Prolonged effect of single carnitine administration on fasted carnitine-deficient JVS mice regarding their locomotor activity and energy expenditure

Carnitine is an essential cofactor for the oxidation of fatty acid in the mitochondria and an efficient therapeutics for primary carnitine deficiency. We herein analyzed the prolonged effects of carnitine on the reduced locomotor activity and energy metabolism of fasted carnitine-deficient juvenile visceral steatosis (jvs(-/-)) mice. We found that a single carnitine administration to 24-h fasted jvs(-/-) mice in the morning increased both the locomotor activity and oxygen consumption at night not only on the same day, but also on the next day, when the carnitine levels in the blood and tissues were already as low as at the original carnitine-deficient state. We also found that fat utilization for energy production significantly increased under fasting even in jvs(-/-) mice and was stimulated in the carnitine-administrated fasted jvs(-/-) mice at night, in comparison to that observed in the saline-administered jvs(-/-) mice, at least for 2 days even under the low plasma and tissue carnitine levels. These results suggest that the low tissue carnitine levels are therefore not the sole rate-limiting factor of general fatty acid oxidation in carnitine-deficient jvs(-/-) mice.     

Short-term fasting leads to inhibition of responsiveness to LH-stimulated testosterone secretion in the adult male bonnet monkey

A variety of stressors including fasting profoundly inhibit reproductive function in mammals. Although the effect of short-term fasting on gonadotropic axis is well established, the direct effects of fasting on gonads have not been reported. The objectives of the present experiments were to examine the effect of short-term fasting on circulating luteinizing hormone (LH) and testosterone (T) secretion, and to determine the responsiveness of testis to exogenous recombinant human (rh) LH treatment in male bonnet monkeys. In addition, an experiment was carried out to examine whether brief inhibition of endogenous LH secretion causes alteration in testicular responsiveness. Adult male monkeys were fasted for 1 day for examining the circulating endocrine hormone concentrations and challenged with rhLH injection 1 day after fasting. Food withdrawal for 1 day resulted in significant (P<0.05) decrease in LH, T and increase in cortisol concentrations. Surprisingly, T secretion in response to direct stimulation of Leydig cells by LH was not observed in fasted monkeys. In fed monkeys, treatment with Antide (a specific gonadotropin releasing hormone receptor antagonist to inhibit pituitary LH secretion) for 1 day did not compromise T secretion stimulated by rhLH, suggesting that loss of responsiveness of testis to exogenous LH treatment in fasted monkeys was not because of interruption in pituitary LH stimulation of the testis. The results indicate that short-term fasting in adult male monkeys cause inhibition of LH and T secretion, and inhibition of responsiveness of testis to LH stimulation.     

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     

Altered distribution and toxicity of digitoxigenin in fasted mice

Intravenous administration of digitoxigenin (DTXGN) evokes seizure episodes in mice which may be dependent on brain biogenic amines such as serotonin (5-HT). Fasting is known to have effects on both drug toxicity and brain 5-HT synthesis. The purpose of this study was to assess the effects of overnight fasting on DTXGN toxicity. The i.v. LD-50 of DTXGN was increased by 61% in fasted mice. Adjustment of DTXGN dose for the decrease in body weight of fasted mice did not alter the fasting induced protection. A loading dose of 1-tryptophan (25 mg/kg, i.p.) did not alter mortality rates in either fed or fasted mice. Cortical levels of 3H-DTXGN were decreased significantly by 25% in fasted mice. Liver and blood levels were elevated significantly. These data suggest that decreased DTXGN toxicity is associated with a decrease in its distribution to the cerebral cortex and emphasize the importance of acute dietary status in the expression of drug toxicity.     

Absence of effects of short-term fasting on plasma ghrelin and brain expression of ghrelin receptors in the tilapia, Oreochromis mossambicus

Ghrelin is an important endocrine peptide that links the gastrointestinal system and brain in the regulation of food intake and energy expenditure. In human, rat, and goldfish plasma levels of ghrelin and GH are elevated in fasted animals, suggesting that ghrelin is an orexigenic signal and a driving force behind the elevated plasma levels of GH during fasting. Ghrelin's orexigenic action is mediated by the ghrelin receptor (GHS-R1a and GHS-R1b) which is localized on neuropeptide Y (NPY) neurons in the brain. Studies were undertaken to investigate the effect of short-term fasting on plasma ghrelin and brain expression of GHS-R1a, GHS-R1b, and NPY in the tilapia. Fasting for 7 days had no effect on plasma ghrelin concentrations, whereas significant increases in plasma levels of GH were observed on day 3. Fasting significantly reduced plasma levels of IGF-I on days 3 and 7, and of glucose on days 3, 5, and 7. Brain expression of ghrelin and GHS-R1b were significantly elevated in fasted fish on day 3, but were significantly reduced on day 5. This reduction was likely due to a significant increase in the expression in the fed controls on day 5 compared to day 0. No change was detected in the expression of GHS-R1a or NPY in the brain. These results indicate that ghrelin is not acting as a hunger signal in short-term fasted tilapia and is not responsible for the elevated levels of plasma GH.     

Forkhead Box O1 (FOXO1) Protein,but Not p53, Contributes to Robust Induction of p21 Expression in Fasted Mice

Reporter mice that enable the activity of the endogenous p21 promoter to be dynamically monitored in real time in vivo and under a variety of experimental conditions revealed ubiquitous p21 expression in mouse organs including the brain. Low light bioluminescence microscopy was employed to localize p21 expression to specific regions of the brain. Interestingly, p21 expression was observed in the paraventricular, arcuate, and dorsomedial nuclei of the hypothalamus, regions that detect nutrient levels in the blood stream and signal metabolic actions throughout the body. These results suggested a link between p21 expression and metabolic regulation. We found that short-term food deprivation (fasting) potently induced p21 expression in tissues involved in metabolic regulation including liver, pancreas and hypothalamic nuclei. Conditional reporter mice were generated that enabled hepatocyte-specific expression of p21 to be monitored in vivo. Bioluminescence imaging demonstrated that fasting induced a 7-fold increase in p21 expression in livers of reporter mice and Western blotting demonstrated an increase in protein levels as well. The ability of fasting to induce p21 expression was found to be independent of p53 but dependent on FOXO1. Finally, occupancy of the endogenous p21 promoter by FOXO1 was observed in the livers of fasted but not fed mice. Thus, fasting promotes loading of FOXO1 onto the p21 promoter to induce p21 expression in hepatocytes.     

Effect of 2-Amino-7-Phosphonoheptanoic Acid on Regional Brain Amino Acid Levels in Fed and Fasted Rodents

Abstract: 2-Amino-7-phosphonoheptanoic acid, an antagonist of excitation caused by dicarboxylic amino acids with a selective action on N -methyl-d-aspartate receptors, has been administered in an anticonvulsant dose (1 mmol/kg i.p.) to fed or fasted rats and mice. The drug impaired motor activity in fasted mice. Glucose and amino acids were determined in dissected regions of brain fixed by microwave irradiation. Glucose content was low in the brains of fasted rats and mice but was restored to normal (fed) concentration 45 min after the administration of 2-amino-7-phosphonoheptanoic acid in fasted mice. In fed animals, 2-amino-7-phosphonoheptanoic acid did not change brain aspartate concentration. In fasted animals, aspartate concentration was raised in most brain regions. In fasted rats and mice, 2-amino-7-phosphonoheptanoic acid significantly increased glutamine in rat cortex and mouse striatum, decreased glutamate content in rat striatum, and decreased aspartate concentration in all regions except mouse cortex and striatum. GABA levels were significantly decreased in rat striatum and hippocampus. These changes are consistent with an increased synaptic release of glutamate and aspartate following blockage of their post-synaptic action at selected sites.     

Effect of fasting on islet lysosomal enzyme activities and the in vivo insulin response to different secretagogues

The effect of a 24 hr starvation period on islet lysosomal enzyme activities and the in vivo insulin response to glucose, glibenclamide and L-isopropyl-noradrenaline (L-IPNA) was studied in mice. It was observed that fasting induced a significant decrease of islet acid amyloglucosidase activity, whereas the activities of acid phosphatase, beta-N-acetyl-glucosaminidase, and beta-glucuronidase in islet tissue were unaffected by the fasting period studied. Starvation markedly reduced the acute insulin response to a maximal dose of glucose or glibenclamide. However, the insulin response to a maximal dose of L-IPNA was of similar magnitude in both fed and fasted animals. Pretreatment of fasted mice with purified fungal acid amyloglucosidase could restore the impaired insulin response to glucose to the normal level seen in fed mice. It is suggested that islet acid amyloglucosidase activity is of importance for glucose-stimulated insulin secretion, and that reduced levels of islet amyloglucosidase may contribute to the impairment of glucose-induced insulin release seen after fasting.     

Activation of glycogen synthase in myocardium induced by intermittent hypoxia is much lower in fasted than in fed rats

Obstructive sleep apnea is characterized by intermittent obstruction of the upper airway, which leads to intermittent hypoxia. Myocardial glycogen is a major energy resource for heart during hypoxia. Previous studies have demonstrated that intermittent hypoxia rapidly degrades myocardial glycogen and activates glycogen synthase (GS). However, the underlying mechanisms remain undefined. Because sleep apnea/intermittent hypoxia usually happens at night, whether intermittent hypoxia leads to GS activation in the postabsorptive state is not known. In the present study, male adult rats were studied after either an overnight fast or ad libitum feeding with or without intermittent ventilatory arrest (3 90-s periods at 10-min intervals). Hearts were quickly excised and freeze-clamped. Intermittent hypoxia induced a significant decrease in myocardial glycogen content in fed rats and stimulated GS in both fasted and fed rats. However, the portion of GS in the active form increased by approximately 38% in fasted rats compared with a larger, approximately 130% increase in fed rats. The basal G-6-P content was comparable in fasted and fed animals and increased approximately threefold after hypoxia. The basal phosphorylation states of Akt and GSK-3beta and the activity of protein phosphatase 1 (PP1) were comparable between fasted and fed control rats. Hypoxia significantly increased Akt phosphorylation and PP1 activity only in fed rats. In contrast, hypoxia did not induce significant change in GSK-3beta phosphorylation in either fasted or fed rats. We conclude that hypoxia activates GS in fed rat myocardium through a combination of rapid glycogenolysis, elevated local G-6-P content, and increased PP1 activity, and fasting attenuates this action independent of local G-6-P content.     

Schistosoma mansoni, S. japonicum, S. haematobium: glycogen content and glucose uptake in parasites from fasted and control hosts

The glycogen content of male and female Schistosoma mansoni has been measured in flukes from normally fed hosts and those from fasted hosts. In infections from both the mouse and the hamster, a significant reduction in schistosomal glycogen of males is seen hours after food is withdrawn from the host. Reductions in protein content of the schistosomes were only observed in hamster infections fasted at least 72 hr. The livers of infected mice not only decrease in size during fasting, but there is a concomitant reduction in glycogen per unit wet weight. Comparisons of glycogen:protein ratios of mansonian males, females, and host livers indicate that the fasting-induced loss of liver glycogen is also observed in the male schistosome, but not the female. Studies of both S. mansoni and S. haematobium pairs from fed hosts suggest that the ratio of glycogen:protein contents in the male schistosome correlates with the glycogen:protein ratio of the female partner. Measurements of glucose uptake in vitro suggest that greater uptake rates may be observed in flukes perfused from fasted hosts. In S. japonicum from infected mice, a reduction in male glycogen was also detected as early as after a 6-hr fasting period, but changes in the females were not significant. Unmated male S. japonicum also exhibit a reduction in glycogen levels after fasting, but the quantity of worm glycogen present in these males remains higher than comparable mated males. In mice entrained to a regulated pattern of available food, fluctuations in glycogen content of the male schistosomes were observed, but in the female partners fluctuations were of a smaller magnitude.(ABSTRACT TRUNCATED AT 250 WORDS)     

THE EFFECT OF FASTING ON THE METABOLISM OF 5-HYDROXYTRYPTAMINE AND DOPAMINE IN THE BRAIN OF THE MOUSE

Abstract— The turnover of 5-hydroxytryptamine in the forebrain and of dopamine in the striatum was studied in mice fasted for 20 h. Such mice showed an increased tissue concentration of 5-hydroxyindoleacetic acid in the forebrain and an increased accumulation of this acid after probenecid. Fasted mice also showed a higher concentration of homovanillic acid in the striatum than fed mice. However, the administration of probenecid produced a smaller increase in homovanillic acid concentration in fasted than in fed mice. The decay of dopamine following α-methyl- p -tyrosine was reduced in fasted mice at 2 h, but not at 1 h or 6 h after administration of the inhibitor. The possibility that fasting increases the activity of some dopaminergic neurones while decreasing the activity of others is considered. The existence of a pool of homovanillic acid at a site within the striatum where the probenecid-sensitive transport is not effective is postulated.     

Prolonged fasting in mice: a more sensitive approach to genetic diabetes.

Effects of two different periods of fasting were studied on glucose tolerance and insulin response to glucose in genetically diabetic KK and nondiabetic C57BL/6J mice. Blood sugar levels of the KK mice did not differ markedly from those of the C57BL/6J mice at the fed state or after 8 h fasting. They were, however, significantly higher in the KK mice when fasted for 18 h. The serum IRI levels, which were at least twice as high in the KK mice, decreased more markedly after 18 h fasting. The KK mice showed impaired glucose tolerance after 8 h fasting, which became more pronounced after 18 h fasting. The insulin response to glucose in the KK mice was not altered after an 8-hour fast; it was, however, diminished greatly after an 18-hour fast. These data suggest that prolonged fasting is necessary to detect the diabetic traits in the KK mice. The C57BL/6J mice showed neither impaired glucose tolerance nor diminished insulin response to glucose at both periods of fasting. Studies with the F1 hybrids (KK male X C57BL/6J female), which carry half of the diabetic genes, suggest that the mode of inheritance of diabetes in the KK mice might be polygenic.     

Leptin prevents the fall in plasma osteocalcin during starvation in male mice

Plasma osteocalcin, a marker of osteoblastic activity, is reduced in starvation, malnutrition, and anorexia nervosa, resulting in low bone turnover osteoporosis. Contradictory findings about the role of leptin as a link between nutritional status and bone physiology have been reported. We demonstrate that leptin-deficient ob/ob and leptin-resistant db/db male mice have increased plasma osteocalcin, and that in male ob/ob mice osteocalcin is not decreased by starvation, unlike control mice. Intraperitoneal leptin administration increased plasma osteocalcin in male ob/ob mice, and prevented its fall during 24h fasting and 5 days of food restriction in normal male mice. This effect may be mediated via actions on the hypothalamic-pituitary-testicular or -growth hormone axes, or a direct action on osteoblasts. These studies support the hypothesis that the fall in leptin during starvation and weight loss is responsible for the associated reduction in osteoblast activity, and suggest a role for leptin in regulating bone turnover.     

Effects of hormones, fasting and diabetes on triglyceride lipase activities in rat heart and liver

Male rats were fasted for 3 days, subjected to streptozotocin-diabetes or injected with L-thyroxine, Kenacort-A40 (corticosteroid) and Synacthen (ACTH). Cardiac heparin-releasable lipoprotein lipase (LPL) activity was increased after fasting, experimental diabetes and all hormone treatments. Cardiac neutral lipase activity was decreased during diabetes and enhanced in the fasted state and by L-thyroxine, corticosteroid and ACTH administration. The close correlation between vascular LPL and tissue neutral lipase with cardiac triglyceride content is in agreement with the contention that tissue neutral lipase is similar to LPL (Hülsmann, Stam and Breeman 1982). Myocardial acid lipase activity was reduced during diabetes and L-thyroxine treatment, increased during fasting and corticosteroid administration and not affected by short-term ACTH treatment. Hepatic acid lipase activity was increased during fasting, diabetes and by L-thyroxine and reduced after corticosteroid and ACTH treatment. The alkaline liver lipase activity was depressed by fasting, experimental diabetes, corticosteroid and ACTH treatment, whereas L-thyroxine induced a slight increase in enzyme activity. The possible mechanism underlying the observed changes in acid, neutral, alkaline, and LPL activities in heart and liver are discussed.     

Hyperthermia and increased physical activity in the fasting American mink Mustela Vison

The aim of this study was to investigate the thermoregulatory adaptations to fasting in a medium-sized mustelid with a high metabolic rate and energetic requirements. Sixteen farm-bred female American minks, Mustela vison, were divided into a fed control group and an experimental group fasted for 5 days. The deep body temperature (T(b)) of the minks was registered at 10 min intervals with intraabdominal thermosensitive loggers and the locomotor activity was videotaped continuously for 5 days during the fasting procedure. The T(b) of the fasted animals increased during the first day of fasting and decreased during the second day. After 3-4 days of fasting, the levels of physical activity and T(b) of the fasted minks increased above the levels of the fed animals. Significant increases in these parameters were observed at the beginning of the working day on the farm, during the feeding of the fed animals and around midnight. It is concluded that the mink differs from previously studied homeotherms in thermoregulatory and behavioral responses to fasting probably due to its high energy requirements and predatory success.     

The effects of fasting and refeeding on serum parathormone and calcitonin concentrations in young and old male rats.

Although fasting and refeeding reveal the existence of age-related changes in carbohydrate and lipid metabolism, the effects of aging on mineral metabolism in refed animals are unknown. We therefore investigated hormonal regulation of calcium metabolism in young (4 months) and old (26 months) male rats fasted for 48 hours and then refed for 4 or 24 hours. Serum concentrations of total and ionized calcium and parathormone were similar in control young and old rats. Serum calcitonin level was higher, and the concentrations of albumin and inorganic phosphate and alkaline phosphatase activity were lower in fed old rats. In young fasted rats, the serum ionized and total calcium was decreased, and phosphate concentration was increased. In old rats, fasting resulted in the increase of serum parathormone level. Fasting reduced serum alkaline phosphatase activity to a similar extent in both age groups. In young rats, refeeding for 24h normalized serum calcium and phosphate levels and alkaline phosphatase activity, and decreased serum concentrations of PTH and calcitonin. In old refed rats, serum calcitonin concentration was raised by 77% compared to fed or fasted animals, whereas parathormone levels were normalized. Our results indicate that old fasted or refed rats maintain normal serum calcium concentration in a different way than young animals, possibly through the increase in serum levels of parathormone and/or calcitonin. Thus, dietary manipulations such as fasting and refeeding constitute an interesting model for the investigation of the effects of aging on the hormonal regulation of serum calcium level.     

The effect of nutritional state and allopurinol on nucleotide formation in enterocytes from the guinea pig small intestine

The uptake of purine nucleosides (guanosine and hypoxanthine) and bases (guanine, hypoxanthine and adenine) and their incorporation into nucleotides were studied in enterocytes isolated from fed and 3-day fasted guinea pig jejunum. Both total uptake and synthesis of nucleotides were greater for these purines in the fasted, as compared to the fed state for the first 5 min, when the initial substrate concentration in the medium was 10 microM. Increased uptake did not result from a change in the relative distribution of synthesized nucleotides between the fed and fasted states. Reduced catabolism was observed in the medium by enterocytes from fasted as compared to fed animals after 1 min of incubation with both inosine and guanosine. Preincubation of enterocytes with allopurinol (a xanthine oxidase inhibitor) decreased total uptake but increased the formation of IMP from hypoxanthine. Xanthine oxidase activity measured in mucosa from fasted guinea pigs was lower than that from fed animals (6.29 vs. 9.30 nmol/min per mg protein, respectively). However, activities of the salvage enzymes adenine phosphoribosyltransferase and hypoxanthine-guanine phosphoribosyltransferase were not significantly different between the fed and fasted states. These data show that allopurinol treatment, and mucosal atrophy resulting from fasting, decrease xanthine oxidase activity and increase nucleotide synthesis from exogenous substrates in enterocytes from the guinea-pig small intestine, suggesting a regulatory function of mucosal xanthine oxidase in purine salvage by the small intestine.     

Estrogen induces hyperlipidemia in fasted chicks

To determine whether the estrogen-induced hyperlipidemia is affected by fasting, male growing chicks were administered subcutaneously a single dose of 17 beta-estradiol (25 mg/kg body wt), and the hormone treatment lasted for 2 days with or without feed (Experiment 1). In the second experiment, chicks were initially fasted for 1 or 3 days, and then treated with the same dosage of 17 beta-estradiol as in Experiment 1 for 2 days without feed. Plasma and liver lipids, and the activities of hepatic malic enzyme, glucose-6-phosphate dehydrogenase, and hormone-sensitive lipase in the adipose tissue were determined. Compared with fed control chicks, estrogen treatment in fed birds resulted in a marked elevation of plasma lipids, especially triglyceride during the 2-day period (137 vs 2263 mg/dl). In fasted chicks, the present finding that estrogen also induced a marked hyperlipidemia is noteworthy. Upon estrogen treatment (Experiment 1), the level of plasma triglyceride in fasted birds increased about 16 times over that of the fasted control group (133 vs 2093 mg/dl). Even in chicks fasted for 5 days (Experiment 2), estrogen treatment resulted in a persistent hypertriglyceridemia (75 vs 1369 mg/dl). In fed chicks, estrogen treatment also induced a fatty liver with massive accumulation of triglyceride, but the liver of estrogen-treated/fasted chicks appeared to be normal. In both fed and fasted chicks, malic enzyme was found to be the major NADPH producing enzyme in the liver. Upon fasting, both malic enzyme and glucose-6-phosphate dehydrogenase activities decreased significantly (P less than 0.05). In fed chicks, the total activities of both enzymes increased with estrogen treatment, whereas the effect of hormone on these enzymes was less obvious in fasted chicks. The hormone-sensitive lipase activity in the adipose tissue was much lower in fed chicks compared with that of fasted birds (0.15 vs 0.33 nmol of oleic acid released/min/mg protein). Estrogen treatment in fed chicks had no effect on the hormone-sensitive lipase activity, but its activity was enhanced by the hormone treatment in fasted chicks. The present finding that hyperlipidemia persisted in estrogenized chicks during the fasting seems to indicate the complex nature of this hormonal influence on lipid metabolism.     

Dietary regulation of adenosine deaminase activity in stomach, small intestine and spleen of mice

Activity of adenosine deaminase (ADA) and its regulation by dietary restriction were studied in the stomach, small intestine and spleen of mice. ADA activity (U/mg protein) was highest in the stomach, followed by small intestine and spleen of mice on normal diet. The activity decreased significantly in the stomach (41%) and small intestine (45%) of 24 hr fasted mice, when compared to mice fed ad-libitum. However, ADA activity in spleen did not show any change by dietary intervention. Refeeding of fasted mice for 24 hr restored the activity of ADA in tissues. In addition, dietary restriction (alternate days of feeding for three months) had a cumulative effect, whereby ADA activity decreased significantly in the stomach (53% on the day of feeding and 60% on the day of fasting) and small intestine (50% and 54% on the day of feeding and fasting, respectively) without any change in activity in spleen. These findings indicate that dietary restriction reduces ADA activity in a tissue-specific manner. Long-term dietary restriction leads to a cumulative adaptation in lowering the ADA activity of GIT, but not in spleen.