Jaume Fernández-Borràs

Plasma glucose kinetics and tissue uptake in brown trout in vivo: effect of an intravascular glucose load

The object of the present study was to elucidate whether a glucose load modifies glucose uptake by tissues in brown trout in vivo. By the use of 2-[1,2-3H]-deoxyglucose, plasma glucose disappearance rate and tissue glucose uptake were measured after an intraaortic glucose load of 500 mg·kg-1 (glucose load group) and under normoglycemic conditions (control). We also attempted to determine whether fasting modifies the glucose load disposal (fasted glucose load group). The procedure used to calculate 2-deoxyglucose uptake by tissues was evaluated, and the levels of 2-deoxyglucose uptake were compared with those of 2-deoxyglucose phosphorylation. Uptake and phosphorylation rates were similar in all tissues, except in brain and heart. In all the groups glucose uptake rates were highest in spleen, kidney, brain and gills, and lowest in red muscle, heart and white muscle. However, white muscle was the main site of glucose uptake on a whole tissue basis. The glucose load led to strong, long-lasting hyperglycemia, in spite of the increases observed in plasma insulin levels and in glucose uptake rate by the whole body (control: 4.9 μmol·min-1·kg-1; glucose load group: 6.5 μmol·min-1·kg-1). This higher rate was due to the higher glucose uptake only in white and red muscles (four- and threefold, respectively). Fasting halved the uptake of glucose by both red and white muscles in the load condition. In consequence the use of exogenous glucose decreased with fasting (fasted glucose load group: 5.1 μmol·min-1·kg-1), causing still longer hyperglycemia.

Fate of plasma glucose in tissues of brown trout in vivo: effects of fasting and glucose loading

We report the fate of glucose, both as a source of energy and as a temporary store, in the tissues of brown trout (Salmo trutta) in control, fasted and glucose-loaded fish. Tissue glucose utilization (3H-2-deoxyglucose phosphorylation) and storage (conversion of 14C-glucose into glycogen, protein, and lipid) were measured in immature brown trout, and the oxidation rate was calculated as glucose utilization minus storage and 14C-ionic metabolites remaining in the tissue. The glucose utilization rate is tissue-specific, the highest values being found in spleen, kidney, hindgut, brain, and gill. All these tissues also showed a highly active glycolytic pathway. The lowest utilization indices were observed in white and red muscles, skin, stomach and caeca, which also presented the largest proportion of glucose converted into stores (mainly protein and glycogen). Fasting reduced the glucose disappearance rate by 24%, although there were no significant variations in glucose utilization indices or distribution profile. After a glucose load, plasma glucose and insulin levels rose and the rates of glucose utilization, storage, and oxidation also increased in all tissues (from 1.5- to 4-fold). The relative importance of each tissue in glucose disposal was similar to that in normoglycaemia. In liver, only glucose storage was measured reliably; the conversion of glucose to glycogen was higher than in other tissues, and rose markedly (35-fold) in glucose-loaded fish. In most tissues glucose flux into lipids, glycogen and protein increased. The distribution of glucose may not be a merely substrate-mediated process because fasting in glucose-loaded fish caused lower tissue glucose utilization, particularly in gut, red muscle and gills. Conversion of glucose to tissue stores was reduced, lipids being the most affected.

Low-temperature challenges to gilthead sea bream culture: review of cold-induced alterations and ‘Winter Syndrome’

Although gilthead sea bream have been cultured successfully for the last two decades they are particularly sensitive to low temperature. Especially in the northern Mediterranean area, cold affects fish health and decreases fish-farm production, and may even cause mortality through what is known as ‘Winter Disease’ or ‘Winter Syndrome’. This paper reviews the diagnosis and physiological effects of this disease, focusing on recent studies of cold-induced alterations in gilthead sea bream physiology. ‘Winter Syndrome’ is characterised by multi-organ dysfunction entailing hyposensitivity, erratic swimming, pale and friable livers, necrotic muscles, atrophy of the exocrine pancreas, and distended digestive tract. Its complex aetiology involves several factors such as thermal stress, metabolic depression, immune suppression, and occasional opportunistic pathogens. Low temperatures may be the initial cause of all these factors, except pathogen action. Indoor studies have demonstrated that a drop in temperature causes cold-induced fasting, thermal stress and metabolic depression. These immediate effects are related to an ionic imbalance caused by malfunctions of the gills and digestive system. They are also related to a fatty liver, which appeared steatotic and affected hepatic metabolism and blood composition. The result is a lower immune capacity and fish that are more susceptible to infection. There is no significant thermal compensation under cold conditions and in this situation any additional stress factors can cause fish to suffer metabolic collapse. This study reviews the physiological and zootechnical origins of the disease and, where possible, recommends ways of improving culture conditions during pre-cold, cold and recovery periods.

Tracing metabolic routes of dietary carbohydrate and protein in rainbow trout ( Oncorhynchus mykiss ) using stable isotopes ([ 13 C]starch and [ 15 N]protein): effects of gelatinisation of starches and sustained swimming

Here we examined the use of stable isotopes, [¹³C]starch and [¹⁵N]protein, as dietary tracers to study carbohydrate assimilation and distribution and protein utilisation, respectively, by rainbow trout (Oncorhynchus mykiss). The capacity of glucose uptake and use by tissues was studied, first, by varying the digestibility of carbohydrate-rich diets (30 % carbohydrate), using raw starch and gelatinised starch (GS) and, second, by observing the effects of two regimens of activity (voluntary swimming, control; sustained swimming at 1·3 body lengths/s, exercise) on the GS diet. Isotopic ratio enrichment (¹³C and ¹⁵N) of the various tissue components (protein, lipid and glycogen) was measured in the liver, muscles, viscera and the rest of the fish at 11 and 24 h after a forced meal. A level of 30 % of digestible carbohydrates in the food exceeded the capacity of rainbow trout to use this nutrient, causing long-lasting hyperglycaemia that raises glucose uptake by tissues, and the synthesis of glycogen and lipid in liver. Total 13C recovered 24 h post-feeding in the GS group was lower than at 11 h, indicating a proportional increase in glucose oxidation, although the deposition of lipids in white muscle (WM) increased. Prolonged hyperglycaemia was prevented by exercise, since sustained swimming enhances the use of dietary carbohydrates, mainly through conversion to lipids in liver and oxidation in muscles, especially in red muscle (RM). Higher recoveries of total 15N for exercised fish at 24 h, mainly into the protein fraction of both RM and WM, provide evidence that sustained swimming improves protein deposition, resulting in an enhancement of the protein-sparing effect.

The effects of a temperature rise on oxygen consumption and energy budget in gilthead sea bream

The rates of daily average, minimum and maximum oxygen consumption, specific dynamic action (SDA), ingestion and digestibility in gilthead sea bream, Sparus auratus L., were measured before, during and after a gradual rise of water temperature from 20 to 28 °C at a rate of 1 °C day-1. The relationship between energy gain and the metabolic costs has also been investigated. Oxygen consumption of juvenile sea bream had a clear daily rhythmicity, with low levels during darkness and higher levels during the light period. Both light and feeding processes raised the metabolism of sea bream, but fish fasted for 2 days also showed an increase in oxygen consumption during the light period. Acclimation to 28 °C was complete for the minimum oxygen consumption rate in 12 days, but daily average and maximum oxygen consumption rates on the 19th day after the change were still 33% higher than at 20 °C mainly due to increased SDA. The digestibility of food components did not change significantly throughout the experiment. These relationships are discussed with respect to the potential for growth when fish are reared at 28 °C, although the energy costs are high.

Oxygen consumption and feeding rates of gilthead sea bream (Sparus aurata) reveal lack of acclimation to cold

Low temperature has been implicated in inducing outbreaks of ‘winter syndrome’ or ‘winter disease’ in farmed gilthead sea bream (Sparus aurata). The responses of gilthead sea bream to reduced temperature followed by maintenance at low temperaturewere studied. In a first experiment, oxygen consumptionwas measured when water temperature was reduced from 18°C to 8 °C at either a rate of 1 °C· day-1 or as two ‘sharp drops’ (from 18 °C to 12 °C, and from 12 °C to 8 °C). In a second experiment, the water temperature was reduced from 16 °C to 8 °C or 12 °C and then maintained for 20 days to study the fish acclimation to these temperatures. In both experiments, fish stopped feeding below 13 °C and did not resume feeding when maintained at low temperatures. The decrease in metabolic activity, expressed by the oxygen consumption rate, was directly related to the fall in water temperature: the Q10(18 °C-8 °C) values were between 2.2–2.5, independently of the descend rate in water temperature. However, we observed a more reduced metabolic rate when the water temperature was below 12 °C. Fish maintained at low temperatures showed only a partial recovery in oxygen consumption (15% at 8 °C and 20% at 12 °C) after 20 days. A higher metabolic rate together with a fasting-temperature condition meant that maintenance at 12 °C was more aggressive than at 8 °C, as revealed by the condition factor and energy needs. Data suggest that 12 .C could be a threshold temperature for the metabolic activity of gilthead sea bream. The relationship between low temperatures and their possible implication in the appearance of ‘winter disease’ in gilthead sea bream is also discussed.

New Insights into Fish Swimming: A Proteomic and Isotopic Approach in Gilthead Sea Bream

Moderate exercise enhances fish growth, although underlying physiological mechanisms are not fully known. Here we performed a proteomic and metabolic study in white (WM) and red (RM) muscle of gilthead sea bream juveniles swimming at 1.5 body lengths per second. Continuous swimming for four weeks enhanced fish growth without increasing food intake. Exercise affected muscle energy stores by decreasing lipid and glycogen contents in WM and RM, respectively. Protein synthesis capacity (RNA/protein), energy use (estimated by lipid-δ(13)C and glycogen-δ(13)C), and enzymatic aerobic capacity increased in WM, while protein turnover (expressed by δ(15)N-fractionation) did not change. RM showed no changes in any of these parameters. 2D-PAGE analysis showed that almost 15% of sarcoplasmic protein spots from WM and RM differed in response to exercise, most being over-expressed in WM and under-expressed in RM. Protein identification by MALDI-TOF/TOF-MS and LC-MS/MS revealed exercise-induced enhancement of several pathways in WM (carbohydrate catabolism, protein synthesis, muscle contraction, and detoxification) and under-expression of others in RM (energy production, muscle contraction, and homeostatic processes). The mechanism underpinning the phenotypic response to exercise sheds light on the adaptive processes of fish muscles, being the sustained-moderate swimming induced in gilthead sea bream achieved mainly by WM, thus reducing the work load of RM and improving swimming performance and food conversion efficiency.

Energy reserves and metabolic status affect the acclimation of gilthead sea bream (Sparus aurata) to cold.

During winter, low temperatures induce a direct metabolic depression in gilthead sea bream, without any significant compensatory effect below 13 degrees C. The present study therefore focused on how to improve response to cold in these fish, looking specifically at the two factors of diet (high energy, HiE, and low energy, LoE) and activity (normal, -SW, and sustained activity, +SW) prior to exposure to cold. Following a preparatory period of 75 days water was adjusted to 10 degrees C and kept for 40 days. Enzymatic activities and store deposition revealed that the HiE-SW group had acquired an energy surplus whilst the LoE+SW group exhibited an energy deficit. Liver enzyme activities evidenced diet dependence: LoE groups showed greater glucose-6-phosphate dehydrogenase activity and HiE groups showed greater lipoprotein lipase and hepatic lipase activities. Moreover, the HiE-SW groups lower citrate synthase/cytochrome-c-oxidase ratio reflected the energy surplus available. Perivisceral fat mobilisation caused by cold stress affected liver integrity, resulting in a pre-steatotic condition for the HE-SW group. The differences in liver enzyme activities produced by pre-cold conditions disappeared at low temperatures and enzymatic activities did not compensate. Therefore any improvement that would enable gilthead sea bream to face up to winter must be achieved prior to the appearance of low temperatures.

Naturally Occurring Stable Isotopes Reflect Changes in Protein Turnover and Growth in Gilthead Sea Bream (Sparus aurata) Juveniles under Different Dietary Protein Levels

Ideal nutritional conditions are crucial to sustainable aquaculture due to economic and environmental issues. Here we apply stable isotope analysis as an indicator of fish growth and feeding balance, to define the optimum diet for efficient growing conditions. Juveniles of gilthead sea bream were fed with six isoenergetic diets differing in protein to lipid proportion (from 41/26 to 57/20). As protein intake increased, δ¹⁵N and Δδ¹⁵N of muscle and Δδ¹⁵N and Δδ¹³C of its protein fraction decreased, indicating lower protein turnover and higher protein deposition in muscle. This is reflected in the inverse relationship found between Δδ¹⁵N and growth rate, although no differences were observed in either parameter beyond the protein/lipid proportion 47/23. Principal component analysis (PCA) also signaled 47/23 diet as the pivotal point with the highest growing efficiency, with isotopic parameters having the highest discrimination load. Thus, muscle isotope composition, especially ¹⁵N, can be used to evaluate nutritional status in farmed fish.

Insulin, IGF-I, and muscle MAPK pathway responses after sustained exercise and their contribution to growth and lipid metabolism regulation in gilthead sea bream

Herein, we studied whether sustained exercise positively affects growth of gilthead sea bream by alterations in a) plasma concentrations of insulin and IGF-I, b) signaling pathways in muscle, or c) regulation of lipid metabolism. Specifically, we evaluated the effects of moderated swimming (1.5 body lengths per second; BL/s) on the circulating concentrations of insulin and IGF-I, morphometric parameters, and expression of genes related to lipid metabolism in gilthead sea bream (80-90 g BW). Exercise increased the specific growth rate (P < 0.05) and reduced the hepatosomatic index (P = 0.006). Plasma IGF-I concentrations increased in exercised fish (P = 0.037), suggesting a role for this endocrine factor in the control of muscular growth and metabolic homeostasis during swimming. The observed decrease in plasma insulin concentrations (P = 0.016) could favor the mobilization of tissue reserves in exercised fish. In this sense, the increase in liver fatty acid content (P = 0.041) and the changes in expression of peroxisome proliferator-activated receptors PPARα (P = 0.017) and PPARγ (P = 0.033) indicated a hepatic lipid mobilization. Concentration of glycogen in both white and red muscles was decreased (P = 0.021 and P = 0.017, respectively) in exercised (n = 12) relative to control (n = 12) gilthead sea bream, whereas concentrations of glucose (P = 0.016) and lactate (P = 0.0007) were decreased only in red muscle, indicating the use of these substrates. No changes in the glucose transporter and in lipoprotein lipase mRNA expression were found in any of the tissues studied. Exercised sea bream had decreased content of PPARβ mRNA in white and red muscle relative to control sea bream expression (P = 0.001 and P = 0.049, respectively). Mitogen-activated protein kinase phosphorylation was significantly down-regulated in both white and red muscles of exercised sea bream (P = 0.0374 and P = 0.0371, respectively). Tumor necrosis factor-α expression of white muscle was down-regulated in exercised gilthead sea bream (P = 0.045). Collectively, these results contribute to the knowledge base about hormonal regulation of growth and lipid metabolism in exercised gilthead sea bream.