Antiketogenic action of fructose, glyceraldehyde, and sorbitol in the rat in vivo.

The purpose of this study was to compare the metabolism and antiketogenic properties of fructose, glyceraldehyde, and sorbitol. Fructose, glyceraldehyde, and sorbitol were readily metabolized and exhibited an antiketogenic effect in both blood and liver when injected intramuscularly to starved (forty-eight hours) rats. Sorbitol had the most pronounced antiketogenic effect and produced an 80 to 90 per cent decrease in the blood ketone bodies sixty minutes after administration. Fructose and glyceraldehyde were equally effective and produced about a 60 to 70 per cent decrease in ketone bodies. Fructose, glyceraldehyde, and sorbitol caused a significant decrease in the concentration of hepatic ketone bodies. In liver, sorbitol was found to be most effective in its antiketogenic action. The concentration of plasma free fatty acids remained unchanged after injection of all three antiketogenic substrates. Fructose, glyceraldehyde, or sorbitol caused increased blood lactate and pyruvate concentrations, and fructose was the most effective of the three substrates. Fructose administration resulted in a significant decrease in hepatic lactate/pyruvate and beta-OH-butyrate/acetoacetate concentration ratios, whereas sorbitol caused an increase in the concentration ratio of these two substrat pairs. Decreases in blood and liver ketone body levels were associated with lowering of liver acetyl-CoA concentration . However, the decrease in hepatic acetyl-CoA produced upon the administration of antiketogenic substrates was not pronounced. Sorbitol administration resulted in the most pronounced increase in hepatic alpha-glycerophosphate concentration. Fructose or glyceraldehyde also caused an increase in alpha-glycerophosphate content. Administration of each of the three antiketogenic substrates produced an increase in hepatic dihydroxyacetone phosphate concentration. All three antiketogenic compounds increased liver glycogen and blood glucose concentrations. No significant changes were observed in hepatic ATP, ADP, or AMP concentrations sixty minutes after the injections of any of the antiketogenic substrates. Although decreased liver acetyl-CoA levels were associated with the antiketogenic effects of the compounds tested, the increased liver alpha-glycerophosphate content best explains the differences between fructose or glyceraldehyde and sorbitol.

Plasma free tryptophan, brain serotonin, and an endocrine profile of the genetically obese hyperglycemic mouse at 4--5 months of age.

Genetically obese hyperglycemic mice (ob/ob) were compared with their nonlittermate lean controls at 4-5 months of age with regard to brain serotonin, pituitary ACTH content, and circulating levels of glucose, glucagon, insulin, TSH, T3, T4, total tryptophan and free tryptophan. Brain serotonin pituitary ACTH content, and plasma insulin, glucose, total tryptophan, and free tryptophan were all significantly higher in obese mice than in the controls. TSH, T3, and T4 were not significantly different in obese mice vs. controls, suggesting that the obese mouse is euthyroid. Fasting improved but failed to normalize the glucose and insulin levels or insulin to glucagon ratios. Since serotonin is an important neurotransmitter with regard to hypothalamic-pituitary function and since its levels in the brain are dependent on the availability of tryptophan, the findings of elevated levels of free tryptophan in the plasma and serotonin in the brain of the obese hyperglycemic mouse may help to explain some of the previously observed abnormalities of pituitary hormone secretion in these animals.

A longitudinal hormonal profile of the genetically obese mouse.

Obese mice (C57BL/6J ob/ob) and their lean littermates were studied at various ages from immediately post weaning until 62 weeks of age, at which mortality increased markedly. Several age-related changes were noted. 1) Plasma glucose levels were elevated in obese mice 5-20 weeks and 62 weeks of age, but were similar to those in the lean mice at 20-60 weeks of age. Plasma insulin levels were elevated in obese mice, and there were no age-related differences. 2) Brain serotonin was elevated in obese mice at all ages and increased with age in both obese and lean animals. 3) Pituitary contents of ACTH and beta-endorphin were elevated in young obese mice and increased further as these mice approached their life expectancy. 4) The ratios of ACTH to beta-endorphin immunoreactivities were similar in obese and lean mice, except in obese mice over 50 weeks of age where this ratio was increased. We conclude that: 1) the obese mouse is characterized by hyperinsulinemia and hyperadrenocorticism throughout its life; 2) the insulin resistance of the obese mouse improves at 20 weeks of age, yet deteriorates as its life expectancy is approached; 3) the obese mouse has an elevated brain serotonin content similar to previously described elevations of the putative neurotransmitters dopamine and norepinephrine in these mice; and 4) as the obese mouse approaches its life expectancy, abnormalities may occur in the synthesis, processing, or secretion of ACTH and/or beta-endorphine.

Age-related changes in liver and adipose tissue pyruvate dehydrogenase of genetically obese mice.

Changes of the pyruvate dehydrogenase complex in liver and epididymal fat pad were examined longitudinally in obese mice (C57BL/6J-ob/ob) and their lean controls as a function of age. Total pyruvate dehydrogenase in liver was expressed on several reference bases because of differences in hepatic cellularity and protein content between obese mice and their age-matched lean controls. When total hepatic pyruvate dehydrogenase was expressed on a protein basis, the enzyme activity was elevated in obese mice older than 28 weeks in age when compared to lean controls of a similar age. However, when expressed on a DNA basis, total pyruvate dehydrogenase activity in livers of obese mice up to 10 weeks in age was increased when compared to the age-matched lean control. The proportion of hepatic pyruvate dehydrogenase in the active form was also augmented significantly in obese mice from 5 to 28 weeks of age. In 18-week-old obese mice, the proportion of total pyruvate dehydrogenase in the active form of adipose tissue was significantly higher than that of the lean controls. When expressed on a DNA basis, total pyruvate dehydrogenase in the fat pad was also increased in obese mice up to 10 weeks in age when compared to age-matched controls. Total pyruvate dehydrogenase activity in the epididymal fat pad was higher in obese mice than the lean controls in animals as old as 32 weeks in age when the enzyme activity was expressed per 100 g body weight. The increase in the active form and total activity of pyruvate dehydrogenase in both liver and epididymal fat pad during the dynamic early phase of obesity would augment the capacity for acetyl-coenzyme A formation necessary in the support of an accelerated lipogenesis and fat deposition.

Age-related changes in 5'-nucleotidase and alkaline phosphatase activities in mouse thymocytes.

Enzymatic activities of thymocytes isolated from Swiss albino mice were studied at various ages from immediately post weaning until 100 weeks of age, approaching the life expectancy of these animals. Between 5 and 10 weeks of age, the activities of lactate dehydrogenase and alkaline phosphatase decreased to a level that was maintained throughout the remainder of the aging profile. Neutral beta-glycerophosphatase (pH 7.5) activity in a thymus membrane preparation was similar in all age groups. The activity of membrane-bound 5'-nucleotidase, that is, AMP-hydrolyzing activity inhibited by 100 microM alpha, beta-methyleneadenosine 5'-diphosphate, progressively increased as a function of age, indicating thymocyte population changes occurring very late in life. In thymocytes of the oldest mice examined (100 weeks of age), 5'-nucleotidase specific activity was approximately ten-fold greater than the activity found in 5-week-old mice. Thus, membrane-bound 5'-nucleotidase activity in thymocytes increased markedly as a function of age in Swiss albino mice; yet several other enzymatic activities, including alkaline phosphatase, remained relatively unchanged in mature mice.

A longitudinal metabolic profile of Swiss albino mice during aging.

The consequence of aging on carbohydrate and lipid metabolism of the Swiss albino mouse was examined in a complete aging profile. The profile was developed by obtaining groups of at least 10 weanling mice at 2-3 month intervals for approximately a 2-year period. Body weight plateaued at approximately 10 months-of-age. Maximal heart and liver weights were found in 3-month-old mice. Plasma glucose concentration was highest in 1-month-old mice, declined rapidly during the first 3 months-of-age and then gradually throughout the profile. Liver glycogen content spiked sharply in 2-month-old mice, decreasing sharply to a level maintained in mature animals. A second peak in hepatic glycogen occurred in 15-month-old mice with a precipitous decrease taking place in older animals. Plasma concentrations of glycerol, 3-hydroxybutyrate and triacylglycerols, indices of lipid metabolism, were remarkably constant and showed no consistent trend with age. The level of triacylglycerols in the heart peaked in 2-month-old mice and then decreased gradually in older animals. The maximal content of triacylglycerols in the liver was found in 10- to 20-month-old mice. Hepatic levels of triacylglycerols decreased precipitously in senescent Swiss albino mice.

Hormonal, metabolic and morphologic studies of aged C57BL/6J obese mice.

Genetically obese mice (C57BL/6J-ob/ob), fed ad libitum, demonstrated a precipitous increase in the spontaneous death rate after 50 weeks. The first signs of morbidity were a ruffled hair coat and a progressive motor ataxia. Necropsy revealed that obese mice had pale and fatty livers, urolithiasis and grossly distended bladders. Microscopically, the hepatocellular changes observed in all aged obese mice included: a loss of orientation of hepatocytes, an enormous variability in the size of both hepatocytes and their nuclei, and an extensive deposition of both large and small lipid droplets, confirmed by an increase content of triacylglycerols. A subacute-to-chronic, multifocal, necrotizing hepatitis was also present. Kidneys from aged obese mice contained hypertrophied glomeruli and increased PAS-stained material. Tubular dilation with compaction of the tubular cells was also seen. There were no significant alterations in the microanatomy or mineralization of femurs from obese mice, yet there was a significant increase in plasma alkaline phosphatase activity. In obese mice at 62-63 weeks of age, hyperglycemia was present even in spite of hyperinsulinemia. Pituitary immunoreactive ACTH and its molar ratio to pituitary immunoreactive beta-endorphin were also increased in obese mice at this age. Even though the etiology of the decreased lifespan of genetically obese mice remains uncertain, the possibility is discussed that an overall defect in the central nervous system may be involved.

Hepatic indices of thyroid status in rats treated with 2,3,7,8-tetrachlorodibenzo-p-dioxin.

The functional thyroid status of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD)-treated rats is unknown. Therefore, activities of certain thyroid-responsive enzymes were examined in the livers of adult male Sprague-Dawley rats 1 week after treatment with TCDD (6.25, 25 or 100 micrograms/kg). Activity of the thyroid-responsive flavin L-glycerol-3-phosphate dehydrogenase (per mg mitochondrial protein) was decreased slightly in livers of TCDD-treated rats, while that of succinate dehydrogenase remained unchanged. In contrast, activities (per mg supernatant protein) of three thyroid-responsive NADP-dependent cytosolic enzymes, malic enzyme, glucose-6-phosphate dehydrogenase and 6-phosphogluconate dehydrogenase, were increased by TCDD treatment in a dose-dependent manner. Lactate dehydrogenase (activity per mg supernatant protein) was also augmented slightly 1 week after TCDD administration. Liver mass was increased by TCDD treatment in a dose-dependent manner, but DNA content per liver was similar at all doses examined. Total hepatic protein, expressed per liver or mg hepatic DNA, was increased in TCDD-treated rats when compared to their pair-fed counterparts. The decreased activity of the mitochondrial L-glycerol-3-phosphate dehydrogenase, in contrast to the increased activities of the supernatant enzymes, malic enzyme, glucose-6-phosphate dehydrogenase and 6-phosphogluconate dehydrogenase, is not consistent with a shift in functional thyroid status following TCDD treatment.

Effects of perfluorodecanoic acid on hepatic indices of thyroid status in the rat.

Perfluorodecanoic acid (PFDA) alters the circulating level of thyroid hormones, but the physiological significance of this change at the target tissue remains to be defined. To this end, the activities of thyroid-responsive hepatic enzymes were examined in adult male rats 1 week after treatment with a single dose of PFDA (20, 40 or 80 mg/kg). Since PFDA treatment caused a dose-related reduction in feed intake, vehicle-treated rats pair-fed to their counterparts receiving PFDA were used to determine if any of the PFDA-induced alterations in enzyme activity were secondary to hypophagia. Following the administration of PFDA, L-glycerol-3-phosphate dehydrogenase, a liver mitochondrial enzyme sensitive to thyroid status, exhibited a modest increase in activity, whereas that of succinate dehydrogenase, a constitutive mitochondrial marker enzyme, was similar in both PFDA-treated rats and their pair-fed counterparts at all dose levels examined. Activity of cytosolic lactate dehydrogenase was also augmented modestly in livers of rats receiving PFDA. In contrast, activity of cytosolic malic enzyme, a thyroid-responsive enzyme, was increased markedly in PFDA-treated rats. Hepatic activity of glucose-6-phosphate dehydrogenase, which also responds to alterations in thyroid status, exhibited a modest increase with 20 and 40 mg/kg PFDA but was similar in both PFDA-treated rats and their pair-fed counterparts at the 80 mg/kg dose level. Absolute and relative liver mass was elevated in PFDA-treated rats at all dose levels in comparison to the appropriate vehicle-treated pair-fed animals. Total hepatic content of DNA was maintained in PFDA-treated rats at all dose levels, whereas a significant decrease in liver DNA was found in the vehicle-treated rats pair-fed to animals receiving 80 mg/kg PFDA. Following administration of PFDA, protein content per total liver was similar to that of their pair-fed counterparts. Thus, the pattern of activity of thyroid-responsive hepatic enzymes was not compatible with a functional shift toward a lessened thyroid status in rats treated with PFDA.

Age-related changes in lipid and carbohydrate metabolism of the genetically obese mouse.

Obese mice (C57BL/6J ob/ob) and their lean controls were studied longitudinally from immediately post-weaning until 62 wk of age, at which time the experiment was terminated. The dynamic nature of the metabolic aberrations of the obese mouse syndrome was clearly demonstrated. Obese mice were hyperinsulinemic at all ages yet the concentration of glucose in plasma was elevated only at 5-20 wk and 63 wk of age, but was similar to that of lean mice at 20-60 wk of age. Triacylglycerols accumulated in the liver of obese mice between 5 and 18 wk of age to a level that was 20-fold greater than that found in the age-matched lean control. A decreased concentration of DNA/g of liver was also found in 5-18 wk-old obese mice, indicative of an enlarged hepatocyte. With the exception of 5-wk-old animals, total DNA per liver was increased in obese mice when compared to the lean control throughout the profile. Following the peak in 18-wk-old mice, the hepatic content of triacylglycerols precipitously fell so that at 45 wk of age its concentration in obese mice was similar to that of the lean control. Plasma free fatty acid levels as well as liver glycogen content were comparable in obese mice and their lean controls throughout the profile. In obese mice older than 45 wk of age, the content of triacylglycerols in plasma was significantly lower than that of the age-matched lean control while an accumulation of liver triacylglycerols was again found in obese mice. Myocardial triacylglycerols were elevated in obese mice when compared to the lean control at all ages. The longitudinal metabolic profile of the obese mouse developed in the present study clearly demonstrates the dynamic nature of the deviations in carbohydrate and lipid metabolism in this animal model of human obesity and insulin resistance.

Characterization of elevated plasma alkaline phosphatase activity in genetically obese mice.

Alkaline phosphatase activity in obese mice (C57BL/6J ob/ob) was significantly increased from 18 to 63 weeks of age when compared to that of their lean controls (C57BL/6J +/?). In 5 week old animals, the earliest age examined, the circulating activity of alkaline phosphatase was similar in both obese mice and their lean counterparts. To characterize the circulating alkaline phosphatase activity in the obese mouse and its lean counterpart, the response of the enzyme to fasting, various inhibitors, heat inactivation, and urea denaturation was examined and compared. L-homoarginine and L-p-bromotetramisole inhibited to a large extent the circulating activity of alkaline phosphatase in both obese mice and their lean controls in the fed state, while L-phenylalanine had essentially no effect. Even though the response of alkaline phosphatase in plasma to several inhibitors was similar, the rate of denaturation by urea of enzyme activity in plasma was significantly slower in obese mice than in their lean controls in the fed state. While the rate of inactivation of alkaline phosphatase activity in plasma for the initial two minutes at 56 degrees C was similar in obese mice and their lean counterparts, the subsequent rate of heat inactivation was significantly slower in the plasma from obese mice. Thus, both obese and lean mice in the fed state have a circulating activity of alkaline phosphatase in plasma with a greater contribution from a skeletal isoenzyme and a lesser one of intestinal origin.(ABSTRACT TRUNCATED AT 250 WORDS)

Glucose-dependent lactate production by homogenates of neuronal tissues prepared from rats treated with 2,4-dithiobiuret, acrylamide, p-bromophenylacetylurea and 2,5-hexanedione.

Chronic treatment of rats with dithiobiuret (DTB) produces a delayed onset muscle weakness and, as suggested by a preliminary study, a distal axonopathy. An inhibition of glycolysis resulting in an energy deficit has been suggested as a possible mechanism of neurotoxin-induced distal axonopathies. To determine whether chronic DTB administration (1 mg/kg/day X 7 days; sacrificed on Day 7) might be associated with a perturbation of glucose metabolism in neuronal tissues, lactate production was measured in brain and spinal cord homogenates prepared from rats which exhibited hindlimb skeletal muscle weakness. Kinetic and statistical analysis showed that glucose-dependent lactate production in these homogenates was not different from that of controls. Lactate production was also determined in brain homogenates prepared from rats intoxicated with acrylamide (50 mg/kg/day X 9 days; sacrificed on Day 9), p-bromophenylacetylurea (200 mg/kg/day X 2 days; sacrificed on Day 14) or 2,5-hexanedione (400 mg/kg/day X 21 days; sacrificed on Day 21). The results indicated that whereas 2,5-hexanedione produced a slight decrease in Vmax, acrylamide and p-bromophenylacetylurea did not cause changes in glucose-dependent lactate production. These findings do not support the suggestion that an inhibition of glycolysis represents a common biochemical lesion associated with neurotoxins which cause delayed onset muscle weakness and distal axonopathy.

Interrelationships between energy and fat metabolism and hypophagia in rats treated with perfluorodecanoic acid.

Energy metabolism and body composition were investigated in perfluorodecanoic acid (PFDA)-treated rats (single i.p. dose of 20, 40 or 80 mg/kg) and their respective pair-fed counterparts 7 days after dosing. Cumulative feed intake and body weight were decreased in a dose-dependent manner. However, PFDA-treated rats either gained less weight or lost more weight (dependent on the dose administered) than their pair-fed, vehicle-treated counterparts, even though feed intake in these two treatment groups was similar. Energy expenditure, determined indirectly by quantifying oxygen consumption and carbon dioxide production, decreased in a dose-dependent fashion, yet it was similar in PFDA-treated rats and their pair-fed counterparts at a given dose. Body composition analysis indicated a dose-dependent decrease in carcass water and protein content in both PFDA-treated rats and pair-fed partners, while total amount of ash remained unchanged in all treatment groups. These alterations in body composition are compatible with a negative energy balance. Even though PFDA-treated rats had a lower body weight than their pair-fed counterparts at each dose level, they were found to have a greater carcass fat content. Thus, at maintenance (i.e., zero change of body weight) PFDA-treated rats require a higher caloric intake associated with a greater body fat content than vehicle-treated animals.

Perfluorodecanoic acid and lipid metabolism in the rat.

Alterations in lipid metabolism were examined in adult male Sprague-Dawley rats seven days after a single intraperitoneal injection of perfluorodecanoic acid (PFDA; 20, 40 or 80 mg/kg). Because PFDA treatment caused a dose-related reduction in feed intake, the response of vehicle-treated rats pair-fed to those receiving PFDA was monitored to distinguish direct effects of the perfluorinated fatty acid from those secondary to hypophagia. Carcass content of lipid phosphorus and free cholesterol decreased in dose-dependent fashion in both PFDA-treated and pair-fed rats. Carcass triacylglycerols diminished in a similar manner, yet PFDA-treated rats at each dose had a higher concentration of neutral acylglycerols than their vehicle-treated, pair-fed counterparts. In vehicle-treated, pair-fed rats at the 80 mg/kg dose level, lipid phosphorus and free cholesterol as a proportion of carcass fat increased, whereas the share of the triacylglycerols declined. Because of the higher concentration of triacylglycerols in the carcass of rats treated with 80 mg/kg PFDA, enrichment of lipid phosphorus and free cholesterol in carcass fat was less than in their pair-fed partners. The amount of lipid phosphorus and free cholesterol per hepatocyte was similar in both PFDA-treated rats and their pair-fed partners. Liver triacylglycerols were markedly increased in PFDA-treated rats. A similar but less extensive augmentary effect of PFDA on hepatic esterified cholesterol was found. Concentration of triacylglycerols in plasma was not elevated in PFDA-treated rats, in spite of hepatic accumulation of esterified compounds. Also, the plasma level of free fatty acids and 3-hydroxybutyrate was similar in all treatment groups, including those receiving PFDA.(ABSTRACT TRUNCATED AT 250 WORDS)

Isolation and purification of perfluorodecanoic and perfluorooctanoic acids from rat tissues.

A procedure for the extraction, separation, and isolation of perfluorodecanoic and perfluorooctanoic acids from biological samples is described. The use of conventional lipid extraction procedures leads to substantial loss of the perfluorinated fatty acids added to tissue. The presence of sulfuric acid in aqueous saline during phase partitioning is essential for the recovery of perfluorodecanoic and perfluorooctanoic acids in the organic phase following their extraction from tissue. The perfluorinated fatty acids are co-eluted with simple lipids from silica gel columns using diethyl ether/trifluoroacetic acid (100:1, v/v). Simple lipids are separated by thin layer chromatography. By substituting trifluoroacetic acid for acetic acid in the developing solvents, perfluorodecanoic and perfluorooctanoic acids migrate with other free fatty acids.