Mobilization and recovery of energy stores in traíra, Hoplias malabaricus Bloch (Teleostei, Erythrinidae) during long-term starvation and after re-feeding

In some neotropical environments, fishes often experience periods of poor food supply, especially due to extreme fluctuations in rainfall regime. The fish species that experience periods of drought such as the traíra Hoplias malabaricus (Bloch 1794), may stand up to long-term food deprivation. In this study, experiments were performed in order to determine the dynamic of utilization of endogenous reserves in this species during starvation. Adult traíra were both fasted for 30–240 days and re-fed for 30 days following 90 and 240 days of fasting. Glycogen and perivisceral fat were primary energy substrates consumed. During the first 30 days, fish consumed hepatic and muscular glycogen, without exhausting these reserves, and used lipids from perivisceral fat. Hepatic lipids were an important energy source during the first 60 days of starvation and perivisceral fat were consumed gradually, being exhausted after 180 days. Protein mobilization was noticeable after 60 days of fasting, and became the major energy source as the lipid reserves were decreased (between 90 and 180 days). Following the longest periods of food deprivation, fish had utilized hepatic glycogen again. Fish re-fed for 30 days after 90 and 240 days of fasting were able to recover hepatic glycogen stores, but not the other energy reserves.     

The effects of long-term food deprivation on respiration and haematology of the neotropical fish Hoplias malabaricus

Growth of adult traíras Hoplias malabaricus ceased and body mass ( M ) decreased during starvation periods of 30, 60, 90, 150, 180 and 240 days. Hepatic reserves were mobilized in fish starved for 30 days, but liver mass of fish starved for longer periods was not significantly different from those starved for 30 days. Perivisceral fat bodies were consumed gradually, being completely exhausted after 240 days of food deprivation. Length of starvation was associated with a significant decrease in the oxygen uptake ( V o₂). In spite of this reduction, the respiratory frequency ( f _R) was kept nearly constant during the starvation periods. The haematocrit and the number of red blood cells decreased after 150 and 240 days of starvation, respectively. These parameters did not recover after refeeding (after 90 and 240 days of starvation). This hypometabolic state in response to food deprivation contributed to energy conservation during these periods. Traíras can survive food deprivation for periods of up to 180 days without reductions in metabolism and when they do become hypometabolic, normal metabolic rates are rapidly restored upon refeeding.     

The regulation of endogeneous energy stores during starvation and refeeding in the somatic tissues of the golden perch

Adult golden perch Macquaria ambigua were fed to satiety, starved for up to 210 days, or starved for 150 days then fed to satiety for 60 days to investigate the utilization of energy stores in response to food deprivation and re-feeding. Golden perch sequentially mobilize energy from hepatic tissue, extra-hepatic lipid, and finally muscle components in response to food deprivation. The relative size of the liver was significantly reduced by 30 days after the onset of food deprivation due to the simultaneous mobilization of lipid, protein and glycogen reserves. These stores were renewed rapidly within 30 days by satiety feeding. Mobilization of lipid stores in perivisceral fat bodies occurred between 30 and 60 days of food deprivation. These deposits were also renewed upon re-feeding, although not as rapidly as liver reserves. The glycogen content of the epaxial muscle was reduced by the 60th day of food deprivation but subsequently increased indicating the mobilization of other energy reserves. The concentration of muscle lipid decreased after 90 days of food deprivation. The only significant response in body composition observed in the fish fed to satiety throughout the study was an increase in the relative size of the perivisceral fat bodies. The results of this study suggest that golden perch are well adapted to cope with extended periods of food deprivation, storing energy as perivisceral fat when food is readily available and having a clearly sequential process for mobilizing energy when food is scarce which largely protects the integrity of the musculature.     

Long‐term starvation in cave salamander effects on liver ultrastructure and energy reserve mobilization

The morphological alterations of hepatocytes of cave‐dwelling salamander Proteus anguinus anguinus after food deprivation periods of one and 18 months were investigated and the concentrations of glycogen, lipids, and proteins in the liver were determined. Quantitative analyses of the hepatocyte size, the lipid droplets, the number of mitochondria, and volume densities of M and P in the hepatocytes were completed. After one month of food deprivation, the cytological changes in the hepatocytes are mainly related to the distribution and amount of glycogen, which was dispersed in the cytoplasm and failed to form clumps typical of normal liver tissue. After 18 months of food deprivation hepatocytes were reduced in size, lipid droplets were less numerous, peroxisomes formed clusters with small, spherical mitochondria, and specific mitochondria increased in size and lost cristae. Lysosomes, autophagic vacuoles, and clear vacuoles were numerous. The liver integrity was apparently maintained, no significant loss of cytoplasmic constituents have been observed. Biochemical analysis revealed the utilization of stored metabolic reserves in the liver during food deprivation. Glycogen is rapidly utilized at the beginning of the starvation period, whereas lipids and proteins are utilized subsequently, during prolonged food deprivation. In the Proteus liver carbohydrates are maintained in appreciable amounts and this constitutes a very important energy depot, invaluable in the subterranean environment. J. Morphol. 274:887–900, 2013. © 2013 Wiley Periodicals, Inc.     

The ultrastructure of the parietal cells of the stomach and their functional activity]

Submicroscopic changes in the parietal cells of the fundus were compared with the data of spot-film intragastric pH-metry of the same mucosal areas in 18 patients treated at the hospital in the department of therapeutic food deprivation. There were seen no changes in the ultrastructure of the parietal cells after a brief (36-hour) fasting when the pH of the mucous membrane failed to differ from the normal. With increased duration of food deprivation there occurred a gradual alkalinization of the gastric secretion and the pH of the gastric wall shifted from 2.02 after a 3-day food deprivation to 4.8-5.0 after fasting for 20 to 30 days. In accordance, there was a change in the ultrastructure of the parietal cells consisting in flattening and subsequent disappearance of tubulovesicles, a reduction of the lumina of the intracellular channels and shortening of microvilli. The results of comparison of the submicroscopic changes with the data of pH-metry indicated that such submicroscopic shifts corresponded co depression of the functional activity of the parietal cells.     

Adaptations to fasting in the American mink (Mustela vison): carbohydrate and lipid metabolism

The aim of this study was to investigate whether the actively wintering American mink Mustela vison is strictly dependent on continuous food availability or if it has evolved physiological adaptations to tolerate nutritional scarcity. Fifty farm-bred male minks were divided into a fed control group and four experimental groups fasted for 2, 3, 5 or 7 days. The rate of weight loss was several-fold higher (1.5-3.2% day(-1)) in the mink than recorded previously in larger carnivores utilizing passive wintering strategies. The minks remained normoglycaemic, although their liver glycogen stores and glucose-6-phosphatase activities decreased during fasting. Adipose tissue constituted approximately 36% of their body mass after 7 days of food deprivation. Intra-abdominal fat, especially retroperitoneal but also mesenteric adipose tissue, were the most important fat depots to be hydrolyzed, but the ability of the mink to utilize its body lipids during fasting may be limited. The increased liver size, hepatic triacylglycerol accumulation and increases in the activities of plasma aminotransferases indicated liver dysfunction. Food deprivation also affected the red blood cell indices, and the blood monocyte and lymphocyte counts decreased suggesting immunosuppression during fasting. The results of the present study suggest that the mink has not evolved sophisticated adaptations to wintertime fasting.     

Effect of fasting on carbohydrate metabolism in frugivorous bats (Artibeus lituratus and Artibeus jamaicensis)

The compensatory changes of carbohydrate metabolism induced by fasting were investigated in frugivorous bats, Artibeus lituratus and Artibeus jamaicensis. For this purpose, plasma levels of glucose and lactate, liver and muscle glycogen content, rates of liver gluconeogenesis and the activity of related enzymes were determined in male bats. After a decrease during the first 48 h of fasting, plasma glucose levels remained constant until the end of the experimental period. Plasma lactate levels, extremely high in fed bats, decreased after 48 h of fasting. Similarly, liver glycogen content, markedly high in fed animals, was reduced to low levels after 24 h without food. Muscle glycogen was also reduced in fasted bats. The expected increase in liver gluconeogenesis during fasting was observed after 48 h of fasting. The activities of liver glucose-6-phosphatase and fructose-1,6-bisphosphatase were not affected by food withdrawn. On the other hand, fasting for 24 h induced an increase in the activity of liver cytosolic phosphoenolpyruvate carboxykinase. The data indicate that liver gluconeogenesis has an important role in the glucose homeostasis in frugivorous bats during prolonged periods of food deprivation. During short periods of fasting liver glycogenolysis seems to be the main responsible for the maintenance of glycemia.     

The effect of starvation and subsequent feeding on the hepatocytes of Chanos chanos (Forsskal) fingerlings and fry

Excised liver sections of the milkfish, Chanos chanos , fry and fingerlings were studied by transmission electron microscopy. The hepatocytes underwent marked ultrastructural alterations in response to food deprivation of 10-day starvation for fry and 2 months for the fingerlings. The prominent features characterizing the hepatocytes of starved fish were: a reduction of cell and nucleus size; apparent loss of nucleoli; condensation of chromatin material in fry; loss of stored glycogen; reduction of ER profiles; increase in the number of electron-dense bodies containing large amounts of iron in fingerlings; and an increase in mitochondrial size. These changes were reversible following short periods of re-feeding, i.e. 2 days for fry and 4 days for fingerlings, using natural food for the fry and formulated diet for the fingerlings.     

Combined effects of fasting and vinblastine treatment on serum insulin level, the size of autophagic-lysosomal compartment, protein content and lysosomal enzyme activities of liver and exocrine pancreatic cells of the mouse

1. The volume fraction of autophagic vacuoles in liver parenchymal and exocrine pancreatic cells was smallest and the serum insulin level highest in the 24 hr prestarved mouse immediately after 3 hr feeding period. 2. The size of the autophagic vacuole and lysosome (dense body) compartments increased in both types of cells during 2-72 hr fasting parallel with decreasing serum insulin levels. 3. The protein content of the cells decreased and the DNA-based activity of acid phosphatase showed little change throughout fasting. The activity of cathepsin D increased during days 2 and 3 of food deprivation. 4. Vinblastine (50 mg/kg body wt) applied for the last 2 hr of different periods (2, 12, 24, 48 and 72 hr) of fasting decreased serum insulin level and increased the fractional cytoplasmic volume of autophagic vacuoles and dense bodies. This increase was smaller when the drug was applied shortly after feeding and much larger after prolonged fasting. The increase was more pronounced in the pancreatic than in the liver cells. 5. Our data show that the effect of vinblastine on the size of the autophagic-lysosomal compartment depends on the feeding status of the animals.     

Energy reserves during food deprivation and compensatory growth in juvenile roach: the importance of season and temperature

The effect of 21 days of starvation, followed by a period of compensatory growth during refeeding, was studied in juvenile roach Rutilus rutilus during winter and summer, at 4, 20 and 27° C acclimation temperature and at a constant photoperiod (12L : 12D). Although light conditions were the same during summer and winter experiments and fish were acclimated to the same temperatures, there were significant differences in a range of variables between summer and winter. Generally winter fish were better prepared to face starvation than summer fish, especially when acclimated at a realistic cold season water temperature of 4° C. In winter, the cold acclimated fish had a two to three‐fold larger relative liver size with an approximately double fractional lipid content, in comparison to summer animals at the same temperature. Their white muscle protein and glycogen concentration, but not their lipid content, were significantly higher. Season, independent of photoperiod or reproductive cycle, was therefore an important factor that determined the physiological status of the animal, and should generally be taken into account when fish are acclimated to different temperature regimes. There were no significant differences between seasons with respect to growth. Juvenile roach showed compensatory growth at all three acclimation temperatures with maximal rates of compensatory growth at 27° C. The replenishment of body energy stores, which were utilized during the starvation period, was responsible for the observed mass gain at 4° C. The contribution of the different energy resources (protein, glycogen and lipid) was dependent on acclimation temperature. In 20 and 27° C acclimated roach, the energetic needs during food deprivation were met by metabolizing white muscle energy stores. While the concentration of white muscle glycogen had decreased after the fasting period, the concentrations of white muscle lipid and protein remained more or less constant. The mobilization of protein and fat was revealed by the reduced size of the muscle after fasting, which was reflected in a decrease in condition factor. At 20° C, liver lipids and glycogen were mobilized, which caused a decrease both in the relative liver size and in the concentration of these substrates. Liver size was also decreased after fasting in the 4° C acclimated fish, but the substrate concentrations remained stable. This experimental group additionally utilized white muscle glycogen during food deprivation. Almost all measured variables were back at the control level within 7 days of refeeding.     

Food deprivation in the common vole (<Emphasis Type="Italic">Microtus arvalis</Emphasis>) and the tundra vole (<Emphasis Type="Italic">Microtus oeconomus</Emphasis>)

Arvicolinae voles are small herbivores relying on constant food availability with weak adaptations to tolerate prolonged food deprivation. The present study performed a comparative analysis on the responses to 4–18 h of food deprivation in the common vole (Microtus arvalis) and the tundra vole (Microtus oeconomus). Both species exhibited rapid decreases in the plasma and liver carbohydrate concentrations during phase I of fasting and the decline in the liver glycogen level was more pronounced in the tundra vole. The plasma thyroxine concentrations of the common vole decreased after 4 h. Lipid mobilization (phase II of fasting) was indicated by the increased plasma free fatty acid levels after 8–18 (the common vole) or 4–18 h (the tundra vole) and by the elevated lipase activities. In the tundra vole, the plasma ghrelin concentrations increased after 12 h possibly to stimulate appetite. Both species showed increased liver lipid concentrations after 4 h and plasma aminotransferase and creatine kinase activities after 12–18 h of food deprivation implying liver dysfunction and skeletal muscle damage. No signs of stimulated protein catabolism characteristic to phase III of fasting were present during 18 h without food.     

Metabolic response to starvation in late pregnant rats. II. Fetal response

Effect of prolonged maternal fasting on the fetal liver and heart glycogen and triglyceride content and on concentration of glucose, urea, uric acid and alpha amino-nitrogen in the amniotic fluid has been studied in rats. The animals were divided into four groups: fed (control), fasted for one day (from 20 to 21 day of pregnancy), fasted for two days (from 19 to 21 day) and fasted for three days (from 18 to 21 day). Maternal fasting for two and three days resulted in reduction in fetal growth. The fetal liver glycogen content was reduced already after one day of fasting, stabilized after two days and then further decreased after three days. The fetal heart glycogen content was reduced only after three days of fasting. The fetal liver triglyceride content increased gradually during the first two days of fasting and then stabilized. The content of triglycerides in the heart was elevated after two and three days of food deprivation. The amniotic fluid glucose concentration decreased after one day of fasting and then stabilized. Fasting did not effect the concentration of the nitrogenous compounds in the amniotic fluid. It is concluded that maternal fasting affects markedly metabolism of energy substrates stored in the fetal liver and the heart and the composition of the amniotic fluid.     

Structural and functional changes after the administration of tetrachlormethane in the liver of rats fed on diets with different protein contents

Morphological and biochemical changes characterizing the degree of liver damage and the development of liver repair were studied in rats fed 21 days on a low protein diet (LPD), a standard diet (SLD) and a high protein diet (HPD) and then given a single i.p. injection of tetrachlormethane (CCl4) in a dose of 0.75 ml/kg body weight. The HPD was found to increase sensitivity to CCl4, but it also promoted the liver repair process, as seen from the increment in liver DNA synthesis and the total DNA content of the liver, increased ploidy of the hepatocytes and growth of the size of their nuclei and of the hepatocytes themselves. An increase in the total surface area of the membranes of the granular endoplasmic reticulum and the inner and outer membrane of the mitochondria, but a decrease in the surface area of the membranes of the smooth endoplasmic reticulum, were also observed after the administration of CCl4. The LPD raised liver resistance to CCl4, but the development of liver repair activity differed from the process after the SLD and HPD, since polyploidy of the hepatocytes (especially the growth of octaploid cells) predominated and there was also an increase in the number of binuclear hepatocytes. Cell hypertrophy was expressed less in rats fed on the LPD than in animals given the HPD. As far as liver repair was concerned, the HPD showed no explicit advantages over the SLD.     

Short-term fasting induces intra-hepatic lipid accumulation and decreases intestinal mass without reduced brush-border enzyme activity in mink (<Emphasis Type="Italic">Mustela vison</Emphasis>) small intestine

For many mammalian species short-term fasting is associated with intestinal atrophy and decreased digestive capacity. Under natural conditions, strictly carnivorous animals often experience prey scarcity during winter, and they may therefore be particularly well adapted to short-term food deprivation. To examine how the carnivorous gastrointestinal tract is affected by fasting, small-intestinal structure, brush-border enzyme activities and hepatic structure and function were examined in fed mink (controls) and mink that had been fasted for 1–10 days. During the first 1–2 days of fasting, intestinal mass decreased more rapidly than total body mass and villus heights were reduced 25–40%. In contrast, tissue-specific activity of the brush-border enzymes sucrase, maltase, lactase, aminopeptidase A and dipeptidylpeptidase IV increased 0.5- to1.5-fold at this time, but returned to prefasting levels after 6 days of fasting. After 6–10 days of fasting there was a marked increase in the activity of hepatic enzymes and accumulation of intra-hepatic lipid vacuoles. Thus, mink may be a useful model for studying fasting-induced intestinal atrophy and adaptation as well as mechanisms involved in accumulation of intra-hepatic lipids following food deprivation in strictly carnivorous domestic mammals, such as cats and ferrets.Communicated by I.D. Hume     

Metabolic activity in isolated hepatocytes from cold exposed rats subjected to 24-hour fasting.

The aim of this work was to study the metabolic activity in isolated hepatocytes from control rats and rats exposed for 15 or 30 days to cold, all subjected to 24-h fasting. Hepatocyte oxygen consumption was used as an index of metabolic activity. The results show that 24-h fasting induces a decrease in energy expenditure at the level of the liver in cold-exposed rats but not in control animals.     

Correlation between plasma membrane surface area and transferrin secretion rate in isolated hepatocytes

It is generally considered that in exocytosis the size of the secreting cells does not increase when the membranes of exocytosis vesicles fuse with the plasma membrane. As the factors involved in the regulation of this phenomenon are poorly understood, we thought it worthwhile to investigate the relationship between the plasma membrane surface area and secretory activity. Isolated rat hepatocytes were prepared by liver collagenase perfusion. Secretion of the plasma protein, transferrin (Tf) was detected at the single cell level with specific anti-rat transferrin antibodies using the reverse hemolytic plaque test. Hepatocyte surface and hemolytic ring surface areas were calculated from diameters of hepatocyte and hemolytic plaque measured after 5h of incubation. A highly significant correlation was established between the plaque-forming hepatocyte surface areas and the corresponding hemolytic surface areas. This result was confirmed using an automatic image analysis method. Two-month-old rats were compared to 4-month-old rats. We observed that the ratio of the quantity of transferrin secreted by hepatocytes to the hepatocyte surface area was constant for a given incubation time, whatever the size of the hepatocytes. These results suggest that the plasma membrane surface area of hepatocytes may constitute a limiting factor in Tf secretion.     

Effect of a short-term fasting and altitude hypoxia on the cytochrome oxidase activity of rat brain mitochondria

The effect of short-term fasting and thirst, prolonged fasting and hypoxic hypoxia upon the activity of cytochrome oxidase was studied in mitochondrial fractions obtained from the brain and the liver. The investigation was carried out in two groups of rats, 5 and 60 days old. a) The activity of cytochrome oxidase in mitochondria isolated from the brain cortex, subcortical regions and the medulla oblongata rises, while the changes in liver mitochondrial fractions are reverse. b) A significant increase of mitochondrial cytochrome oxidase was found in 5-day-old rats after both types of fasting and hypoxia in all regions of the brain, as well as in the liver. c) The cytochrome oxidase activity in brain and liver mitochondria of 60-day-old rats was not affected appreciably after 24 h nutritional deprivation, with the exception of a significant rise of activity in the medulla oblongata. Prolonged fasting and hypoxia again markedly increased the activity of this enzyme in all regions of the brain and in the liver.     

Cytological analysis of the intact and regenerating liver of different strains of rats]

Comparative cytological studies were conducted on control and regenerating liver of two strains (August and Cotton) of rats, 2/3 of the liver was resected; 5 or 6 animals were sacrificed at each of the following postoperative periods: in 30 hours, 3, 8, 42 and 120 days. The number of binuclear cells, the size of mononuclear hepatocytes and their nuclei, the mitotic activity, and ploidity of hepatocytes were determined. The intact and regenerating liver of the August rats differed from the intact and regenerating liver of the Cotton rats by a number of cytological indices, excluding the mitotic activity. A conclusion was drawn that the observed interstrain differences in the cytological indices providing regeneration of the liver after resection in the August and Cotton rats depended on the genotype of the given strain.     

Metabolic responses to prolonged starvation, food restriction, and refeeding in the brown trout, Salmo trutta: oxidative stress and antioxidant defenses

The effects of long-term starvation and food restriction (49 days), followed by refeeding (21 days) have been studied with respect to antioxidant defense in the liver and gills (branchial tissues) of the brown trout, Salmo trutta. Malondialdehyde levels in both tissues increased in parallel with starvation and food restriction and these values did not return to normal after the refeeding period. The activities of superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx), and glutathione reductase (GR) in liver and gills increased during the 49 days of starvation, but glucose-6-phosphate dehydrogenase (G6PD) activities decreased. Glutathione S-transferase (GST) activity decreased in the liver at the 49th day of starvation, but increased in the branchial tissues. Some of the antioxidant enzyme activities (such as hepatic GST and branchial G6PD) returned to control values of fed fish after the refeeding period, but others (e.g. hepatic SOD and branchial GPx) did not return to normal values. In conclusion, our study indicates that total or partial food deprivation induces oxidative stress in brown trout.