Aquaporin (AQP) channels in the spiny dogfish, Squalus acanthias I: Characterization of AQP3 and AQP15 function and expression, and localization of the proteins in gill and spiral valve intestine.

Complementary DNAs (cDNAs) for two aquaporin water channel genes (AQP3 and AQP15) were amplified cloned and sequenced to initiate this study. Northern blot analysis was carried out to confirm the mRNA sizes of these AQP genes with AQP3 mRNA bands exhibiting sizes of 1.2 and 1.

Myo-inositol phosphate synthase expression in the European eel (Anguilla anguilla) and Nile tilapia (Oreochromis niloticus): effect of seawater acclimation.

A single MIPS gene (Isyna1/Ino1) exists in eel and tilapia genomes with a single myo-d-inositol 3-phosphate synthase (MIPS) transcript identified in all eel tissues, although two MIPS spliced variants [termed MIPS(s) and MIPS(l)] are found in all tilapia tissues. The larger tilapia transcript [MIPS(l)] results from the inclusion of the 87-nucleotide intron between exons 5 and 6 in the genomic sequence. In most tilapia tissues, the MIPS(s) transcript exhibits much higher abundance (generally >10-fold) with the exception of white skeletal muscle and oocytes, in which the MIPS(l) transcript predominates.

In absence of local adaptation, plasticity and spatially varying selection rule: a view from genomic reaction norms in a panmictic species (Anguilla rostrata).

Background: American eel (Anguilla rostrata) is one of the few species for which panmixia has been demonstrated at the scale of the entire species. As such, the development of long term local adaptation is impossible. However, both plasticity and spatially varying selection have been invoked in explaining how American eel may cope with an unusual broad scope of environmental conditions.

Seawater acclimation and inositol monophosphatase isoform expression in the European eel (Anguilla anguilla) and Nile tilapia (Orechromis niloticus).

Inositol monophosphatase (IMPA) is responsible for the synthesis of inositol, a polyol that can function as an intracellular osmolyte helping re-establish cell volume when exposed to hypertonic environments. Some epithelial tissues in euryhaline teleosts such as the eel and tilapia encounter considerable hyperosmotic challenge when fish move from freshwater (FW) to seawater (SW) environments; however, the roles played by organic osmolytes, such as inositol, have yet to be determined. Syntenic analysis has indicated that, as a result of whole genome- and tandem-duplication events, up to six IMPA isoforms can exist within teleost genomes.

Surviving in a toxic world: transcriptomics and gene expression profiling in response to environmental pollution in the critically endangered European eel.

Background: Genomic and transcriptomic approaches have the potential for unveiling the genome-wide response to environmental perturbations. The abundance of the catadromous European eel (Anguilla anguilla) stock has been declining since the 1980s probably due to a combination of anthropogenic and climatic factors. In this paper, we explore the transcriptomic dynamics between individuals from high (river Tiber, Italy) and low pollution (lake Bolsena, Italy) environments, which were measured for 36 PCBs, several organochlorine pesticides and brominated flame retardants and nine metals.

Cloning, tissue distribution and sub-cellular localisation of phospholipase C X-domain containing protein (PLCXD) isoforms.

Phosphatidylinositol-specific phospholipase C (PI-PLC) enzymes comprise a small family of receptor-regulated phosphodiesterases that control many cellular processes by the regulation of cytosolic calcium and/or the activity of several protein kinases. To date, six distinct classes of PI-PLC are known to exist in mammals. Here we characterise a seventh class of PI-PLC, which contains only the catalytic X domain in its structure, termed phospholipase C X-domain containing protein (PLCXD).

Differential timing of gene expression regulation between leptocephali of the two Anguilla eel species in the Sargasso Sea.

The unique life-history characteristics of North Atlantic catadromous eels have long intrigued evolutionary biologists, especially with respect to mechanisms that could explain their persistence as two ecologically very similar but reproductively and geographically distinct species. Differential developmental schedules during young larval stages have commonly been hypothesized to represent such a key mechanism. We performed a comparative analysis of gene expression by means of microarray experiments with American and European eel leptocephali collected in the Sargasso Sea in order to test the alternative hypotheses of (1) differential timing of gene expression regulation during early development versus (2) species-specific differences in expression of particular genes.

Aquaporin 4 is a Ubiquitously Expressed Isoform in the Dogfish (Squalus acanthias) Shark.

The dogfish ortholog of aquaporin 4 (AQP4) was amplified from cDNA using degenerate PCR followed by cloning and sequencing. The complete coding region was then obtained using 5' and 3' RACE techniques. Alignment of the sequence with AQP4 amino acid sequences from other species showed that dogfish AQP4 has high levels (up to 65.

Expression and Localization of Aquaporin 1a in the Sea-Bass (Dicentrarchus labrax) during Ontogeny.

The successful establishment of a species in a given habitat depends on the ability of each of its developing stages to adapt to the environment. In order to understand this process we have studied the adaptation of a euryhaline fish, the sea-bass Dicentrarchus labrax, to various salinities during its ontogeny. The expression and localization of Aquaporin 1a (AQP1a) mRNA and protein were determined in different osmoregulatory tissues.

Phospholipase C-η2 is activated by elevated intracellular Ca(2+) levels.

Phospholipase C-η2 (PLCη2) is a novel enzyme whose activity in a cellular context is largely uncharacterised. In this study the activity of PLCη2 was examined via [(3)H]inositol phosphate release in COS7 cells expressing the enzyme. PLCη2 activity increased approximately 5-fold in response to monensin, a Na(+)/H(+) antiporter.

A role for inositol monophosphatase 1 (IMPA1) in salinity adaptation in the euryhaline eel (Anguilla anguilla).

This study investigated the expression and tissue distribution of inositol monophosphatase (IMPA1) and characterized its role in salinity adaptation in the eel. The coding sequence of eel IMPA1 was determined and confirmed to be orthologous to the mammalian gene/enzyme by phylogenetic analysis and structural modeling. Quantitative real-time PCR and Western blot techniques indicated up to 17-fold increases in mRNA expression and 2-fold increases in protein abundance in major osmoregulatory tissues following transfer of fish to seawater (SW).

Identification of functionally segregated sarcoplasmic reticulum calcium stores in pulmonary arterial smooth muscle.

In pulmonary arterial smooth muscle, Ca(2+) release from the sarcoplasmic reticulum (SR) via ryanodine receptors (RyRs) may induce constriction and dilation in a manner that is not mutually exclusive. We show here that the targeting of different sarcoplasmic/endoplasmic reticulum Ca(2+)-ATPases (SERCA) and RyR subtypes to discrete SR regions explains this paradox. Western blots identified protein bands for SERCA2a and SERCA2b, whereas immunofluorescence labeling of isolated pulmonary arterial smooth muscle cells revealed striking differences in the spatial distribution of SERCA2a and SERCA2b and RyR1, RyR2, and RyR3, respectively.

Regulation of expression of the myo-inositol monophosphatase 1 gene in osmoregulatory tissues of the European eel Anguilla anguilla after seawater acclimation.

Previous microarray studies in our laboratory identified a number of genes that were differentially expressed in "silver" eels after transfer from freshwater (FW) to seawater (SW). A group of genes, which are related to the synthesis, processing, and transport of certain known osmolytes in mammalian cells, have been identified. One gene implicated with osmolyte production is myo-inositol monophosphatase (IMPA1).

Guanylin-like peptides, guanylate cyclase and osmoregulation in the European eel (Anguilla anguilla).

Three guanylin-like peptides, guanylin, uroguanylin and renoguanylin and two guanylate cyclase type C (GC-C) receptor isoforms were cloned and sequenced from the European eel (Anguilla anguilla). All peptides and both receptors (GC-C1 and GC-C2) were predominantly expressed within the intestine and kidney of both sexually immature yellow, and sexually maturing, migratory silver eels. The derived amino acid sequences for the pre-prohormones and guanylate cyclase isoforms had structural features in common with sequences previously reported for guanylin-like peptides and guanylate cyclases from teleost fish and other species in general.

'Nobody likes a back bore'--exploring lay perspectives of chronic pain: revealing the hidden voices of nonservice users.

Chronic pain is pain that persists beyond 12 weeks or that lasts beyond the expected duration of healing. The chronic pain sufferer also often experiences associated anxiety, depression and stress. An exploratory qualitative approach was adopted by the authors to explore what it is like to live with chronic pain from a sample of people who had not accessed secondary or tertiary health services.

Differential expression of absorptive cation-chloride-cotransporters in the intestinal and renal tissues of the European eel (Anguilla anguilla).

Duplicate pairs of isoforms of each of the NKCC2 and the NCC absorptive cation-chloride-cotransporters have been isolated from the European eel. As with mammalian NKCC2, NKCC2alpha isoform mRNA expression was restricted to renal tissues, whereas NKCC2beta isoform expression was present in intestine and urinary bladder. Similar to mammalian NCC, NCCalpha mRNA expression was also found in the kidney, whereas, expression of NCCbeta mRNA was found at low levels in a number of tissues but particularly in intestine.

Transcriptomic approach to the study of osmoregulation in the European eel Anguilla anguilla.

In euryhaline teleosts, osmoregulation is a fundamental and dynamic process that is essential for the maintenance of ion and water balance, especially when fish migrate between fresh water (FW) and sea water (SW) environments. The European eel has proved to be an excellent model species to study the molecular and physiological adaptations associated with this osmoregulatory plasticity. The life cycle of the European eel includes two migratory periods, the second being the migration of FW eels back to the Sargasso Sea for reproduction.

Aquaporin molecular characterization in the sea-bass (Dicentrarchus labrax): the effect of salinity on AQP1 and AQP3 expression.

Euryhaline fish possess the ability to compensate for environmental salinity changes through hydro-mineral regulation. A number of proteins have been studied in order to understand water and ion exchanges, known as fish osmoregulation. Sea-bass (Dicentrarchus labrax) cDNA sequences encoding a homologue of mammalian aquaporin (termed AQP1) and a homologue of mammalian aquaglyceroporin (termed AQP3) have been isolated and sequenced.

Cortisol regulates eel (Anguilla anguilla) aquaporin 3 (AQP3) mRNA expression levels in gill.

Previous studies in eel (Anguilla anguilla) gill have shown that the expression of the aquaporin 3 (AQP3) water and small solute channel is dramatically decreased (mRNA abundance decreased by up to 97%) when these euryhaline fish are acclimated from freshwater (FW) to seawater (SW). However, AQP3 mRNA expression levels in the intestine following SW-acclimation do not change. The SW-acclimating corticosteroid hormone, cortisol has previously been shown to regulate the expression of aquaporins (particularly AQP1) in eel osmoregulatory tissues in a tissue-specific and isoform-specific fashion.

Salinity adaptation and gene profiling analysis in the European eel (Anguilla anguilla) using microarray technology.

The life cycle of the European eel (Anguilla anguilla) includes two long migratory periods, when the newly hatched leptocephali larvae drift on ocean currents from the Sargasso Sea to the shores of Western Europe and then again up to 30 years later when adult eels swim back to their place of birth for reproductive purposes. Prior to the migration from fresh water (FW) to sea water (SW) adult yellow eels undergo various anatomical and physiological adaptations (silvering) which promote sexual development and aid the transition to increased environmental salinities. The aim of this study was to identify and characterise changes in gene expression within the major osmoregulatory tissues of the eel which enable these fish to make the physiological adaptations required for transfer to SW environments.

The role of aquaporin 3 in teleost fish.

The aquaporin isoform, AQP3 has now been identified in a number of different teleost fish species, with additional DNA sequence information on AQP3 genes in further fish species available in genome databases. In zebrafish (Danio rerio), the AQP3 gene is present as two duplicate isoforms resulting from a teleostean fish genome-wide duplication. A further splicoform/isoform has also been identified in rainbow trout (Oncorhynchus mykiss).

Functional significance of the shark Na,K-ATPase N-terminal domain. Is the structurally variable N-Terminus involved in tissue-specific regulation by FXYD proteins?

The proteolytic profile after mild controlled trypsin cleavage of shark rectal gland Na,K-ATPase was characterized and compared to that of pig kidney Na,K-ATPase, and conditions for achieving N-terminal cleavage of the alpha-subunit at the T(2) trypsin cleavage site were established. Using such conditions, the shark enzyme N-terminus was much more susceptible to proteolysis than the pig enzyme. Nevertheless, the maximum hydrolytic activity was almost unaffected for the shark enzyme, whereas it was significantly decreased for the pig kidney enzyme.

Sequence, circulating levels, and expression of C-type natriuretic peptide in a euryhaline elasmobranch, Carcharhinus leucas.

The present study has examined expression and circulating levels of C-type natriuretic peptide (CNP) in the euryhaline bull shark, Carcharhinus leucas. Complementary DNA and deduced amino acid sequence for CNP in C. leucas were determined by RACE methods.

Effect of cortisol on aquaporin expression in the esophagus of the European eel, Anguilla anguilla.

Long-term cortisol infusion into freshwater (FW)-adapted eels induced a significant increase in aquaporin-1 (AQP1) mRNA expression within the esophageal epithelium of migratory "silver" eels, but not in nonmigratory, immature "yellow" eels. Cortisol treatment had no significant effect on the mRNA abundance of a second aquaporin-1 isoform, termed AQP1dup, which exhibited a highly variable expression profile among individual members of all fish groups. These results suggest that cortisol, at plasma concentrations similar to that found during FW/seawater (SW) acclimation, induces upregulation in AQP1 expression and thus increases esophageal water permeability during the migration of eels to the SW environment.

Cloning and expression of guanylin-like peptides in teleost fish.

Homologues of the guanylin peptide family are expressed in teleost fish. Using the eel as a model euryhaline species, cDNAs for three peptides, guanylin, uroguanylin, and renoguanylin, were cloned and found to be expressed within both renal and intestinal epithelia. Seawater (SW) acclimation resulted in upregulation of uroguanylin mRNA expression in the intestinal epithelia of both immature "yellow" and sexually mature "silver" eels.