Glycogen synthesis in rat liver from a pool of free glucose.

Glycogen synthesis in isolated perfused livers or livers of anesthesized rats (in situ), was studied using radioactively labelled fructose, lactate, and inositol as substrates. The specific radioactivity of glucose and glycogen was measured at various times and compared with that of some intermediates. The results suggest that liver glycogen is formed from the pool of free glucose which in turn is fed by the so-called "direct and indirect pathway" of glycogen synthesis. This points to an important role of glucose-6-phosphatase, an enzyme complex subject to regulation by glucocorticoids, well known promoters of hepatic glycogen synthesis.

Biochemical effects and zonal heterogeneity of peroxisome proliferation induced by perfluorocarboxylic acids in rat liver.

Rats were treated for 5 to 14 days with perfluoroacetate, perfluorobutyrate and perfluorooctanoate. Alterations in hepatic morphology with special reference to the peroxisomal compartment were investigated by light and electron microscopy following cytochemical staining of catalase activity with the alkaline 3,3'-diaminobenzidine medium. All three compounds induced hepatomegaly and peroxisome proliferation. Perfluorobutyrate and perfluorooctanoate were found to be more active than perfluoroacetate. Perfluorooctanoate-induced peroxisome proliferation was more prevalent in centrilobular than in periportal hepatocytes. Peroxisomes in centrilobular liver cells frequently were of round shape, exhibited diameters of up to 1.5 microns and were predominantly located within smooth endoplasmic reticulum-glycogen areas. In periportal cells, however, clusters of polymorphous peroxisomes ranging from 250 to 1,100 nm in diameter were observed at the periphery of smooth endoplasmic reticulum-glycogen regions. Peroxisome proliferation was accompanied by a change of peroxisomal and mitochondrial enzyme activities, in particular an increase in peroxisomal palmitoyl-CoA oxidation. Significant alterations in the concentration of peroxisomal matrix and membrane polypeptides were also noted. Within the first 2 days, perfluorooctanoate treatment exerted a strong hypolipidemic activity and both compounds perfluorooctanoate and perfluorobutyrate raised the level of hepatic free acid-soluble CoA nearly 10-fold as compared with control livers. The results suggest perfluorinated carboxylic acids to be model substances suitable to correlate biochemical and morphological parameters with the zonal heterogeneity of the peroxisomal compartment in rat liver. Due to the manifold hepatic effects, contact of humans with perfluorinated carboxylic acids or their metabolic precursors may represent a severe health risk.

Improved isolation and purification of rat liver peroxisomes by combined rate zonal and equilibrium density centrifugation.

Two different peroxisome preparations were isolated from male rat liver by using total homogenate (TH) as the starting material for one and the light mitochondrial (L) fraction for the other. The technique worked out is based on rate zonal (RZ) centrifugation in a sucrose gradient and subsequent isopycnic centrifugation in a Nycodenz gradient. The peroxisome fraction isolated from the L fraction consisted of 97-98% peroxisomal protein with catalase activity 49-fold enriched over TH. The peroxisome preparation isolated directly from TH represented about 55% of the total liver peroxisome population and had catalase activity 43-fold enriched compared with TH. The contribution of peroxisome protein to the liver protein was calculated to be in the range 1.82-2.02%. Peroxisomes isolated from TH were considerably more heterogeneous in size than peroxisomes isolated from the L fraction. Comparison of the polypeptide patterns of both preparations by sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed some quantitative differences. Several major polypeptides were found to be exclusively located in the peroxisome membrane. These polypeptides migrated in the gel with apparent molecular masses of 69, 42.5, 36, 26, 21, and 15 kDa.

Rat liver peroxisomes. I. New peroxisome population induced by thyroid hormones in the liver of male rats.

The paper presents results that a hyperthyroid state for 5 to 10 days induces a new peroxisome population in the liver of male rats. This assumption is based on the following observations: 1. Treatment of male rats for five consecutive days with 25 micrograms thyroxine per day resulted in an increase of catalase and D-amino acid oxidase activities by 32 and 61%, respectively. On the other hand, urate oxidase activity declined by 20%. Prolonged treatment for ten days raised catalase activity by 68% and lowered urate oxidase activity by 35%. 2. Morphological examinations on the light and electron microscopic level revealed that the increase in peroxisomal enzyme activities was accompanied by an augmentation of peroxisomal profiles. "Microperoxisomes" and peroxisomes lacking the electron dense core were frequently observed. 3. The peroxisomes of the livers of thyroxine-treated animals exhibited an altered centrifugal behaviour in sucrose density gradients. The mean equilibrium density of normal liver peroxisomes was 1.23, whereas peroxisomes of thyroxine-treated livers equilibrated at the density of 1.20. It is discussed that peroxisomal metabolism may contribute considerably to the late biochemical effects of thyroid hormones, that means enhancement of lipid oxidation and heat production. Up to now these late effects were attributed to a stimulated mitochondrial metabolism only.

Lack of ornithine decarboxylase activity in isolated rat liver parenchymal cells.

Polyamines are associated with fundamental metabolic and functional steps in cell metabolism. The activity of ornithine decarboxylase, the key enzyme in polyamine metabolism, was followed during the preparation of rat liver parenchymal cells and in the isolated cells during incubation. In experiments in which ornithine decarboxylase was not induced in vivo, enzyme activity dropped to barely measurable values during the preparation. An even more drastic loss of enzyme activity was noted in livers in which ornithine decarboxylase activity was stimulated in vivo 20-40fold by previous injection of bovine growth hormone, or thioacetamide or elevated because of circadian rhythmical changes of the enzyme activity. Within the first 20 min of liver perfusion to disintegrate the tissue, ornithine decarboxylase activity decreased by up to 80%. The presence of bovine growth hormone during cell preparation cannot prevent the loss of enzyme activity. Incubation of the isolated cells for periods of up to 240 min did not restore the enzyme activity. Furthermore, incubation of the cells with bovine growth hormone did not induce ornithine decarboxylase, even though the medium was supplemented with amino acids in physiological concentrations. During normal liver perfusion and in contrast to the situation with isolated cells, there is no loss of enzyme activity but a small rise. Following pretreatment of the animals with bovine growth hormone or thioacetamide the highly stimulated activity of ornithine decarboxylase declined slowly during liver perfusion, but never dropped to values lower than normal for perfusion periods of up to 240 min. Moreover, in the intact perfused organ ornithine decarboxylase remains responsive to bovine growth hormone. The experiments demonstrate that enzymatic tissue dispersion by collagenase in particular or the preparation of isolated cells in general drastically alters the metabolic and functional state of rat liver parenchymal cells.

Ornithine decarboxylase activity in rat liver during phalloidin poisoning.

Rat liver ornithine decarboxylase activity was measured during phalloidin poisoning of 17 day-old rats, adult rats, and adult rats protected against phalloidin toxicity by pretreatment with somatotropin. The results show: 1. In adult rats, after a short period of activation, ornithine decarboxylase activity decreases to basal levels after phalloidin administration and remains so until death. 2. In somatotropin-pretreated rats, the enzyme activity increases with a peak 4 hours after injection of somatotropin but then drops rapidly to control levels. All rats survive poisoning. In somatotropin-pretreated rats that has also been given the ornithine decarboxylase inhibitor, 1,3-diaminopropane, phalloidin poisoning causes a decrease in the enzyme activity to basal levels and death of all animals. 3. In 17 day-old rats, ornithine decarboxylase activity decreased initially after phalloidin poisoning but then returned to unpoisoned levels. All rats survived poisoning, even those pretreated with 1,3-diaminopropane.

[Effects of phalloidin in liver cells: binding, morphological changes, and elimination (author's transl)]. / Wirkung von Phalloidin Auf Leberzellen:Bindung, Strukturveränderung und Elimination.

1. The uptake, binding and elimination of phalloidin in liver is compared in adult (180 to 240 g) and "baby" (17 to 19 days old) rats in vivo and in vitro. 2. In both groups there is no relation between the concentration of the poison in the liver and the toxicity. 3. Although baby rats show a significantly higher tolerance against phalloidin than the adult animals, the concentration of the poison in the liver of baby rats is higher, and the elimination is significantly slower than in adult rats. 4. The very tight binding and concentration of phalloidin in the liver is explained by an extremely low dissociation constant. 5. Furthermore, the morphological differences between the poisoning of the liver cells in the entire organ and of isolated liver cells are discussed.

[Somatotropin, antidot against phalloidin (author's transl)]. / Somatotropin, Antidot gegen Phalloidin.

1. Somatotropin protects rats against a lethal dose of phalloidin (1.3 mg/kg). 2. Scanning electron microscopy has shown that 2 hours after phalloidin injection the liver from a somatotropin-pretreated rat is not significantly different to that from an untreated rat. Phalloidin alone caused complete destruction of the structure of the liver lobules. 3. Somatotropin does not prevent phalloidin uptake by the liver but slows down elimination. 4. The findings are discussed with respect to their therapeutic possibilities as somatropin protects rats against death also after phalloidin poisoning.