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Papers by Juan Fuentes

Research paper thumbnail of Increased intestinal carbonate precipitate abundance in the sea bream (Sparus aurata L.) in response to ocean acidification

PLOS ONE, 2019

Marine fish contribute to the carbon cycle by producing mineralized intestinal precipitates gener... more Marine fish contribute to the carbon cycle by producing mineralized intestinal precipitates generated as by-products of their osmoregulation. Here we aimed at characterizing the control of epithelial bicarbonate secretion and intestinal precipitate presence in the gilthead sea bream in response to predicted near future increases of environmental CO 2. Our results demonstrate that hypercapnia (950 and 1800 μatm CO 2) elicits higher intestine epithelial HCO 3 secretion ex vivo and a subsequent parallel increase of intestinal precipitate presence in vivo when compared to present values (440 μatm CO 2). Intestinal gene expression analysis in response to environmental hypercapnia revealed the up-regulation of transporters involved in the intestinal bicarbonate secretion cascade such as the basolateral sodium bicarbonate co-transporter slc4a4, and the apical anion transporters slc26a3 and slc26a6 of sea bream. In addition, other genes involved in intestinal ion uptake linked to water absorption such as the apical nkcc2 and aquaporin 1b expression, indicating that hypercapnia influences different levels of intestinal physiology. Taken together the current results are consistent with an intestinal physiological response leading to higher bicarbonate secretion in the intestine of the sea bream paralleled by increased luminal carbonate precipitate abundance and the main related transporters in response to ocean acidification.

Research paper thumbnail of 海水ティラピア(Oreochromis mossambicus)における腸重炭酸塩分泌の高頻度【Powered by NICT】

Comparative Biochemistry and Physiology A-molecular & Integrative Physiology, 2017

Research paper thumbnail of Intestinal response to ocean acidification in the European sea bass (Dicentrarchus labrax)

Comparative Biochemistry and Physiology Part A: Molecular & Integrative Physiology, 2020

Background: Right hepatectomy (RH) is the most common type of major hepatectomy and can be achiev... more Background: Right hepatectomy (RH) is the most common type of major hepatectomy and can be achieved without portal triad clamping (PTC) in non-cirrhotic liver. The present study reviews our standardized policy of performing RH without systematic PTC. Methods: One hundred and eighty-one consecutive RH were performed in non-cirrhotic patients, with division of the right afferent and efferent blood vessels prior to transection, without systematically using PTC. Prospectively collected data were analysed, focusing on the following endpoints: need for salvage PTC, ischaemic time, blood loss and post-operative outcome. Results: Extra-hepatic division of the right hepatic vessels was feasible in all patients, but was ineffective in 48 patients (26.5%) who required salvage PTC during transection. In those patients, the median ischaemic time was 20 min. The median blood loss was 500 ml (50-3000). Six patients (3.3%) experienced post-operative liver failure. Overall morbidity, severe morbidity and mortality were 42%, 12.1% and 1.6%, respectively, with peri-operative transfusion rate (16.6%) being the only factor associated with morbidity. Discussion: By performing RH with extra-hepatic vascular division prior to transection, PTC can be safely avoided in the majority of patients.

Research paper thumbnail of Control of Calcium Balance in Fish

Fish Osmoregulation, 2007

Research paper thumbnail of Tissue and salinity specific Na+/Cl− cotransporter (NCC) orthologues involved in the adaptive osmoregulation of sea lamprey (Petromyzon marinus)

Scientific Reports, 2021

Two orthologues of the gene encoding the Na+-Cl− cotransporter (NCC), termed ncca and nccb, were ... more Two orthologues of the gene encoding the Na+-Cl− cotransporter (NCC), termed ncca and nccb, were found in the sea lamprey genome. No gene encoding the Na+-K+-2Cl− cotransporter 2 (nkcc2) was identified. In a phylogenetic comparison among other vertebrate NCC and NKCC sequences, the sea lamprey NCCs occupied basal positions within the NCC clades. In freshwater, ncca mRNA was found only in the gill and nccb only in the intestine, whereas both were found in the kidney. Intestinal nccb mRNA levels increased during late metamorphosis coincident with salinity tolerance. Acclimation to seawater increased nccb mRNA levels in the intestine and kidney. Electrophysiological analysis of intestinal tissue ex vivo showed this tissue was anion absorptive. After seawater acclimation, the proximal intestine became less anion absorptive, whereas the distal intestine remained unchanged. Luminal application of indapamide (an NCC inhibitor) resulted in 73% and 30% inhibition of short-circuit current (Is...

Research paper thumbnail of Adenosine receptor blockade and hypoxia-tolerance in rainbow trout and Pacific hagfish. I. Effects on anaerobic metabolism

Journal of Experimental Biology, 1996

The physiological properties of adenosine may be essential in the control of energy metabolism fo... more The physiological properties of adenosine may be essential in the control of energy metabolism for the survival of animals exposed to oxygen shortages. Accordingly, we tested the hypothesis that adenosine modulates metabolic regulation in rainbow trout and Pacific hagfish exposed to acute hypoxia. Treatment of hypoxic rainbow trout (PwO2=3.33 or 4.00 kPa) with the adenosine receptor (AR) blocker theophylline was associated with greater increases in plasma [lactate], more rapid and pronounced metabolic acidosis, higher tissue [lactate], and lower heart creatine charge and glycogen content than in the hypoxic controls. The recruitment of anaerobic metabolism in hypoxic trout treated with enprofylline, an AR blocker with very weak affinity, was intermediate to that of the hypoxic theophylline-injected and control groups. In hagfish, plasma [lactate] increased following exposure to a PwO2 of 1.33 kPa but did not increase following exposure to 3.33 kPa and, like plasma acidosis, it was g...

Research paper thumbnail of Marine fish intestine responds to ocean acidification producing more carbonate aggregates

ABSTRACTMarine fish contribute to the carbon cycle by producing mineralized intestinal aggregates... more ABSTRACTMarine fish contribute to the carbon cycle by producing mineralized intestinal aggregates generated as by-products of their osmoregulation. Here we aimed at characterizing the control of intestinal aggregate production in the gilthead sea bream in response to near future increases of environmental CO2. Our results demonstrate that hypercapnia (800 and 1200 μatm CO2) elicits higher intestine epithelial HCO3−secretion and the subsequent parallel increase of intestinal aggregate production when compared to present values (400 μatm CO2). Intestinal gene expression analysis revealed the up-regulation of crucial transport mechanisms involved not only in the intestinal secretion cascade (Slc4a4, Slc26a3 and Slc26a6) of sea bream, but also in other mechanisms involved in intestinal ion uptake linked to water absorption such asNKCC2and theAquaporin 1b.These results highlight the important role of fish in the marine carbon cycle, and their potential growing impact of intestinal biomin...

Research paper thumbnail of Disruption of gut integrity and permeability contributes to enteritis in a fish-parasite model: a story told from serum metabolomics

Parasites & Vectors, 2019

Background In the animal production sector, enteritis is responsible for serious economic losses,... more Background In the animal production sector, enteritis is responsible for serious economic losses, and intestinal parasitism is a major stress factor leading to malnutrition and lowered performance and animal production efficiency. The effect of enteric parasites on the gut function of teleost fish, which represent the most ancient bony vertebrates, is far from being understood. The intestinal myxozoan parasite Enteromyxum leei dwells between gut epithelial cells and causes severe enteritis in gilthead sea bream (Sparus aurata), anorexia, cachexia, growth impairment, reduced marketability and increased mortality. Methods This study aimed to outline the gut failure in this fish-parasite model using a multifaceted approach and to find and validate non-lethal serum markers of gut barrier dysfunction. Intestinal integrity was studied in parasitized and non-parasitized fish by immunohistochemistry with specific markers for cellular adhesion (E-cadherin) and tight junctions (Tjp1 and Cldn3...

Research paper thumbnail of Author for correspondence

A PTH/PTHrP receptor antagonist blocks the hypercalcemic response to estradiol-17.

Research paper thumbnail of Water calcium concentration modifies whole-body calcium uptake in sea bream larvae during short-term adaptation to altered salinities

Journal of Experimental Biology, 2004

SUMMARYWhole-body calcium uptake was studied in gilthead sea bream larvae(9–83 mg) in response to... more SUMMARYWhole-body calcium uptake was studied in gilthead sea bream larvae(9–83 mg) in response to changing environmental salinity and[Ca2+]. Calcium uptake increased with increased fish size and salinity. Fish exposed to calcium-enriched, diluted seawater showed increased calcium uptake compared with fish in diluted seawater alone. Calcium uptake was unchanged in Na+-enriched, diluted seawater. Overall,[Ca2+], and not salinity/osmolarity per se, appears to be the main factor contributing to calcium uptake. By contrast, drinking was reduced by a decrease in salinity/osmolarity but was little affected by external [Ca2+]. Calculations of the maximum contribution from drinking-associated calcium uptake showed that it became almost insignificant(less than 10%) through a strong decrease in drinking rate at low salinities(0–8‰). Diluted seawater enriched in calcium to the concentration present in full-strength seawater (i.e. constant calcium, decreasing salinity)restored intestinal calcium...

Research paper thumbnail of Expression of pituitary prolactin, growth hormone and somatolactin is modified in response to different stressors (salinity, crowding and food-deprivation) in gilthead sea bream Sparus auratus

General and Comparative Endocrinology, 2009

Prolactin (PRL), growth hormone (GH) and somatolactin (SL) expression was studied in gilthead sea... more Prolactin (PRL), growth hormone (GH) and somatolactin (SL) expression was studied in gilthead sea bream (Sparus auratus) in response to several different stressors (salinity, food deprivation or stocking density). In the first experiment, specimens were acclimated during 100 days at three different environmental salinities: low salinity water (LSW, 6 ppt), brackish water (BW, 12 ppt) and seawater (SW, 38 ppt). Osmoregulatory parameters corresponded to those previously reported for this species under similar osmotic conditions. Pituitary PRL expression increased with decreasing environmental salinity, and was significantly different between SW- and LSW-acclimated fish. Pituitary GH expression was similar between SW- and BW-acclimated fish but decreased in LSW-acclimated specimens. Pituitary SL expression had a "U-shaped" relationship to environmental salinity with the lowest expression in BW-acclimated fish. In a second experiment SW-acclimated specimens were randomly assigned to one of four treatments and maintained for 14 days: (1) fed fish under low density (LD, 4 kg m(-3)); (2) fed fish under high density (HD, 70 kg m(-3)); (3) food deprived fish under LD; and (4) food deprived fish under HD. Plasma glucose and cortisol levels corresponded to those previously reported in S. auratus under similar experimental conditions. Pituitary PRL and SL expression increased in fish maintained under HD and decreased in food deprived fish. In conclusion, an effect of environmental salinity on pituitary PRL and GH expression has been demonstrated. In addition, crowding stress seems to interact with food deprivation in S. auratus and this is reflected by changes in pituitary PRL, GH and SL expression levels.

Research paper thumbnail of 16.P15. Branchial osmoregulatory response to salinity challenge in the Lusitanian toadfish

Comparative Biochemistry and Physiology Part A: Molecular & Integrative Physiology, 2007

concentration in the blood. Seasonality and the changes in the quality of the water can influence... more concentration in the blood. Seasonality and the changes in the quality of the water can influence in the activity of this enzyme which is related with the active or mature branchial chloride cells differentiation (increase in the activity of the enzyme); or still young or in apoptosis and necrosis (reduction of the enzymatic activity). Furthermore, the increase use of pesticides cause general deterioration of the environment and the widespread use of pesticides may affect the natural population of aquatic fauna. The present study was carried out to evaluated the activity of the gill Na + /K +-ATPase in Pimelodus maculatus collected in five upstream sites of Furnas Power Plant, MG, Brazil, in June and December/2006. The gills of the fish were removed and the Na + /K +-ATPase activity determined. The results showed great variability of Na + /K +-ATPase in the fish collected in different sites of the Furnas Lake. These differences may be due to seasonality and possible water quality. Financial support: Furnas Centrais Elétricas S.A., FAPESP, CNPq, CAPES.

Research paper thumbnail of PACAP system evolution and its role in melanophore function in teleost fish skin

Molecular and Cellular Endocrinology, 2015

Pituitary adenylate cyclase-activating polypeptide (PACAP) administered to tilapia melanophores e... more Pituitary adenylate cyclase-activating polypeptide (PACAP) administered to tilapia melanophores ex-vivo causes significant pigment aggregation and this is a newly identified function for this peptide in fish. The G-protein coupled receptors (GPCRs), adcyap1r1a (encoding Pac1a) and vipr2a (encoding Vpac2a), are the only receptors in melanophores with appreciable levels of expression and are significantly (p < 0.05) down-regulated in the absence of light. Vpac2a is activated exclusively by peptide histidine isoleucine (PHI), which suggests that Pac1a mediates the melanin aggregating effect of PACAP on melanophores. Paradoxically activation of Pac1a with PACAP caused a rise in cAMP, which in fish melanophores is associated with melanin dispersion. We hypothesise that the duplicate adcyap1ra and vipr2a genes in teleosts have acquired a specific role in skin and that the melanin aggregating effect of PACAP results from the interaction of Pac1a with Ramp that attenuates cAMP-dependent PKA activity and favours the Ca(2+)/Calmodulin dependent pathway.

Research paper thumbnail of PRL and GH synthesis and release from the sea bream (Sparus auratus L.) pituitary gland in vitro in response to osmotic challenge

General and Comparative Endocrinology, 2010

The endocrine factors prolactin (PRL) and growth hormone (GH) are believed to have counteracting ... more The endocrine factors prolactin (PRL) and growth hormone (GH) are believed to have counteracting effects in the adaption of fish to changes in environmental salinity. In order to further investigate this interaction sea bream were challenged with full seawater (SW) or freshwater (FW) for 7 days and the response of pituitary glands cultured in vitro to an osmotic challenge (230, 275 and 320 mOsm/kg) was assessed. In vitro PRL secretion from pituitaries of SW-adapted fish was unaltered in response to an osmotic challenge, while GH secretion increased in the lowest osmolality (230 mOsm/kg). In contrast, both GH and PRL secretion by pituitaries from FW challenged fish was significantly increased (p<0.01) over that of pituitaries from SW fish at the highest osmolality (320 mOsm/kg). After FW challenge pituitary PRL content and de novo synthesised and released PRL were significantly increased (p<0.01), while total PRL secretion was not different from SW animals. GH pituitary content decreased in FW animals while total secretion and secretion of de novo synthesised protein were significantly increased (p<0.01). In addition, after transfer of fish to FW expression of PRL and GH increased 3- and 2-fold, respectively. Despite the increase in PRL expression, no increase in total PRL secretion occurred and although in gills a 2-fold increase in the osmoregulatory marker, Na(+)/K(+)-ATPase activity was detected, profound haemodilution and a cumulative mortality of 40% occurred in sea bream placed in FW. Taken together the results suggest that the sea bream pituitary gland fails to respond appropriately to the osmotic challenge caused by low salinity and the physiological response evoked in vivo is not enough to allow this species to withstand and adapt to FW.

Research paper thumbnail of Ca2+-Calmodulin regulation of testicular androgen production in Mozambique tilapia (Oreochromis mossambicus)

General and Comparative Endocrinology, 2009

The Ca(2+)-Calmodulin (CaM) signaling pathway has previously been shown to be involved in the reg... more The Ca(2+)-Calmodulin (CaM) signaling pathway has previously been shown to be involved in the regulation of teleost fish ovarian steroidogenesis. However, a putative role of CaM in testicular steroidogenesis and potential targets has not been examined. To examine whether basal steroidogenesis is modulated by Ca(2+) and CaM levels in the testis of Mozambique tilapia (Oreochromis mossambicus) we have incubated testicular fragments in vitro under different conditions and analyzed steroid output. Calcium-free medium with or without EGTA did not affect testicular basal 11-ketotestosterone (11-KT) and testosterone (T) secretion. However, addition of 80microM the CaM inhibitor W7 significantly reduced basal 11-KT, T and androstenedione secretion. Interestingly, the decreased androgen production by 80microM of W7 was accompanied by increased 11-desoxicortisol output and by the activation of cortisol synthesis in the testis, the latter undetected in untreated tissues. However, production of 17,20alpha-dihydroxy-4-pregnen-3-one was unaltered by W7. This suggests that C17,20 desmolase, 21-hydroxylase and possibly 11beta-hydroxysteroid dehydrogenase are targets for CaM. In addition, androgen production was also found to be regulated by the level of cAMP since incubations with forskolin (FK) significantly increased 11-KT and T output. A cross-talk between the cAMP and Ca(2+)-CaM signaling pathways was detected since W7 administration also decreased FK stimulated androgen production. Altogether, these data show that both basal and cAMP stimulated androgen levels were modulated by intracellular Ca(2+)-dependent CaM and that possibly Ca(2+)-CaM determines the shift in steroidogenesis from C21 steroids to androgens.

Research paper thumbnail of Integument structure and function in juvenile Xenopus laevis with disrupted thyroid balance

General and Comparative Endocrinology, 2011

The skin is the largest organ in the body and is a barrier between the internal and external envi... more The skin is the largest organ in the body and is a barrier between the internal and external environment. The present study evaluates how PTU, a goitrogen, that is used to treat hyperthyroidism affects the structure and electrical properties of the frog (Xenopus laevis) skin. The results are considered in the context of the two-membrane model established in the seminal work of Ussing and collegues in the 1940s and 1950s. In vitro experiments with skin from Xenopus adults revealed that PTU can act directly on skin and causes a significant increase (p<0.05, One-way ANOVA) in short circuit current (Isc) via an amiloride-insensitive mechanism. Juvenile Xenopus exposed to waterborne PTU (5 mg/L) had a significantly bigger and more active thyroid gland (p<0.01, Student's t-test) than control Xenopus. The bioelectric properties of skin taken from Xenopus juveniles treated with PTU in vivo had a lower Isc, (3.05±0.4, n=13) and Rt (288.2±39.5) than skin from control Xenopus (Isc, 4.19±1.14, n=14; Rt, 343.3±43.3). A histological assessment of skin from PTU treated Xenopus juveniles revealed the epidermis was significantly thicker (p<0.01, Student's t-test) and had a greater number of modified exocrine glands (p<0.01,…

Research paper thumbnail of DAX1 regulatory networks unveil conserved and potentially new functions

Gene, 2013

DAX1 is an orphan nuclear receptor with actions in mammalian sex determination, regulation of ste... more DAX1 is an orphan nuclear receptor with actions in mammalian sex determination, regulation of steroidogenesis, embryonic development and neural differentiation. Conserved patterns of DAX1 gene expression from mammals to fish have been taken to suggest conserved function. In the present study, the European sea bass, Dicentrarchus labrax, DAX1 promoter was isolated and its conserved features compared to other fish and mammalian DAX1 promoters in order to derive common regulators and functional gene networks. Fish and mammalian DAX1 promoters share common sets of transcription factor frameworks which were also present in the promoter region of another 127 genes. Pathway analysis clustered these into candidate gene networks associated with the fish and mammalian DAX1. The networks identified are concordant with described functions for DAX1 in embryogenesis, regulation of transcription, endocrine development and steroid production. Novel candidate gene network partners were also identified, which implicate DAX1 in ion homeostasis and transport, lipid transport and skeletal development. Experimental evidence is provided supporting roles for DAX1 in steroid signalling and osmoregulation in fish. These results highlight the usefulness of the in silico comparative approach to analyse gene regulation for hypothesis generation. Conserved promoter architecture can be used also to predict potentially new gene functions. The approach reported can be applied to genes from model and non-model species.

Research paper thumbnail of Novel bioactive parathyroid hormone and related peptides in teleost fish

FEBS Letters, 2005

We report the identification, gene expression and biological activity of two parathyroid hormones... more We report the identification, gene expression and biological activity of two parathyroid hormones (PTH; PTHA and PTHB), two PTH-related peptides (PTHrP; PTHrPA and PTHrPB) and a PTH-like ligand (PTH-L) with hybrid characteristics in puffer fishes (Takifugu rubripes and Tetraodon fluviatilis). Experimental data are consistent with PTH-L and PTHrPA having calciotropic activities equivalent, respectively, to tetrapod PTH and PTHrP. We hypothesise on the basis of phylogenetic and functional analysis that PTH-L could be a fish relic of an ancestral PTH/PTHrP gene.

Research paper thumbnail of PTHrP regulates water absorption and aquaporin expression in the intestine of the marine sea bream (Sparus aurata, L.)

General and Comparative Endocrinology, 2015

Water ingestion by drinking is fundamental for ion homeostasis in marine fish. However, the fluid... more Water ingestion by drinking is fundamental for ion homeostasis in marine fish. However, the fluid ingested requires processing to allow net water absorption in the intestine. The formation of luminal carbonate aggregates impacts on calcium homeostasis and requires epithelial HCO 3 À secretion to enable water absorption. In light of its endocrine importance in calcium handling and the indication of involvement in HCO 3 À secretion the present study was designed to expose the role of the parathyroid hormone-related protein (PTHrP) in HCO 3 À secretion, water absorption and the regulation of aqp1 gene expression in the anterior intestine of the sea bream. HCO 3 À secretion rapidly decreased when PTHrP(1-34) was added to anterior intestine of the sea bream mounted in Ussing chambers. The effect achieved a maximum inhibition of 60% of basal secretion rates, showing a threshold effective dose of 0.1 ng ml À1 compatible with reported plasma values of PTHrP. When applied in combination with the adenylate cyclase inhibitor (SQ 22.536, 100 lmol l À1) or the phospholipase C inhibitor (U73122, 10 lmol l À1) the effect of PTHrP(1-34) on HCO 3 À secretion was reduced by about 50% in both cases. In parallel, bulk water absorption measured in intestinal sacs was sensitive to inhibition by PTHrP. The inhibitory action conforms to a typical doseresponse curve in the range of 0.1-1000 ng ml À1 , achieves a maximal effect of 60-65% inhibition from basal rates and shows threshold significant effects at hormone levels of 0.1 ng ml À1. The action of PTHrP in water absorption was completely abolished in the presence of the adenylate cyclase inhibitor (SQ 22.536, 100 lmol l À1) and was insensitive to the phospholipase C inhibitor (U73122, 10 lmol l À1). In vivo injections of PTHrP(1-34) or the PTH/PTHrP receptor antagonist PTHrP(7-34) evoked respectively, a significant decrease or increase of aqp1ab, but not aqp1a. Overall the present results suggest that PTHrP acts as a key regulator of carbonate aggregate formation in the intestine of marine fish via its actions on water absorption, calcium regulation and HCO 3 À secretion.

Research paper thumbnail of Low dietary inclusion of nutraceuticals from microalgae improves feed efficiency and modifies intermediary metabolisms in gilthead sea bream (Sparus aurata)

Scientific Reports

The aim of this work was to evaluate two functional feeds for the gilthead seabream, Sparus aurat... more The aim of this work was to evaluate two functional feeds for the gilthead seabream, Sparus aurata, containing low inclusion of two microalgae-based products (LB-GREENboost, LBGb; and LB-GUThealth, LBGh). Fish (12–13 g) were fed for 13 weeks a control diet or one of the four diets supplemented with both products at 0.5% or 1%. LBGb and LBGh did not affect specific growth rate or survival, but increased feed efficiency by decreasing feed intake and enlarging the intestines. LBGb increased hepatosomatic index and reduced cortisol levels in plasma, while both products lowered plasma lactate. Extensive metabolite and metabolic enzyme profiling revealed that microalgae supplementations, especially 1% LBGh: (i) decrease plasma lactate and increase hepatic glycogen, (ii) reduce hepatic gluconeogenesis, (iii) enhance hepatic lipogenic activity and lipid secretion, (iv) led fish to double triglyceride content in muscle and to stimulate its lipid oxidative capacity, and (v) increase the conte...

Research paper thumbnail of Increased intestinal carbonate precipitate abundance in the sea bream (Sparus aurata L.) in response to ocean acidification

PLOS ONE, 2019

Marine fish contribute to the carbon cycle by producing mineralized intestinal precipitates gener... more Marine fish contribute to the carbon cycle by producing mineralized intestinal precipitates generated as by-products of their osmoregulation. Here we aimed at characterizing the control of epithelial bicarbonate secretion and intestinal precipitate presence in the gilthead sea bream in response to predicted near future increases of environmental CO 2. Our results demonstrate that hypercapnia (950 and 1800 μatm CO 2) elicits higher intestine epithelial HCO 3 secretion ex vivo and a subsequent parallel increase of intestinal precipitate presence in vivo when compared to present values (440 μatm CO 2). Intestinal gene expression analysis in response to environmental hypercapnia revealed the up-regulation of transporters involved in the intestinal bicarbonate secretion cascade such as the basolateral sodium bicarbonate co-transporter slc4a4, and the apical anion transporters slc26a3 and slc26a6 of sea bream. In addition, other genes involved in intestinal ion uptake linked to water absorption such as the apical nkcc2 and aquaporin 1b expression, indicating that hypercapnia influences different levels of intestinal physiology. Taken together the current results are consistent with an intestinal physiological response leading to higher bicarbonate secretion in the intestine of the sea bream paralleled by increased luminal carbonate precipitate abundance and the main related transporters in response to ocean acidification.

Research paper thumbnail of 海水ティラピア(Oreochromis mossambicus)における腸重炭酸塩分泌の高頻度【Powered by NICT】

Comparative Biochemistry and Physiology A-molecular & Integrative Physiology, 2017

Research paper thumbnail of Intestinal response to ocean acidification in the European sea bass (Dicentrarchus labrax)

Comparative Biochemistry and Physiology Part A: Molecular & Integrative Physiology, 2020

Background: Right hepatectomy (RH) is the most common type of major hepatectomy and can be achiev... more Background: Right hepatectomy (RH) is the most common type of major hepatectomy and can be achieved without portal triad clamping (PTC) in non-cirrhotic liver. The present study reviews our standardized policy of performing RH without systematic PTC. Methods: One hundred and eighty-one consecutive RH were performed in non-cirrhotic patients, with division of the right afferent and efferent blood vessels prior to transection, without systematically using PTC. Prospectively collected data were analysed, focusing on the following endpoints: need for salvage PTC, ischaemic time, blood loss and post-operative outcome. Results: Extra-hepatic division of the right hepatic vessels was feasible in all patients, but was ineffective in 48 patients (26.5%) who required salvage PTC during transection. In those patients, the median ischaemic time was 20 min. The median blood loss was 500 ml (50-3000). Six patients (3.3%) experienced post-operative liver failure. Overall morbidity, severe morbidity and mortality were 42%, 12.1% and 1.6%, respectively, with peri-operative transfusion rate (16.6%) being the only factor associated with morbidity. Discussion: By performing RH with extra-hepatic vascular division prior to transection, PTC can be safely avoided in the majority of patients.

Research paper thumbnail of Control of Calcium Balance in Fish

Fish Osmoregulation, 2007

Research paper thumbnail of Tissue and salinity specific Na+/Cl− cotransporter (NCC) orthologues involved in the adaptive osmoregulation of sea lamprey (Petromyzon marinus)

Scientific Reports, 2021

Two orthologues of the gene encoding the Na+-Cl− cotransporter (NCC), termed ncca and nccb, were ... more Two orthologues of the gene encoding the Na+-Cl− cotransporter (NCC), termed ncca and nccb, were found in the sea lamprey genome. No gene encoding the Na+-K+-2Cl− cotransporter 2 (nkcc2) was identified. In a phylogenetic comparison among other vertebrate NCC and NKCC sequences, the sea lamprey NCCs occupied basal positions within the NCC clades. In freshwater, ncca mRNA was found only in the gill and nccb only in the intestine, whereas both were found in the kidney. Intestinal nccb mRNA levels increased during late metamorphosis coincident with salinity tolerance. Acclimation to seawater increased nccb mRNA levels in the intestine and kidney. Electrophysiological analysis of intestinal tissue ex vivo showed this tissue was anion absorptive. After seawater acclimation, the proximal intestine became less anion absorptive, whereas the distal intestine remained unchanged. Luminal application of indapamide (an NCC inhibitor) resulted in 73% and 30% inhibition of short-circuit current (Is...

Research paper thumbnail of Adenosine receptor blockade and hypoxia-tolerance in rainbow trout and Pacific hagfish. I. Effects on anaerobic metabolism

Journal of Experimental Biology, 1996

The physiological properties of adenosine may be essential in the control of energy metabolism fo... more The physiological properties of adenosine may be essential in the control of energy metabolism for the survival of animals exposed to oxygen shortages. Accordingly, we tested the hypothesis that adenosine modulates metabolic regulation in rainbow trout and Pacific hagfish exposed to acute hypoxia. Treatment of hypoxic rainbow trout (PwO2=3.33 or 4.00 kPa) with the adenosine receptor (AR) blocker theophylline was associated with greater increases in plasma [lactate], more rapid and pronounced metabolic acidosis, higher tissue [lactate], and lower heart creatine charge and glycogen content than in the hypoxic controls. The recruitment of anaerobic metabolism in hypoxic trout treated with enprofylline, an AR blocker with very weak affinity, was intermediate to that of the hypoxic theophylline-injected and control groups. In hagfish, plasma [lactate] increased following exposure to a PwO2 of 1.33 kPa but did not increase following exposure to 3.33 kPa and, like plasma acidosis, it was g...

Research paper thumbnail of Marine fish intestine responds to ocean acidification producing more carbonate aggregates

ABSTRACTMarine fish contribute to the carbon cycle by producing mineralized intestinal aggregates... more ABSTRACTMarine fish contribute to the carbon cycle by producing mineralized intestinal aggregates generated as by-products of their osmoregulation. Here we aimed at characterizing the control of intestinal aggregate production in the gilthead sea bream in response to near future increases of environmental CO2. Our results demonstrate that hypercapnia (800 and 1200 μatm CO2) elicits higher intestine epithelial HCO3−secretion and the subsequent parallel increase of intestinal aggregate production when compared to present values (400 μatm CO2). Intestinal gene expression analysis revealed the up-regulation of crucial transport mechanisms involved not only in the intestinal secretion cascade (Slc4a4, Slc26a3 and Slc26a6) of sea bream, but also in other mechanisms involved in intestinal ion uptake linked to water absorption such asNKCC2and theAquaporin 1b.These results highlight the important role of fish in the marine carbon cycle, and their potential growing impact of intestinal biomin...

Research paper thumbnail of Disruption of gut integrity and permeability contributes to enteritis in a fish-parasite model: a story told from serum metabolomics

Parasites & Vectors, 2019

Background In the animal production sector, enteritis is responsible for serious economic losses,... more Background In the animal production sector, enteritis is responsible for serious economic losses, and intestinal parasitism is a major stress factor leading to malnutrition and lowered performance and animal production efficiency. The effect of enteric parasites on the gut function of teleost fish, which represent the most ancient bony vertebrates, is far from being understood. The intestinal myxozoan parasite Enteromyxum leei dwells between gut epithelial cells and causes severe enteritis in gilthead sea bream (Sparus aurata), anorexia, cachexia, growth impairment, reduced marketability and increased mortality. Methods This study aimed to outline the gut failure in this fish-parasite model using a multifaceted approach and to find and validate non-lethal serum markers of gut barrier dysfunction. Intestinal integrity was studied in parasitized and non-parasitized fish by immunohistochemistry with specific markers for cellular adhesion (E-cadherin) and tight junctions (Tjp1 and Cldn3...

Research paper thumbnail of Author for correspondence

A PTH/PTHrP receptor antagonist blocks the hypercalcemic response to estradiol-17.

Research paper thumbnail of Water calcium concentration modifies whole-body calcium uptake in sea bream larvae during short-term adaptation to altered salinities

Journal of Experimental Biology, 2004

SUMMARYWhole-body calcium uptake was studied in gilthead sea bream larvae(9–83 mg) in response to... more SUMMARYWhole-body calcium uptake was studied in gilthead sea bream larvae(9–83 mg) in response to changing environmental salinity and[Ca2+]. Calcium uptake increased with increased fish size and salinity. Fish exposed to calcium-enriched, diluted seawater showed increased calcium uptake compared with fish in diluted seawater alone. Calcium uptake was unchanged in Na+-enriched, diluted seawater. Overall,[Ca2+], and not salinity/osmolarity per se, appears to be the main factor contributing to calcium uptake. By contrast, drinking was reduced by a decrease in salinity/osmolarity but was little affected by external [Ca2+]. Calculations of the maximum contribution from drinking-associated calcium uptake showed that it became almost insignificant(less than 10%) through a strong decrease in drinking rate at low salinities(0–8‰). Diluted seawater enriched in calcium to the concentration present in full-strength seawater (i.e. constant calcium, decreasing salinity)restored intestinal calcium...

Research paper thumbnail of Expression of pituitary prolactin, growth hormone and somatolactin is modified in response to different stressors (salinity, crowding and food-deprivation) in gilthead sea bream Sparus auratus

General and Comparative Endocrinology, 2009

Prolactin (PRL), growth hormone (GH) and somatolactin (SL) expression was studied in gilthead sea... more Prolactin (PRL), growth hormone (GH) and somatolactin (SL) expression was studied in gilthead sea bream (Sparus auratus) in response to several different stressors (salinity, food deprivation or stocking density). In the first experiment, specimens were acclimated during 100 days at three different environmental salinities: low salinity water (LSW, 6 ppt), brackish water (BW, 12 ppt) and seawater (SW, 38 ppt). Osmoregulatory parameters corresponded to those previously reported for this species under similar osmotic conditions. Pituitary PRL expression increased with decreasing environmental salinity, and was significantly different between SW- and LSW-acclimated fish. Pituitary GH expression was similar between SW- and BW-acclimated fish but decreased in LSW-acclimated specimens. Pituitary SL expression had a "U-shaped" relationship to environmental salinity with the lowest expression in BW-acclimated fish. In a second experiment SW-acclimated specimens were randomly assigned to one of four treatments and maintained for 14 days: (1) fed fish under low density (LD, 4 kg m(-3)); (2) fed fish under high density (HD, 70 kg m(-3)); (3) food deprived fish under LD; and (4) food deprived fish under HD. Plasma glucose and cortisol levels corresponded to those previously reported in S. auratus under similar experimental conditions. Pituitary PRL and SL expression increased in fish maintained under HD and decreased in food deprived fish. In conclusion, an effect of environmental salinity on pituitary PRL and GH expression has been demonstrated. In addition, crowding stress seems to interact with food deprivation in S. auratus and this is reflected by changes in pituitary PRL, GH and SL expression levels.

Research paper thumbnail of 16.P15. Branchial osmoregulatory response to salinity challenge in the Lusitanian toadfish

Comparative Biochemistry and Physiology Part A: Molecular & Integrative Physiology, 2007

concentration in the blood. Seasonality and the changes in the quality of the water can influence... more concentration in the blood. Seasonality and the changes in the quality of the water can influence in the activity of this enzyme which is related with the active or mature branchial chloride cells differentiation (increase in the activity of the enzyme); or still young or in apoptosis and necrosis (reduction of the enzymatic activity). Furthermore, the increase use of pesticides cause general deterioration of the environment and the widespread use of pesticides may affect the natural population of aquatic fauna. The present study was carried out to evaluated the activity of the gill Na + /K +-ATPase in Pimelodus maculatus collected in five upstream sites of Furnas Power Plant, MG, Brazil, in June and December/2006. The gills of the fish were removed and the Na + /K +-ATPase activity determined. The results showed great variability of Na + /K +-ATPase in the fish collected in different sites of the Furnas Lake. These differences may be due to seasonality and possible water quality. Financial support: Furnas Centrais Elétricas S.A., FAPESP, CNPq, CAPES.

Research paper thumbnail of PACAP system evolution and its role in melanophore function in teleost fish skin

Molecular and Cellular Endocrinology, 2015

Pituitary adenylate cyclase-activating polypeptide (PACAP) administered to tilapia melanophores e... more Pituitary adenylate cyclase-activating polypeptide (PACAP) administered to tilapia melanophores ex-vivo causes significant pigment aggregation and this is a newly identified function for this peptide in fish. The G-protein coupled receptors (GPCRs), adcyap1r1a (encoding Pac1a) and vipr2a (encoding Vpac2a), are the only receptors in melanophores with appreciable levels of expression and are significantly (p < 0.05) down-regulated in the absence of light. Vpac2a is activated exclusively by peptide histidine isoleucine (PHI), which suggests that Pac1a mediates the melanin aggregating effect of PACAP on melanophores. Paradoxically activation of Pac1a with PACAP caused a rise in cAMP, which in fish melanophores is associated with melanin dispersion. We hypothesise that the duplicate adcyap1ra and vipr2a genes in teleosts have acquired a specific role in skin and that the melanin aggregating effect of PACAP results from the interaction of Pac1a with Ramp that attenuates cAMP-dependent PKA activity and favours the Ca(2+)/Calmodulin dependent pathway.

Research paper thumbnail of PRL and GH synthesis and release from the sea bream (Sparus auratus L.) pituitary gland in vitro in response to osmotic challenge

General and Comparative Endocrinology, 2010

The endocrine factors prolactin (PRL) and growth hormone (GH) are believed to have counteracting ... more The endocrine factors prolactin (PRL) and growth hormone (GH) are believed to have counteracting effects in the adaption of fish to changes in environmental salinity. In order to further investigate this interaction sea bream were challenged with full seawater (SW) or freshwater (FW) for 7 days and the response of pituitary glands cultured in vitro to an osmotic challenge (230, 275 and 320 mOsm/kg) was assessed. In vitro PRL secretion from pituitaries of SW-adapted fish was unaltered in response to an osmotic challenge, while GH secretion increased in the lowest osmolality (230 mOsm/kg). In contrast, both GH and PRL secretion by pituitaries from FW challenged fish was significantly increased (p<0.01) over that of pituitaries from SW fish at the highest osmolality (320 mOsm/kg). After FW challenge pituitary PRL content and de novo synthesised and released PRL were significantly increased (p<0.01), while total PRL secretion was not different from SW animals. GH pituitary content decreased in FW animals while total secretion and secretion of de novo synthesised protein were significantly increased (p<0.01). In addition, after transfer of fish to FW expression of PRL and GH increased 3- and 2-fold, respectively. Despite the increase in PRL expression, no increase in total PRL secretion occurred and although in gills a 2-fold increase in the osmoregulatory marker, Na(+)/K(+)-ATPase activity was detected, profound haemodilution and a cumulative mortality of 40% occurred in sea bream placed in FW. Taken together the results suggest that the sea bream pituitary gland fails to respond appropriately to the osmotic challenge caused by low salinity and the physiological response evoked in vivo is not enough to allow this species to withstand and adapt to FW.

Research paper thumbnail of Ca2+-Calmodulin regulation of testicular androgen production in Mozambique tilapia (Oreochromis mossambicus)

General and Comparative Endocrinology, 2009

The Ca(2+)-Calmodulin (CaM) signaling pathway has previously been shown to be involved in the reg... more The Ca(2+)-Calmodulin (CaM) signaling pathway has previously been shown to be involved in the regulation of teleost fish ovarian steroidogenesis. However, a putative role of CaM in testicular steroidogenesis and potential targets has not been examined. To examine whether basal steroidogenesis is modulated by Ca(2+) and CaM levels in the testis of Mozambique tilapia (Oreochromis mossambicus) we have incubated testicular fragments in vitro under different conditions and analyzed steroid output. Calcium-free medium with or without EGTA did not affect testicular basal 11-ketotestosterone (11-KT) and testosterone (T) secretion. However, addition of 80microM the CaM inhibitor W7 significantly reduced basal 11-KT, T and androstenedione secretion. Interestingly, the decreased androgen production by 80microM of W7 was accompanied by increased 11-desoxicortisol output and by the activation of cortisol synthesis in the testis, the latter undetected in untreated tissues. However, production of 17,20alpha-dihydroxy-4-pregnen-3-one was unaltered by W7. This suggests that C17,20 desmolase, 21-hydroxylase and possibly 11beta-hydroxysteroid dehydrogenase are targets for CaM. In addition, androgen production was also found to be regulated by the level of cAMP since incubations with forskolin (FK) significantly increased 11-KT and T output. A cross-talk between the cAMP and Ca(2+)-CaM signaling pathways was detected since W7 administration also decreased FK stimulated androgen production. Altogether, these data show that both basal and cAMP stimulated androgen levels were modulated by intracellular Ca(2+)-dependent CaM and that possibly Ca(2+)-CaM determines the shift in steroidogenesis from C21 steroids to androgens.

Research paper thumbnail of Integument structure and function in juvenile Xenopus laevis with disrupted thyroid balance

General and Comparative Endocrinology, 2011

The skin is the largest organ in the body and is a barrier between the internal and external envi... more The skin is the largest organ in the body and is a barrier between the internal and external environment. The present study evaluates how PTU, a goitrogen, that is used to treat hyperthyroidism affects the structure and electrical properties of the frog (Xenopus laevis) skin. The results are considered in the context of the two-membrane model established in the seminal work of Ussing and collegues in the 1940s and 1950s. In vitro experiments with skin from Xenopus adults revealed that PTU can act directly on skin and causes a significant increase (p<0.05, One-way ANOVA) in short circuit current (Isc) via an amiloride-insensitive mechanism. Juvenile Xenopus exposed to waterborne PTU (5 mg/L) had a significantly bigger and more active thyroid gland (p<0.01, Student's t-test) than control Xenopus. The bioelectric properties of skin taken from Xenopus juveniles treated with PTU in vivo had a lower Isc, (3.05±0.4, n=13) and Rt (288.2±39.5) than skin from control Xenopus (Isc, 4.19±1.14, n=14; Rt, 343.3±43.3). A histological assessment of skin from PTU treated Xenopus juveniles revealed the epidermis was significantly thicker (p<0.01, Student's t-test) and had a greater number of modified exocrine glands (p<0.01,…

Research paper thumbnail of DAX1 regulatory networks unveil conserved and potentially new functions

Gene, 2013

DAX1 is an orphan nuclear receptor with actions in mammalian sex determination, regulation of ste... more DAX1 is an orphan nuclear receptor with actions in mammalian sex determination, regulation of steroidogenesis, embryonic development and neural differentiation. Conserved patterns of DAX1 gene expression from mammals to fish have been taken to suggest conserved function. In the present study, the European sea bass, Dicentrarchus labrax, DAX1 promoter was isolated and its conserved features compared to other fish and mammalian DAX1 promoters in order to derive common regulators and functional gene networks. Fish and mammalian DAX1 promoters share common sets of transcription factor frameworks which were also present in the promoter region of another 127 genes. Pathway analysis clustered these into candidate gene networks associated with the fish and mammalian DAX1. The networks identified are concordant with described functions for DAX1 in embryogenesis, regulation of transcription, endocrine development and steroid production. Novel candidate gene network partners were also identified, which implicate DAX1 in ion homeostasis and transport, lipid transport and skeletal development. Experimental evidence is provided supporting roles for DAX1 in steroid signalling and osmoregulation in fish. These results highlight the usefulness of the in silico comparative approach to analyse gene regulation for hypothesis generation. Conserved promoter architecture can be used also to predict potentially new gene functions. The approach reported can be applied to genes from model and non-model species.

Research paper thumbnail of Novel bioactive parathyroid hormone and related peptides in teleost fish

FEBS Letters, 2005

We report the identification, gene expression and biological activity of two parathyroid hormones... more We report the identification, gene expression and biological activity of two parathyroid hormones (PTH; PTHA and PTHB), two PTH-related peptides (PTHrP; PTHrPA and PTHrPB) and a PTH-like ligand (PTH-L) with hybrid characteristics in puffer fishes (Takifugu rubripes and Tetraodon fluviatilis). Experimental data are consistent with PTH-L and PTHrPA having calciotropic activities equivalent, respectively, to tetrapod PTH and PTHrP. We hypothesise on the basis of phylogenetic and functional analysis that PTH-L could be a fish relic of an ancestral PTH/PTHrP gene.

Research paper thumbnail of PTHrP regulates water absorption and aquaporin expression in the intestine of the marine sea bream (Sparus aurata, L.)

General and Comparative Endocrinology, 2015

Water ingestion by drinking is fundamental for ion homeostasis in marine fish. However, the fluid... more Water ingestion by drinking is fundamental for ion homeostasis in marine fish. However, the fluid ingested requires processing to allow net water absorption in the intestine. The formation of luminal carbonate aggregates impacts on calcium homeostasis and requires epithelial HCO 3 À secretion to enable water absorption. In light of its endocrine importance in calcium handling and the indication of involvement in HCO 3 À secretion the present study was designed to expose the role of the parathyroid hormone-related protein (PTHrP) in HCO 3 À secretion, water absorption and the regulation of aqp1 gene expression in the anterior intestine of the sea bream. HCO 3 À secretion rapidly decreased when PTHrP(1-34) was added to anterior intestine of the sea bream mounted in Ussing chambers. The effect achieved a maximum inhibition of 60% of basal secretion rates, showing a threshold effective dose of 0.1 ng ml À1 compatible with reported plasma values of PTHrP. When applied in combination with the adenylate cyclase inhibitor (SQ 22.536, 100 lmol l À1) or the phospholipase C inhibitor (U73122, 10 lmol l À1) the effect of PTHrP(1-34) on HCO 3 À secretion was reduced by about 50% in both cases. In parallel, bulk water absorption measured in intestinal sacs was sensitive to inhibition by PTHrP. The inhibitory action conforms to a typical doseresponse curve in the range of 0.1-1000 ng ml À1 , achieves a maximal effect of 60-65% inhibition from basal rates and shows threshold significant effects at hormone levels of 0.1 ng ml À1. The action of PTHrP in water absorption was completely abolished in the presence of the adenylate cyclase inhibitor (SQ 22.536, 100 lmol l À1) and was insensitive to the phospholipase C inhibitor (U73122, 10 lmol l À1). In vivo injections of PTHrP(1-34) or the PTH/PTHrP receptor antagonist PTHrP(7-34) evoked respectively, a significant decrease or increase of aqp1ab, but not aqp1a. Overall the present results suggest that PTHrP acts as a key regulator of carbonate aggregate formation in the intestine of marine fish via its actions on water absorption, calcium regulation and HCO 3 À secretion.

Research paper thumbnail of Low dietary inclusion of nutraceuticals from microalgae improves feed efficiency and modifies intermediary metabolisms in gilthead sea bream (Sparus aurata)

Scientific Reports

The aim of this work was to evaluate two functional feeds for the gilthead seabream, Sparus aurat... more The aim of this work was to evaluate two functional feeds for the gilthead seabream, Sparus aurata, containing low inclusion of two microalgae-based products (LB-GREENboost, LBGb; and LB-GUThealth, LBGh). Fish (12–13 g) were fed for 13 weeks a control diet or one of the four diets supplemented with both products at 0.5% or 1%. LBGb and LBGh did not affect specific growth rate or survival, but increased feed efficiency by decreasing feed intake and enlarging the intestines. LBGb increased hepatosomatic index and reduced cortisol levels in plasma, while both products lowered plasma lactate. Extensive metabolite and metabolic enzyme profiling revealed that microalgae supplementations, especially 1% LBGh: (i) decrease plasma lactate and increase hepatic glycogen, (ii) reduce hepatic gluconeogenesis, (iii) enhance hepatic lipogenic activity and lipid secretion, (iv) led fish to double triglyceride content in muscle and to stimulate its lipid oxidative capacity, and (v) increase the conte...