Effect of rubomycin (daunorubicin) and its nitroxyl analog on the functioning of rat heart mitochondria

Changes in the functional parameters of the rat heart mitochondria were studied in time after a single intraperitoneal administration of rubomycin, the rubomycin combination with 4-hydroxy-2,2,6,6-tetramethylpiperidine-N-oxyl (TEMPO-OH) and ruboxyl, a nitroxyl derivative of rubomycin. The administration of rubomycin resulted in inhibition of the heart mitochondria bioenergetic functions (a decrease in the respiration control coefficient, RCC, and the respiration rate, RR, on phosphorylation) during respiration in the presence of NAD(+)-dependent substrates 6 to 24 hours after the administration. Later the mitochondria functions recovered while in 2 to 3 weeks a secondary decrease in the RCC and RR was observed. During respiration in the presence of succinate the inhibitory effect on the antibiotics was higher. The combined administration of rubomycin and TEMPO-OH eliminated the primary inhibition. In the presence of ruboxyl the inhibitory effect in regard to the NAD(+)-dependent substrates was not detected. The mechanisms of the toxic action of the anthracycline antibiotics are discussed.     

Effects of daunorubicin and its nitroxyl analog on the synthesis of nucleic acids]

The impact of daunorubicin, emoksil in sublethal doses and daunorubicin mixtures with a nitroxyl radical of 2,2,6,6-tetramethyl-4-oxopiperidine-1-oxyl (NR) on synthesis of DNA and RNA in some organs of rats was studied. Daunorubicin and emoksil induced marked (by 80 to 90%) inhibition of DNA synthesis in all the examined organs even within the first hours of administration. In the heart, DNA synthesis remained lower by the end of the experiment. In the spleen its partial recovery up to 40 to 60% of the control level was observed. In the liver the synthesis was stimulated in 24 hours. Its highest stimulation was recorded with the use of emoksil and daunorubicin in combination with NR. After administration of daunorubicin the maximum synthesis of DNA exceeded the control level 2.5 times. Addition of NR lowered 2-fold inhibition of DNA synthesis by daunorubicin within the first hours. It was of interest that the anthracyclines appeared to markedly stimulate RNA synthesis in the spleen, the fact not described in the literature. The stimulation of DNA synthesis in the liver was supposed to be one of the components of the compensatory mechanisms engaged by the cell in response to the drug's damaging effect.     

In vitro effects of microwave radiation on rat liver mitochondria

Liver mitochondria were exposed in vitro at 30°C to microwave radiation (2.45 GHz) during the following states of respiraton: resting, state 1; substrate dependent, state 2; ADP stimulated, state 3; and ADP depleted, state 4. At 10 or 100 mW/g, with succinate as substrate, no effect of exposure was observed on states 1–4 or the respiratory control index (state 3/state 4) of either tightly or loosely coupled mitochondria. When glutamate was used as substrate, no effects were observed at 10 mW/g. However, in the loosely coupled mitochondria the 100 mW/g exposure produced an increase in states 2 and 4 and a decrease in the respiratory control index. The results suggest that the function of loosely coupled mitochondria can be affected at high power levels of microwave radiation.     

Comparative study on uncoupling effects of laurate and lauryl sulfate on rat liver and skeletal muscle mitochondria

Uncoupling effects of laurate and lauryl sulfate have been studied in the isolated rat liver and skeletal muscle mitochondria. In the oligomycin-treated liver mitochondria, 0.02 mM laurate or 0.16 mM lauryl sulfate caused a two-fold stimulation of respiration, accompanied by a membrane potential decrease. Carboxyatractylate (CAtr) and glutamate (or aspartate) strongly decrease the effect of laurate and lauryl sulfate on respiratory rate and membrane potential (the recoupling effect). With both uncouplers, this effect is maximal for CAtr and glutamate (aspartate) at pH 7.8 and 7.0, respectively. Tetraphenyl phosphonium cations, which decrease negative membrane charges, cause an alkaline shift of these pH dependences. Small amounts of lauryl sulfate, which increase the membrane negative charge, induce the opposite shift when laurate is used as an uncoupler. ADP, but not GDP, partially recouple with both laurate and lauryl sulfate. We conclude that lauryl sulfate-induced uncoupling in rat liver, like the uncoupling induced by laurate, is mediated by the ATP/ ADP and glutamate/aspartate antiporters. In skeletal muscle mitochondria uncoupled by laurate, 200 microM GDP causes partial recoupling which can be enhanced by a subsequent additions of CAtr, glutamate and serum albumin. CAtr added before GDP promotes a larger recoupling than when added after GDP and prevents the subsequent effect of GDP. ADP is effective as recoupler at lower concentrations that GDP, whereas CDP is without influence. Lauryl sulfate uncoupling of skeletal muscle mitochondria is GDP-resistant but is sensitive to ADP, CAtr, glutamate and serum albumin. Our data suggest that in skeletal muscle mitochondria a GDP-sensitive mechanism is involved in uncoupling induced by laurate. This mechanism is absent in liver mitochondria. Possible mechanisms of laurate and lauryl sulfate-induced uncoupling are discussed.     

Effect of thyroxine in vitro on the transmembrane potential of rat liver mitochondria

It has been shown in in vitro experiments that a certain latent period after the addition of thyroxine (T4) and triiodothyronine (T3) was necessary for the manifestation of their effects on transmembrane potential (TMP) of the rat liver mitochondria. The duration of the lag-period decreased upon an increase in the concentrations of these hormones, and T4 at a dose of 2.10(-4) M produced a fall in TMP immediately after its addition. The rate of TMP fall was in proportion with the concentrations of thyroid hormones introduced into the cell, with T3 30-40% more effective than T4. It was established that the action of I2 resembled that of thyroid hormones, namely, a fall in TMP, mitochondrial swelling, activation of transhydrogenase Kl was ineffective. It is suggested that the appearance of the lag-period upon the action of thyroid hormones might be explained by the period of time necessary for the formation of the active iodine forms, as well as by the formation of fatty acids (donators of H+) by mitochondrial phospholipases. All these factors lead to TMP fall resulting in decreased formation of sufficient ATP quantities in mitochondria.     

Substantial quantities of the high energy derivative oligophosphoglyceroyl-ATP are located in mitochondria in rat liver

Perfusion with [8-14C]adenosine demonstrated the likely existence in rat liver of oligophosphoglyceroyl-ATP (OPG-ATP). Purification followed by assay with a new specific 3' phosphodiesterase confirmed this. The quantities present were 5-10-fold those found previously and comparable to total soluble nucleotides. OPG-ATP was also purified from the mitochondrial fraction, shown to co-distribute with succinate dehydrogenase and can be co-purified with an enzyme confined to intermembrane space.     

Effect of morphine in vitro on the oxidative phosphorylation in rat liver mitochondria]

The rates of respiration in the presence of ADP and of phosphorylation as an ATP-ase activity of rat liver mitochondria was inhibited was in vitro by morphine with Ki=6.5 mM. The uncoupler-stimulated respiration of the mitochondria and the activity of ATP-ase and synthesis of ATP in the submitochondrial particles were not altered in the presence of morphine. It is suggested that morphine inhibited the adenine nucleotide transport through the mitochondrial membrane     

The action of defoliants on the energetics of rat liver mitochondria in vitro]

The effect of defoliants butyphos (I), dropp (II), butylcaptacs (III), hinazopin (IV), syhat (V), tetra-n-butylammonium bromide (VI), etrel (VII), gemetrel (VIII), allyl-4-methylpyridinium bromide (IX), 1-amino-cyclopropan-1-carbonate (ACPC) (X) at various concentrations (1 x 10(-5)-2 x 10(-4) M) on respiration, oxidative phosphorylation (OP) and permeability of the inner mitochondrial membrane from rat liver has been studied. It has been established that some of the compounds uncouple OP by increasing the inner mitochondrial membrane permeability for H+ (II) inhibit the respiration in V3 condition and induce less selective permeability for a number of ions (I, III). The other defoliants either induce respiration generally in metabolic states 3 and 4 (IV, VI, IX) or have no effect on the respiration and OP (V, VII, VIII, X). On the whole a good correlation between the common toxicity of the studied preparation (LD50) and their mitochondrial effect has been revealed, therefore the latter can be considered as intracellular "targets" involved in the realization of pesticide action.     

A study on actin-like protein biosynthesis in rat liver mitochondria

The in vitro experiments revealed no incorporation of amino acids into actin-like protein of isolated rat liver mitochondria. The method of pulse label showed the presence of [14C]actin-like protein in mitochondria of intact animals which were not administered cycloheximide. A new synthesized actin-like protein is identified in mitochondria as a labelled polypeptide with apparent molecular weight 42 kDa. The data obtained may evidence for cytoplasmic localization of mitochondrial actin-like protein biosynthesis.     

Rhodamine 123 inhibits bioenergetic function in isolated rat liver mitochondria

Rhodamine 123 accumulates in the mitochondria of living cells and exhibits selective anticarcinoma activity. The biochemical basis of toxicity was investigated by testing the effect of the dye on isolated rat liver mitochondria. Much lower concentrations of rhodamine 123 were required to inhibit ADP-stimulated respiration and ATP synthesis in well-coupled energized mitochondria than were required to inhibit uncoupled respiration and uncoupler-stimulated ATP hydrolysis. The amount of rhodamine 123 associated with the mitochondria was several-fold greater under energized as compared to non-energized conditions, which may explain why coupled functions appeared to be more sensitive than uncoupled functions to inhibition at low concentrations of rhodamine 123. It was concluded that the site of rhodamine 123 inhibition is most likely the F0F1 ATPase complex and possibly electron transfer reactions as well.