oligomycins and malonic-acid

We investigated the mechanism by which 3,5-diiodo-l-thyronine (T2) affects skeletal muscle mitochondrial bioenergetic parameters following its acute administration to hypothyroid rats. One hour after injection, T2 increased both coupled and uncoupled respiration rates by +27% and +42%, respectively. Top-down elasticity analysis revealed that these effects were the result of increases in the substrate oxidation and mitochondrial uncoupling. Discriminating between proton-leak and redox-slip processes, we identified an increased mitochondrial proton conductance as the "pathway" underlying the effect of T2 on mitochondrial uncoupling. As a whole, these results may provide a mechanism by which T2 rapidly affects energy metabolism in hypothyroid rats.

Topics: Animals; Carbonyl Cyanide p-Trifluoromethoxyphenylhydrazone; Cell Respiration; Diiodothyronines; Energy Metabolism; Hypothyroidism; Kinetics; Male; Malonates; Membrane Potential, Mitochondrial; Mitochondria, Muscle; Muscle, Skeletal; Oligomycins; Oxidation-Reduction; Phosphorylation; Protons; Rats; Rats, Wistar

We have studied the effect of blockade of mitochondrial respiration on the binding of human 125I-TNF alpha to L929 cell receptors. Specific TNF alpha binding was decreased to about 20-40% of controls by blocking mitochondrial respiration. This effect was dose- and time-related and was observed independently of the level at which the respiration was blocked (respiratory chain, proton backflow, ATPase, anaerobiosis). This blockade had no effect on the half-life of the specific TNF alpha binding, the internalization or degradation of TNF alpha-receptor complexes, or the number of TNF alpha-binding sites. Scatchard analysis of TNF alpha binding data indicated a 2-4-fold decrease in the affinity of these binding sites. These effects did not appear to be related to the protein kinase C activity or to reactive oxygen radicals, since they were not antagonized by pretreatment of cells with oxygen radical scavengers, deferoxamine, or inhibitors of protein kinase C. Decrease in TNF alpha binding capacity correlated significantly with cellular ATP content (r = 0.94; p < 0.01) and with the cytocidal activity of TNF alpha against L929 cells. These findings suggest that blockade of mitochondrial respiration down-regulates the binding of TNF alpha to cells, most likely by changing the affinity of receptors for this cytokine. This down-regulation may increase the resistance of cells to TNF alpha cytotoxicity.

Topics: 2,4-Dinitrophenol; Anaerobiosis; Animals; Antimycin A; Dinitrophenols; Down-Regulation; Free Radical Scavengers; Humans; Kinetics; L Cells; Malonates; Mice; Mitochondria; Oligomycins; Oxygen Consumption; Potassium Cyanide; Receptors, Tumor Necrosis Factor; Recombinant Proteins; Rotenone; Sodium Fluoride; Thenoyltrifluoroacetone; Uncoupling Agents

It is well established that calcium ionophore A 23187 induces acrosome reaction (AcR) of uncapacitated spermatozoa in the presence of extracellular Ca2+ ions. In the present study, we have investigated how extracellular energy substrates (glucose, pyruvate, and lactate) affect the ionophore-induced AcR of guinea pig spermatozoa. It was found that 0.3 microM concentration of A 23187 had the maximum effect to initiate AcR of guinea pig spermatozoa. Virtually no spermatozoa underwent their AcR when incubated in substrate-free modified Tyrode's medium containing 0.3 microM A 23187 and 2 mM Ca2+. At least one exogenous substrate is essential for the ionophore-induced AcR of spermatozoa. As for efficacy of the substrates, lactate was more effective than pyruvate and glucose. However, a better result was observed when lactate was added along with pyruvate. Malonate inhibited the ionophore-induced AcR but not the hyperactivated motility of spermatozoa. The mitochondrial electron transport chain blockers rotenone, antimycin, and oligomycin failed to inhibit AcR, although in the presence of these blockers spermatozoa were unable to show hyperactivated motility. These results suggest that the mitochondrial citric acid cycle, not the electron transport chain, is probably the energy source for ionophore-induced AcR of guinea pig spermatozoa.

Topics: Acrosome; Animals; Antimycin A; Calcimycin; Energy Metabolism; Glucose; Guinea Pigs; Lactates; Male; Malonates; Mitochondria; Oligomycins; Pyruvates; Rotenone

We have determined the relationship between rate of respiration and protonmotive force in oligomycin-inhibited liver mitochondria isolated from euthyroid, hypothyroid and hyperthyroid rats. Respiration rate was titrated with the respiratory-chain inhibitor malonate. At any given respiration rate mitochondria isolated from hypothyroid rats had a protonmotive force greater than mitochondria isolated from euthyroid controls, and mitochondria isolated from hyperthyroid rats had a protonmotive force less than mitochondria isolated from euthyroid controls. In the absence of malonate mitochondrial respiration rate increased in the order hypothyroid less than euthyroid less than hyperthyroid, while protonmotive force increased in the order hyperthyroid less than euthyroid less than hypothyroid. These findings are consistent with a thyroid-hormone-induced increase in the proton conductance of the inner mitochondrial membrane or a decrease in the H+/O ratio of the respiratory chain at any given protonmotive force. Thus the altered proton conductance or H+/O ratio of mitochondria isolated from rats of different thyroid hormone status controls the respiration rate required to balance the backflow of protons across the inner mitochondrial membrane. We discuss the possible relevance of these findings to the control of state 3 and state 4 respiration by thyroid hormone.

Topics: Animals; Hyperthyroidism; Hypothyroidism; Male; Malonates; Membrane Potentials; Mitochondria, Liver; Oligomycins; Oxygen Consumption; Phosphorylation; Protons; Rats; Rats, Inbred Strains; Thyroid Hormones

In brain, heart and kidney, cell work in the absence of oxygen has been thought to precipitate anoxic damage by increasing the rate of depletion of cellular energy stores. In the medullary thick ascending limb of isolated perfused rat kidneys, however, reduction of ATP synthesis by a variety of mitochondrial or metabolic inhibitors caused ATP depletion comparable to that produced by oxygen deprivation but did not reproduce the lesions of anoxia. In these cells, unrestrained mitochondrial activity may be an important source of anoxic injury.

Topics: Adenosine Triphosphate; Animals; Antimycin A; Cyanides; Deoxyglucose; Energy Metabolism; Hypoxia; Kidney Medulla; Malonates; Mitochondria; Oligomycins; Oxygen Consumption; Rats; Rotenone

The relationship between the respiration rate and the magnitude of the electrochemical proton potential (delta mu H+) in rat liver mitochondria was investigated. (1) Under the active-state conditions, the action of inhibitors of either phosphorylation (oligomycin) or respiration (rotenone, malonate) on the respiration and delta mu H+ was measured. Both inhibitors diminished the respiration, whereas rotenone resulted in a decrease of delta mu H+, and oligomycin produced an increase of this potential. The effect of the inhibitors was much more pronounced on the respiration rate than on delta mu H+; for example, the excess of oligomycin produced a 90% inhibition of the respiration while delta mu H+ was changed only by 9%. (2) Under the resting-state conditions, small concentrations of the uncoupler stimulated the respiration while changing delta mu H+ to a relatively small extent. The uncoupler concentrations which doubled and tripled the respiration rate produced only 5 and 9% decrease of delta mu H+, respectively. (3) The present results enabled us to propose a model describing the interrelationship between respiration and delta mu H+.

Topics: Animals; Homeostasis; Malonates; Mitochondria, Liver; Oligomycins; Oxygen Consumption; Phosphorylation; Protons; Rats; Rotenone

In oxidative phosphorylation and ATP-driven uphill electron transfer from succinate to NAD, double-reciprocal plots of rates vs. substrate concentrations of the energy-driven reactions are a family of parallel lines at several fixed subsaturating concentrations of the substrates or at several moderate concentrations of the inhibitors of the energy-yielding reactions. Thus, as shown elsewhere [Hatefi, Y., Yagi, T., Phelps, D. C., Wong, S.-Y., Vik, S. B., & Galante, Y. M. (1982) Proc. Natl. Acad. Sci. U.S.A. 79, 1756-1760], partial uncoupling decreases the Vappmax and increases the Kappm of the substrates of the energy-driven reactions, resulting in a decrease of Vmax/Km as a function of increased uncoupling. However, partial limitation of the flow rates of the energy-yielding reactions decreases both the Vappmax and the Kappm of the substrates of the energy-driven reactions, resulting in no change in Vmax/Km. This is true as long as the rate limitation is moderate (e.g., less than 60%), under which conditions the steady-state membrane potential (delta psi) remains essentially unchanged. At high inhibition of the energy-yielding reactions, or at moderate inhibition in the presence of low levels of an uncoupler to cause partial uncoupling, then the family of double-reciprocal plots is no longer parallel and tends to converge toward the left. Under these conditions, steady-state delta psi and Vmax/Km also decrease as inhibition is increased. The relationship between the magnitude of steady-state delta psi and the rate of the energy-driven reaction was studied in oxidative phosphorylation, ATP-driven electron transfer from succinate to NAD, and respiration-driven uniport calcium transport by intact mitochondria.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Adenosine Triphosphate; Animals; Azides; Calcium; Carbonyl Cyanide m-Chlorophenyl Hydrazone; Cattle; Kinetics; Malonates; Mitochondria, Heart; NAD; Oligomycins; Oxidation-Reduction; Oxidative Phosphorylation; Oxygen Consumption; Rotenone; Uncoupling Agents