Trout Muscle Research Articles

Direct involvement of tumor necrosis factor-α in the regulation of glucose uptake in rainbow trout muscle cells

The proinflammatory cytokine TNF-α is known to have a direct action on skeletal muscle in mammals. However, little is known regarding the potential effects of cytokines on nonimmune tissues, particularly in skeletal muscle, in fish. The aim of this study was to investigate the effects of recombinant trout TNF-α (rtTNF-α) on skeletal muscle carbohydrate metabolism in rainbow trout (Oncorhynchus mykiss). We used a primary cell culture of muscle cells from rainbow trout to show that rtTNF-α stimulates glucose uptake in myoblasts and myotubes at concentrations that do not affect the viability of the cells, requiring de novo protein synthesis as shown by the impairment of rtTNF-α-stimulated glucose uptake by cycloheximide. With the use of specific inhibitors, we show that rtTNF-α-stimulated glucose uptake is mediated by the p38MAPK, NF-κB, and JNK pathways. Additionally, we provide evidence that the stimulatory effects of rtTNF-α on glucose uptake in trout skeletal muscle cells may be caused, at least in part, by an increase in the amount of GLUT4 at the plasma membrane. Incubation of trout muscle cells with conditioned medium from LPS-stimulated trout macrophages, enriched in TNF-α, increased glucose uptake. Our results indicate that recombinant, as well as native trout TNF-α, directly stimulates glucose uptake in trout muscle cells and provide evidence, for the first time in nonmammalian vertebrates, for a potential regulatory role of TNF-α in skeletal muscle metabolism.

Carbohydrate energy reserves and effects of food deprivation in male and female rainbow trout

We investigated the effects of nutritional state on carbohydrate, lipid, and protein stores in the heart, liver, and white skeletal muscle of male and female rainbow trout. For fed animals we also partitioned glycogen into fractions based on acid solubility. Fish (10–14months-old, ~400–500g) were held at 14°C and either fed (1% of body weight, every other day) or deprived of food for 14days. Under fed conditions, glycogen was increased 54% in ventricles from males compared with females, and elevated in the liver (87%) and white muscle (70%) in sexually-maturing versus immature males. Acid soluble glycogen predominated over the acid insoluble fraction in all tissues and was similar between sexes. Food deprivation 1) selectively reduced glycogen and free glucose in male ventricles by ~30%, and 2) did not change glycogen in the liver or white muscle, or triglyceride, protein or water levels in any tissues for both sexes. These data highlight sex differences in teleost cardiac stores and the metabolism of carbohydrates, and contrast with mammals where cardiac glycogen increases during fasting and acid insoluble glycogen is a significant fraction. Increased glycogen in the hearts of male rainbow trout appears to pre-empt sex-specific cardiac growth while storage of acid soluble glycogen may reflect a novel strategy for efficient synthesis and mobilization of glycogen in fishes.

Acute and long-term genotoxicity of deltamethrin to insulin-like growth factors and growth hormone in rainbow trout

We report here the acute and long-term influences of deltamethrin on the expression of IGF-I, IGF-II and GH-I in rainbow trout muscles. We treated rainbow trouts with different concentrations of deltamethrin (0.25 μg/L, 1 μg/L and 2.5 μg/L) and observed the alterations in mRNA expression levels of IGF-I, IGF-II and GH-I at different time intervals (at 6th, 12th, 24th, 48th, 72nd hours and 30th day). The mRNA levels significantly decreased with increasing deltamethrin concentrations for acute administration. Interestingly, a significant recovery in GH-I expression was seen after the 72nd hour up to 30th day while no significant differences were observed for IGF-I and IGF-II between the same time intervals. Here we demonstrate that deltamethrin exposure decreases the expression of IGF-I, IGF-II and GH-I in rainbow trout which might cause undesirable outcomes not only in growth, but also in development and reproduction.

Sequence and expression analysis of two T helper master transcription factors, T-bet and GATA3, in rainbow trout Oncorhynchus mykiss and analysis of their expression during bacterial and parasitic infection

The polarization of naïve CD4+ T cells to T helper (Th)1 or Th2 cells is specified by two master transcription factors, T-bet and GATA3, and is an essential feature of mammalian adaptive immune responses to pathogens and the development of long-lasting immunity. We report here the cloning of rainbow trout Oncorhynchus mykiss T-bet and GATA3, to allow the future evaluation of the existence of Th1 and Th2 cells in salmonid fish. The trout T-bet translation shares high amino acid identities to other fish T-bet molecules (71–72%) but low identities to mammalian T-bet genes (41–42%), although the middle T-box DNA binding domain is highly conserved among all the T-bet proteins from fish and mammals. The trout GATA3 has high amino acid sequence identities (73–88%) to all known vertebrate molecules, with two highly conserved zinc finger motifs. The identity of the trout T-bet and GATA3 molecules was confirmed by phylogenetic tree analysis. A comparable expression level of T-bet and GATA3 was seen in the spleen, head kidney and muscle in healthy trout, but a higher expression level of GATA3 was seen in the gills, brain, skin and intestine relative to that of T-bet. T-bet and GATA3 expression was modulated by different stimulants. The T cell stimulant PHA up-regulated the expression of both T-bet and GATA3 in splenocytes, suggesting that they may be mainly expressed by activated T cells. The expression of T-bet and GATA3 in the spleen was increased by acute stress, but their expression was inhibited by bacterial ( Yersinia ruckeri) infection. In a parasitic infection model, Tetracapsuloides bryosalmonae infection induced a biased gene expression profile where a large increase in the expression of T-bet, IFN-γ and IL-2 was seen, suggesting that a Th1-like response is likely induced by this disease. A better understanding of pathogen modulated expression of T-bet and GATA3, and the potential underlying host immune responses elicited as a consequence of their expression, may allow novel future control measures against disease in fish to be developed.

Biochemical and physiological responses in liver and muscle of rainbow trout after long-term exposure to propiconazole

In this study, the chronic toxic effects of PCZ, a triazole fungicide commonly present in surface and ground water, on morphological indices, ROS generation and RNA/DNA ratio in liver and white muscle of rainbow trout were investigated. Fish were exposed at sublethal concentrations of PCZ (0.2, 50 and 500μgL−1) for 7, 20 and 30d. Compared with the control, there were significant lower CF and HSI in fish exposed at the highest concentration of PCZ. ROS levels in both tissues increased significantly at higher PCZ concentrations (50 and 500μgL−1) after 20d and above, as well as in muscle of fish exposed at lowest PCZ concentration (0.2μgL−1) after 30d. The hepatic antioxidant enzymes (SOD and CAT) activities were induced significantly at higher concentrations (50, 500μgL−1) of PCZ after 20d and at 50μgL−1 after 30d. Additional, hepatic SOD activity was significantly induced at 0.2μgL−1 after 30d. Compared with the hepatic antioxidant enzymes activities in fish exposed to 50μgL−1 of PCZ, there was a decreasing trend in those exposed to 500μgL−1 after 30d exposure. However, both the antioxidant enzymes activities were significantly inhibited in muscle of fish exposed to 500μgL−1 PCZ after 30d. Moreover there was significant lower RNA/DNA ratio in both tissues after long-term exposure to higher concentration of PCZ. In short, environmental concentrations of PCZ could not induce obvious impacts on fish, but long-term exposure to higher concentrations of CBZ could affect seriously the health status of fish.

An in vivo and in vitro assessment of autophagy-related gene expression in muscle of rainbow trout (Oncorhynchus mykiss)

In mammals, new evidence has demonstrated the important role of the autophagic/lysosomal pathway in regulating muscle mass and identified the transcription factor FoxO3 as a key factor of the control of this proteolytic system by inducing several autophagy-related genes. In contrast, the mechanisms responsible for the regulation of autophagy have not been investigated in teleosts, known to exhibit different muscle growth dynamics. The present work aimed to characterize both in vivo and in vitro the transcriptional regulation of several major genes involved in autophagy (LC3B, gabarapl1, atg12l, atg4b) in the white skeletal muscle of rainbow trout. We found that fasting fish for 14days or serum depletion of trout myocytes strongly induces the expression of all studied genes. Our in vitro study on trout myocytes indicated that IGF1 induces FoxO3 phosphorylation but has a low or no effect on autophagy-related gene expression, suggesting a moderate role for this transcription factor on the autophagic/lysosomal pathway in this species. Data reported here show for the first time in a lower vertebrate, the existence and the regulation of several major genes involved in the autophagy, opening a new area of research on the molecular bases of muscle protein degradation in teleosts.

Toxicokinetics and the related metabolites in rainbow trout (Oncorhynchus mykiss) after exposure to decabromodiphenyl ether

Decabromodiphenyl ether (BDE209) is poorly absorbed by mammals, and little information is available on the toxicokinetics of BDE209 and its metabolites in fish. In the present study, rainbow trout (Oncorhynchus mykiss) were administered to 100 ng/g and 500 ng/g body wet weight of BDE209 via a single intraperitoneal injection and parent BDE209 and its metabolites were sequentially monitored for 28 days. The results showed that toxicokinetic profiles of BDE209 could be described by the one-compartment model. In the higher dose group (500 ng/g wet weight), the calculated half-life (t 1/2) and elimination rate (k e) were 17.7 d and 0.039/d in the liver, and 100.3 d and 0.007/d in the muscle, respectively. Three major methoxylated brominated diphenyl ethers (MeO-BDEs) were detected with 2,2′,4,4′-tetrabromo-5-methoxydiphenyl ether (5-MeO-BDE47) being detected in all tissue samples. There was no significant temporal change of 5-MeO-BDE47 concentration in the muscle, whereas an exponential increase was observed in the liver. Therefore, the metabolism rate of BDE209 depended on the administered dose. BDE209 was hardly accumulated in the muscle of rainbow trout, while the liver was a primary metabolic organ. MeO-BDEs were formed via metabolism of BDE209, which probably played a significant role in fish toxicology as a potential indicator.

EXPERIMENTAL INVESTIGATION OF HEAVY METAL ACCUMULATION IN TISSUES OF STONE LOACH NOEMACHEILUS BARBATULUS (L.) AND RAINBOW TROUT ONCORHYNCHUS MYKISS (WALBAUM) EXPOSED TO A MODEL MIXTURE (CU, ZN, NI, CR, PB, CD)

During experiment, fish were exposed to a heavy metal model mixture (Cu, Zn, Ni, Cr, Pb, Cd) for 14 days. Heavy metal concentrations corresponded to Maximum Permitted Concentrations (MPC) in surface waters. The amount of heavy metals in fish tissues was determined using atomic absorption spectrophotometry (AAS). Though both investigated fish species accumulated heavy metals with similar general intensity, the stone loach did it in the sequence gills > liver > muscle, and the rainbow trout ‐ conversely: muscle > liver > gills. Ni concentration in the stone loach muscle made 0.607 mg/kg and Pb concentration was 0.21 mg/kg, whereas Cd concentration in the rainbow trout muscle reached 0.45 mg/kg and exceeded the MPC indicated in the Lithuanian hygiene standard. These results coincide with the data of the heavy metal research into fish species from natural water bodies. Pb and Cd concentrations in fish tissues are the highest and frequently exceed the MPC. These findings urge for the constant control of the amount of heavy metals in fish. Santrauka Bandomosios žuvys 14 paru buvo veikiamos modeliniu sunkiuju metalu mišiniu (Cu, Zn, Ni, Cr, Pb, Cd). Metalu koncentracijos atitiko ju didžiausias leidžiamas koncentracijas (DLK) paviršiniuose vandenyse. Sunkiuju metalu kiekis audiniuose buvo nustatomas atomines absorbcijos spektrofotometriniu (AAS) būdu. Nors abieju tirtu rūšiu žuvu audiniuose sunkieji metalai kaupesi vienodu intensyvumu, šlyžio audiniuose jie kaupesi tokiu eiliškumu: žiaunos > kepenys > raumenys, o vaivorykštinio upetakio ‐ atvirkščiai: raumenys > kepenys > žiaunos. Šlyžio raumenyse Ni koncentracija buvo 0,607 mg/kg, o Pb ‐ 0,21 mg/kg ,vaivorykštinio upetakio raumenyse Cd koncentracija sieke 0,45 mg/kg ir viršijo Lietuvos higienos normoje nurodytaja DLK. Šie rezultatai sutampa su sunkiuju metalu nuolatines stebesenos kontroliuojamu rūšiu žuvyse iš natūraliu vandens telkiniu tyrimu duomenimis. Žuvu audiniuose Pb ir Cd koncentracijos yra didžiausios ir dažnai viršija DLK. Tokie rezultatai ipareigoja nuolat kontroliuoti sunkiuju metalu kieki žuvyse. Резюме Подопытные рыбы в течение 14 суток подвергались воздействию смеси тяжёлых металлов (Cu, Zn, Ni, Cr, Pb, Cd) в предельно допустимых концентрациях (ПДК), предусмотренных для внутренних водоёмов. Количество тяжелых металлов в тканях рыб устанавливали при помощи атомной абсорбции спектрофотометрическим методом. Несмотря на то, что оба вида рыб накапливали металлы с одинаковой общей интенсивностью, голец их накапливал в следующей последовательности: жабры > печень> мышцы, а радужная форель – наоборот: мышцы > печень> жабры. В мышцах гольца концентрация Ni составила 0,607 мг/кг, Pb – 0,21 мг/кг, а в тканях форели концентрация Cd составила 0,45 мг/кг и превышала ПДК. Экспериментальные данные совпадают с данными, полученными в мониторинговых водоемах. Установлено также, что концентрация Pb и Cd в тканях зачастую прeвышаeт ПДК. Полученные результаты обязывают постоянно контролировать уровень тяжелых металлов в тканях рыб.

Dietary protein differentially regulates the kinetic behaviour of serine dehydratase and tyrosine aminotransferase of liver and white muscle of rainbow trout (Oncorhynchus mykiss)

We have determined the kinetic behaviour of serine dehydratase (SerDH) and tyrosine aminotransferase (TyrAT) in the liver and white muscle of juvenile rainbow trout (Oncorhynchus mykiss) fed on a low-protein/high-fat (LP/HF) and control diets. The relationship between the kinetic parameters and tissue-DNA concentration has also been determined. SerDH and TyrAT showed hyperbolic kinetics in all cases. The partial replacement of protein with fat significantly increased hepatic SerDH-specific activity, maximum velocity (V(max)) and Michaelis constant (K(m)) while no significant changes were detected in the values of these parameters in white muscle. Nevertheless, hepatic and white-muscle TyrAT specific activity and V(max) decreased in trout fed on LP/HF diet with respect to control. The K(m) of hepatic TyrAT was also lower in trout fed on LP/HF diet than in control. The TyrAT kinetic parameters expressed per cell unit also changed in the same sense as previously described. These results show that, in trout, SerDH and TyrAT are two enzymes regulated by the partial replacement of dietary protein by fat.

Endocrine control of oleic acid and glucose metabolism in rainbow trout (Oncorhynchus mykiss) muscle cells in culture

The effects of insulin and IGF-I on fatty acid (FA) and glucose metabolism were examined using oleic acid or glucose as tracers in differentiated rainbow trout (Oncorhynchus mykiss) myotubes. Insulin and IGF-I significantly reduced the production of CO(2) from oleic acid with respect to the control values. IGF-I also significantly reduced the production of acid-soluble products (ASP) and the concentration of FA in the medium, while cellular triacylglycerols (TAG) tended to increase. Only insulin produced a significant accumulation of glycogen inside the cells in glucose distribution experiments. Incubation with catecholamines did not affect oleic acid metabolism. Cells treated with rapamycin [a target of rapamycin (TOR) inhibitor] significantly increased the oxidation of oleic acid to CO(2) and ASP, while the accumulation of TAG diminished. Rosiglitazone (a peroxisome proliferator-activated receptor gamma agonist) and etomoxir (a CPT-1 inhibitor) produced a severe and significant reduction in the production of CO(2) and ASP. Rosiglitazone and etomoxir also produced a significant accumulation of FA outside and inside the cells, respectively. No significant effects of these drugs on glucose distribution were observed. These data indicate that insulin and IGF-I act as anabolic hormones in trout myotubes in both oleic acid and glucose metabolism, although glucose oxidation appears to be less sensitive than FA oxidation to insulin and IGF-I. The use of rapamycin, etomoxir, and rosiglitazone may help us to understand the mechanisms of regulation of lipid metabolism in fish.

The production of fluorescent transgenic trout to study in vitro myogenic cell differentiation

BackgroundFish skeletal muscle growth involves the activation of a resident myogenic stem cell population, referred to as satellite cells, that can fuse with pre-existing muscle fibers or among themselves to generate a new fiber. In order to monitor the regulation of myogenic cell differentiation and fusion by various extrinsic factors, we generated transgenic trout (Oncorhynchus mykiss) carrying a construct containing the green fluorescent protein reporter gene driven by a fast myosin light chain 2 (MlC2f) promoter, and cultivated genetically modified myogenic cells derived from these fish.ResultsIn transgenic trout, green fluorescence appeared in fast muscle fibers as early as the somitogenesis stage and persisted throughout life. Using an in vitro myogenesis system we observed that satellite cells isolated from the myotomal muscle of transgenic trout expressed GFP about 5 days post-plating as they started to fuse. GFP fluorescence persisted subsequently in myosatellite cell-derived myotubes. Using this in vitro myogenesis system, we showed that the rate of muscle cell differentiation was strongly dependent on temperature, one of the most important environmental factors in the muscle growth of poikilotherms.ConclusionsWe produced MLC2f-gfp transgenic trout that exhibited fluorescence in their fast muscle fibers. The culture of muscle cells extracted from these trout enabled the real-time monitoring of myogenic differentiation. This in vitro myogenesis system could have numerous applications in fish physiology to evaluate the myogenic activity of circulating growth factors, to test interfering RNA and to assess the myogenic potential of fish mesenchymal stem cells. In ecotoxicology, this system could be useful to assess the impact of environmental factors and marine pollutants on fish muscle growth.

무지개 송어(Oncorhynchus mykiss)의 영양 성분에 대한 astaxanthin 첨가사료의 영향

This study investigated the effects of astaxanthin, which is used to improve the muscle color of fish, on the nutritional composition of rainbow trout (Oncorhynchus mykiss). The trout were fed extruded pellets containing astaxanthin (pigmented rainbow trout) or without astaxanthin (non-pigmented rainbow trout). No significant differences in the contents of crude protein and crude ash between the two muscles of the two groups were observed. However, the crude fat composition of the muscle of pigmented rainbow trout was about two times higher than that of control muscle. In the muscle of pigmented rainbow trout, the contents of docosahexaenoic acid (DHA, 22:6n-3) and eicosapentaenoic acid (EPA, 20:5n-3), which are functional polyunsaturated fatty acids, were 1.6 fold higher than in non-pigmented muscle. In addition, the contents of minerals such as Zn and Fe, vitamins (B group, C, and E), free amino acids, carotenoids and astaxanthin were increased in the muscle of rainbow trout fed pellets supplemented with astaxanthin. Specifically, the content of the bioactive compound astaxanthin, was increased six times in the pigmented muscle, as compared to the control muscle. The results suggest that a diet supplemented with astaxanthin improves the nutritional composition of rainbow trout muscle as well as improving the muscle color.

Skeletal muscle cellularity and expression of myogenic regulatory factors and myosin heavy chains in rainbow trout (Oncorhynchus mykiss): Effects of changes in dietary plant protein sources and amino acid profiles

The nutritional regulation of skeletal muscle growth is very little documented in fish. The aim of the study presented here was to determine how changes in dietary plant protein sources and amino acid profiles affect the muscle growth processes of fish. Juvenile rainbow trout (Oncorhynchys mykiss) were fed two diets containing fish meal and a mixture of plant protein sources either low (control diet) or rich in soybean meal (diet S). Both diets were supplemented with crystalline indispensable amino acids (IAA) to match the rainbow trout muscle IAA profile. Diet S was also supplemented with glutamic acid, an AA present in high quantities in trout muscle. Rainbow trout fed diets C and S were not significantly different in terms of overall somatic growth or daily nitrogen gain, although their parameters of dietary protein utilisation differed. Distribution of skeletal white muscle fibre diameter and expression of certain selected muscle genes were also affected by dietary changes. In the white muscle, diet S led to a significant decrease (×0.9) in the mean and median diameters of muscle fibres, to a significant decrease (×0.6) in the expression of MyoD and to a significant increase (×1.7) in the expression of fast-MHC, with no significant changes in myogenin expression. There was no change in the expression of the genes analysed in lateral red muscle (MyoD, MyoD2, myogenin and slow-MHC). These results demonstrated that changes occurred in skeletal white muscle cellularity and expression of MyoD and fast-MHC, although overall growth and protein accretion were not modified, when a diet rich in soybean meal and glutamic acid was ingested. Present findings also indicated that the white and red muscles of rainbow trout are differently affected by nutritional changes.

Glass Transition of Rainbow Trout and Its Oxidation Stability During Storage

Glass transition of rainbow trout muscle was measured by differential scanning calorimetry (DSC) as −13°C. Sucrose and sorbitol (2, 2), sucrose and mannitol (2, 2), sucrose and gum arabic (2, 0.15), sucrose and carrageenan (2, 0.15), sorbitol and mannitol (2, 2), sorbitol and gum arabic (2, 0.15), sorbitol and carrageenan (2, 0.15), mannitol and gum arabic (2, 0.15) and mannitol and carrageenan (2, 0.15) were blended with ground rainbow trout as g/100 g fish and stored for 6 months separately at −9°C, −13°C and −18°C. Total volatile basic nitrogen (TVB-N) and Thiobarbituric acid-reactive substances (TBARS) were determined at 1st, 3rd and 6th months of storage periods. Biopolymers blends, storage temperature and storage period had a significant effect (p < 0.05) on the TVB-N and TBARS values.

Skeletal muscle growth dynamics and expression of related genes in white and red muscles of rainbow trout fed diets with graded levels of a mixture of plant protein sources as substitutes for fishmeal

Juvenile rainbow trout were fed to visual satiation for 12 weeks with four diets containing fishmeal as the primary protein source (FM diet) or FM substituted with graded levels of a mix of plant proteins to replace 50% (PP50 diet), 75% (PP75) or 100% (PP100) of fishmeal with addition of crystalline amino acids to meet the requirements in indispensable amino acids. Growth performance, feed efficiency and protein efficiency were not affected by 50% fishmeal substitution but all decreased with higher substitution levels (FM = PP50 > PP75 > PP100). Ammonia excretion was inversely changed with FM substitution level (FM = PP50 < PP75 < PP100). Trout fed the FM diet had the highest level of nitrogen gain and those fed diet PP100 the lowest. Distribution of white muscle fibre diameter and expression of selected muscle genes were not affected by 50% fishmeal substitution. At higher substitution levels (75% and 100%) there was a significant decrease in the median diameter of white muscle fibres. Total substitution of fishmeal by plant protein sources (PP100) significantly increased (× 2.4) cathepsin D expression in white muscle but not in red muscle. Expression of MyoD, myogenin, PCNA, myosin heavy chains and calpain 2 was not modified by dietary treatments in either muscle type. This study thus demonstrates that the decrease in growth of trout fed a high level of plant protein sources is linked to changes in the dynamics of white muscle growth and correlated with changes in expression of a gene involved in lysosomal proteolysis in the white muscle.

Effects of training on lipid metabolism in swimming muscles of sea trout (Salmo trutta)

We examined the effect of exercise intensity and endurance training on plasma free fatty acid (FFA) kinetics and lipid metabolism in swimming muscles of reared sea trout. In both training groups [water current velocities 1 and 2 body lengths per second (bl s(-1))] the plasma level of FFAs decreased significantly (P < 0.001) compared to the control group. Similar significant (P < 0.01) post-exercise decrease was observed also in the lipase-esterase activity in the red muscle, but not in white. Moreover, in the group swimming with higher intensity a significantly higher (P < 0.05) lipase-esterase activity in the red muscle was found compared with the group on moderate exercise. As with cytochrome c oxidase activity, a significant elevation in the enzyme activity was also observed after training in the 1 bl s(-1) group in red and white muscle (P < 0.05 and P < 0.01, respectively). No changes were observed in beta hydroxyacyl CoA dehydrogenase activity. The lipid content was on average nine times higher in red compared to white muscle being 16.7, 21.1, and 24.9% in the red muscle of the control, 1 and 2 bl s(-1) groups, respectively, with a significant (P < 0.05) increase after training. We conclude that (1) unlike in mammals, plasma FFA kinetics and oxidation are not linearly related to exercise intensity in reared sea trout, (2) training enhances the capacity to uptake FFA from plasma, and (3) high intensity training shifts the proportion of energy derived from fat oxidation to carbohydrate-derived energy.

Molecular regulation of lipid metabolism in liver and muscle of rainbow trout subjected to acute and chronic insulin treatments

Although the metabolic actions of insulin in fish have been investigated widely in the past several years, lipid metabolism has received little attention, especially in tissues like the liver or white muscle. In the present study, rainbow trout received insulin treatments both acutely (intraperitoneal injection) and chronically (through mino-osmotic pumps) to elucidate hormone metabolic actions at molecular levels on the 2 main insulin target tissues in trout, namely, liver and muscle. Plasma and free fatty acid concentrations in plasma, as well as mRNA measurements of some key enzymes involved in lipid metabolism, were assessed in these tissues after 6 h and 4 d of acute and chronic insulin treatments, respectively. Our results showed that although fish received the same final total amount of hormone in both treatments, the actions of insulin on lipid metabolism were both time and tissue dependent. After the acute insulin treatment, the main anabolic role of insulin was reflected in decreased plasma free fatty acid concentrations linked to enhanced hepatic lipogenesis. We also found that insulin increased the mRNA levels of enzymes involved in lipid oxidation, perhaps to counteract insulin-induced hypoglycemia. In contrast, our data show that after chronic insulin treatment, liver and muscle exhibit different metabolic strategies: whereas in the liver chronic insulin-induced hypoglycemia may stimulate lipolytic processes to spare glucose stores, the muscle responds directly to the anabolic hormone action by increasing its lipogenic capacity and by inhibiting pathways of lipid oxidation.

Cytokine regulation of glucose entry into the skeletal muscle of the rainbow trout (Oncorhynchus mykiss): role of tumour necrosis factor alpha (TNFα)

Event Abstract Back to Event Cytokine regulation of glucose entry into the skeletal muscle of the rainbow trout (Oncorhynchus mykiss): role of tumour necrosis factor alpha (TNFα) Yoryia Vraskou1*, Nerea Roher2, Simon Mackenzie2 and Josep Planas1 1 University of Barcelona, Department of Physiology, Spain 2 Universitat Autonoma de Barcelona, Departamento de Biologia Cellular, Immunologia i Fisiologia Animal, Spain Tumour necrosis factor alpha (TNFα) is one of the most studied cytokines for its catabolic effects in target cells during excessive production. TNFα is known to have a direct action on skeletal muscle in mammals. However, little is known regarding the potential effects of cytokines on non-immune tissues, and particularly in skeletal muscle, in fish. The aim of this study was to investigate the effects of recombinant trout TNFα (rtTNFα), on skeletal muscle function in rainbow trout (Oncorhynchus mykiss). We used a primary cell culture of muscle cells from rainbow trout to show that rtTNFα stimulates glucose uptake in myoblasts and myotubes at concentrations that do not affect the viability of the cells and requiring de novo protein synthesis, as shown by the impairment of rtTNFα-stimulated glucose uptake by cycloheximide. With the use of specific inhibitors we show that rtTNFα-stimulated glucose uptake is mediated by the p38-MAPK, NF-kB and JNK pathways . Additionally, we provide evidence that the stimulatory effects of rtTNFα on glucose uptake in trout skeletal muscle cells may be caused, at least in part, by an increase in the amount of GLUT4 at the plasma membrane. Incubation of trout muscle cells with conditioned medium from LPS-stimulated trout macrophages stimulated glucose uptake. Our results indicate that recombinant as well as native trout TNFα directly stimulates glucose uptake in trout muscle cells and provides evidence, for the first time in non mammalian vertebrates, for a potential regulatory role of TNFα in skeletal muscle function. Keywords: comparative endocrinology Conference: 25th Conference of the European Comparative Endocrinologists, Pécs, Hungary, 31 Aug - 4 Sep, 2010. Presentation Type: Conference Presentation Topic: Comparative endocrinology Citation: Vraskou Y, Roher N, Mackenzie S and Planas J (2010). Cytokine regulation of glucose entry into the skeletal muscle of the rainbow trout (Oncorhynchus mykiss): role of tumour necrosis factor alpha (TNFα). Front. Endocrinol. Conference Abstract: 25th Conference of the European Comparative Endocrinologists. doi: 10.3389/conf.fendo.2010.01.00069 Copyright: The abstracts in this collection have not been subject to any Frontiers peer review or checks, and are not endorsed by Frontiers. They are made available through the Frontiers publishing platform as a service to conference organizers and presenters. The copyright in the individual abstracts is owned by the author of each abstract or his/her employer unless otherwise stated. Each abstract, as well as the collection of abstracts, are published under a Creative Commons CC-BY 4.0 (attribution) licence (https://creativecommons.org/licenses/by/4.0/) and may thus be reproduced, translated, adapted and be the subject of derivative works provided the authors and Frontiers are attributed. For Frontiers’ terms and conditions please see https://www.frontiersin.org/legal/terms-and-conditions. Received: 27 Aug 2010; Published Online: 29 Aug 2010. * Correspondence: Dr. Yoryia Vraskou, University of Barcelona, Department of Physiology, Barcelona, Spain, yoryiavrascou@ub.edu Login Required This action requires you to be registered with Frontiers and logged in. To register or login click here. Abstract Info Abstract The Authors in Frontiers Yoryia Vraskou Nerea Roher Simon Mackenzie Josep Planas Google Yoryia Vraskou Nerea Roher Simon Mackenzie Josep Planas Google Scholar Yoryia Vraskou Nerea Roher Simon Mackenzie Josep Planas PubMed Yoryia Vraskou Nerea Roher Simon Mackenzie Josep Planas Related Article in Frontiers Google Scholar PubMed Abstract Close Back to top Javascript is disabled. Please enable Javascript in your browser settings in order to see all the content on this page.

Kinetic properties of lactate dehydrogenase from trout muscle

The kinetics of pyruvate reduction by LDH isozymes in the presence of NADH is said to be an ordered bisubstrate reaction characterized by Michaelian concentration dependence (Borgmann et al., 1975). If the reaction is ordered, no cooperativity can occur between the two substrates (NADH and pyruvate) (Neet, 1995). Experiments with fish LDH showed, however, that this enzyme might display two types of saturation profile, hyperbolic and sigmoidal, depending on the pH in the reaction medium. At pH 9, for example, the initial velocity of pyruvate reduction by epaxial muscle LDH from goldfish showed a sigmoidal dependence upon substrate concentration, whereas at pH 8 the kinetics were of the hyperbolic type (Hochachka, 1965). Our experiments showed, in addition, that LDH from trout muscle (Onchorhynchus mykiss) could display both types of saturation profile at one and the same pH, but in the presence of different concentrations of the LDH preparation. Rainbow trout (Onchorhyncus mykiss) were obtained in October from the Goc Fishpond of the School of Forestry, University of Belgrade, Serbia. White skeletal muscle was immediately excised and kept in liquid nitrogen until use. After thawing, muscle samples were deprived of skin and spine, cut into small pieces, and homogenized in 5 volumes (w/v) of Tris-EDTA buffer. The homogenate was centrifuged for 10 min at 10 000× g. Partial purification with ammonium sulfate was performed according to Place and Powers, 1984. After centrifugation, solid ammonium sulfate was added to the supernatant to give 30% saturation at 0oC. The suspension was stirred for 1 hour and centrifuged for 30 min at 10 000× g. The pellet was discarded, and the supernatant brought to 70% saturation. The suspension was then stirred again for 1 h at 0oC, and centrifuged as already described. This time, the supernatant was discarded and the pellet redissolved in Tris-EDTA buffer and dialyzed overnight at 4oC against four 1liter replacements of the same buffer. The concentration of protein, determined by the method of Lowry (Lowry et al., 1951), amounted to 5.3 mg/ml of the final enzyme suspension.

Agonistic encounters and cellular angst: social interactions induce heat shock proteins in juvenile salmonid fish

Juvenile salmonid fish readily form dominance hierarchies when faced with limited resources. While these social interactions may result in profound behavioural and physiological stress, it is unknown if this social stress is evident at the level of the cellular stress response--specifically, the induction of stress or heat shock proteins (Hsps). Thus, the goal of our study was to determine if Hsps are induced during hierarchy formation in juvenile rainbow trout (Oncorhynchus mykiss). To this end, we measured levels of three Hsps, Hsp70, Hsc (heat shock cognate)70 and Hsp90 in the white muscle, liver and brain of trout that had been interacting for 36 h, 72 h or 6 days. Our data indicate that Hsps are induced in both dominant and subordinate fish in a time- and tissue-specific manner. In further mechanistic experiments on fasted and cortisol-treated fish, we demonstrated that high plasma cortisol does not affect Hsp induction in trout white muscle or liver, but both conditions may be part of the mechanism for Hsp induction with social stress in the brain. We conclude that the behavioural and physiological stress experienced by juvenile rainbow trout in dominance hierarchies can be extended to the induction of Hsps.