Metabolism Of Rat Liver Mitochondria Research Articles

Effects of the Long-Term Administration of Uridine on the Functioning of Rat Liver Mitochondria in Hyperthyroidism

The effect of uridine (30 mg/kg for 7 days; intraperitoneally) on the functions of liver mitochondria in rats with experimentally induced hyperthyroidism (HT) (200 µg/100 g for 7 days, intraperitoneally) is studied in this paper. An excess of thyroid hormones (THs) led to an intensification of energy metabolism, the development of oxidative stress, a significant increase in the biogenesis, and changes in the content of proteins responsible for the fusion and fission of mitochondria. The injection of uridine did not change the concentration of THs in the blood of hyperthyroid rats (HRs) but normalized their body weight. The exposure to uridine improved the parameters of oxidative phosphorylation and corrected the activity of some complexes of the electron transport chain (ETC) in the liver mitochondria of HRs. The analysis of ETC complexes showed that the level of CI–CV did not change by the action of uridine in rats with the condition of HT. The application of uridine caused a significant increase in the activity of superoxide dismutase and lowered the rate of hydrogen peroxide production. It was found that uridine affected mitochondrial biogenesis by increasing the expression of the genes Ppargc1a and NRF1 and diminishing the expression of the Parkin gene responsible for mitophagy compared with the control animals. In addition, the mRNA level of the OPA1 gene was restored, which may indicate an improvement in the ETC activity and oxidative phosphorylation in the mitochondria of HR. As a whole, the results obtained demonstrate that uridine has a protective effect against HT-mediated functional disorders in the metabolism of rat liver mitochondria.

Ecdysterone prevents negative effect of acute immobilization stress on energy metabolism of rat liver mitochondria

Ecdysterone is a naturally occurring steroid hormone, which presents in arthropods and in a number of plants as an insect defence tool. There are many studies showing that application of ecdysterone can alter mitochondrial functions of mammalian cells, however it is not clear whether its effects are direct or mediated by activation of other cellular processes. In our study, we have shown how ecdysterone acts at the mitochondrial level in normal conditions and in certain pathology. We have demonstrated that application of immobilization stress to male rats causes uncoupling of mitochondrial oxidative phosphorylation, the preliminary application of ecdysterone prevents negative effect of immobilization stress on mitochondria. In-vitro experiments with isolated mitochondria have shown that ecdysterone can increase mitochondrial coupling and hyperpolarise mitochondria but without a noticeable effect on ADP/O ratio. Molecular docking experiments revealed that ecdysterone has high binding energy with mitochondrial FOF1 ATP synthase, but further biochemical analysis have not revealed either stimulatory or inhibitory effect of ecdysterone on FOF1 ATPase activity of the enzyme. Thus, ecdysterone can directly affect mitochondrial bioenergetics, though we assume that its preventive effect on mitochondria during immobilization stress is also coupled with the activation of some other cellular processes.

The photodynamic and direct actions of methylene blue on mitochondrial energy metabolism: A balance of the useful and harmful effects of this photosensitizer

According to the literature, methylene blue (MB) is a photosensitizer (PS) with a high affinity for mitochondria. Therefore, several studies have explored this feature to evaluate its photodynamic effects on the mitochondrial apoptotic pathway under normoxic conditions. We are aware only of limited reports regarding MB's photodynamic effects on mitochondrial energy metabolism, especially under hypoxic conditions. Thus, the purposes of this study were to determine the direct and photodynamic acute effects of MB on the energy metabolism of rat liver mitochondria under hypoxic conditions and its direct acute effects on several parameters linked to energy metabolism in the isolated perfused rat liver. MB presented a high affinity for mitochondria, irrespective of photostimulation or proton gradient formation. Upon photostimulation, MB demonstrated high in vitro oxidizing species generation ability. Consequently, MB damaged the mitochondrial macromolecules, as could be evidenced by the elevated levels of lipid peroxidation and protein carbonyls. In addition to generating a pro-oxidant environment, MB also led to a deficient antioxidant defence system, as indicated by the reduced glutathione (GSH) depletion. Bioenergetically, MB caused uncoupling of oxidative phosphorylation and led to photodynamic inactivation of complex I, complex II, and F1FO–ATP synthase complex, thus decreasing mitochondrial ATP generation. Contrary to what is expected for an ideal PS, MB displayed appreciable dark toxicity on mitochondrial energy metabolism. The results indicated that MB acted via at least three mechanisms: direct damage to the inner mitochondrial membrane; uncoupling of oxidative phosphorylation; and inhibition of electron transfer. Confirming the impairment of mitochondrial energy metabolism, MB also strongly inhibited mitochondrial ATP production. In the perfused rat liver, MB stimulated oxygen consumption, decreased the ATP/ADP ratio, inhibited gluconeogenesis and ureogenesis, and stimulated glycogenolysis, glycolysis, and ammoniagenesis, fully corroborating its uncoupling action in intact cells, as well. It can be concluded that even under hypoxic conditions, MB is a PS with potential for photodynamic effect-induced mitochondrial dysfunction. However, MB disrupts the mitochondrial energy metabolism even in the dark, causing energy-linked liver metabolic changes that could be harmful in specific circumstances.

Stimulation of oxidative energy metabolism in liver mitochondria from old and young rats by treatment with dehydroepiandrosterone (DHEA). A comparative study

Effects of treatment with DHEA (0.2 or 1.0 mg/kg body weight for 7 days) on oxidative energy metabolism of rat liver mitochondria from old (18-24 month old) and young (8-10 weeks old) male albino rats belonging to Charles-Foster strain were examined. Treatment with 1.0 mg DHEA resulted in increased body weights of the young rats without change in the liver weight. In the old animals the liver weight increased progressively with increasing dose of DHEA without affecting body weight. The state 3 respiration rates in liver mitochondria from old animals were, in general, lower than those in the young rats. The state 3 and state 4 respiration rates increased following DHEA treatment in dose-dependent manner bringing them close to values for young animals or beyond that with the effect being more pronounced at 1.0 mg dose. Treatment with DHEA also stimulated state 3 and state 4 respiration rates in young rats in dose-dependent manner. Contents of cytochrome aa(3), b and c + c(1) increased significantly in old animals in dose-dependent manner. In the young rats the lower dose (0.2 mg) of DHEA was more effective in bringing about a maximum increase in the contents of the cytochromes; the effect declined at the higher dose (1.0 mg). DHEA treatment also stimulated the mitochondrial ATPase activity in the old as well as in the young rats. The dehydrogenases activities were considerably low in the old rats compared to the values for the young animals. Treatment with DHEA stimulated dehydrogenases activities in old rats in dose-dependent manner bringing them close to values for the young animals or beyond. Treatment with lower dose (0.2 mg) of DHEA maximally stimulated dehydrogenases activities in young animals.

Effect of local anesthetic ropivacaine on isolated rat liver mitochondria

Ropivacaine is a new long-acting aminoamide local anesthetic with a reduced systemic and cardiac toxicity. Since the latter seems to be related, at least partially, to an interference with mitochondrial energy transduction, the effect of ropivacaine on the metabolism of rat liver mitochondria was studied. Ropivacaine alone exhibited little effect on mitochondrial metabolism, whereas effects were strongly enhanced by tetraphenylboron (TPB −) anion. At low drug concentrations, state 4 respiration was stimulated and mitochondrial membrane potential collapsed. At higher concentrations, state 4 and uncoupled respiration were inhibited by impairment of electron transfer from NAD- and flavine adenine dinucleotide-linked substrates to the respiratory chain. The fact that TPB − increased drug effects indicated that stimulation of respiration was due to dissipation of the electrochemical proton gradient caused by its electrophoretic uptake, although a classical uncoupling mechanism cannot be excluded. The mechanism for the lower toxicity of ropivacaine in vivo was ascribed to low liposolubility leading to reduced access to the mitochondrial membrane, resulting in a minimal perturbation of mitochondrial metabolism.

Role of glutathione and nitric oxide in the energy metabolism of rat liver mitochondria

Previous studies have suggested that nitric oxide (NO) inhibited mitochondrial respiration. NO and/or its intermediate(s) react with various molecules, such as hemeproteins and free SH groups. The inhibitory effect of NO on mitochondrial respiration was decreased by exogenously added glutathione (GSH). However, a decrease of intramitochondrial GSH by pretreating animals with l-buthionine sulfoximine had no appreciable effect on the inhibitory effect of isolated mitochondria. Furthermore, the effect of NO was not affected by depleting free SH residues in mitochondria by N-ethylmaleimide. These results suggest that cytosolic but not intramitochondrial GSH might be an important factor that determines the NO-dependent regulation of mitochondrial energy metabolism.

Effect of fluoxetine on rat liver mitochondria

The in vitro and In vivo effects of fluoxetine (and its active metabolite norfluoxetine) on mitochondrial respiration and F 0F 1-ATPase were studied, respectively, in mitochondria and submitochondrial particles isolated from rat liver. Fluoxetine in vitro inhibited state 3 mitochondrial respiration for α-ketoglutarate and succinate oxidations (50% of effect at 0.25 and 0.35 mM drug concentrations, respectively); stimulated state 4 for succinate; and induced a decrease in the respiratory control ratio (RCR) for both oxidizable substrates. The F 0F 1-ATPase activity was determined at various pH levels in the absence and presence of Triton X-100. The solubilized form was not affected markedly, but an inhibition, apparently non-competitive, was observed for the membrane-bound enzyme, with 50% of the effect at a 0.06 mM drug concentration in pH7.4. These results suggest that fluoxetine in vitro acts on F 0F 1ATPase through direct interaction with the membrane F 0 component (similar to oligomycin), or first with mitochondrial membrane and then affecting F 0. A very similar behavior concerning the respiratory parameters and F 0F 1-ATPase properties was observed with norfluoxetine. The in vivo studies with fluoxetine showed stimulation of mitochondrial respiration in state 4 for α-ketoglutarate or succinate oxidations in acute or prolonged treatments (1 hr after a single i.p. dose of 20 mg of drug/kg of body weight, and 22 hr after 12 days of treatment with a daily dose of 10 mg/kg of body weight, respectively), indicating uncoupling of oxidative phosphorylation. Pronounced changes were not observed in the K 0.5 values of F 0F 1-ATPase catalytic sites, but the V max decreased during the prolonged treatment. The results show that fluoxetine (as well as norfluoxetine) has multiple effects on the energy metabolism of rat liver mitochondria, being potentially toxic in high doses. The drug effects seem to be a consequence of the drug and/or metabolite solubilization in the inner membrane of the mitochondria.

Effect of Tordon 2,4-D 64/240 triethanolamine BR on the energy metabolism of rat liver mitochondria.

Tordon herbicide, which is a mixture of 4-amino-3,5,6-trichloropicolinic acid (picloram) and 2,4-dichlorophenoxyacetic acid (2,4-D), depresses the phosphorylation efficiency of the rat liver mitochondria, as inferred from the decrease of the respiratory control coefficient and the ADP/O ratios when NAD(+)-dependent substrates were used; NADH oxidase and NADH cytochrome c reductase were also inhibited, without any effect on the other enzymatic complexes of the respiratory chain. Tordon (66.2 nmol picloram + 270 nmol 2,4-D mg-1 protein) affected the amplitude of swelling induced by glutamate, succinate, (N,N,N',N'-tetramethyl-p-phenyldiamine + sodium ascorbate and ATP. These results characterize an interaction of Tordon with complex I of the respiratory chain and also a partial collapse of the proton motive force of the mitochondrial inner membrane without affecting its elasticity.

Effect of styrene and other alkyl benzene derivatives on oxidation of FAD- and NAD- linked substrates in rat liver mitochondria

The effect on energetic metabolism of rat liver mitochondria (RLM) of styrene and other aliphatic benzene derivatives, i.e. toluene, ethylbenzene, α-methylstyrene and butylbenzene, is studied. It is shown that these compounds uncouple oxidative phosphorlyation and this effect is connected with the stimulation of passive entry of protons into mitochondria. The relationship between hydrophobicity of these compounds and their biological activity and mechanism of uncoupling effect are discussed.

Studies on the effects of saturated and unsaturated short-chain monocarboxylic acids on the energy metabolism of rat liver mitochondria.

The effect of eight branched-chain amino acid metabolites, four metabolites from the beta-oxidation, and the unphysiologic acid 4-pentenoic acid on the oxygen consumption rate of liver mitochondria oxidizing pyruvate, 2-oxo-glutarate, and L-palmitoylcarnitine has been investigated. The 12 metabolites are: propionic, isobutyric, 2-Me-butyric, isovaleric, acrylic, Me-acrylic, tiglic, Me-crotonic, butyric, hexanoic, crotonic, and 2-hexenoic acids. The oxidation rate of pyruvate was strongly inhibited by propionic, 4-pentenoic, and isovaleric acids at 0.1, 0.1, and 1.0 mM, respectively. With 2-oxo-glutarate as substrate, the oxygen consumption rate was strongly inhibited at 0.1 mM of propionic, 4-pentenoic, and isovaleric acids. The L-palmitoyl-carnitine oxidation rate was very strongly inhibited by 0.1 mM 4-petenoic acid, whereas butyric and hexanoic acids exerted a moderate inhibition at 0.1 mM. Propionic acid inhibited L-palmitoylcarnitine oxidation slightly at 1.0 mM. It is argued that propionyl-CoA and isovaleryl-CoA inhibit pyruvate and 2-oxo-glutarate dehydrogenases directly, and the significance of the results for ketotic episodes in organic acidurias is discussed.

Nutritional and hormonal regulation of pyruvate metabolism in the liver.

The effect of fasting, glucose, and glucagon injection on pyruvate metabolism of rat liver mitochondria was studied. Fasting for 24 h caused a) a twofold increase in mitochondrial pyruvate uptake, b) fivefold increase in CO2 fixation, and c) no change in pyruvate decarboxylation. Injection of glucose to fasted rats 2 h prior to preparation suppressed by one-half the increase in mitochondrial pyruvate uptake and CO2 fixation and increased hepatic pyruvate content. Injection of glucagon together with glucose abolished the depression of pyruvate uptake by glucose but did not prevent the decrease in mitochondrial CO2 fixation or hepatic ketone content caused by glucose alone. The effects of insulin injection resembled that of glucose in decreasing hepatic ketone content, but differed by increasing pyruvate uptake without much change in CO2 fixation. It is concluded that the increase in gluconeogenesis induced by fasting is due to an increase in pyruvate uptake and carboxylation by hepatic mitochondria. The latter is due to the increased mobilization and oxidation of fatty acids induced by reciprocal changes in insulin and glucagon.

Defective oxidative metabolism of rat liver mitochondria in hemorrhagic and endotoxin shock.

Defective oxidative metabolism of rat liver mitochondria in hemorrhagic and endotoxin shock.

Ion specific electrochemical behavior of macrotetrolides in membranes

Electrochemical cells using macrotetrolides (nonactin homologs) on inert supports as membranes show a specificity for cations comparable to the ion specific behavior observed in the metabolism of rat liver mitochondria.

Action of diethylstilbestrol and related compounds on the metabolism of rat liver mitochondria

The artificial estrogens stilbestrol, dienestrol, and hexestrol inhibit the oxidation of l-malate- l-glutamate by phosphorylating rat liver mitochondria in metabolic State 3. The inhibition takes place mostly at the NAD-flavoprotein region of the electron transport chain, as with nonphosphorylating particles. Succinate oxidation is also inhibited in State 3 but relatively higher concentrations of estrogens are required to produce the same inhibition as with the NAD-linked substrates. With succinate or ascorbate-TMPD ( N,N,N′,N′-tetramethyl- p-phenylenediamine) substrates, artificial estrogens uncouple oxidative phosphorylation as shown by the diminution of P:O quotients, the release of oligomycin inhibition, and the release of respiratory control. In State 4, stimulation of succinate oxidation by stilbestrol is lesser when the estrogen is added before succinate. This relative inhibition is counteracted by addition of ATP, and also when amytal or orthophosphate are present during preincubation with stilbestrol. According to the estrogen concentrations required to inhibit electron transfer and respiratory control, the latter effect is of minor metabolic importance. At variance with other typical uncouplers (DNP, dicoumarol), the estrogens relieve the inhibition of energy transfer by phenetylbiguanidine (DBI) much less and do not induce significant activation of latent ATPase activity even in the presence of Mg ++. Estrogens may induce physical changes of mitochondria, as revealed by swelling. However, the alteration of mitochondrial structure does not cause per se the metabolic responses to estrogens since stilbestrol monomethyl ether, a relatively powerful swelling agent, scarcely affects respiration. Unsubstituted phenolic groups are essential for the estrogens metabolic actions because stilbestrol dimethyl ether, stilbestrol dipropionate, and hexestrol dimethyl ether do not affect mitochondrial respiration.

Mitochondrial formation and hydrolysis of adenosine triphosphate in the presence of some rare-earth ions

Investigations were made to determine whether the toxic effect of rare earths and the changes in liver lipid metabolism are due to an alteration in adenosine triphosphate (ATP) metabolism in mitochondria. In order to test the possibility that oxidative phosphorylation is impaired by the parenteral administration of lanthanide metal ions prior to manifestation of fatty infiltration of the liver, the metabolism of rat-liver mitochondria was studied 8 hr after the intravenous injection of LaCl/sub 3/ or Pr(NO/sub 3/)/sub 3/. However, the P/O ratios of the liver mitochondria isolated after such pretreatment were found to be normal. The effects of the rare earth ions on non- enzymic hydrolysis of ATP at pH 7.5 to 7.5 were also tested. The data show that the trivalent ions of the rare earths tested do not stimulate the enzymic splitting of ATP by mitochondrial digitonin fragments. The data presented then do not support the view that the ions of La, Pr, Nd, and Sm have a direct toxic effect on the synthesis or breakdown of ATP in mitochondria. (P.C.H.)

Influence of some steroids on the metabolism of rat liver mitochondria and of hepatoma ascites cells in vitro

The effects of some steroids of the testosterone and 19-nortestosterone groups on the phosphorylative oxidation of ketoglutarate by isolated rat liver mitochondria and on the O 2 uptake and aerobic glycolysis of hepatoma AH 130 ascites cells have been compared. The ratios between oxidation inhibiting activity and oxidative phosphorylation uncoupling efficiency of some of the steroids tested are widely different. Both inhibitors of oxidations and uncouplers of oxidative phosphorylations cause an increase of aerobic glycolysis in intact hepatoma ascites cells. Only the oxidation of DPN-dependent substrates is influenced by these compounds. The steroids inhibiting mitochondrial oxidations decrease the rate of external DPNH reoxidation by mitochondria pretreated with hypotonic solutions, some steroids which do not inhibit mitochondrial oxidations have the same effect in a lesser degree. 16α-Hydroxy-17α-methyltestosterone has the highest uncoupling activity that has been so far described for a steroid compound.