microarray analysis. Enrichment analysis revealed that targets were significantly enriched in smoltification-associated biological process groups. These included lipid and cholesterol synthesis, carbohydrate metabolism, protein metabolism and protein transport, immune system genes, circadian rhythm and stress response. The results indicate that gDE-miRNAs may regulate many of the changes associated with this developmental transition in liver. The results pave the way for validation of the predicted target genes and further study of gDE-miRNA and their targets by functional assays. [ABSTRACT FROM AUTHOR]

Abstract Adaptation to a hypertonic marine environment is one of the major topics in animal physiology research. Marine teleosts lose water osmotically from the gills and compensate for this loss by drinking surrounding seawater and absorbing water from the intestine. This situation is in contrast to that in mammals, which experience a net osmotic loss of water after drinking seawater. Water absorption in fishes is made possible by (1) removal of monovalent ions (desalinization) by the esophagus, (2) removal of divalent ions as carbonate (Mg/CaCO3) precipitates promoted by HCO3 − secretion, and (3) facilitation of NaCl and water absorption from diluted seawater by the intestine using a suite of unique transporters. As a result, 70–85% of ingested seawater is absorbed during its passage through the digestive tract. Thus, the digestive tract is an essential organ for marine teleost survival in the hypertonic seawater environment. The eel is a species that has been frequently used for osmoregulation research in laboratories worldwide. The eel possesses many advantages as an experimental animal for osmoregulation studies, one of which is its outstanding euryhalinity, which enables researchers to examine changes in the structure and function of the digestive tract after direct transfer from freshwater to seawater. In recent years, the molecular mechanisms of ion and water transport across epithelial cells (the transcellular route) and through tight junctions (the paracellular route) have been elucidated for the esophagus and intestine. Thanks to the rapid progress in analytical methods for genome databases on teleosts, including the eel, the molecular identities of transporters, channels, pumps and junctional proteins have been clarified at the isoform level. As 10 y have passed since the previous reviews on this subject, it seems relevant and timely to summarize recent progress in research on the molecular mechanisms of water and ion transport in the digestive tract in eels and to compare the mechanisms with those of other teleosts and mammals from comparative and evolutionary viewpoints. We also propose future directions for this research field to achieve integrative understanding of the role of the digestive tract in adaptation to seawater with regard to pathways/mechanisms including the paracellular route, divalent ion absorption, metabolon formation and cellular trafficking of transporters. Notably, some of these have already attracted practical attention in laboratories.

Abstract Most Atlantic salmon (Salmo salar L.) populations follow an anadromous life cycle, spending early life in freshwater, migrating to the sea for feeding, and returning to rivers to spawn. At the end of the last ice age ~10,000 years ago, several populations of Atlantic salmon became landlocked. Comparing their genomes to their anadromous counterparts can help identify genetic variation related to either freshwater residency or anadromy. The objective of this study was to identify consistently divergent loci between anadromous and landlocked Atlantic salmon strains throughout their geographical distribution, with the long‐term aim of identifying traits relevant for salmon aquaculture, including fresh and seawater growth, omega‐3 metabolism, smoltification, and disease resistance. We used a Pool‐seq approach (n = 10–40 individuals per population) to sequence the genomes of twelve anadromous and six landlocked Atlantic salmon populations covering a large part of the Northern Hemisphere and conducted a genomewide association study to identify genomic regions having been under different selection pressure in landlocked and anadromous strains. A total of 28 genomic regions were identified and included cadm1 on Chr 13 and ppargc1a on Chr 18. Seven of the regions additionally displayed consistently reduced heterozygosity in fish obtained from landlocked populations, including the genes gpr132, cdca4, and sertad2 on Chr 15. We also found 16 regions, including igf1 on Chr 17, which consistently display reduced heterozygosity in the anadromous populations compared to the freshwater populations, indicating relaxed selection on traits associated with anadromy in landlocked salmon. In conclusion, we have identified 37 regions which may harbor genetic variation relevant for improving fish welfare and quality in the salmon farming industry and for understanding life‐history traits in fish.

Adaptation to a hypertonic marine environment is one of the major topics in animal physiology research. Marine teleosts lose water osmotically from the gills and compensate for this loss by drinking surrounding seawater and absorbing water from the intestine. This situation is in contrast to that in mammals, which experience a net osmotic loss of water after drinking seawater. Water absorption in fishes is made possible by (1) removal of monovalent ions (desalinization) by the esophagus, (2) removal of divalent ions as carbonate (Mg/CaCO3) precipitates promoted by HCO3− secretion, and (3) facilitation of NaCl and water absorption from diluted seawater by the intestine using a suite of unique transporters. As a result, 70–85% of ingested seawater is absorbed during its passage through the digestive tract. Thus, the digestive tract is an essential organ for marine teleost survival in the hypertonic seawater environment. The eel is a species that has been frequently used for osmoregulation research in laboratories worldwide. The eel possesses many advantages as an experimental animal for osmoregulation studies, one of which is its outstanding euryhalinity, which enables researchers to examine changes in the structure and function of the digestive tract after direct transfer from freshwater to seawater. In recent years, the molecular mechanisms of ion and water transport across epithelial cells (the transcellular route) and through tight junctions (the paracellular route) have been elucidated for the esophagus and intestine. Thanks to the rapid progress in analytical methods for genome databases on teleosts, including the eel, the molecular identities of transporters, channels, pumps and junctional proteins have been clarified at the isoform level. As 10 y have passed since the previous reviews on this subject, it seems relevant and timely to summarize recent progress in research on the molecular mechanisms of water and ion transport in the digestive tract in eels and to compare the mechanisms with those of other teleosts and mammals from comparative and evolutionary viewpoints. We also propose future directions for this research field to achieve integrative understanding of the role of the digestive tract in adaptation to seawater with regard to pathways/mechanisms including the paracellular route, divalent ion absorption, metabolon formation and cellular trafficking of transporters. Notably, some of these have already attracted practical attention in laboratories. [ABSTRACT FROM AUTHOR]

Most Atlantic salmon (Salmo salar L.) populations follow an anadromous life cycle, spending early life in freshwater, migrating to the sea for feeding, and returning to rivers to spawn. At the end of the last ice age ~10,000 years ago, several populations of Atlantic salmon became landlocked. Comparing their genomes to their anadromous counterparts can help identify genetic variation related to either freshwater residency or anadromy. The objective of this study was to identify consistently divergent loci between anadromous and landlocked Atlantic salmon strains throughout their geographical distribution, with the long‐term aim of identifying traits relevant for salmon aquaculture, including fresh and seawater growth, omega‐3 metabolism, smoltification, and disease resistance. We used a Pool‐seq approach (n = 10–40 individuals per population) to sequence the genomes of twelve anadromous and six landlocked Atlantic salmon populations covering a large part of the Northern Hemisphere and conducted a genomewide association study to identify genomic regions having been under different selection pressure in landlocked and anadromous strains. A total of 28 genomic regions were identified and included cadm1 on Chr 13 and ppargc1a on Chr 18. Seven of the regions additionally displayed consistently reduced heterozygosity in fish obtained from landlocked populations, including the genes gpr132, cdca4, and sertad2 on Chr 15. We also found 16 regions, including igf1 on Chr 17, which consistently display reduced heterozygosity in the anadromous populations compared to the freshwater populations, indicating relaxed selection on traits associated with anadromy in landlocked salmon. In conclusion, we have identified 37 regions which may harbor genetic variation relevant for improving fish welfare and quality in the salmon farming industry and for understanding life‐history traits in fish. [ABSTRACT FROM AUTHOR]

Optimal smoltification is crucial for normal development, growth, and health of farmed Atlantic salmon in seawater. Here, we characterize miRNA expression in liver to reveal whether miRNAs regulate gene expression during this developmental transition. Expression changes of miRNAs and mRNAs was studied by small-RNA sequencing and microarray analysis, respectively. This revealed 62 differentially expressed guide miRNAs (gDE-miRNAs) that could be divided into three groups with characteristic dynamic expression patterns. Three of miRNA families are known as highly expressed in liver. A rare arm shift was observed during smoltification in the Atlantic salmon-specific novel-ssa-miR-16. The gDE-miRNAs were predicted to target 2804 of the genes revealing expression changes in the microarray analysis. Enrichment analysis revealed that targets were significantly enriched in smoltification-associated biological process groups. These included lipid and cholesterol synthesis, carbohydrate metabolism, protein metabolism and protein transport, immune system genes, circadian rhythm and stress response. The results indicate that gDE-miRNAs may regulate many of the changes associated with this developmental transition in liver. The results pave the way for validation of the predicted target genes and further study of gDE-miRNA and their targets by functional assays.

Salinity adaptation and sea water tolerance are vital, challenging phenomena in the life history of anadromous species such as kuttum fish. Evaluation on feed deprivation in gill chloride cell development of kuttum fish is the aim of this study. Kuttum juveniles that weighed an average weight of 0.5 grams were divided into two groups. One group (control) was fed ad libitum during the experiment. Fish in the other group were feed deprived. Both groups were exposed to the salinity challenge for 7 days. Gill sampling was performed on the second, third, fourth, and seventh days after salinity challenge initiation. Consequently, we evaluated Na+ K+ ATPase activity, gill chloride cell numbers and size. Results showed a significant decrease in gill Na+ K+ ATPase activity, chloride cell diameter, perimeter and area due tofasting. Based on the obtained results we concluded that feed deprivation might have a disruptive effect on chloride cell development during seawater adaptation.

Taxon-specific de novo protein-coding sequences are thought to be important for taxon-specific environmental adaptation. A recent study revealed that bottlenose dolphins acquired a novel isoform of aquaporin 2 generated by alternative splicing (alternative AQP2), which helps dolphins to live in hyperosmotic seawater. The AQP2 gene consists of four exons, but the alternative AQP2 gene lacks the fourth exon and instead has a longer third exon that includes the original third exon and a part of the original third intron. Here, we show that the latter half of the third exon of the alternative AQP2 arose from a non-protein-coding sequence. Intact ORF of this de novo sequence is shared not by all cetaceans, but only by delphinoids. However, this sequence is conservative in all modern cetaceans, implying that this de novo sequence potentially plays important roles for marine adaptation in cetaceans. [ABSTRACT FROM AUTHOR]

Background: During the course of evolution, fishes have acquired adaptability to various salinity environments, and acquirement of seawater (SW) adaptability has played important roles in fish evolution and diversity. However, little is known about how saline environments influence the acquirement of SW adaptability. The Japanese medaka Oryzias latipes is a euryhaline species that usually inhabits freshwater (FW), but is also adaptable to full-strength SW when transferred through diluted SW. In the present study, we examined how past SW experience affects hyposmoregulatory ability in Japanese medaka. Results: For the preparation of SW-experienced fish, FW medaka were acclimated to SW after pre-acclimation to 1/2 SW, and the SW-acclimated fish were transferred back to FW. The SW-experienced fish and control FW fish (SW-inexperienced fish) were transferred directly to SW. Whereas control FW fish did not survive direct transfer to SW, 1/4 of SW-experienced fish adapted successfully to SW. Although there were no significant differences in blood osmolality and plasma Na+ and Cl- concentrations between SW-experienced and control FW medaka in FW, increments in these parameters following SW transfer were lower in SW-experienced fish than in control FW fish. The gene expression of SW-type Na+, K+-ATPase (NKA) in the gills of SW-experienced medaka increased more quickly after direct SW transfer compared with the expression in control FW fish. Prior to SW transfer, the density of NKA-immunoreactive ionocytes in the gills was higher in SW-experienced fish than in control FW fish. Ionocytes expressing CFTR Cl- channel at the apical membrane and those forming multicellular complexes, both of which were characteristic of SW-type ionocytes, were also increased in SW-experienced fish. Conclusion: These results indicate that past SW experience enhances the capacity of Na+ and Cl- secretion in ionocytes and thus hypoosmoregulatory ability of Japanese medaka, suggesting the presence of epigenetic mechanisms involved in seawater adaptation. [ABSTRACT FROM AUTHOR]

Na+,K+-ATPase (NKA) α-subunit 1a (α1a) and 1b (α1b) gene expressions in the gills are changeable in response to ambient salinity in a few salmonids. In this study, the expressions were compared among ambient salinities and used to infer sea entry migration of chum salmon Oncorhynchus keta fry. The expression of α1a decreased from the 2 days after seawater (SW) transfer from freshwater (FW) and was significantly lower in SW-acclimated fry than that in FW-fry. On the other hand, the expression of α1b peaked on the first to second day after SW transfer and then settled to a level 2-fold higher than in FW-fry. In fry caught in the waterfronts of the beaches, the expression levels were quite similar to those on the first and second days after SW transfer, whereas, in fry caught off beach, the expressions were identical to those of SW-acclimated fry. These suggest that fry adapt to SW with moving along the shoal in the bay, and move to off beach after completing SW adaptation. One of the physiological significances in a usage of waterfront may be to transform the gills to SW type. Only fry on the 2 days after SW transfer failed to exhibit condition factor-dependency of burst swimming, probably due to physiological perturbation, which may be related to poor predation avoidance. The physiological approach used in this study inferred sea entry migration of fry; furthermore, it shows the possible significance of adaptation to SW in the shoal is to reduce predation risk. [Display omitted] • Na+,K+-ATPase (NKA) gene expressions clarify the functional transformation to the seawater-resident type of the gills. • The migratory pathway inferred from physiological insights suggest the physiological and ecological significance. • NKA expressions suggest that fry stay in shallows of waterfront to complete the transformation (physiological significance). • Staying in shallows may reduce predation risk during seawater adaptation (ecological significance). [ABSTRACT FROM AUTHOR]

Guanylin (GN) inhibited water absorption and short-circuit current (Isc) in seawater eel intestine. Similar inhibition was observed after bumetanide, and the effect of bumetanide was abolished by GN or vice versa, suggesting that both act on the same target, Na+-K+-2Cl- cotransporter (NKCC), which is a key player for the Na+-K+-Cl- transport system responsible for water absorption in marine teleost intestine. However, effect of GN was always greater than that of bumetanide: 10% greater in middle intestine (MI) and 40% in posterior intestine (PI) for Isc, and 25% greater in MI and 34% in PI for water absorption. After treatment with GN, Isc decreased to zero, but 20-30% water absorption still remained. The remainder may be due to the Cl-/HCO3- exchanger and Na+-Cl- cotransporter (NCC), since inhibitors for these transporters almost nullified the remaining water absorption. Quantitative PCR analysis revealed the presence of major proteins involved in water absorption; the NKCC2β and AQP1 genes whose expression was markedly upregulated after seawater acclimation. The SLC26A6 (anion exchanger) and NCCβ genes were also expressed in small amounts. Consistent with the inhibitors' effect, expression of NKCC2β was MI > PI, and that of NCCβ was MI << PI. The present study showed that GN not only inhibits the bumetanide-sensitive Na+-K+-Cl- transport system governed by NKCC2β, but also regulates unknown ion transporters different from GN-insensitive SLC26A6 and NCC. A candidate is cystic fibrosis transmembrane conductance regulator Cl- channel, as demonstrated in mammals, but its expression is low in eel intestine, and its role may be minor, as indicated by the small effect of its inhibitors. [ABSTRACT FROM AUTHOR]

This study aimed to test the hypothesis that branchial osmoregulatory acclimation involved increased apoptosis and replacement of mitochdonrion-rich cells (MRCs) in the climbing perch, Anabas testudineus, during a progressive acclimation from freshwater to seawater. A significant increase in branchial caspase-3/-7 activity was observed on day 4 (salinity 20), and an extensive TUNEL-positive apoptosis was detected on day 5 (salinity 25), indicating salinity-induced apoptosis had occurred. This was further supported by an up-regulation of branchial mRNA expression of p53, a key regulator of cell cycle arrest and apoptosis, between day 2 (salinity 10) and day 6 (seawater), and an increase in branchial p53 protein abundance on day 6. Seawater acclimation apparently activated both the extrinsic and intrinsic pathways, as reflected by significant increases in branchial caspase-8 and caspase-9 activities. The involvement of the intrinsic pathway was confirmed by the significant increase in branchial mRNA expression of bax between day 4 (salinity 20) and day 6 (seawater). Western blotting results revealed the presence of a freshwater Na+/K+-ATPase (Nka) α-isoform, Nka α1a, and a seawater isoform, Nka α1b, the protein abundance of which decreased and increased, respectively, during seawater acclimation. Immunofluorescence microscopy revealed the presence of two types of MRCs distinctly different in sizes, and confirmed that the reduction in Nka α1a expression, and the prominent increases in expression of Nka α1b, Na+:K+:2Cl- cotransporter 1, and cystic fibrosis transmembrane conductance regulator Cl- channel coincided with the salinity-induced apoptotic event. Since modulation of existing MRCs alone could not have led to extensive salinity-induced apoptosis, it is probable that some, if not all, freshwater-type MRCs could have been removed through increased apoptosis and subsequently replaced by seawater-type MRCs in the gills of A. testudineus during seawater acclimation. [ABSTRACT FROM AUTHOR]

In previous studies in freshwater tilapia ( Oreochromis mossambicus), dietary supplementation with arachidonic acid (ArA; 20:4 n − 6) had considerable, opposing effects on the main ion-transporting enzyme Na/K-ATPase in gills and kidneys and changed the release of osmoregulatory hormones, such as cortisol. The present study was performed to assess the influence of dietary ArA on (1) the osmoregulatory capacity of tilapia acclimated to seawater (SW) (34‰) and (2) the osmoregulatory imbalance associated with acute stress. The increased ambient salinity was associated with significant alterations in the tissue fatty acid composition, particularly the n − 6 polyunsaturated fatty acids (PUFAs). Tissue levels of ArA were further increased as a result of dietary supplementation, whereas docosahexaenoic acid (DHA, 22:6 n − 3) and eicosapentaenoic acid (EPA, 20:5 n − 3) decreased in gills and kidneys. Basal plasma cortisol as well as lactate levels were elevated in the ArA-supplemented SW-acclimated tilapia compared with the control group. The 5 min of confinement (transient stress) increased plasma cortisol, glucose, and lactate levels with significantly higher levels in ArA-supplemented tilapia. Confinement was also associated with significantly elevated plasma osmolality, sodium, chloride, and potassium levels. ArA-supplemented tilapia showed markedly lower ionic disturbances after confinement, suggesting that dietary ArA can attenuate the hydromineral imbalance associated with acute stress. These results emphasize the involvement of ArA and/or its metabolites in the endocrine and osmoregulatory processes and the response to confinement stress. [ABSTRACT FROM AUTHOR]

This study aimed to clone and sequence the cystic fibrosis transmembrane conductance regulator ( cftr) from, and to determine the effects of seawater acclimation or exposure to 100 mmol l NHCl in freshwater on its mRNA and protein expressions in, the gills of Anabas testudineus. There were 4,530 bp coding for 1,510 amino acids in the cftr cDNA sequence from A. testudineus. The branchial mRNA expression of cftr in fish kept in freshwater was low (<50 copies of transcript per ng cDNA), but significant increases were observed in fish acclimated to seawater for 1 day (92-fold) or 6 days (219-fold). Branchial Cftr expression was detected in fish acclimated to seawater but not in the freshwater control, indicating that Cl excretion through the apical Cftr of the branchial epithelium was essential to seawater acclimation. More importantly, fish exposed to ammonia also exhibited a significant increase (12-fold) in branchial mRNA expression of cftr, with Cftr being expressed in a type of Na/K-ATPase-immunoreactive cells that was apparently different from the type involved in seawater acclimation. It is probable that Cl excretion through Cftr generated a favorable electrical potential across the apical membrane to drive the excretion of $$ {\text{NH}}_{4}^{ + } $$ against a concentration gradient through a yet to be determined transporter, but it led to a slight loss of endogenous Cl. Since ammonia exposure also resulted in significant decreases in blood pH, [HCO] and [total CO] in A. testudineus, it can be deduced that active $$ {\text{NH}}_{4}^{ + } $$ excretion could also be driven by the exit of HCO through the apical Cftr. Furthermore, A. testudineus uniquely responded to ammonia exposure by increasing the ambient pH and decreasing the branchial bafilomycin-sensitive V-type H-ATPase activity, which suggests that its gills might have low NH permeability. [ABSTRACT FROM AUTHOR]

The freshwater climbing perch, Anabas testudineus, is an obligatory air-breathing teleost which can acclimate to seawater, survive long period of emersion, and actively excrete ammonia against high concentrations of environmental ammonia. This study aimed to clone and sequence the Na:K:2Cl cotransporter ( nkcc) from the gills of A. testudineus, and to determine the effects of seawater acclimation or exposure to 100 mmol l NHCl in freshwater on its branchial mRNA expression. The complete coding cDNA sequence of nkcc from the gills of A. testudineus consisted of 3,495 bp, which was translated into a protein with 1,165 amino acid residues and an estimated molecular mass of 127.4 kDa. A phylogenetic analysis revealed that the translated Nkcc of A. testudineus was closer to fish Nkcc1a than to fish Nkcc1b or Nkcc2. After a progressive increase in salinity, there were significant increases in the mRNA expression and protein abundance of nkcc1a in the gills of fish acclimated to seawater as compared with that of the freshwater control. Hence, it can be concluded that similar to marine teleosts, Cl excretion through basolateral Nkcc1 of mitochondrion-rich cells (MRCs) was essential to seawater acclimation in A. testudineus. Exposure of A. testudineus to 100 mmol l NHCl for 1 or 6 days also resulted in significant increases in the mRNA expression of nkcc1a in the gills, indicating a functional role of Nkcc1a in active ammonia excretion. It is probable that NH enter MRCs through basolateral Nkcc1a before being actively transported across the apical membrane. Since the operation of Nkcc1a would lead to an increase in the intracellular Na concentration, it can be deduced that an upregulation of basolateral Na/K-ATPase (Nka) activity would be necessary to compensate for the increased influx of Na into MRCs during active NH excretion. This would imply that the main function of Nka in active NH excretion is to maintain intracellular Na and K homeostasis instead of transporting NH directly into MRCs as proposed previously. In conclusion, active salt secretion during seawater acclimation and active NH excretion during exposure to ammonia in freshwater could involve similar transport mechanisms in the gills of A. testudineus. [ABSTRACT FROM AUTHOR]

Abstract: The Mozambique tilapia (Oreochromis mossambicus) is a euryhaline species that does not survive direct seawater exposure. Cortisol is involved in re-establishing electrolyte homeostasis in seawater and is thought to play a role in allowing tilapia to cope with abrupt seawater exposure, but the mechanism(s) are far from clear. Recently, osmotic stress transcription factor1 (OSTF1) was identified as a key signaling molecule involved in hyperosmotic stress adaptation in tilapia. Consequently, we tested the hypothesis that upregulation of OSTF1 expression by cortisol is a key response for hyperosmotic stress adaptation in tilapia. Fish were exposed to different salinities over a 24h period, while a major electrolyte disturbance and mortality was observed only with full-strength seawater exposure. Therefore, we administered cocoa butter implants of cortisol (50mg/kg) intraperitoneally to tilapia maintained in fresh water and after three days exposed these fish to full-strength seawater. There was 50% mortality in the control fish upon seawater exposure, but this was abolished by cortisol treatment. Abrupt seawater exposure did not affect plasma cortisol levels, while, as expected, exogenous administration of this steroid elevated plasma cortisol levels both in fresh water and seawater. Cortisol treatment significantly induced OSTF1 gene expression in fresh water tilapia, and also enhanced further the seawater-induced OSTF1 mRNA abundance. Plasma osmolality decreased, while gill Na+/K+-ATPase activity was suppressed in the cortisol group in seawater compared to the sham group. This corresponded with a significant reduction in gill ionocyte size and Na+/K+-ATPase activity and protein expression after seawater exposure. Cortisol did not modify liver metabolism, but significantly suppressed gill metabolic capacity in seawater. Overall, cortisol adapts tilapia to a hyperosmotic shock associated with abrupt seawater exposure. This involves upregulation of OSTF1 gene expression and a concomitant suppression of branchial metabolism in tilapia. [Copyright &y& Elsevier]

1. About 25 % of juvenile Atlantic salmon ( Salmo salar) migrating downstream in the River Frome in southern England do so in the autumn rather than in the spring. Here, we examine the physiological status of these fish with regard to those features that adapt them to sea water during the parr–smolt transformation (i.e. gill Na+K+ ATPase activity; the number, size and type of chloride cells on the gill lamellae; salinity tolerance and relative plasma thyroid levels). 2. Autumn migrants, and those fish which subsequently reside in the tidal reaches during the winter, are not sufficiently physiologically adapted to permit permanent or early, entry into the marine environment. 3. It is not known what proportion of autumn migrating fish survive and return to spawn as adults. If significant numbers do return, however, the production from tidal reach habitats must be taken into account in the development of salmon stock management strategies, especially monitoring and assessment programmes, and in the evaluation of factors affecting stocks. [ABSTRACT FROM AUTHOR]

Abstract: Guanylyl cyclase C (GC-C) is a single transmembrane receptor for a family of intestinal hormones, guanylins. In the eel, we previously identified three guanylins, whose gene expression was enhanced in the intestine after transfer from fresh water to seawater. However, only limited information is available about the structure and function of their receptor(s). In the present study, we cloned full-length cDNAs encoding two isoforms of GC-C, named GC-C1 and GC-C2, from eel intestine. The predicted GC-C proteins consisted of extracellular ligand-binding domain, membrane-spanning domain, kinase-like domain and cyclase catalytic domain, in which GC-C-specific sequences were largely conserved. Phylogenetic analyses showed that the cloned membrane GCs are grouped with the GC-C of other vertebrates but not with GC-A and GC-B. However, eel GC-Cs appear to have undergone unique structural evolution compared with other GC-Cs. The three eel guanylins (guanylin, uroguanylin and renoguanylin), but not eel atrial natriuretic peptide, stimulated cGMP production dose-dependently in COS cells expressing either of the cloned cDNAs, providing functional support for assignment as eel guanylin receptors. The potency order for cGMP production was uroguanylin>guanylin⩾renoguanylin for GC-C1; guanylin⩾renoguanylin>uroguanylin for GC-C2. The distinctive ligand selectivity was consistent with the low homology (53%) of the extracellular domain of the two GC-Cs compared with that observed for other domains (74–90%). Both GC-C genes were expressed in the alimentary tract (esophagus, stomach and intestine) and kidney, and their expression was higher in the intestine of seawater-adapted eels than that of freshwater eels just as observed with the guanylin genes. However, the expression of the receptor genes was unchanged for 24h after transfer of eels from fresh water to seawater or vice versa, showing slower response of the receptors to salinity changes than their ligands. Collectively, the multiple guanylin—GC-C system may be involved as a paracrine factor in seawater adaptation at the intestine and kidney of the eel. [Copyright &y& Elsevier]

Abstract: Juvenile steelhead (Oncorhynchus mykiss) acclimated to freshwater (FW) were subjected for 3h to confinement stress in FW, and subsequently saltwater (SW, 25ppt) was introduced to all tanks. Fish were sampled immediately after the stress treatment, and 1, 7, and 14 days after introduction of SW. Electrolytes, cortisol, glucose, and lactate showed the typical stress response expected after stress treatment in FW. Fish regained osmotic balance within 24h. Glucose concentrations were increasing throughout the experiment and lactate levels stayed elevated during the time spent in SW. Insulin-like growth factor-1 (IGF-1) did not show an immediate response to stress but after transfer to SW we detected significantly higher concentrations for control fish at days 1 and 14. The differences in IGF-1 levels between stressed and control fish are not reflected in SW adaptability but positive correlations between IGF-1 and electrolyte levels in control fish may indicate its role for osmoregulation. Confinement stress did not impair feed intake subsequently in SW, but our results suggest that feed intake was suppressed by the change of the media from FW to SW. [Copyright &y& Elsevier]

Abstract: Salinity is a major environmental factor that strongly influences cellular and organismal function. We have used the euryhaline fish Oreochromis mossambicus to identify and annotate immediate hyperosmotic stress responsive molecular mechanisms and biological processes in gill epithelial cells. Using a suppression subtractive hybridization (SSH) approach, we have identified and cloned 20 novel immediate early genes whose mRNAs are induced in gill epithelial cells 4 h after transfer of fish from freshwater (FW) to seawater (SW). Full-length or partial sequences of open reading frames (ORFs) were obtained using the rapid amplification of cDNA ends (RACE) technique. Kinetics of induction was analyzed for all hyperosmotic stress-induced genes. Most genes show a robust transient increase in mRNA abundance characteristic of immediate early stress response genes with peak levels observed between 2 and 8 h after seawater transfer. The newly identified genes were classified according to their sequence similarity with other vertebrate homologs and based on their predicted functions. Pathway analysis revealed that more than half of the identified immediate hyperosmotic stress genes interact closely within a cellular stress response signaling network. Moreover, the 20 genes cluster together in six molecular processes that are rapidly activated in tilapia gills upon salinity transfer. These processes are (1) stress response signal transduction, (2) compatible organic osmolyte accumulation, (3) energy metabolism, (4) lipid transport and cell membrane protection, (5) actin-based cytoskeleton dynamics, and (6) protein and mRNA stability. Our identification and analysis of a set of novel osmo-responsive tilapia genes provides insight into critical physiological processes and pathways constituting the hyperosmotic stress adaptation program in gill epithelial cells of euryhaline fishes. [Copyright &y& Elsevier]

Abstract: Atrial natriuretic peptide (ANP) reduces plasma Na+ concentration and promotes seawater (SW) adaptation in SW eels. However, little is known about the mechanisms for the hyponatremic effect of ANP. In order to evaluate the role of ANP in the whole-body Na+ homeostasis of marine teleost, we reviewed previous in vivo experiments using exogenously administered ANP and present additional experiments to assess the role of endogenous ANP in Na+ homeostasis in conscious SW eels. The Na+ influx and efflux rate across the body surfaces including the gills measured with isotopic 22Na were not altered by the hyponatremic dose (5pmolkg−1 min−1) of ANP infusion in SW eels. ANP infusion also had no effect on renal Na+ excretion in SW eels. In contrast, ANP strongly inhibited drinking, and the inhibition was quantitatively correlated with the hyponatremic effect of ANP. Further, intestinal absorption of Na+ was inhibited by ANP as examined in situ using intestinal sac in conscious SW eels. The combined inhibitory actions of ANP on drinking and intestinal absorption were sufficient to explain the decrease in plasma Na+ concentration. In addition, removal of endogenous circulating ANP by immunoneutralization increased plasma Na+ concentration with a concomitant increase in drinking rate in SW eels. These results strongly suggest that endogenous ANP is involved in the hyponatremic regulation through actions on drinking, and probably on intestine, in SW eels. [Copyright &y& Elsevier]

Abstract: Cortisol plays an important role in controlling intestinal water and ion transport in teleosts possibly through glucocorticoid receptor (GR) and/or mineralocorticoid receptor. To better understand the role of GR in the teleost intestine, in a euryhaline tilapia, Oreochromis mossambicus, we examined (1) the intestinal localizations of GR; (2) the effects of environmental salinity challenge and cortisol treatment on GR mRNA expression. The mRNA abundance of GR in the posterior intestinal region of tilapia was found to be higher than that in the anterior and middle intestine. In the posterior intestine, GR appears to be localized in the mucosal layer. GR mRNA levels in the posterior intestine were elevated after exposure of freshwater fish to seawater for 7 days following an increase in plasma cortisol. Similarly, cortisol implantation in freshwater tilapia for 7 days elevated the intestinal GR mRNA. These results indicate that seawater acclimation is accompanied by upregulation of GR mRNA abundance in intestinal tissue, possibly as a consequence of the elevation of cortisol levels. In contrast, a single intraperitoneal injection of cortisol into freshwater tilapia decreased intestinal GR mRNA. This downregulation of the GR mRNA by cortisol suggests a dual mode of autoregulation of GR expression by cortisol. [Copyright &y& Elsevier]

Abstract: Xenoestrogens such as 4-nonylphenol (4-NP) have been shown to affect the parr–smolt transformation, but their mechanisms of action are not known. We therefore examined effects of 4-NP and estradiol-17β (E2) on expression of estrogen receptor (ER) α gene in the liver, gill, pituitary and brain of sockeye salmon to elucidate molecular mechanisms of 4-NP and E2 and developmental differences in response during smolting. Fish were treated twice within a week with 4-NP (15 and 150 mg/kg BW), E2 (2 mg/kg BW) or only vehicle at three stages of smolting, pre-smolting in March, early smolting in April and late smolting in May. The absolute amounts of ERα mRNA were determined by real-time PCR. The basal amounts of ERα mRNA peaked in April in the liver, gill and pituitary. In March, E2 extensively increased the amounts in the liver, while 4-NP had no effects at this stage. In contrast, 4-NP (but not E2) decreased liver ERα mRNA in April. 4-NP also decreased the amount of ERα mRNA in the gill in April. In the pituitary, 4-NP increased ERα mRNA in March but decreased it in May. There were no significant effects in the brain. Changes in basal ERα mRNA observed in this study indicate that estrogen responsiveness of tissues may change during salmon smolting. Furthermore, 4-NP and E2 have different effects on expression of ERα gene in the liver and gill during smolting, and the response is dependent on smolt stage. [Copyright &y& Elsevier]

Glycine betaine has been suggested to improve the maintenance of ionic and osmotic homeostasis during seawater adaptation in teleost fish. Arsenobetaine may also behave as an osmolyte, due to its structural similarity to glycine betaine. The influence of seawater adaptation on intestinal uptake and muscle accumulation of arsenobetaine in the teleost Atlantic salmon (Salmo salar L.) was investigated. Atlantic salmon (freshwater and seawater adapted) were given a single oral dose of arsenobetaine, which was absorbed over the intestine within 6 h after exposure. Seawater adapted Atlantic salmon had significantly higher levels of accumulated arsenobetaine in blood compared to the freshwater adapted salmon. However, seawater adaptation had no effect on the levels of accumulated arsenobetaine in muscle tissue. Similar retention of the administered dose was found in muscle tissue in both freshwater and seawater adapted salmon, with 49±6% and 50±10% retention after 144 h, respectively. Results indicate that muscle retention was not influenced by salinity in seawater adapting teleosts. [Copyright &y& Elsevier]

Cortisol is an important seawater (SW) osmoregulatory hormone in the Mozambique tilapia (Oreochromis mossambicus), a highly euryhaline cichlid able to live in environments ranging from fresh water (FW) to salinities well in excess of full-strength seawater. Previous studies indicate that cortisol may promote SW adaptation by increasing gill chloride cell differentiation, Na+/K+-ATPase activity and subsequent excretion of excess salt following seawater acclimation. Despite cortisol’s widely accepted role as a SW-adapting hormone, cortisol receptor regulation during SW acclimation is not well understood. The purpose of these studies was to determine whether the intracellular glucocorticoid receptor (GR) might be regulated in a manner consistent with cortisol’s actions in SW adaptation. Saturation radioligand binding assays were conducted on gill cytoplasm preparations from fish sampled 4 and 24 h and 4 and 14 days after transfer from FW to 2/3 SW or FW (control). Affinity (Kd) of the gill GR remained constant over the timecourse, while numbers of receptors (Bmax) in SW fish were significantly elevated compared with controls at 24 h and 4 days after transfer. Plasma osmolality was higher in fish transferred to SW for 24 h, 4 days, and 14 days compared with those animals moved to FW. Plasma cortisol levels and hepatic cortisol binding remained constant between SW and FW fish throughout the timecourse of the salinity challenge. These studies indicate that seawater acclimation is accompanied by a specific upregulation of intracellular GR numbers in gill tissue. The lack of increase in circulating cortisol following SW adaptation may reflect enhancement of clearance of the steroid. It appears that an increase in cortisol receptors, which is closely associated with the rise in blood osmotic pressure that accompanies SW exposure, is an important component of cortisol’s ability to promote SW adaptation in the tilapia. [Copyright &y& Elsevier]

Morphological changes in the chloride cells (CCs) in the yolk-sac membrane of euryhaline tilapia ( Oreochromis mossambicus) embryos and larvae were examined in relation to environmental salinity. Half of a brood of embryos spawned in fresh water (FW) were transferred directly to seawater (SW) 1 day before hatching; the other half was maintained in FW. The embryos and larvae in both FW and SW contained a rich population of CCs in the yolk-sac membrane; the CCs were visualized by whole-mount immunocytochemistry with an antiserum specific for Na+,K+-ATPase. The sectional areas of CCs increased markedly following SW transfer, whereas they remained small in the embryos and larvae maintained in FW. Scanning electron microscopy showed that the apical opening of CCs was enlarged in the fish transferred to SW. Transmission electron microscopy revealed enhanced cellular activity in SW, as evidenced by well-developed mitochondria and tubular systems. The CCs in SW frequently formed a multicellular complex, consisting of a main CC and one or two accessory cells. Accessory cells interdigitated with the main cells and extended their cytoplasmic processes to the apex of the main cell. The three-dimensional arrangement of the cells participating in the complex was identified by confocal laser scanning microscopy. Such complexes were rarely observed in FW fish. The activated CCs in the yolk-sac membrane in the SW fish probably function as ion-extruding sites during embryonic and larval stages until gill CCs become functional. [ABSTRACT FROM AUTHOR]

Thin sections and freeze-fracture replicas have been used to study the structure of the zonulae occludentes of the branchial chloride cells in young adults of the anadromous lamprey Geotria australis, caught during their downstream migration to the sea and after acclimation to full-strength seawater (35‰). The chloride cells in the epithelium of the gill filaments of both freshwater- and seawater-acclimated animals form extensive multicellular complexes. In freshwater animals, the majority of chloride cells (64%) are covered by pavement cells and are thus not exposed to the external environment. Most of the other chloride cells are separated from each other by pavement cells or their processes. The zonulae occludentes between chloride cells and pavement cells and between adjacent chloride cells are extensive and characterised by a network of 4 (range 3-7) superimposed strands. In seawater-acclimated animals, the pavement cells cover only 30% of the chloride cells and their processes no longer occur between chloride cells. Whereas the zonulae occludentes between chloride cells and pavement cells are still extensive, those between chloride cells are shallow and comprise only a single strand or two parallel strands. The zonulae occludentes between the chloride cells of lampreys acclimated to seawater are similar to those in the gills of teleosts in seawater, and are thus considered to be leaky and to provide a low-resistance paracellular pathway for the passive transepithelial movement of Na. [ABSTRACT FROM AUTHOR]

Slices from the kidneys of the rainbow trout which were exposed to 10 or 10 M angiotensin II (AII) and isolated glomeruli exposed to 10 or 10 M AII showed ultrastructural changes compared to control tissues incubated without AII. The studies indicate that angiotensin II has a direct action on glomerular ultrastructure, flattening the epithelial podocytes and broadening the primary processes with fusion of pedicels in extreme cases. These changes suggest a probable effect of AII on water permeability of the trout glomerulus, an intrarenal action which is believed to form an essential part of the antidiuretic adaptation to increased environmental salinities. [ABSTRACT FROM AUTHOR]

EM studies on the coronet cells of the freshwater stickleback showed that the cells were characterised by abundant SER, perinuclear fibrils, microtubules, free ribosomes and mitochondria. Acid phosphatase-positive dense bodies were present, but there was no alkaline phosphatase activity nor were there any glycogen granules in the coronet cells. On the surface of the coronet cell and its globules was a layer of Ruthenium red-stainable material which almost completely disappeared when the fish were kept in 70% seawater for 1 week. Quantitative EM of fish kept in freshwater or 70% seawater for 1 week showed that the SER tubules of seawater-maintained fish had a wider lumen than those of freshwater-maintained fish. The volume occupied by the Golgi apparatus also increased but there was no change in the volume of the cytoplasm occupied by mitochondria. Both the morphology of the coronet cells and their response to 70% seawater support the hypothesis that the saccus vasculosus is an osmo(iono)regulatory structure. [ABSTRACT FROM AUTHOR]

Growth hormone (GH) effectively promotes seawater (SW) adaptation in salmonids, but little is known of its effect in tilapias. Experiments were performed to investigate the effects of recombinant eel GH (reGH) on osmoregulatory actions and ultrastructural features of gill chloride cells in juvenile tilapia, Oreochromis niloticus. Tilapia showed a markedly improved SW survival, when directly transferred from freshwater (FW) to 62.5% SW 24h after a single reGH injection (0.25 or 2.5 µg g-1) or 3 reGH injections (0.25 µg g-1 every other day). Plasma Na+ and Mg2+ levels were significantly reduced by reGH (0.25 and 2.5 µg g-1) compared with saline injections; Ca2+ concentrations were reduced significantly by high dose of reGH (2.5 µg g-1) after SW transfer. However, fish failed to survive more than 24h when directly transferred to 70% SW, although the fish treated with reGH could survive longer than the controls. When examined by electron microscopy, the chloride cells were identified as mitochondrion-rich and an extensive tubular system was induced by GH treatment. The results of the present study suggest that, similar to its effect on salmonids, GH also exerts acute osmoregulatory actions and enhances SW adaptation in juvenile tilapia. GH also stimulates the differentiation of chloride cells toward SW adaptation. [ABSTRACT FROM AUTHOR]

The relative volume of the RPD region of the pituitary gland of Gasterosteus aculeatus form leiurus was greater in animals collected in the spring than in winter. A morphometric analysis of the prolactin cells from spring animals showed only slight changes in cell ultrastructure compared with winter animals. However, in spring the prolactin cells apparently formed and released more secretory granules. Three distinct sites for the release of secretory granules are described. A preliminary study of the response of prolactin cells from spring animals to 70% sea water did not reveal any large scale changes in cell ultrastructure. The significance of this finding is discussed. It is concluded that although the freshwater stickleback cannot be regarded as 'physiologically hypophysectomised:' with regard to prolactin secretion in the winter, it still shows a seasonal change in prolactin cell ultrastructure. [ABSTRACT FROM AUTHOR]

Atlantic salmon may return to the sea after spawning in fresh water. These fish, known as kelts, reportedly show a limited ability to hypoosmoregulate. However, this study shows that fresh-water-adapted kelts exposed to seawater demonstrate rapid adaptation (within 48 h) in osmoregulatory parameters to values characteristic of seawater-adapted salmonids. The urine flow rate falls from 1.2 to 0.2 ml·kg·h within 24 h. Over the same period, urine osmolality increases from 48 mosmol·kg to become isosmotic with the plasma, and Mg secretion by the kidney tubules elevates the urine concentration from 0.5 to 100 mmol·l. As is characteristic for marine teleosts, kelts drink seawater and process the ingested water in the gut to replace body water lost by osmosis to the hyperosmotic medium. Seawater exposure causes a marked hypoxia, arterial oxygen tension falling by 43% within minutes and persisting for at least 4 days at this low level. This is associated with large changes in blood pH and acid-base balance. The physiological mechanisms involved in adaptation to a hyperosmotic external medium are discussed, and the osmoregulatory capacity of kelts is compared with that of salmon at other stages of the life cycle. [ABSTRACT FROM AUTHOR]

To clarify the role of growth hormone (GH) in salmonids during seawater (SW) adaptation, we examined GH kinetics in chronically cannulated rainbow trout, weighing about 1 kg. When trout were transferred from fresh water (FW) to 75% SW, plasma chloride concentration was normalized within 1 week. Plasma GH concentration increased significantly 2 days after transfer and decreased to the initial level thereafter. Metabolic clearance rate (MCR) and secretion rate (SR) of GH were calculated from the plasma levels of GH measured by radioimmunoassay after intra-arterial injection of recombinant chum salmon GH. 4 days after transfer to 75% SW, both MCR and SR increased to levels five times higher than those in FW, and returned to the FW levels after 3-4 weeks. In rainbow trout GH seems to be involved in the development of hypoosmoregulatory mechanisms, especially during the early phase of adaptation. [ABSTRACT FROM AUTHOR]

The epithelial ultrastructure of the glomerulus of fresh- and sea-water adapted rainbow trout, Salmo gairdneri was examined by scanning and transmission electron microscopy after in vivo-perfusion fixation. Adaptation to seawater for a period of up to 9 weeks was not associated with a change in glomerular size, although after one year in seawater both glomeruli and their capsules were significantly smaller than the glomeruli of freshwater adapted fish. In freshwater adapted trout the epithelial ultrastructure conformed to the general vertebrate pattern, with rounded podocytes, well-defined primary processes and interdigitating pedicels. Adaptation to seawater was associated with a marked increase in the frequency of cytoplasmic microprojections. In many glomeruli there was a flattening of the podocytes, broadening of the primary processes, and areas of closely-packed pedicels were common. These observations are discussed in the light of previous studies on the mammalian kidney and the changes of single nephron function known to be associated with adaptation of trout to increased environmental salinities. [ABSTRACT FROM AUTHOR]

In sexually mature male sticklebacks, the renal tubular cells are transformed from ion reabsorbing to mucus secreting cells and in these fish concomitant changes take place in the glomeruli. The present study compares glomerular structure of immature males in fresh water (controls) to those of mature males in fresh water and to immature male sticklebacks in seawater. Glomerular structure is markedly altered in the latter two groups and the changes are similar to a large extent. In these two groups the renal capsules and glomeruli are smaller and the lumina of the glomerular capillaries decrease in diameter, while the number and size of the endothelial fenestrations are reduced. Mesangial cells proliferate and the mesangial matrix greatly expands in both the centrolobular region and the subendothelial space around the capillaries. The secretory activity of the podocytes is enhanced and is responsible for the observed increase in thickness of the outer layer of the basal lamina, the lamina rara externa. The area covered by the filtration slit membranes is reduced, probably as a consequence of fusion of the pedicels of the podocytes. The permeability characteristics of the glomerular filtration barrier for macromolecules, as studied with ferritin injections, remain unaltered. However, the observed differences point to a reduction of the glomerular filtration rate (GFR) during maturation in male sticklebacks, as well as during adaptation of sticklebacks to seawater. This conclusion is in line with physiological evidence. [ABSTRACT FROM AUTHOR]

The esophageal epithelium of the Japanese eel, Anguilla japonica, was studied by light and electron microscopy. In freshwater-adapted eels, longitudinal folds of the mucosal surface are simple in form and lined by a stratified epithelium composed of mucous cells, filament and ribosome-rich cells. Mucous cells are numerous. The filament-rich cells form the outermost and the basal layers of the stratified epithelium and are scattered in the middle zone among the mucous cells. They are firmly bound to one another by many desmosomes and prominent interdigitations of plasma membrane. The distal free surface of the filament-rich cell has a fingerprint-like pattern of microridges. A small number of columnar cells occur at the apices of the folds. They are rich in mitochondria and their distal surfaces bear short microvilli. In seawater-adapted eels, irregularly meandering folds increase the surface area of the mucosa. The stratified epithelium is extensively replaced by a simple columnar epithelium free of mucous cells. The columnar cells resemble in many respects those found in freshwater-adapted eels. They are rich in mitochondria and their distal free surface were provided with short microvilli. However, prominent lateral intercellular spaces and elaborate interdigitations of cytoplasmic processes in the distal zone distinguish the former from the latter. Results are considered in connection with the changes in ion and water permeability of the epithelium after seawater adaptation. [ABSTRACT FROM AUTHOR]

Two types of chloride cells were recognized in the gill epithelium of the fresh-water eel. Following transfer of the eel to sea water the number of strongly acidophilic cells (type A cells) increased markedly during the first week of adaptation. Simultaneously the cell size increased and cell inclusions such as mitochondria and smooth endoplasmic reticulum were also developed. On the other hand, the number of weakly acidophilic cells (type B cells) increased only during the initial 3 days of adaptation and thereafter decreased. This decrease in number of type B cells showed a mirror image of the increase in number of type A cells. Considering the changes in number and fine structures of type B cells, it is likely that the type B cell is a transitional stage of the typical chloride cell (type A cell). When seawater adapted eels were transferred to fresh water, the chloride cell degenerated slowly during 10 weeks. The involvement of the chloride cell in salt secretion of the gill of the seawater eel is discussed. [ABSTRACT FROM AUTHOR]

Smoltification and early seawater phase are critical developmental periods with physiological and biochemical changes in Atlantic salmon that facilitates survival in saltwater. MicroRNAs (miRNAs) are known to have important roles in development, but whether any miRNAs are involved in regulation of gene expression during smoltification and the adaption to seawater is largely unknown. Here, small RNA sequencing of materials from head kidney before, during smoltification and post seawater transfer were used to study expression dynamics of miRNAs, while microarray analysis was applied to study mRNA expression dynamics. Comparing all timepoints, 71 miRNAs and 2709 mRNAs were identified as differentially expressed (DE). Hierarchical clustering analysis of the DE miRNAs showed three major clusters with characteristic expression changes. Eighty-one DE mRNAs revealed negatively correlated expression patterns to DE miRNAs in Cluster I and III. Furthermore, 42 of these mRNAs were predicted as DE miRNA targets. Gene enrichment analysis of negatively correlated target genes showed they were enriched in gene ontology groups hormone biosynthesis, stress management, immune response, and ion transport. The results strongly indicate that post-transcriptional regulation of gene expression by miRNAs is important in smoltification and sea water adaption, and this study identifies several putative miRNA-target pairs for further functional studies. [ABSTRACT FROM AUTHOR]

Most Atlantic salmon ( Salmo salar L.) populations follow an anadromous life cycle, spending early life in freshwater, migrating to the sea for feeding, and returning to rivers to spawn. At the end of the last ice age ~10,000 years ago, several populations of Atlantic salmon became landlocked. Comparing their genomes to their anadromous counterparts can help identify genetic variation related to either freshwater residency or anadromy. The objective of this study was to identify consistently divergent loci between anadromous and landlocked Atlantic salmon strains throughout their geographical distribution, with the long-term aim of identifying traits relevant for salmon aquaculture, including fresh and seawater growth, omega-3 metabolism, smoltification, and disease resistance. We used a Pool-seq approach ( n  = 10-40 individuals per population) to sequence the genomes of twelve anadromous and six landlocked Atlantic salmon populations covering a large part of the Northern Hemisphere and conducted a genomewide association study to identify genomic regions having been under different selection pressure in landlocked and anadromous strains. A total of 28 genomic regions were identified and included cadm1 on Chr 13 and ppargc1a on Chr 18. Seven of the regions additionally displayed consistently reduced heterozygosity in fish obtained from landlocked populations, including the genes gpr132 , cdca4, and sertad2 on Chr 15. We also found 16 regions, including igf1 on Chr 17, which consistently display reduced heterozygosity in the anadromous populations compared to the freshwater populations, indicating relaxed selection on traits associated with anadromy in landlocked salmon. In conclusion, we have identified 37 regions which may harbor genetic variation relevant for improving fish welfare and quality in the salmon farming industry and for understanding life-history traits in fish., Competing Interests: None declared., (© 2020 The Authors. Evolutionary Applications published by John Wiley & Sons Ltd.)

Seawater adaptability of steelhead trout increases along with the increase in the size of the fish, independent of parr-smolt transformation. Three 96 h seawater challenge tests were conducted to determine the size at which seawater adaptability of steelhead trout develops. Plasma Na + and Cl - levels, moisture content, gill Na + /K + ATPase activity, and mortality during the 96 h after direct transfer to seawater (32 ppt) were determined. Plasma Na + and Cl - levels in 50 g fish continuously increased during the 96 h after the transfer to seawater ( p <0.05), but the levels in 100 and 150 g fish leveled off after 24 h ( p <0.05). Both 100 and 150 g size steelhead trout maintained muscle moisture content (%) better than 50 g size fish ( p <0.05). Gill Na + /K + ATPase activity in the 100 g size group increased in a time-dependent manner after transfer to seawater ( p <0.05), whereas activity in the 50 and 150 g sizes did not increase ( p >0.05), for which a possible explanation was discussed. A mere 2.6% mortality in both the 50 and 150 g size groups was observed. In conclusion, the current results indicate that 50 g size steelhead trout did not show development of a high level of hypoosmoregulatory capacity, whereas fish in the 100 and 150 g size groups showed a high level in our experimental conditions. Therefore, the steelhead trout larger than a 100 g size is recommended for transfer to seawater culture., Competing Interests: The authors declare no potential conflict of interest., (© Copyright 2020 The Korean Society of Developmental Biology.)