Energy metabolism of hyperthyroid gilthead sea bream Sparus aurata L.

Thyroid hormones, in particular 3,5,3'-triiodothyronine or T3, are involved in multiple physiological processes in mammals such as protein, fat and carbohydrate metabolism. However, the metabolic actions of T3 in fish are still not fully elucidated. We therefore tested the effects of T3 on Sparus aurata energy metabolism and osmoregulatory system, a hyperthyroid-induced model that was chosen. Fish were implanted with coconut oil depots (containing 0, 2.5, 5.0 and 10.0µg T3/g body weight) and sampled at day 3 and 6 post-implantation. Plasma levels of free T3 as well as glucose, lactate and triglyceride values increased with increasing doses of T3 at days 3 and 6 post-implantation. Changes in plasma and organ metabolite levels (glucose, glycogen, triglycerides, lactate and total α amino acid) and enzyme activities related to carbohydrate, lactate, amino acid and lipid pathways were detected in organs involved in metabolism (liver) and osmoregulation (gills and kidney). Our data implicate that the liver uses amino acids as an energy source in response to the T3 treatment, increasing protein catabolism and gluconeogenic pathways. The gills, the most important extruder of ammonia, are fuelled not only by amino acids, but also by lactate. The kidney differs significantly in its substrate preference from the gills, as it obtained metabolic energy from lactate but also from lipid oxidation processes. We conclude that in S. aurata lipid catabolism and protein turnover are increased as a consequence of experimentally induced hyperthyroidism, with secondary osmoregulatory effects.

Maternal Effects and Trophodynamics Drive Interannual Larval Growth Variability of Atlantic Bluefin Tuna (Thunnus thynnus) from the Gulf of Mexico.

Two cohorts of Atlantic bluefin tuna (Thunnus thynnus) larvae were sampled in 2017 and 2018 during the peak of spawning in the Gulf of Mexico (GOM). We examined environmental variables, daily growth, otolith biometry and stable isotopes and found that the GOM18 cohort grew at faster rates, with larger and wider otoliths. Inter and intra-population analyses (deficient vs. optimal growth groups) were carried out for pre- and post-flexion developmental stages to determine maternal and trophodynamic influences on larval growth variability based on larval isotopic signatures, trophic niche sizes and their overlaps. For the pre-flexion stages in both years, the optimal growth groups had significantly lower δ15N, implying a direct relationship between growth potential and maternal inheritance. Optimal growth groups and stages for both years showed lower C:N ratios, reflecting a greater energy investment in growth. The results of this study illustrate the interannual transgenerational trophic plasticity of a spawning stock and its linkages to growth potential of their offsprings in the GOM.

Evidence of isotopic maternal transmission influence on bluefin tuna (Thunnus thynnus) larval growth.

Pre-flexion stages of Atlantic bluefin tuna (Thunnus thynnus) larvae were collected in 2014 during the peak of spawning in the two main spawning areas: Gulf of Mexico (GOM) and Mediterranean Sea (MED). We examined daily growth, otolith biometry, and stable isotopes and found that the GOM grew at a faster rate, had larger otoliths, wider daily increments, and significantly lower values of δ15N when compared to the MED. In addition, an intra-population comparative analysis between slow- and fast-growing individuals (deficient vs. optimal growth groups, respectively) showed that optimal growth groups had significantly lower δ15N within each spawning area, implying a direct relationship between growth potential, development, and maternal transmission of isotopic signatures. A third pre-flexion larval group that was aquaculture-reared also exhibited the same pattern to the wild larval groups. In addition, for the first time, we estimated the maternal trophic niches using models developed with field-captured pre-flexion larvae. The estimated maternal trophic niches for the GOM were narrower than the MED, implying differences in the maternal trophodynamics from each nursery area. Overall, the inter-population (GOM vs. MED) and intra-population growth groups (deficient vs. optimal) grew faster and had narrower maternal niches. This study shows the advantages that larval SIA research can aid in the understanding of the trophodynamics of their breeders by examining the trophic relationship of a spawning stock jointly with the development of growth potential in offspring within the same breeding season.

Plankton food webs in the oligotrophic Gulf of Mexico spawning grounds of Atlantic bluefin tuna.

We used linear inverse ecosystem modeling techniques to assimilate data from extensive Lagrangian field experiments into a mass-balance constrained food web for the Gulf of Mexico open-ocean ecosystem. This region is highly oligotrophic, yet Atlantic bluefin tuna (ABT) travel long distances from feeding grounds in the North Atlantic to spawn there. Our results show extensive nutrient regeneration fueling primary productivity (mostly by cyanobacteria and other picophytoplankton) in the upper euphotic zone. The food web is dominated by the microbial loop (>70% of net primary productivity is respired by heterotrophic bacteria and protists that feed on them). By contrast, herbivorous food web pathways from phytoplankton to metazoan zooplankton process <10% of the net primary production in the mixed layer. Nevertheless, ABT larvae feed preferentially on podonid cladocerans and other suspension-feeding zooplankton, which in turn derive much of their nutrition from nano- and micro-phytoplankton (mixotrophic flagellates, and to a lesser extent, diatoms). This allows ABT larvae to maintain a comparatively low trophic level (~4.2 for preflexion and postflexion larvae), which increases trophic transfer from phytoplankton to larval fish.

Trade-offs between risks of predation and starvation in larvae make the shelf break an optimal spawning location for Atlantic bluefin tuna.

Atlantic bluefin tuna (ABT) (Thunnus thynnus) travel long distances to spawn in oligotrophic regions of the Gulf of Mexico (GoM) which suggests these regions offer some unique benefit to offspring survival. To better understand how larval survival varies within the GoM a spatially explicit, Lagrangian, individual-based model was developed that simulates dispersal and mortality of ABT early life stages within realistic predator and prey fields during the spawning periods from 1993 to 2012. The model estimates that starvation is the largest cumulative source of mortality associated with an early critical period. However, elevated predation on older larvae is identified as the main factor limiting survival to late postflexion. As a result, first-feeding larvae have higher survival on the shelf where food is abundant, whereas older larvae have higher survival in the open ocean with fewer predators, making the shelf break an optimal spawning area. The modeling framework developed in this study explicitly simulates both physical and biological factors that impact larval survival and hence could be used to support ecosystem based management efforts for ABT under current and future climate conditions.

Genetic connectivity between Atlantic bluefin tuna larvae spawned in the Gulf of Mexico and in the Mediterranean Sea.

The highly migratory Atlantic bluefin tuna (ABFT) is currently managed as two distinct stocks, in accordance with natal homing behavior and population structuring despite the absence of barriers to gene flow. Larval fish are valuable biological material for tuna molecular ecology. However, they have hardly been used to decipher the ABFT population structure, although providing the genetic signal from successful breeders. For the first time, cooperative field collection of tuna larvae during 2014 in the main spawning area for each stock, the Gulf of Mexico (GOM) and the Mediterranean Sea (MED), enabled us to assess the ABFT genetic structure in a precise temporal and spatial frame exclusively through larvae. Partitioning of genetic diversity at nuclear microsatellite loci and in the mitochondrial control region in larvae spawned contemporarily resulted in low significant fixation indices supporting connectivity between spawners in the main reproduction area for each population. No structuring was detected within the GOM after segregating nuclear diversity in larvae spawned in two hydrographically distinct regions, the eastern GOM (eGOM) and the western GOM (wGOM), with the larvae from eGOM being more similar to those collected in the MED than the larvae from wGOM. We performed clustering of genetically characterized ABFT larvae through Bayesian analysis and by Discriminant Analysis of Principal Components (DAPC) supporting the existence of favorable areas for mixing of ABFT spawners from Western and Eastern stocks, leading to gene flow and apparent connectivity between weakly structured populations. Our findings suggest that the eastern GOM is more prone for the mixing of breeders from the two ABFT populations. Conservation of this valuable resource exploited for centuries calls for intensification of tuna ichthyoplankton research and standardization of genetic tools for monitoring population dynamics.

Early life trophodynamic influence on daily growth patterns of the Alboran Sea sardine (Sardina pilchardus) from two distinct nursery habitats (bays of málaga and almería) in the western Mediterranean Sea.

The Alborán Sea sardine (Sardina pilchardus) is the most abundant small pelagic fish in the Mediterranean Sea. Along the northern coast of the Alborán Sea, Málaga and Almería Bay are two important nursery grounds with contrasting oceanographic characteristics. Post-larval stages of S. pilchardus were collected during the late autumn and late winter along with the mesozooplankton community. We examined the influence of feeding behavior on larval growth using environmental variables, daily growth metrics, otolith biometry, stable isotope analysis and trophodynamics. Trophic positions were calculated using mesoozooplankton as potential prey for S. pilchardus. During both seasons, we observed faster growth in larger otoliths that also corresponded to higher trophic positions. Our results indicate that the isotopic signature of larvae is highly influenced by the mesozooplankton community structure. Moreover, the trophic variables showed a significant linear relationship with the estimated ages for all larvae indicating dietary changes throughout ontogenic development.

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.

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.

Trophic ecology of Atlantic Bluefin Tuna (Thunnus thynnus) [corrected] larvae from the Gulf of Mexico and NW Mediterranean spawning grounds: A Comparative Stable Isotope Study.

The present study uses stable isotopes of nitrogen and carbon (δ15Nandδ13C) as trophic indicators for Atlantic bluefin tuna larvae (BFT) (6-10 mm standard length) in the highly contrasting environmental conditions of the Gulf of Mexico (GOM) and the Balearic Sea (MED). These regions are differentiated by their temperature regime and relative productivity, with the GOM being significantly warmer and more productive. MED BFT larvae showed the highest δ15N signatures, implying an elevated trophic position above the underlying microzooplankton baseline. Ontogenetic dietary shifts were observed in the BFT larvae from the GOM and MED which indicates early life trophodynamics differences between these spawning habitats. Significant trophic differences between the GOM and MED larvae were observed in relation to δ15N signatures in favour of the MED larvae, which may have important implications in their growth during their early life stages.These low δ15N levels in the zooplankton from the GOM may be an indication of a shifting isotopic baseline in pelagic food webs due to diatrophic inputs by cyanobacteria. Lack of enrichment for δ15N in BFT larvae compared to zooplankton implies an alternative grazing pathway from the traditional food chain of phytoplankton-zooplankton-larval fish. Results provide insight for a comparative characterization of the trophic pathways variability of the two main spawning grounds for BFT larvae.

17beta-Estradiol affects osmoregulation in Fundulus heteroclitus.

The effect of 17beta-estradiol (E(2)) on osmoregulatory performance was examined in the euryhaline killifish, Fundulus heteroclitus. Fish were injected once with 1, 2 and 5 microg g(-1) E(2) and, 6 h after injection, transferred from 1 ppt seawater (SW) to full strength SW (40 ppt) or from SW to 1 ppt SW. In another set of experiments, fish were injected four times on alternate days with 2 microg g(-1) E(2) and then, 6 h after the last injection, transferred from 1 ppt SW to SW or from SW to 1 ppt SW. Fish were sampled 18 h after transfer (i.e., 24 h post-injection), and plasma osmolality, Na(+) and Cl(-) concentration and gill K(+)-pNPPase activity (a reflection of the sodium pump) were examined. Transfer from 1 ppt SW to SW resulted in significantly increased plasma osmolality, but did not affect gill K(+)-pNPPase activity. A single dose of E(2) (1, 2 and 5 microg g(-1)) prior to transfer from 1 ppt SW to SW increased plasma osmolality and decreased gill K(+)-pNPPase activity in a dose-dependent manner. Prolonged treatment with E(2) increased plasma osmolality and decreased gill K(+)-pNPPase activity in 1 ppt SW-adapted fish. Transfer of fish thus treated from 1 ppt SW to SW increased plasma osmolality and did not alter gill K(+)-pNPPase activity. Transfer from SW to 1 ppt SW had no significant effect on plasma osmolality or gill K(+)-pNPPase activity. Only the highest single dose of E(2) (5 microg g(-1)) prior to transfer from SW to 1 ppt SW decreased gill K(+)-pNPPase activity. Prolonged treatment with 2 microg g(-1) E(2) decreased gill K(+)-pNPPase activity only following transfer from SW to 1 ppt SW. The results substantiate an inhibitory action of E(2) on hypoosmoregulatory capacity in this euryhaline teleost.

Interactive effects of high stocking density and food deprivation on carbohydrate metabolism in several tissues of gilthead sea bream Sparus auratus.

The influence of high stocking density (HSD) and food deprivation was assessed on carbohydrate metabolism of several tissues of gilthead sea bream Sparus auratus for 14 days. Fish were randomly assigned to one of four treatments: (1) fed fish under normal stocking density (NSD) (4 kg m(-3)); (2) fed fish under HSD (70 kg m(-3)); (3) food-deprived fish under NSD; and (4) food-deprived fish under HSD. After 14 days, samples were taken from the plasma, liver, gills, kidney and brain for the assessment of plasma cortisol, levels of metabolites and the activity of several enzymes involved in carbohydrate metabolism. HSD conditions alone elicited important changes in energy metabolism of several tissues that in some cases were confirmatory (5-fold increase in plama cortisol, 20% increase in plasma glucose, 60% decrease in liver glycogen and 20% increase in gluconeogenic potential in the liver) whereas in others provided new information regarding metabolic adjustments to cope with HSD in the liver (100% increase in glucose phosphorylating capacity), gills (30% decrease in capacity for phosphorylating glucose), kidney (80% increase in the capacity of phosphorylating glucose) and brain (2.5-fold increase in ATP levels). On the other hand, food deprivation alone resulted in increased plasma cortisol, and metabolic changes in the liver (enhanced gluconeogenic and glycogenolytic potential of 13% and 18%, respectively) and brain (10% increase in glycolytic capacity), confirmatory of previous studies, whereas new information regarding metabolic adjustments during food deprivation was obtained in the gills and kidney (decreased lactate levels in both tissues of 45% and 55%, respectively). Furthermore, the results obtained provided, for the first time in fish, information indicating that food deprivation increased the sensitivity of gilthead sea bream to the stress induced by HSD compared with the fed controls, as demonstrated by increased plasma cortisol levels (50% increase vs. fed fish) and a further increase in the capacity to export glucose mobilized from liver glycogen stores (70% decrease vs. fed fish). These results lend support for a cumulative effect of both stressors on plasma cortisol and parameters assessed on carbohydrate metabolism in the present experiments, and provide information regarding reallocation of metabolic energy to cope with simultaneous stressors in fish.

Branchial osmoregulatory response to salinity in the gilthead sea bream, Sparus auratus.

The branchial osmoregulatory response of gilthead sea bream (Sparus auratus L.) to short-term (2-192 hr) and long-term (2 weeks) exposure to different environmental salinities (5 per thousand, 15 per thousand, 25 per thousand, 38 per thousand and 60 per thousand) was investigated. A "U-shaped" relationship was observed between environmental salinity and gill Na+,K+ -ATPase activity in both long- and short-term exposure to altered salinity, with the increase in activity occurring between 24 and 96 hr after the onset of exposure. Plasma osmolality and plasma ions (sodium, chloride, calcium and potassium) showed a tendency to increase in parallel with salinity. These variables only differed significantly (P<0.05) in fish adapted to 60 per thousand salinity with respect to fish adapted to full-strength sea-water (SW). Plasma glucose remained unchanged whereas plasma lactate was elevated at 5 per thousand and 60 per thousand. Muscle water content (MWC) was significantly lower in fish adapted to 60 per thousand. Chloride cells (CC) were only present on the surface of the gill filaments and absent from the secondary lamellae. CC distribution was not altered by external salinity. However, the number and size of CC were significantly increased at salinity extremes (5 per thousand and 60 per thousand), whereas fish exposed to intermediate salinities (15 per thousand and 25 per thousand) had fewer and smaller cells. Furthermore, the CC of fish exposed to diluted SW became rounder whereas they were more elongated in fish in full-strength and hypersaline SW. This is consistent with previous reports indicating the existence of two CC types in euryhaline fish. At likely environmental salinities, gilthead sea bream show minor changes in plasma variables and the effective regulation of gill Na+,K+ -ATPase. However, at very low salinities both haemodilution and up-regulation of gill Na+,K+ -ATPase predict a poor adaptation most likely related to deficiency or absence of specific components of the CC important for ion xuptake.

Ontogeny of adenohypophyseal cells in the pituitary of the American shad (Alosa sapidissima).

The distribution and ontogeny of adenohypophyseal cells have been studied in the pituitary gland of embryos, larvae, and juveniles of the clupeid American shad (Alosa sapidissima) using immunocytochemical techniques. In juvenile specimens, adenohypophysis was composed of rostral pars distalis (RPD), formed by cavities lined by prolactin (PRL), adrenocorticotropic hormone (ACTH), and gonadotropic hormone (GTH) cells; proximal pars distalis (PPD), containing growth hormone (GH), GTH, and putative thyroid stimulating hormone (TSH) cells; and pars intermedia (PI) with somatolactin (SL) and melanophore stimulating hormone (MSH) cells. At 3 days post-fertilization (3 days pre-hatching) the pituitary of embryos consisted of an oval mass of cells, close to the ventral margin of the diencephalon, divided in rostral and caudal regions. At this time PRL and ACTH cells appeared in the rostral region of the adenohypophysis, while SL cells were observed in the caudal region where MSH cells showed reactivity 1 day before hatching. At variance, GH cells showed a weak immunoreactivity in the rostral portion at hatching that increased 2 days latter. GTH cells also showed weak immunoreactivity in the rostral region of the adenohypophysis at hatching time. Two days later GTH cells were located in the rostral and central regions of the adenohypophysis. At hatching, the neurohypophysis was very small and no nerve processes were seen to penetrate the adenohypophysis tissue. After hatching, the pituitary gland elongated and in 7 days old larvae, the RPD showed a small lumen surrounded by a palisade of PRL, ACTH, and GHT cells; the PPD showed GH and GTH cells while the PI contained SL and MSH cells. The adenohypophysis and neural lobe increased in size with development and, in 42 days old larvae, they were similar to those of juvenile specimens.

Influence of cortisol on osmoregulation and energy metabolism in gilthead seabream Sparus aurata.

Gilthead seabream Sparus aurata were injected intraperitoneally with slow-release implants of coconut oil alone or containing cortisol (50 and 100 microg x g(-1) body weight), and sampled after two, five, and seven days to assess the simultaneous effects of cortisol on both osmoregulation and energy metabolism. Plasma cortisol levels increased in treated fish to 50-70 ng x ml(-1). An enhanced hypoosmoregulatory capacity of cortisol-implanted fish is suggested by the increase observed in gill Na+, K+-ATPase activity, and the decrease observed in plasma ion concentration (Na+ and Cl-) and osmolality. Cortisol also elicited metabolic changes in liver (increased gluconeogenic potential suggested by elevated FBPase activity, and decreased potential of glycolysis and pentose-phosphate shunt, suggested by the decreased activities of both PK and G6PDH) supporting changes in levels of plasma metabolites suitable for use in other tissues. Thus in this study, we demonstrate for the first time in fish that cortisol treatments elicit changes in the use of exogenous glucose in gills (decreased HK activity) and an increased glycolytic and glycogenic potential in brain (increased GPase, PK and PFK activities).

Osmoregulatory action of 17beta-estradiol in the gilthead sea bream Sparus auratus.

The osmoregulatory action of 17beta-estradiol (E2) was examined in the euryhaline teleost Sparus auratas. In a first set of experiments, fish were injected once with vegetable oil containing E2 (1, 2 and 5 microg/g body weight), transferred 12h after injection from sea water (SW, 38 ppt salinity) to hypersaline water (HSW, 55 ppt) or to brackish water (BW, 5 ppt salinity) and sampled 12h later (i.e. 24 h post-injection). In a second experiment, fish were injected intraperitoneally with coconut oil alone or containing E2 (10 microg/g body weight) and sampled after 5 days. In the same experiment, after 5 days of treatment, fish of each group were transferred to HSW, BW and SW and sampled 4 days later (9 days post-implant). Gill Na+,K+ -ATPase activity, plasma E2 levels, plasma osmolality, and plasma levels of ions (sodium and calcium), glucose, lactate, protein, triglyceride, and hepatosomatic index were examined. Transfer from SW to HSW produced no significant effects on any parameters assessed. E2 treatment did not affect any parameter. Transfer from SW to BW resulted in a significant decrease in plasma osmolality and plasma sodium but did not affect gill Na+,K+ -ATPase activity. A single dose of E2 attenuated the decrease in these parameters after transfer from SW to BW, but was without effect on gill Na+,K+ -ATPase activity. An implant of E2 (10 microg/g body weight) for 5 days significantly increased plasma calcium, hepatosomatic index, plasma metabolic parameters, and gill Na+,K+ -ATPase activity. In coconut oil-implanted (sham) fish, transfer from SW to HSW or BW during 4 days significantly elevated gill Na+,K+ -ATPase. Gill Na+,K+ -ATPase activity remained unaltered after transfer of E2-treated fish to HSW or BW. However, in E2-treated fish transferred from SW to SW (9 days in SW after E2-implant), gill Na+,K+ -ATPase activity decreased with respect to HSW- or BW-transferred fish. Shams transferred to HSW showed increased levels of lactate, protein, and trygliceride in plasma, while those transferred to BW only displayed increased trygliceride levels. E2-treated fish transferred to HSW showed higher protein levels without any change in other plasmatic parameters, while those transferred to BW displayed elevated plasma glucose levels but decreased osmolality and protein levels. These results substantiate a chronic stimulatory action of E2 on gill Na+,K+ -ATPase activity in the euryhaline teleost Sparus auratas.

Acclimation of S aurata to various salinities alters energy metabolism of osmoregulatory and nonosmoregulatory organs.

The impact of different environmental salinities on the energy metabolism of gills, kidney, liver, and brain was assessed in gilthead sea bream (Sparus aurata) acclimated to brackish water [BW, 12 parts/thousand (ppt)], seawater (SW, 38 ppt) and hyper saline water (HSW, 55 ppt) for 14 days. Plasma osmolality and levels of sodium and chloride presented a clear direct relationship with environmental salinities. A general activation of energy metabolism was observed under different osmotic conditions. In liver, an enhancement of glycogenolytic and glycolytic potential was observed in fish acclimated to BW and HSW compared with those in SW. In plasma, an increased availability of glucose, lactate, and protein was observed in parallel with the increase in salinity. In gills, an increased Na+-K+-ATPase activity, a clear decrease in the capacity for use of exogenous glucose and the pentose phosphate pathway, as well as an increased glycolytic potential were observed in parallel with the increased salinity. In kidney, Na+-K+-ATPase activity and lactate levels increased in HSW, whereas the capacity for the use of exogenous glucose decreased in BW- and HSW- acclimated fish compared with SW-acclimated fish. In brain, fish acclimated to BW or HSW displayed an enhancement in their potential for glycogenolysis, use of exogenous glucose, and glycolysis compared with SW-acclimated fish. Also in brain, lactate and ATP levels decreased in parallel with the increase in salinity. The data are discussed in the context of energy expenditure associated with osmotic acclimation to different environmental salinities in fish euryhaline species.

Osmoregulatory action of PRL, GH, and cortisol in the gilthead seabream (Sparus aurata L).

The osmoregulatory actions of ovine prolactin (oPRL), ovine growth hormone (oGH), and cortisol were tested in the euryhaline gilthead seabream Sparus aurata. Acclimated to sea water (SW, 40 ppt salinity, 1000 mOsm/kg H(2)O) or brackish water (BW, 5 ppt, salinity, 130 mOsm/kg H(2)O), injected every other day for one week (number of injections, 4) with saline (0.9% NaCl), oPRL (4 microg/g body weight), oGH (4 microg/g body weight) or cortisol (5 microg/g body weight), and transferred from SW to BW or from BW to SW 24h after the last injection. Fish were sampled before and 24h after transfer. Gill Na(+), K(+)-ATPase activity, plasma osmolality, plasma ions (sodium and chloride), plasma glucose, and muscle water moisture were examined. SW-adapted fish showed higher gill Na(+), K(+)-ATPase activity, plasma osmolality, and plasma ions levels than BW-adapted fish. Transfer from SW to BW decreased plasma osmolality and ions levels after 24h, while transfer from BW to SW increased these parameters, whereas gill Na(+),K(+)-ATPase activity was unaffected. oPRL treatment significantly decreased gill Na(+),K(+)-ATPase activity and increased plasma osmolality and ions in SW- and BW-adapted fish. This treatment minimizes loss of osmolality and ions in plasma after transfer to BW and increased these values after transfer to SW. No significant changes were observed in gill Na(+),K(+)-ATPase activity, plasma osmolality, and plasma ions in oGH-treated group with respect to saline group before or after transfer from SW to BW or from BW to SW. Treatment with cortisol induced, in SW-adapted fish, a significant increase of gill Na(+),K(+)-ATPase activity and decrease of plasma osmolality and plasma ions. In BW-adapted fish this treatment induced a significant increases in gill Na(+),K(+)-ATPase activity, plasma osmolality, and plasma ions. After transfer to SW cortisol-treated fish had higher plasma osmolality than the saline group. Our results support the osmoregulatory role of PRL in the adaptation to hypoosmotic environment in the gilthead seabream S. aurata. Further studies will be necessary to elucidate the osmoregulatory role of GH in this species. Cortisol results suggest a "dual osmoregulatory role" of this hormone in S. aurata.