Long-term Fasting Research Articles

Cloning, distribution and effects of fasting status of melanocortin 4 receptor (MC4R) in Schizothorax prenanti

Melanocortin 4 receptor (MC4R) has an important role in the regulation of energy homeostasis in both mammals and fish. In this study, MC4R was characterized in S. prenanti (Schizothorax prenanti) and designated as SpMC4R. SpMC4R cDNA is composed of 1004 nucleotides with a 978 nucleotide open reading frame encoding a protein of 326 amino acids. The SpMC4R contained predicted regions that were structural features of MCR subtypes of vertebrates. In addition, phylogenetic analyses suggested that S. prenanti MC4R was closely related to fish MC4Rs. The SpMC4R mRNA was detected in embryos at developmental stages. Further, its mRNA was detectable in unfertilized eggs. Using real-time RT-PCR, MC4R is widely expressed, with highest levels of expression in brain and ovary. An experiment was conducted to determine the expression profile of MC4R during short-term and long-term fasting of the brain. The expression level of MC4R in unfed fish was significantly increased at 6, 9 and 24h post-fasting (hpf) and 14days fasting than in fed fish, this suggests that MC4R is conserved peptide that might be involved in the regulation of food intake and other physiological function in S. prenanti.

Changes in expression of appetite-regulating hormones in the cunner (Tautogolabrus adspersus) during short-term fasting and winter torpor

Feeding in vertebrates is controlled by a number of appetite stimulating (orexigenic, e.g. orexin and neuropeptide Y, NPY) and appetite suppressing (anorexigenic, e.g. cholecystokinin, CCK and cocaine- and amphetamine-regulated transcript, CART) hormones. Cunners (Tautogolabrus adspersus) survive the winter in shallow coastal waters by entering a torpor-like state, during which they forgo feeding. In order to better understand the mechanisms regulating appetite/fasting in these fish, quantitative real-time PCR was used to measure transcript expression levels of four appetite-regulating hormones: NPY, CART, orexin and CCK in the forebrain (hypothalamus and telencephalon) and CCK in the gut of fed, short-term summer fasted, and natural winter torpor cunners. Summer fasting induced a decrease in hypothalamic orexin levels and telencephalon NPY, CART and CCK mRNA levels. All brain hormone mRNA levels decreased during natural torpor as compared to fed summer fish. In the gut, CCK expression levels decreased during summer fasting. These results indicate that, in cunner, orexin, NPY, CART and CCK may play a role in appetite regulation and might mediate different physiological responses to short-term summer fasting and torpor-induced long-term fasting.

Genes involved in fatty acid metabolism: Molecular characterization and hypothalamic mRNA response to energy status and neuropeptide Y treatment in the orange-spotted grouper Epinephelus coioides

As in mammals, fatty acid (FA) metabolism plays diverse and vital roles in regulating food intake in fish. Multiple lines of evidence suggest that the effect of FA metabolism on food intake is linked to changes in the level of neuropeptide Y (NPY) in the hypothalamus of the rainbow trout. In mammals, the evidence suggests that FA metabolism regulates feeding via hypothalamic NPY. NPY is therefore considered an important factor that mediates the modulation of food intake by FA metabolism in vertebrates. The stimulatory effect of NPY on food intake is well known. However, to the best of our knowledge, the effect of NPY on FA metabolism in the hypothalamus has not been examined. In this study, we cloned the cDNA of four key enzymes involved in FA metabolism and assessed the effect of energy status and NPY on their mRNA expression in the hypothalamus of grouper. The full-length cDNAs of UCP2 and CPT1a and the partial coding sequence (CDS) of ACC1 and FAS were isolated from the grouper hypothalamus. These genes are expressed in the hypothalamus and during the organogenetic stage of embryogenesis. A feeding rhythm study showed that the hypothalamic expression level of NPY and CPT1a was highly correlated with feeding rhythm. Long-term fasting was found to significantly induce the hypothalamic mRNA expression of NPY, CPT1a and UCP2. An in vitro study demonstrated that NPY strongly stimulated CPT1a and UCP2 mRNA expression in a time- and dose-dependent manner. Collectively, these results suggest that these four genes related to FA metabolism may play a role in regulating food intake in grouper and, that NPY modulates FA metabolism in the grouper hypothalamus. This study showed, for the first time in vertebrates, the effect of NPY on the gene expression of FA metabolism-related enzymes.

Characterization, tissue distribution and regulation of agouti-related protein (AgRP) in a cyprinid fish (Schizothorax prenanti)

Agouti-related protein (AgRP) is an important neuropeptide involved in the regulation of feeding in both mammals and fish. In this study, we have cloned the full-length cDNA sequence for AgRP in a cyprinid fish (Schizothorax prenanti). The AgRP gene, encoding 126-amino acids, was strongly expressed in the brain. The AgRP gene was detected in embryos at developmental stages. Further, its mRNA was detectable in unfertilized eggs. An experiment was conducted to determine the expression profile of AgRP during short-term and long-term fasting of the hypothalamus. The expression level of AgRP in unfed fish was significantly increased at 3 and 4h post-fasting than in fed fish but did not affect AgRP mRNA expression after 14days fasting. Overall, our results suggest that AgRP is a conserved peptide that might be involved in the regulation of short-term feeding and other physiological function in Schizothorax prenanti.

Long-term fasting in the anadromous Arctic charr is associated with down-regulation of metabolic enzyme activity and up-regulation of leptin A1 and SOCS expression in the liver.

The life strategy of the anadromous Arctic charr (Salvelinus alpinus) includes several months of voluntary fasting during overwintering in freshwater, leading to emaciation prior to seawater migration in spring. In this study we compared changes in condition, substrate utilization and liver metabolism between captive anadromous charr subjected to food deprivation during late winter and spring, and conspecifics fed in excess. In March, nine out of the 10 sampled fed fish had not eaten, indicating that they were in a voluntary anorexic state. In June, the fed fish were eating and all had higher body mass, condition factor and adiposity than in March. In fasted fish there were only small decreases in body mass, condition factor and adiposity between March and May, but all these parameters decreased markedly from May to June. The fasted fish were depleted in fat and glycogen in June, had suppressed activity of hepatic enzymes involved in lipid metabolism (G6PDH and HOAD) and seemed to rely on protein-derived glucose as a major energy source. This was associated with upregulated liver gene expression of leptin A1, leptin A2, SOCS1, SOCS2 and SOCS3, and reduced IGF-I expression. In an in vitro study with liver slices it was shown that recombinant rainbow trout leptin stimulated SOCS1 and SOCS3 expression, but not SOCS2, IGF-I or genes of enzymes involved in lipid (G6PDH) and amino acid (AspAT) metabolism. It is concluded that liver leptin interacts with SOCS in a paracrine fashion to suppress lipolytic pathways and depress metabolism when fat stores are depleted.

Fsp27/CIDEC is a CREB target gene induced during early fasting in liver and regulated by FA oxidation rate

FSP27 [cell death-inducing DFFA-like effector c (CIDEC) in humans] is a protein associated with lipid droplets that downregulates the fatty acid oxidation (FAO) rate when it is overexpressed. However, little is known about its physiological role in liver. Here, we show that fasting regulates liver expression of Fsp27 in a time-dependent manner. Thus, during the initial stages of fasting, a maximal induction of 800-fold was achieved, whereas during the later phase of fasting, Fsp27 expression decreased. The early response to fasting can be explained by a canonical PKA-CREB-CRTC2 signaling pathway because: i) CIDEC expression was induced by forskolin, ii) Fsp27 promoter activity was increased by CREB, and iii) Fsp27 expression was upregulated in the liver of Sirt1 knockout animals. Interestingly, pharmacological (etomoxir) or genetic (Hmgcs2 interference) inhibition of the FAO rate increases the in vivo expression of Fsp27 during fasting. Similarly, CIDEC expression was upregulated in HepG2 cells by either etomoxir or HMGCS2 interference. Our data indicate that there is a kinetic mechanism of autoregulation between short- and long-term fasting, by which free FAs delivered to the liver during early fasting are accumulated/exported by FSP27/CIDEC, whereas over longer periods of fasting, they are degraded in the mitochondria through the carnitine palmitoyl transferase system.

Differences in Mucociliary activity of volunteers undergoing Ramadan versus Nineveh fasting

This study, aimed to evaluate the difference in mucociliary clearance among volunteers who underwent Ramadan versus Nineveh fasting regimens as well as the difference between the fasting period and 4 weeks following the fasting period in both groups. In this study, two different fasting groups were established: Ramadan (fasting for an average of 15 h for 29 consecutive days, n = 40) and Nineveh (60 h of nonstop fasting, n = 26). Subjects in each group underwent saccharin testing twice: at the end of the fasting period prior to resumption of eating and at 4 weeks after the end of Ramadan or Nineveh fasting. Statistical analysis was performed using the Mann-Whitney U-test, Wilcoxon, Chi-square, and paired t test. A p value less than 0.05 was considered statistically significant. Forty subjects who underwent Ramadan fasting and 26 subjects who underwent Nineveh fasting were included in this study. Of the 66 study participants, 34 (51.5 %) were men and 32 (48.5 %) were women. Their median age was 31 years (range 17-70 years) for Nineveh fasting subjects and 40 years (range 17-70 years) for Ramadan fasting subjects. Chi-square tests revealed no significant difference between the Ramadan and Nineveh fasting groups in gender (p = 0.418), and the Mann-Whitney U-test showed no difference in age. A statistically significant difference was found in the mucociliary clearance time between the Nineveh fasting and non-fasting periods (p = 0.013). Using Wilcoxon signed-rank tests, we found no significant difference in the mucociliary clearance time between the Ramadan fasting and control (4 weeks after the fasting period) periods (p = 0.121). The percentage difference between the fasting and control periods was similar between groups and was not statistically significant for the Ramadan and Nineveh fasting groups (p = 0.086). The results of the present study indicated that long-term fasting with hypohydration contributed to the deterioration of nasal mucociliary clearance. Our data indicate that optimal hydration, sleep patterns, and fasting times contribute to proper mucociliary clearance.

Isolation and selection of suitable reference genes for real-time PCR analyses in the skeletal muscle of the fine flounder in response to nutritional status: assessment and normalization of gene expression of growth-related genes

In the present study, different reference genes were isolated, and their stability in the skeletal muscle of fine flounder subjected to different nutritional states was assessed using geNorm and NormFinder. The combinations between 18S and ActB; Fau and 18S; and Fau and Tubb were chosen as the most stable gene combinations in feeding, long-term fasting and refeeding, and short-term refeeding conditions, respectively. In all periods, ActB was identified as the single least stable gene. Subsequently, the expression of the myosin heavy chain (MYH) and the insulin-like growth factor-I receptor (IGF-IR) was assessed. A large variation in MYH and IGF-IR expression was found depending on the reference gene that was chosen for normalizing the expression of both genes. Using the most stable reference genes, mRNA levels of MYH decreased and IGF-IR increased during fasting, with both returning to basal levels during refeeding. However, the drop in mRNA levels for IGF-IR occurred during short-term refeeding, in contrast with the observed events in the expression of MYH, which occurred during long-term refeeding. The present study highlights the vast differences incurred when using unsuitable versus suitable reference genes for normalizing gene expression, pointing out that normalization without proper validation could result in a bias of gene expression.

Preparation technique and rice variety: Modeling the effect on long-term fasting blood glucose

Preparation technique and rice variety: Modeling the effect on long-term fasting blood glucose

Molecular cloning, characterization and expression profiles of multiple leptin genes and a leptin receptor gene in orange-spotted grouper (Epinephelus coioides)

Leptin plays key roles in body weight regulation, energy metabolism, food intake, reproduction and immunity in mammals. However, its function in teleosts is still unclear. In the present study, two leptin genes (gLepA and gLepB) and one leptin receptor gene (gLepR) were cloned and characterized in orange-spotted grouper (Epinephelus coioides). The cDNAs of gLepA and gLepB were 671bp and 684bp in length, encoding for proteins of 161 amino acid (aa) and 158 aa, respectively. The three-dimensional (3D) structures modeling of gLepA and gLepB showed strong conservation of tertiary structure with that of other vertebrates. The total length of gLepR cDNA was 4242bp, encoding a protein of 1169 aa which contained all functionally important domains conserved among vertebrate LEPR. Tissue distribution analysis showed that gLepA was highly expressed in cerebellum, liver and ovary, while gLepB mRNA abundantly in the brain regions, as well as in the ovary with some extend. The gLepR was mainly expressed in kidney, head kidney and most of brain regions. Analysis of expression profiles of gLep and gLepR genes during the embryonic stages showed that high expression of gLepR was observed in the brain vesicle stage, while neither gLepA nor gLepB mRNA was detected during different embryonic stages. Finally, fasting and refeeding experiments were carried out to investigate the possible function of leptin genes in food intake and energy metabolism, and the results showed that a significant increase of gLepA expression in the liver was induced by food deprivation in both short-term (7days) and long-term (3weeks) fasting and gLepA mRNA upregulation was eliminated after refeeding, while gLepB wasn’t detected in the liver of grouper during fasting. No significant differences in hypothalamic leptin and leptin receptor expression were found during short-term fasting and refeeding. Hepatic expression of gLepA mRNA increased significantly 9h after a single meal. These results suggested gLepA, other than gLepB, functioned in the regulation of energy metabolism and food intake in this Perciform fish.

Decreased expression of adipose CD36 and FATP1 are associated with increased plasma non-esterified fatty acids during prolonged fasting in northern elephant seal pups (Mirounga angustirostris)

The northern elephant seal pup (Mirounga angustirostris) undergoes a 2-3 month post-weaning fast, during which it depends primarily on the oxidation of fatty acids to meet its energetic demands. The concentration of non-esterified fatty acids (NEFAs) increases and is associated with the development of insulin resistance in late-fasted pups. Furthermore, plasma NEFA concentrations respond differentially to an intravenous glucose tolerance test (ivGTT) depending on fasting duration, suggesting that the effects of glucose on lipid metabolism are altered. However, elucidation of the lipolytic mechanisms including lipase activity during prolonged fasting in mammals is scarce. To assess the impact of fasting and glucose on the regulation of lipid metabolism, adipose tissue and plasma samples were collected before and after ivGTTs performed on early (2 weeks, N=5) and late (6-8 weeks; N=8) fasted pups. Glucose administration increased plasma triglycerides and NEFA concentrations in late-fasted seals, but not plasma glycerol. Fasting decreased basal adipose lipase activity by 50%. Fasting also increased plasma lipase activity twofold and decreased the expressions of CD36, FAS, FATP1 and PEPCK-C by 22-43% in adipose tissue. Plasma acylcarnitine profiling indicated that late-fasted seals display higher incomplete LCFA β-oxidation. Results suggest that long-term fasting induces shifts in the regulation of lipolysis and lipid metabolism associated with the onset of insulin resistance in northern elephant seal pups. Delineation of the mechanisms responsible for this shift in regulation during fasting can contribute to a more thorough understanding of the changes in lipid metabolism associated with dyslipidemia and insulin resistance in mammals.

Of the Importance of Metabolic Phases in the Understanding of Oxidative Stress in Prolonged Fasting and Refeeding

Life phases such as migration or reproduction may partly (or totally) prohibit food accessibility, making regulation of the energy balance one of the main challenges faced by wild organisms. Although long-term fasting is common in a number of species, it has been reported to be detrimental for the organism, notably because it induces oxidative stress. However, fasting metabolism is characterized by successive metabolic adaptations that are likely to be different in terms of stress, and no previous studies, to our knowledge, have tested the dynamics of changes in oxidative balance in relation to the metabolic phases of fasting. Our study provides a first insight into this relationship by inducing prolonged fasting and subsequent refeeding in captive mallards (Anas platyrhynchos). Both plasmatic antioxidant and oxidative damage levels were observed to be lower during fasting. Oxidative damage levels decreased by 95% during fasting, and they surprisingly remained at low levels even after 3 d of refeeding. In contrast, restoration of the antioxidant barrier was observed from the third day of refeeding onward, and it thus preceded the increase in oxidative damage levels. Therefore, under our experimental conditions, long-term fasting was not associated with marked oxidative stress in mallards. Because data interpretations may be influenced by concomitant metabolic states, we underline the importance of taking metabolic phases into account when studying oxidative stress during fasting.

Fasting induces ketoacidosis and hypothermia in PDHK2/PDHK4-double-knockout mice.

The importance of PDHK (pyruvate dehydrogenase kinase) 2 and 4 in regulation of the PDH complex (pyruvate dehydrogenase complex) was assessed in single- and double-knockout mice. PDHK2 deficiency caused higher PDH complex activity and lower blood glucose levels in the fed, but not the fasted, state. PDHK4 deficiency caused similar effects, but only after fasting. Double deficiency intensified these effects in both the fed and fasted states. PDHK2 deficiency had no effect on glucose tolerance, PDHK4 deficiency produced only a modest effect, but double deficiency caused a marked improvement and also induced lower insulin levels and increased insulin sensitivity. In spite of these beneficial effects, the double-knockout mice were more sensitive than wild-type and single-knockout mice to long-term fasting, succumbing to hypoglycaemia, ketoacidosis and hypothermia. Stable isotope flux analysis indicated that hypoglycaemia was due to a reduced rate of gluconeogenesis and that slightly more glucose was converted into ketone bodies in the double-knockout mice. The findings establish that PDHK2 is more important in the fed state, PDHK4 is more important in the fasted state, and survival during long-term fasting depends upon regulation of the PDH complex by both PDHK2 and PDHK4.

Activating transcription factor 4-dependent induction of FGF21 during amino acid deprivation

Nutrient deprivation or starvation frequently correlates with amino acid limitation. Amino acid starvation initiates a signal transduction cascade starting with the activation of the kinase GCN2 (general control non-derepressible 2) phosphorylation of eIF2 (eukaryotic initiation factor 2), global protein synthesis reduction and increased ATF4 (activating transcription factor 4). ATF4 modulates a wide spectrum of genes involved in the adaptation to dietary stress. The hormone FGF21 (fibroblast growth factor 21) is induced during fasting in liver and its expression induces a metabolic state that mimics long-term fasting. Thus FGF21 is critical for the induction of hepatic fat oxidation, ketogenesis and gluconeogenesis, metabolic processes which are essential for the adaptive metabolic response to starvation. In the present study, we have shown that FGF21 is induced by amino acid deprivation in both mouse liver and cultured HepG2 cells. We have identified the human FGF21 gene as a target gene for ATF4 and we have localized two conserved ATF4-binding sequences in the 5' regulatory region of the human FGF21 gene, which are responsible for the ATF4-dependent transcriptional activation of this gene. These results add FGF21 gene induction to the transcriptional programme initiated by increased levels of ATF4 and offer a new mechanism for the induction of the FGF21 gene expression under nutrient deprivation.

Meal Anticipatory Rise in Acylated Ghrelin at Dark Onset is Blunted After Long-Term Fasting in Rats

Ghrelin is a 28 amino acid acylated peptide originally characterised for its capacity to stimulate growth hormone secretion. Ghrelin is also an orexigenic and adipogenic hormone and is thought to be a signal to increase locomotor activity in anticipation of a scheduled meal. Although ghrelin is considered to be up-regulated during fasting, there are still conflicting data regarding the impact of starvation on ghrelin secretion. To test whether the secretory pattern of acylated ghrelin is altered during fasting, plasma levels were monitored every 20 min for 6 h in freely-behaving rats at the light/dark cycle transition, when animals initiate feeding and activity and use preferentially free fatty acids (FFA) as a source of energy. Rats were fed ad lib. or fasted at dark onset for 24, 48 or 72 h, with or without refeeding rate. The anticipatory rise in ghrelin levels, as well as home-cage activity at the onset of darkness, was significantly reduced from 48 h of fasting compared to ad lib. conditions. A delayed ghrelin peak, sensitive to renutrition, was observed in fasted animals. Although their motivation to eat appeared to be intact, rats fasted for 72 h showed the smallest compensatory refeeding rate after fasting, possibly reflecting altered gut function. Expression of agouti-related protein and neuropeptide Y, was significantly increased in 48- and 72-h fasted animals. Thus, following fasting, a blunted acylated ghrelin secretion at dark onset (i.e. a period when animals depend on FFA as a source of energy) is associated with reduced locomotor activity and refeeding and an up-regulation of anabolic neuropeptides. Such changes could be interpreted as compensatory mechanisms for helping to conserve energy under conditions where food is not available.

Seasonal Feeding Rhythm Associated with Fasting Period ofPangasianodon gigas: Long-Term Monitoring in an Aquarium

The Mekong giant catfish Pangasianodon gigas is endemic to the Mekong River basin, and is recognized as endangered species, largely due to overfishing and development of the river basin. We monitored food intake of P. gigas in a stable environment in an aquarium over a 6-year period and analyzed their feeding rhythm and fasting periods. The daily food intake for each fish was recorded from 18 June 2004 to 17 June 2010. The feeding rhythm or pattern was determined by the fast Fourier transform (FFT) analysis. The FFT analysis revealed that different cycles of feeding rhythm (168.8, 313.1, and 365.3 days) in three catfishes and no observable cycles in two catfishes. However, three catfishes showed subordinate peaks with approximately 365 days (365.3 days for all). These suggest that, at least, four of five catfish had have approximately 365-days feeding cycle. We also showed that all catfish undergo long-term fasting periods (> 20 days). Of note, the feeding/fasting pattern coincides with the wet/dry seasons in Thailand, which also corresponds to the abundance of the catfish food resource (Cladophora spp.). We found that P. gigas exhibit a seasonal feeding rhythm that is synchronized by food availability. Furthermore, we found that the seasonal feeding rhythm was gradually dampened over time, suggesting that the observed seasonal feeding rhythm with long-term fasting of the catfish is likely controlled by an endogenous clock system. To our knowledge, this is the first case of quantification of the seasonal feeding rhythm with fasting periods in teleost fish.

Ligation of the left aorta in alligators affects acid–base balance: A role for the R→L shunt

This study investigated the effects of preventing bulk flow from the right ventricle to the body via the left aorta (LAo; right to left shunt, R→L) on acid-base status in alligators following feeding, during long-term fasting and a cold temperature exposure. Post-feeding pHv and [Formula: see text] were not significantly different between S and C. Post-feeding pHv increased in both groups of alligators, but not significantly. During fasting, all acid-base variables were similar between the two groups of alligators. A 10 °C reduction in environmental temperature resulted in a significant difference in pHv and HCO3- between S and C. Both pHv and HCO3- were significantly higher in C animals. PV(CO2) significantly decreased in both groups during the cold exposure. Preventing the R→L shunt via the LAo had significant effects on acid-base balance in alligators indicating incomplete compensation for its loss and a role for the LAo in metabolic homeostasis.

Exogenous corticosterone mimics a late fasting stage in captive Adélie penguins (Pygoscelis adeliae)

Fasting is part of penguin's breeding constraints. During prolonged fasting, three metabolic phases occur successively. Below a threshold in body reserves, birds enter phase III (PIII), which is characterized by hormonal and metabolic shifts. These changes are concomitant with egg abandonment in the wild and increased locomotor activity in captivity. Because corticosterone (CORT) enhances foraging activity, we investigated the variations of endogenous CORT, and the effects of exogenous CORT on the behavioral, hormonal, and metabolic responses of failed breeder Adélie penguins. Untreated and treated captive male birds were regularly weighed and sampled for blood while fasting, and locomotor activity was recorded daily. Treated birds were implanted with various doses of CORT during phase II. Untreated penguins entering PIII had increased CORT (3.5-fold) and uric acid (4-fold; reflecting protein catabolism) levels, concomitantly with a rise in locomotor activity (2-fold), while prolactin (involved in parental care in birds) levels declined by 33%. In CORT-treated birds, an inverted-U relationship was obtained between CORT levels and locomotor activity. The greatest increase in locomotor activity was observed in birds implanted with a high dose of CORT (C100), locomotor activity showing a 2.5-fold increase, 4 days after implantation to a level similar to that of birds in PIII. Moreover, uric acid levels increased three-fold in C100-birds, while prolactin levels declined by 30%. The experimentally induced rise in CORT levels mimicked metabolic, hormonal, and behavioral changes, characterizing late fasting, thus supporting a role for this hormone in the enhanced drive for refeeding occurring in long-term fasting birds.

Insulin Stimulates Lipogenesis and Attenuates Beta‐Oxidation in White Adipose Tissue of Fed Rainbow Trout

As lipid deposition tissue in fish, the white adipose tissue (WAT) has important functions related to reproduction and the challenges of long-term fasting. In the study reported here, we infused fish fed a high-carbohydrate diet with two doses of insulin for 5 days in order to explore the effects of this hormone on lipogenesis and beta-oxidation-related enzymes. We demonstrated the presence of some of the main lipogenic enzymes at molecular, protein and activity levels (ATP-citrate lyase and fatty acid synthase). However, while ATP-citrate lyase was unexpectedly down-regulated, fatty acid synthase was up-regulated (at protein and activity levels) in an insulin dose-dependent manner. The main enzymes acting as NADPH donors for lipogenesis were also characterized at biochemical and molecular levels, although there was no evidence of their regulation by insulin. On the other hand, lipid oxidation potential was found in this tissue through the measurement of gene expression of enzymes involved in β-oxidation, highlighting two carnitine palmitoyltransferase isoforms, both down-regulated by insulin infusion. We found that insulin acts as an important regulator of trout WAT lipid metabolism, inducing the final stage of lipogenesis at molecular, protein and enzyme activity levels and suppressing β-oxidation at least at a molecular level. These results suggest that WAT in fish may have a role that is important not only as a lipid deposition tissue but also as a lipogenic organ (with possible involvement in glucose homeostasis) that could also be able to utilize the lipids stored as a local energy source.

Oxidative Stress and Antioxidant Defenses after Long-term Fasting in Blood of Chinese Sturgeon (Acipenser sinensis)

The effects of long-term fasting on the oxidative stress and antioxidant defenses in juvenile Chinese sturgeon (Acipenser sinensis) were studied. The oxidative stress biomarkers included lysozyme (LSZ), the ability of inhibiting hydroxyl radicals (AIHR), and total antioxidant capacity (T-AOC). The juvenile Chinese sturgeon were fasted for 24 hours at first. Blood samples were taken from fish a total of 8 times, upon sampling every 6 days during the remaining 48-day starvation period. The LSZ activity of juveniles generally increased as the starvation time increased. Furthermore, a significant difference in the LSZ activity of juveniles starved for 43 days was found compared with four groups starved for 1–19 days. In addition, the AIHR of juveniles first increased and subsequently decreased over the course of starvation. Specifically, the AIHR of juveniles was significantly higher in fish after 19 days of starvation than in fish sampled after 37 days of starvation, although there were no other significant differences among the levels for the other groups. The variation of the T-AOC levels of juveniles was higher during the earlier stage of starvation, and was decreased after 19 days of starvation. However, there were no significant differences among the T-AOC groups, except the T-AOC of juveniles starved for 13 days was significantly higher than that of fish starved for 37 days. It's concluded that long-term fasting influenced the antioxidant capability of juvenile Chinese sturgeon.