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In this work, it was found that the ability of common uncouplers - carbonyl cyanide p-trifluoromethoxyphenylhydrazone (FCCP) and 2,4-dinitrophenol (DNP) - to reduce membrane potential of isolated rat liver mitochondria was diminished in the presence of millimolar concentrations of the known cytochrome c oxidase inhibitor - cyanide. In the experiments, mitochondria were energized by addition of ATP in the presence of rotenone, inhibiting oxidation of endogenous substrates via respiratory complex I. Cyanide also reduced the uncoupling effect of FCCP and DNP on mitochondria energized by succinate in the presence of ferricyanide. Importantly, cyanide did not alter the protonophoric activity of FCCP and DNP in artificial bilayer lipid membranes. The causes of the effect of cyanide on the efficiency of protonophoric uncouplers in mitochondria are considered in the framework of the suggestion that conformational changes of membrane proteins could affect the state of lipids in their vicinity. In particular, changes in local microviscosity and vacuum permittivity could change the efficiency of protonophore-mediated translocation.

Apoptosis or programmed cell death is a gene-controlled process of cell self-destruction. One of the earliest manifestations of apoptosis, which precedes morphological changes, is a decrease of mitochondrial membrane potential. Here we show that neither inhibitors of the mitochondrial respiratory chain (rotenone and antimycin) nor an uncoupler of oxidative phosphorylation (carbonyl cyanide m-chlorophenylhydrazone), agents which decrease the mitochondrial membrane potential, induce DNA internucleosomal fragmentation, but all of them markedly prevent fragmentation induced either by glucocorticoids or the Ca2+ ionophore A23187. A similar effect was also observed in the presence of a mitochondrial ATPase inhibitor (oligomycin). The inhibition of DNA internucleosomal fragmentation can be explained by the ability of inhibitors to prevent the mitochondrial permeability transition--a key event in apoptosis induction.

Spectroscopic evidence is presented which indicates that the anionic uncoupler carbonyl cyanide-4-nitro-2-chloro-phenylhydrazone and the amine local anesthetics form a complex in aqueous solution. The complex formation studies were carried out for several pharmacologically important tertiary amines and some primary amines. Their relative potencies to form a complex with uncoupler have followed the order: procaine < trimecaine < tetracaine < dibucaine < dodecylamine < dicyclohexylamine < hexadecylamine. As to the more lipophilic nature of the complex the emphasized penetration into octanol and reinforced retention into mitochondria was observed. The higher ability of the complex to colapse the mitochondrial membrane potential confirms this fact. The effective concentration of amine local anesthetics to form a complex was correlated with their physicochemical properties namely lipophilicity and acidobasicity. The highest effectivities for complex formation is shown by the most lipophilic and the most ionized molecules of amines. Present results point to the importance of considering the role of amine anesthetic-uncoupler complex in interpreting physiological or ion transport data in which these substances have been used together.

The metabolic consequences of the uncoupling effect of phenylhydrazonopropanedinitrile and others uncouplers of oxidative phosphorylation on Ehrlich ascites carcinoma (EAC) cells were investigated. Upon application of uncouplers in concentrations stimulating the respiration of EAC cells the accelerate glucose uptake and lactate production was observed. The maximal glycolysis stimulation was fourfold in relation to control at the given experimental conditions. Simultaneously the degree of conversion of glucose on lactate was increased. The acceleration of glycolysis was accompanied by stimulation of 14C-labeled adenine and valine incorporation indicating the increased rate of biosynthetic processes. The prolongation of uncoupler action time and application of their higher concentrations cause the inhibition of glycolysis and biosynthetic processes which is evoked with nonspecific effects of the compounds.

Phenylhydrazonopropanedinitrile, a model uncoupler of oxidative phosphorylation, was used in studies of metabolic and physiological consequences of uncoupling at the cellular level in Candida albicans. Concentrations stimulating respiration induce a faster glucose consumption at a practically unchanged respiratory coefficient. The extracellular production of acids is also without significant changes. When applying higher concentrations of the uncoupler respiration was inhibited, similarly to glucose consumption and acid production. This fact is due to nonspecific interactions of the alkylation type with mercapto groups of functional proteins. Phenylhydrazonopropanedinitrile influences energy-generating processes resulting in slowing down or interruption of biosynthetic processes and occasionally even growth of Candida albicans.