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- Intestinal response to ocean acidification in the European sea bass (Dicentrarchus labrax)Publication . Alves, Alexandra; Gregorio, Silvia F.; Ruiz-Jarabo, Ignacio; Fuentes, Juanthe intestine of marine fishes contributes to the ocean carbon cycle producing carbonate aggregates as part of the osmoregulatory process. Therefore, this study aimed to evaluate physiological adjustments of European sea bass (Dicentrarchus labrax) intestine to a higher pCO(2) environment likely in the near future (similar to 1700 mu atm). At the whole-body level, hypercapnia for 5 weeks resulted in fish having a significantly diminished specific growth rate, condition factor and hepatosomatic index. An increase in plasma osmolality and HCO3- concentration was detected, paralleled by decreased metabolites concentrations. In the intestine, high seawater pCO(2) was without effect on ouabain-sensitive ATPase activities, while Bafilomycin A1-sensitive ATPase activity significantly decreased in the anterior intestine. Anterior and mid intestine were mounted in Ussing chambers in order to measure bioelectrical parameters and bicarbonate secretion by pH-Stat ex-vivo. Hypercapnia induced a 2.3 and 2.8-fold increase in bicarbonate secretion rates in the anterior and mid intestine, respectively. In the intestinal fluid, HCO3- concentration increased 2.2-fold, and carbonate precipitates showed a 4.4-fold increase in response to hypercapnia, paralleled by a > 3-fold increase of drinking and a > 2-fold increase of intestinal volume at any given time. At the molecular level, hypercapnia elicited higher intestinal mRNA expression levels for atp6v1b (V-ATPase B subunit), slc4a4, slc26a3, and slc26a6, both in the anterior and mid intestine. As a whole, our results show that the intestine of sea bass responds to high seawater pCO(2), a response that comes at a cost at the whole-body level with an impact in the fish specific growth rate, condition factor, and hepatosomatic index.
- Oral cortisol and dexamethasone intake: Differential physiology and transcriptional responses in the marine juvenile Sparus aurataPublication . Barany, A.; Fuentes, Juan; Valderrama, V.; Broz-Ruiz, A.; Martínez-Rodríguez, G.; Mancera, J. M.This study approached the long-term oral administration of cortisol (F) and dexamethasone (DEX), two synthetic glucocorticoids, compared to a control group (CT) in the juveniles of a marine teleost, the gilthead seabream (Sparus aurata). Physiologically, DEX treatment impaired growth, which appears to be linked to carbohydrate allocation in muscle and liver, hepatic triglycerides depletion, and reduced hematocrit. Hypophyseal gh mRNA expression was 2-fold higher in DEX than in CT or F groups. Similarly, hypothalamic trh and hypophyseal pomcb followed this pattern. Plasma cortisol levels were significantly lower in DEX than in CT, while F presented intermediate levels. In the posterior intestine, measured short circuit-current (Isc) was more anion absorptive in CT and F compared to the DEX group, whereas Isc remained unaffected in the anterior intestine. The derived transepithelial electric resistance (TEER) significantly differed between intestinal regions in the DEX group. These results provide new insights to understand better potential targeted biomarkers indicative of the differential glucocorticoid or mineralocorticoid-receptors activation in fish.
- Photoperiod and dietary treatment in freshwater modulate the short-term intestinal response to seawater in atlantic salmon (salmo salar)Publication . Gaetano, Pasqualina; Fernandes Duarte, Vilma Vanessa; Striberny, Anja; Hazlerigg, David; Jørgensen, Even H.; Campinho, Marco António; Fuentes, JuanStimulation and timing of smoltification are essential for successful Atlantic salmon (Salmo salar) aquaculture. This study investigated intestinal responses during dietary and photoperiod manipulation in freshwater (FW) and after a subsequent seven days residence in seawater (SW). "Small" and "large" Atlantic salmon parr (-40 g and -130 g respectively) were treated in FW for 12 weeks and thereafter transferred to SW for seven days. During the FW phase, fish underwent two different light conditions, 24 L:0D - 24 L ("LL-LL" groups) and 7 L:17D - 24 L ("SP-LL" groups) or fed with either regular feed ("LL-LL C" and "SP-LL C" groups) or feed enriched with a salt mix plus free tryptophan ("LL-LL + diet" and "SP-LL + diet" groups). We analyzed Na+/K+-ATPase (NKA) activity, tissue bioelectrical properties in Ussing chambers, and intestinal fluid composition. The NKA activity showed minor variations in relation to fish size, treatments, or intestinal region (anterior or posterior). Photoperiod modulated epithelial bioelectrical properties (Isc and Rt) of the anterior and posterior intestine, particularly transepithelial resistance (Rt). Pharmacological experiments, targeting apical Na+/K+/2Cl- (NKCC2) and Na+/ Cl (NCC) co-transporters revealed intestinal region-and water salinity-dependent effects. In addition, stimu-lation of the intracellular cAMP with forskolin and IBMX showed intestinal region-, water salinity, and treatment-dependence responses with clear functional specialization of the anterior and posterior intestine. The intestinal fluid composition reflected the ability to process ingested SW and showed little variation in large fish. In sum-mary, our data suggest a better pre-adaptation of the intestine during light-stimulated smoltification (SP-LL groups), and the combination of light and diet might give, in an industrial aquaculture setting, an advantage to smaller, but not larger smolts. Intestinal fluid composition in small fish can be used as an index of intestinal function and may act as a long-term performance proxy in SW Atlantic salmon.
- Molecular responses in the intestine of Atlantic salmon (Salmo salar) following light and diet stimulation of smoltification: Potential molecular markers for a seawater-ready smoltPublication . Gaetano, Pasqualina; Fernandes Duarte, Vilma Vanessa; Striberny, Anja; Hazlerigg, David G.; Jørgensen, Even H.; Campinho, Marco António; Fuentes, JuanThe transfer to seawater (SW) represents a critical stage in the production of Atlantic salmon. The success of the transfer links with the optimal development of hypo-osmoregulatory capacities during smoltification. While various strategies are adopted in aquaculture to stimulate smoltification, considerable fish loss still occurs after transfer to sea cages. Therefore, we investigated the molecular responses in the anterior and posterior intestine of Atlantic salmon, following 1) a photoperiod treatment (24 h light (L):0 h dark (D) → 24 L:0D vs. 7 L:17D → 24 L:0D) and 2) dietary treatment (regular feed or feed enriched with a salt mix/tryptophan), combined with, or without a photoperiodic treatment in freshwater (FW), to evaluate how intestinal osmoregulatory mechanisms are modulated by these treatments, and to identify potential intestinal markers indicative of a SW-ready smolt. Using quantitative real-time PCR (qPCR), we investigated transcript levels of transporters and channels involved in ion movements through the enterocytes, tight junction components, and receptors (i.e., calcium-sensing re ceptor and prolactin receptor). The two intestinal regions showed different gene profiles and responsiveness towards the experimental treatments. In the anterior intestine, the exposure to short photoperiod (7 L:17D) upregulated Na+/K+ − ATPase subunit alpha 1c (nkaα1c), Na+/K+/2Cl− cotransporter 1 (nkcc1), Na+/K+/2Cl− cotransporter 2 (nkcc2), Cl− /HCO− 3 exchanger Slc26a6 (slc26a6), and cystic fibrosis transmembrane conductance regulator I (cftrI), in FW and SW. Also, Na+/K+ − ATPase subunit alpha 1b (nkaα1b), occludin (ocln), and prolactin receptor (prlr) were upregulated in FW and claudin 15 (cldn15) in SW groups exposed to this photo period. The posterior intestine was less responsive to the experimental treatments, although upregulation of nkcc1, nkcc2, slc26a6, and cftrI was observed in FW in the short photoperiod groups. Hence, our findings show that exposure to a winter signal in FW more effectively activates hypo-osmoregulatory mechanisms in the in testine of Atlantic salmon, where a coordinated and complementary role of the anterior and posterior intestine ensures optimal SW processing. Dietary treatment had a positive but more marginal effect on the regulation of the genes investigated, mainly enhancing the impact of short photoperiod when the two treatments were combined. Overall, we propose the apical Na+/K+/2Cl− cotransporter, nkcc2, and the apical Cl− / HCO− 3 exchanger, slc26a6, as potential FW molecular markers in the anterior intestine to assess “SW-readiness” in Atlantic salmon smolts.