Leptin concentrations in plasma and follicular fluid from prepubertal gilts as influenced by fasting, refeeding and insulin.

This study's aim was to examine whether fasting and refeeding would influence leptin levels in both plasma and follicular fluid from prepubertal gilts, and whether insulin affects leptin levels in fasting gilts. In experiment 1, four gilts were fasted for 72 h and then refed. Blood samples were withdrawn during normoalimentation, at the end of fasting, and for 4 h after refeeding. All samples were assayed for leptin; alternate samples were assayed for insulin, glucose and non-esterified fatty acids (NEFA). Fasting caused a decrease in leptin, glucose and insulin levels in plasma, while NEFA concentrations increased. In experiment 2, four gilts were given insulin as a bolus (0.2 IU/kg body weight) after 68 h of fasting. Blood samples were collected every 15 min around insulin administration and were assayed for leptin, insulin and glucose. This experiment shows that insulin administration increases leptin levels during fasting. In experiment 3, gilts were ovariectomized during normal alimentation (n=4), after 48 h of fasting (n=4), and after 48 h of realimentation following 48 h of fasting (n=4). Leptin levels in both plasma and follicular fluid collected after 48 h of fasting were significantly lower than those observed during normoalimentation or refeeding. In conclusion, a transient increase in insulin during fasting is effective in restoring leptin concentrations; in addition, leptin levels in follicular fluid parallel those in plasma.     

Differential time course of liver and kidney glucose-6 phosphatase activity during long-term fasting in rat correlates with differential time course of messenger RNA level

We have studied the role of Glc6Pase mRNA abundance in the time course of Glc6Pase activity in liver and kidney during long-term fasting in rat. Refered to the mRNA level in the fed state, Glc6Pase mRNA abundance was increased by 3.5 ± 0.5 and 3.7 ± 0.5 times (mean ± S.E.M., n = 5) in the 24 h and 48 h-fasted liver, respectively. Then, the liver Glc6Pase mRNA was decreased to the level of the fed liver after 72 and 96 h of fasting (1.0 ± 0.3 and 1.4 ± 0.3). In the kidney, Glc6Pase mRNA abundance was increased by 2.7 ± 1.0 and 5 ± 1.2 times at 24 and 48 h of fasting, respectively. Then, it plateaued at the level of the 48 h fasted kidney after 72 h and 96 h of fasting (4.5 ± 1.0 and 4.3 ± 1.0). After 24 and 48 h-refeeding, the abundance of Glc6Pase mRNA in 48 h-fasted rats was decreased to the level found in the liver and kidney of fed rats. The time course of the activity of Glc6Pase catalytic subunit during fasting and refeeding was strikingly parallel to the time course of Glc6Pase mRNA level in respective tissues. These data strongly suggest that the differential expression of Glc6Pase activity in liver and kidney in the course of fasting may be accounted for by the respective time course of mRNA abundance in both organs.Abbreviations Glc6Pase Glucose-6 phosphatase - GNG Gluconeogenesis     

Requirement of PPARalpha in maintaining phospholipid and triacylglycerol homeostasis during energy deprivation

The peroxisome proliferator-activated receptor alpha (PPARalpha) has been implicated as a key control of fatty acid catabolism during the cellular fasting. However, little is known regarding changes of individual fatty acids in hepatic triacylglycerol (TG) and phospholipid (PL) as a result of starvation. In the present work, the effects of 72 h fasting on hepatic TG and PL fatty acid profiles in PPARalpha-null (KO) mice and their wild-type (WT) counterparts were investigated. Our results indicated that mice deficient in PPARalpha displayed hepatomegaly and hypoketonemia following 72 h starvation. Histochemical analyses revealed that severe fatty infiltration was observed in the livers of KO mice under fasted conditions. Furthermore, 72 h fasting resulted in a 2.8-fold higher accumulation of hepatic TG in KO mice than in WT mice fasted for the same length of time. Surprisingly, the total hepatic PL contents in fasted KO mice decreased by 45%, but no significant change in hepatic PL content was observed in WT mice following starvation. Gas chromatographic analysis indicated that KO mice were deprived of arachidonic (20:4n-6) and docosahexaenoic (22:6n-3) acids during fasting. Taken together, these results show that PPARalpha plays an important role in regulation of fatty acid metabolism as well as phospholipid homeostasis during energy deprivation.     

禁食状态下MED1肝脏特异性敲除鼠呈现高脂血症

为研究MED1如何影响血浆脂质代谢,以MED1肝脏特异性敲除鼠 (MED1ΔLiv) 为动物模型,禁食0、24、48、72 h,通过H&E染色切片观察了MED1ΔLiv鼠肝脏的形态学变化,运用甘油三酯和胆固醇酶试剂盒及FPLC方法分析了MED1ΔLiv和对照鼠 (MED1fl/fl) 血浆甘油三酯和胆固醇的水平以及脂蛋白的分布情况。研究结果显示,禁食72 h,与MED1fl/fl和PPARα?/?对照鼠相比,MED1ΔLiv鼠肝脏无脂肪沉积。禁食24、48、72 h,与MED1fl/fl鼠相比,MED1Δ     

δ-Aminolevulinic acid synthetase: Regulation of activity in various tissues of the aging rat

The basal and ethanol-induced activities of the rate-limiting enzyme of heme biosynthesis, δ-aminolevulinic acid (ALA) synthetase were measured in the liver, heart, kidney, and brain of young, adult, and aged Sprague-Dawley rats. When assayed in whole mitochondria derived from either fed or 24-h fasted animals, the basal levels of hepatic ALA synthetase activity decreased dramatically as a function of age. An equivalent decrease was seen in the ethanol-induced activity although the ratio of induced to basal activities did not change with age. In the heart, ALA synthetase activity also decreased significantly during aging. The activity was not induced by ethanol and was decreased markedly by fasting. By contrast, kidney ALA synthetase activity showed no age-related changes. The activity was unaffected by fasting and showed a variable induction response to ethanol. Brain ALA synthetase activity displayed a significant age-dependent decrease in its activity which was neither affected by fasting nor sensitive to induction by ethanol. The data presented are consistent with the hypothesis that ALA synthetase activity is subject to metabolic regulation. Further, they indicate that while the enzyme activity is regulated in a tissuespecific manner, a time-dependent decrease is a general feature of the aging animal.     

Regulation of acetyl-coenzyme A carboxylase. II. Effect of fasting and refeeding on the activity, phosphate content, and aggregation state of the enzyme

Acetyl-CoA carboxylase isolated from freeze-clamped livers of fed rats has relatively low phosphate content (5.0 mol of Pi/mol of subunit) and high specific activity (3.5 units/mg in the absence of citrate). The enzyme from rats fasted for 12, 18, 24, and 48 h exhibited decreasing specific activities of 2.75, 1.85, 1.7, and 0.9 units/mg, respectively. Citrate activated all preparations of carboxylase, with most activation observed with the least active preparation. There was no significant change in the sensitivity of the enzyme to citrate since half-maximal activation was observed at 0.2 mM for carboxylase from fed as well as fasted rats. With the decrease in activity as a function of fasting, there was a concomitant increase in the phosphate content of carboxylase, with values of 5.3, 5.6, 6.7, and 7.6 mol of Pi/mol of subunit obtained for preparations from rats fasted for 12, 18, 24, and 48 h, respectively. Refeeding the fasted rats resulted in increased specific activity of carboxylase (3.4 units/mg) and decreased phosphate content (5.1 mol of Pi/mol of subunit). Moreover, dephosphorylation by [acetyl-CoA carboxylase]-phosphatase 2 activated the carboxylase from 48-h fasted rats to a value of 2.9 units/mg, assayed in the absence of citrate, indicating that the low activity of carboxylase from fasted rats was due to its increased phosphate content. Superose 6 chromatography showed that the enzyme exists in two polymeric forms, a highly active polymer of greater than or equal to 40 subunits and less active octamer. The former predominates in livers of fed rats, whereas the latter predominates in livers of fasted rats. The octamer could be converted to the highly active polymer by dephosphorylation. These observations indicate that fasting/refeeding results in phosphorylation/dephosphorylation of acetyl-CoA carboxylase with concomitant depolymerization/polymerization of the protein and ultimately decreasing or increasing its specific activity.     

Hormonal and nutritional regulation of muscle carnitine palmitoyltransferase I gene expression in vivo

Transgenic mice carrying the human heart muscle carnitine palmitoyltransferase I (M-CPTI) gene fused to a CAT reporter gene were generated to study the regulation of M-CPTI gene expression. When the mice were fasted for 48 h, CAT activity and mRNA levels increased by more than 2-fold in heart and skeletal muscle, but not liver or kidney. In the diabetic transgenic mice, there was a 2- to 3-fold increase in CAT activity and CAT mRNA levels in heart and skeletal muscle which upon insulin administration reverted to that observed with the control insulin sufficient transgenic mice. Feeding a high fat diet increased CAT activity and mRNA levels by 2- to 4-fold in heart and skeletal muscle of the transgenic mice compared to the control transgenic mice on regular diet. Overall, the M-CPTI promoter was found to be necessary for the tissue-specific hormonal and dietary regulation of the gene expression.     

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.     

Growth hormone stimulates the peripheral conversion of thyroxine into triiodothyronine by increasing the liver 5'-monodeiodinase activity in the fasted and normal fed chicken

Normal fed and 2 days fasted Warren chickens were injected intravenously with 100 micrograms of ovine growth hormone (GH) and ovine prolactin and plasma concentrations of thyroid hormones were assayed prior and up to 2 h after injection. Fasting alone decreases T3, but increases T4. An injection of GH resulted in increases of plasma T3 concentrations in two fasting experiments by 40% (after 3/4 h) and 104% (after 1 h). In normal fed animals no increase is observed in the first experiment, whereas a 35% increase occurs in the second one. An injection of 100 micrograms prolactin does not influence T3 in normal fed or fasting animals. Both GH and prolactin, however, may decrease plasma concentrations of T4. In a separate experiment 50 micrograms and 200 micrograms of GH raised the decreased T3 levels after fasting by 39% and 60% respectively 1 h after injection and by 24 and 61% respectively in normal fed chicken, whereas prolactin was ineffective in this regard. Using Hisex chickens, the influence of an injection of 100 micrograms GH on plasma concentrations of thyroid hormones could be confirmed. At the same time GH increases the liver 5'-monodeiodinase activity by 330% after 1 h and by 147% after 2 h. The peroxidase activity is not influenced in normal fed chickens, but GH decreases this activity in food deprived animals after 1 h and 2 h. It is concluded that ovine GH, but not prolactin, stimulates the peripheral conversion of T4 into T3 in both normal fed and food deprived chicken and that this effect is dose dependent.     

Metabolic changes in Brycon cephalus (Teleostei,Characidae) during post-feeding and fasting

Metabolic changes during the transition from post-feeding to fasting were studied in Brycon cephalus, an omnivorous teleost from the Amazon Basin in Brazil. Body weight and somatic indices (liver and digestive tract), glycogen and glucose content in liver and muscle, as well as plasma glucose, free fatty acids (FFA), insulin and glucagon levels of B. cephalus, were measured at 0, 12, 24, 48, 72, 120, 168 and 336 h after the last feeding. At time 0 h (the moment of food administration, 09.00 h) plasma levels of insulin and glucagon were already high, and relatively high values were maintained until 24 h post-feeding. Glycemia was 6.42+/-0.82 mM immediately after food ingestion and 7.53+/-1.12 mM at 12 h. Simultaneously, a postprandial replenishment of liver and muscle glycogen reserves was observed. Subsequently, a sharp decrease of plasma insulin occurred, from 7.19+/-0.83 ng/ml at 24 h of fasting to 5.27+/-0.58 ng/ml at 48 h. This decrease coincided with the drop in liver glucose and liver glycogen, which reached the lowest value at 72 h of fasting (328.56+/-192.13 and 70.33+/-14.13 micromol/g, respectively). Liver glucose increased after 120 h and reached a peak 168 h post-feeding, which suggests that hepatic gluconeogenesis is occurring. Plasma FFA levels were low after 120 and 168 h and increased again at 336 h of fasting. During the transition from post-feeding to fast condition in B. cephalus, the balance between circulating insulin and glucagon quickly adjust its metabolism to the ingestion or deprivation of food.     

Mitochondrial pyruvate metabolism in liver and kidney during acidosis

Pyruvate transport and carboxylation have been determined in mitochondria from liver and kidney cortex isolated from Wistar rats with acidosis produced by three different treatments: fasting, exercise and ingestion of ammonium chloride. Fasting for 48 h or swimming for 2 h resulted in an increased rate of CO₂ fixation by mitochondria from both organs incubated with pyruvate. This increase was accompanied by a rise in the rate of pyruvate transport in all cases except in mitochondria derived from the kidney of the fasted animals. Acute acidosis produced by the ingestion of ammonium chloride resulted in increases in pyruvate transport and carboxylation in kidney mitochondria, but a drop in pyruvate carboxylation was observed in mitochondria from the liver. The results are discussed in terms of the differential regulation of the mitochondria steps for gluconeogenesis from three carbon precursors in liver and kidney, taking into consideration the hormonal status of the animals and the prevailing available substrates in each condition.     

Turnover of liver pyruvate kinase

The relative rates of synthesis and degradation for liver pyruvate kinase have been determined in rats fed standard lab chow, fasted, and refed a high-carbohydrate-low-protein diet. Relative rates of synthesis and apparent rates of degradation were determined by pulse-labeling the enzyme in vivo with l-[4,5-³H]leucine and by measuring the incorporation of radioactivity into liver pyruvate kinase after quantitative precipitation of the enzyme with anti-liver pyruvate kinase immunoglobulin. The relative rate of synthesis decreased approximately 75% upon fasting and then increased 20- to 30-fold upon refeeding the high-carbohydrate diet. The apparent half-lives for liver pyruvate kinase in fasted, control, and refed animals are very similar (55, 59, and 47 h, respectively). Thus, the nutritional alterations in the levels of liver pyruvate kinase seem to result primarily from alterations in the rate of enzyme synthesis.     

Hormonal and metabolic response to physical exercise, fasting and cold exposure in the rat. Effects on ketogenesis in isolated hepatocytes

Four groups of rats were subjected to the following conditions: (1) 48 h fasting, (2) 48 h of 4 degrees C cold exposure, (3) 5 h treadmill running, (4) 48 h fasting with 4 degrees C cold exposure. The groups were compared to fed control rats in order to study hormonal and metabolic responses in blood and tissue samples. Isolated hepatocytes were used to evaluate the rate of ketogenesis. Decreases in liver glycogen and increases in blood free fatty acids (FFA) confirmed that glycogenolysis and lipolysis occur in these situations of metabolic stress. Increases in the glucagon/insulin plasma ratio were also noted. Plasma catecholamine levels were only enhanced after running and after cold exposure. Production of blood ketone bodies was stimulated more by running and by fasting than by cold exposure. The low ketone body production observed after cold exposure seems to be linked to increases liver glycogen levels and decreased FFA availability. Liver cells isolated after cold exposure exhibited higher ketogenesis than these isolated after running. This difference in ketogenic capacity could result both from the longer hormonal stimulation by high glucagon/insulin plasma ratios and from the metabolic state of the liver.     

Neuropeptide Y content in the hypothalamic paraventricular nucleus responds to fasting and refeeding in broiler chickens

To examine the neural mechanism by which hypothalamic neuropeptide Y (NPY) regulates energy homeostasis and feeding behavior in commercial broilers, we measured NPY content in several hypothalamic regions of birds that were fasted and then refed. After fasting for 48 and 72 h, body weight significantly decreased, and food intake significantly increased during the subsequent refeeding. The lost body weight was not restored to ad libitum feeding levels even after 3 days of refeeding. Plasma glucose concentration and body fat content significantly decreased and plasma non-esterified fatty acid (NEFA) concentration significantly increased after 48- and 72-h fasting. Refeeding for 24 h restored plasma metabolites and body fat content to pre-fasting levels. NPY content in the paraventricular nucleus (PVN) and infundibular nucleus significantly increased during fasting, and NPY content of the PVN was restored to pre-fasting levels after 24-h refeeding. However, there was no significant change in the NPY content of the lateral hypothalamic area during fasting or refeeding. The present results of changes in the hypothalamic NPY content during fasting and refeeding support the hypothesis that NPY plays a central role in regulation of energy homeostasis, with especially important effect on feeding behavior and body weight in broiler chickens.     

Pulmonary responses to acute ozone exposure in fasted mice: effect of leptin administration.

Leptin is a satiety hormone that also has proinflammatory effects, including augmentation of ozone-induced pulmonary inflammation. The purpose of this study was to determine whether reductions in endogenous levels of leptin can attenuate pulmonary responses to ozone. To reduce serum leptin, we fasted mice overnight before ozone exposure. Fasting caused a marked reduction in serum leptin to approximately one-sixth the levels observed in fed mice, and continuous infusion of leptin via Alzet micro-osmotic pumps restored serum leptin to, but not above, fed levels. Ozone exposure (2 ppm for 3 h) caused a significant, approximately 40% increase in pulmonary resistance (P < 0.01) and increased airway responsiveness in fasted but not in fed mice. The increased effect of ozone on pulmonary mechanics and airway responsiveness in fasted mice was not observed when leptin was restored via continuous infusion. Ozone exposure caused pulmonary inflammation, as evident by increases in bronchoalveolar lavage cells, protein, and soluble tumor necrosis factor receptors. There was no effect of fasting status on ozone-induced changes in the bronchoalveolar lavage inflammatory profile, and leptin treatment did not alter these responses. Our results indicate that fasting augments ozone-induced changes in pulmonary mechanics and airway responsiveness in mice. These effects of fasting are the result of declines in serum leptin. The mechanistic basis for this protective effect of leptin in fasted mice remains to be determined but is not related to effects on ozone-induced inflammation.     

The effect of nutritional state and hormones on the number of circulating red cells in Gallus domesticus

When chickens are fasted, the liver becomes dark, whereas the spleen is bleached. Microscopic and spectrophotometric analysis indicated the colour resulted from red blood cells accumulating in enlarged sinusoids of the liver. In birds fasted for 48 hr the hematocrit increases 30%. Blood volume and red cell volume are not altered; the extra cells are not newly synthesised. The extra cells arise from the spleen, possibly under alpha-adrenergic control if given epinephrine by injection and under beta-adrenergic control during fasting as indicated by experiments with alpha and beta-receptor blocking agents.     

Behavior of L-threonine-degrading enzymes during liver regeneration

We examined the effects of a two-thirds hepatectomy in the adult rat on the activities of the three L-threonine-degrading enzymes, L-threonine dehydratase, L-threonine aldolase and L-threonine dehydrogenase. Noticeable variations were observed which did not occur in either sham-operated or turpentine-treated rats and were not linked to food intake. They were considered specific to the regenerating liver. When the reactions were followed in vitro, L-threonine deaminase and L-threonine aldolase were significantly lower for the first 12-24 h: L-threonine dehydrogenase decreased only after 48 h. These results are linked to a decrease in the enzyme concentration in the tissue. L-Serine and L-threonine liver concentrations increased 2-3-fold during the same periods. When the activities were evaluated in vivo, the levels of the first two enzymes remained constant for 24 h, but increased after 48 h; L-threonine dehydrogenase increased between 12 and 48 h. The in vivo activity of the enzymes was reflected by total L-threonine degradation, which had a single sharp peak at 48 h. The asynchronous variations in enzyme activity are related to the differences in protein metabolism which occur in the regenerating liver, and are the consequence of a new transient differential control. The changes observed are significant in liver regeneration; they regulate the consumption and the serum and liver levels of L-serine and L-threonine, setting them aside for protein synthesis. They minutely control the flux of amino acids toward gluconeogenesis, since, during the first 48 h after partial hepatectomy, the production of glucose is ensured principally by lactate; the contribution of L-threonine seems to be more significant only at 48 h. These findings are useful in the study of the regulation of the enzymes involved in amino acid metabolism during liver regeneration.     

Phosphorylase phosphatase activities of rat liver in streptozotocin-diabetes

Protein phosphatase-1 and 2A, accounting for all the hepatic activity regulating phosphorylase, were assayed in streptozotocin-induced (8 weeks) diabetic Wistar rats. Cytosolic protein phosphatase-1 and 2A were distinguished by chromatography on heparin-Sepharose and by inhibition with inhibitor-2. Approx. 25-35% increases in type-1 phosphorylase phosphatase activity measured in cytosols were registered in diabetic rats when compared with control and 24 h fasting animals. The enrichment of protein phosphatase-1 in the cytosol of streptozotocin-treated rat livers could not be attributed to the reduced glycogen content with the onset of diabetes, since this elevated level of type-1 phosphatase was not observed in fasting rats with low glycogen content. The translocation of type-1 phosphatase from the particulate fraction into the cytosol was also recorded in trypsin-treated samples of diabetic rat livers. The apparent molecular weight of type-1 phosphatase in the cytosol of control and fasted rats was 160,000 as judged by gel filtration. The type-1 phosphatase activity that was released from the particulate fraction by streptozotocin-induced diabetes identified a further enzyme species (Mr 110,000) in the cytosol. Our data imply that the higher levels of cytosolic protein phosphatase-1 in diabetic rat liver could be a consequence of the dissociation of the catalytic subunit of protein phosphatase-1 and the glycogen-binding subunit in rat livers.     

The effect of fasting on the activation in vivo of the insulin receptor kinase.

Fasting causes insulin resistance in liver and fat, and increases insulin sensitivity in muscle. We studied the response in vitro and in vivo to insulin of the insulin receptor tyrosine kinase in muscle and liver from 72 h fasted and control rats. Insulin was injected intraperitoneally together with glucose, and blood and tissue samples were obtained 0, 5, 15 and 30 min later. Basal serum glucose and insulin levels were significantly higher in control than in fasting rats. Serum glucose rose to approximately 300 mg/dl at 5 min and then progressively declined without hypoglycaemia. Receptors were prepared from whole tissue by wheat germ lectin affinity chromatography. 125I-insulin binding to purified receptors was increased by fasting in both muscle (18%) and liver (50%). In untreated fasting and control animals, muscle and liver insulin receptor tyrosine kinase activity was stimulated to similar levels by insulin added in vitro. With only insulin treatment in vivo, muscle receptor tyrosine kinase behaved similarly in fasting and control animals with maximal activation at 15 min post injection. In liver, insulin in vivo stimulated receptor tyrosine kinase activity maximally at 5 min post injection in both fasting and control, but in fasting animals the treatment in vivo caused a significantly larger and more prolonged activation of the enzymic activity, possibly due to a decrease in the rate of dephosphorylation and deactivation of the beta subunits.