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PubMed1992

Endothelins Inhibit Junctional Permeability in Cultured Mouse Astrocytes.

Endothelins, a family of potent vasoconstrictor peptides initially characterized in peripheral tissues, have also been reported to be synthesized in the brain. In this structure several cell types, including astrocytes, endothelial cells and certain neurons, are potential targets for these peptides. In astrocytes, endothelins induce changes in the concentration of several second messengers (calcium, diacylglycerol, arachidonic acid, cAMP) known to be involved in the regulation of gap junction channels. Using the scrape loading/dye transfer technique we have observed that two isoforms of endothelin, endothelin-1 and endothelin-3, strongly inhibit the extent of dye-coupling between confluent astrocytes, suggesting that gap junction permeability was reduced. This inhibitory effect on dye coupling was reproduced by the snake venom sarafotoxin. When used at 10-7 M, these three compounds had inhibitory effects on gap junction channels which were comparable to those induced by the well known uncoupling agents octanol and halothane. In the absence of extracellular calcium, the effects of endothelins were largely prevented, suggesting that second messengers linked to the activation of phospholipases C and/or A2, which both are dependent on external calcium, could be involved in the uncoupling mechanism.

Journal Article↗

PubMed1979

Active K+ transport in Mycoplasms mycoides var. Capri. Relationships between K+ distribution, electrical potential and ATPase activity.

The addition of 5 . 10(-5) M or less of dicyclohexylcarbodiimide to Mycoplasma mycoides var. Capri preferentially influences K+ influx rather than efflux and reduces by 30--40% the activity of the membrane-bound Mg2+- ATPase. Adding valinomycin to metabolizing cells does not markedly affect K+ distribution but induces a rapid and complete loss of intracellular K+ in non-metabolizing cells. Uncoupling agents such as dinitrophenol, carbonylcyanide p-trifluoromethoxyphenylhydrazone, dissipate the K+ concentration gradient only when combined with valinomycin. Variations in the merocyanine fluorescence intensity indicate that a transmembrane electrical potential (delta psi) is generated on cell energization. This delta psi, not affected by valinomycin or uncouplers when used alone, is collapsed by a mixture of both. No change in fluorescence intensity can be detected when glucose is added to dicyclohexylcarbodiimide treated organisms. These experiments suggest that the membrane-bound Mg-ATPase activity control K+ distribution in these organisms through the generation of a transmembrane electrical potential difference.

Adenosine Triphosphatases↗

PubMed1991

2-Ethylhexanol uncouples oxidative phosphorylation in rat liver mitochondria.

2-Ethylhexanol (70 microM), a non-genotoxic carcinogen and peroxisome proliferator, stimulated oxygen uptake in the perfused rat liver by about 10% during the first 10 min of infusion. Perfusion with a higher, hepatotoxic dose of ethylhexanol (3 mM) led to a transient increase in oxygen uptake followed by a rapid inhibition of respiration of over 50% in 10 min. Lactate dehydrogenase (LDH) release, indicative of irreversible cell death, was detected in the effluent perfusate after 20 min. After 10 min of perfusion with ethylhexanol, livers were freeze-clamped, acid extracts were prepared and adenine nucleotides were measured by high-pressure liquid chromatography. Ethylhexanol decreased the ATP/ADP ratio from 2.5 to 0.9. Thus, marked decreases in hepatic energy state due to inhibition of respiration preceded cell death. To attempt to understand this phenomenon, the effect of ethylhexanol on isolated mitochondria was studied. Similar to classical uncoupling agents, ethylhexanol stimulated state-4 rates of respiration, diminished coupled rates of respiration, and decreased the P/O ratio in a dose-dependent manner in isolated mitochondria. Ethylhexanol also decreased uptake of radiolabeled 45CaCl2 by isolated mitochondria 4- to 5-fold. Therefore, we hypothesize that ethylhexanol initially uncouples oxidative phosphorylation leading to diminished ATP synthesis and collapse of ion gradients across the mitochondrial membrane.

Adenosine Diphosphate↗

PubMed1975

Ion transport and respiratory control in vesicles formed from reduced nicotinamide adenine dinucleotide coenzyme Q reductase and phospholipids.

NADH-coenzyme Q reductase from bovine heart mitochondria (complex I) was incorporated into phospholipid vesicles by the cholate dialysis procedure. Mixtures of purified phosphatidylcholine and phosphatidylethanolamine were required. Oxidation of NADH by coenzyme Q1 catalyzed by the reconstituted vesicles was coupled to proton translocation, directed inward, with an H+/2e ratio greater than 1.4. Similar experiments measuring proton translocation in submitochondrial particles gave an H+/2e ratio of 1.8. The proton translocation in both systems was not seen in the presence of uncoupling agents and was in addition to the net proton uptake from the reduction of coenzyme Q1 by NADH. Electron transfer in the reconstituted vesicles also caused the uptake of the permeant anion tetraphenylboron. The rate of electron transfer by the reconstituted vesicles was stimulated about 3-fold by uncouplers or by valinomycin plus nigericin and K+ ions. The results indicate that energy coupling can be observed with isolated NADH-coenzyme Q reductase if the enzyme complex is properly incorporated into a phospholipid vesicle.

Animals↗

PubMedDate not supplied

[Mechanism of low intensity ultrasound effect on mitochondria].

Ultrasound of 0,2 Wt/cm2 intensity affects the ionic transport across the mitochondrial membrane in vitro. In the presence of 1 mM EGTA in the incubation medium ultrasound slows down K+ exit into the external medium after the addition of an uncoupling agent (2,4-dinitrophenol). With the addition of 100 divided by 400 mM Ca2+ to the starting medium ultrasound makes the amount of Ca2+ absorbed by mitochondrial decrease and the rate of Ca2+/H+ electroneutral exchange increase. Without Ca2+ ultrasound does not influence the rate of coupled and 2,4-dinitrophenol uncoupled respiration and oxidative phosphorylation, i.e. does not produce strong functional changes.

2,4-Dinitrophenol↗

PubMed1991

Breath-by-breath measurement of oxygen consumption and FIO2-FEO2 with increased oxygen demand.

Continuous on-line breath-by-breath measurement of pulmonary gas exchange was used to monitor the increase in oxygen uptake (VO2) and carbon dioxide excretion (VCO2) induced by the oxidative phosphorylation uncoupling agent 2, 4-dinitrophenol (DNP) in 10 dogs. With incremental doses of DNP totaling 5 mg/kg, the continuously monitored VO2 increased within 2-3 min after the first injection of the drug. VCO2 showed a similar response 4-6 min after the first injection. Temperature increase due to the pharmacological oxidative phosphorylation uncoupling required 20-30 min for a discernible change at this dose. This study also demonstrated a modified and compromised response to the drug in dogs where oxygen delivery was limited by mechanical ventilation.

Animals↗

PubMed2005

Connexin-made channels as pharmacological targets.

Gap junctions are clusters of intercellular channels that provide morphological support for direct diffusion of ions and low-molecular-weight molecules between adjacent coupled cells. Each gap junction channel is made by docking of two hemichannels or connexons, each formed by assembly of six proteins (connexins). 21 members of the connexin gene family are likely to be expressed in the human genome. These ubiquitous gated channels, allowing rapid intercellular communication and synchronisation of coupled cell activities, play critical roles in many signalling processes, including co-ordinated cardiac and smooth muscle contractions, neuronal excitability, neurotransmitter release, insulin secretion, epithelial electrolyte transport, etc. Mutational alterations in the connexin genes are associated with the occurrence of multiple pathologies, such as peripheral neuropathies, cardiovascular diseases, dermatological diseases, hereditary deafness and cataract. But the neuro- and cardioprotective effects of blocking agents of junctional channels show that closure of these channels may also be beneficial in certain pathological situations. Consequently, modulation of gap junctional intercellular communication is a potential pharmacological target. In contrast to most other membrane channels, no natural toxin or specific inhibitor of junctional channels has been identified yet and most uncoupling agents generally also affect other ionic channels and receptors. Future research, based for example on the recent developments in genetics, may clarify gap junction physiology. This will in turn provide promising perspectives for the development of targeted drugs.

Amino Acid Sequence↗

PubMed1987

Escherichia coli mutants resistant to uncouplers of oxidative phosphorylation.

Two mutant strains of Escherichia coli K 12 Doc-S resistant to the uncoupling agents 4,5,6,7-tetrachloro-2-trifluoromethyl benzimidazole and carbonyl cyanide m-chlorophenylhydrazone were isolated. These strains, designated TUV and CUV, were capable of (a) growth, (b) the transport of succinate and L-proline and (c) electron-transport-linked oxidative synthesis of ATP in the presence of titres of uncoupler which inhibited these processes in strain Doc-S. The inhibition of transport of L-proline by a fixed titre of uncoupler was sharply pH dependent in strain Doc-S: uptake was unaffected at pH 7.6 but completely inhibited at pH 5.6. This pH dependence was not shown by the resistant strains. We believe that uncouplers were equally accessible to their site(s) of action in the energy-conserving membrane of the sensitive and resistant strains. We conclude that uncoupler resistance in these strains of E. coli has arisen as a consequence of mutations which directly affect a specific site of uncoupler action within the cytoplasmic membrane, rather than as a consequence of a decrease in the permeability of cells to uncoupler.

Benzimidazoles↗

PubMed1983

Lack of direct antiarrhythmic electrophysiological effects of salicylate on isolated guinea-pig myocardium.

Conventional microelectrode techniques were used to study the influence of Na-salicylate, Na-benzoate, Na-2,6-dihydroxybenzoate and 2,4-dinitrophenol (2,4-DNP) on the action potential (AP) of guinea-pig papillary muscles and atria. In papillary muscles, Na-salicylate (0.19-1.87 mmol/l) concentration-dependently shortened the AP duration and the functional refractory period. The AP amplitude decreased slightly with the largest concentration, while the resting potential and the maximum depolarisation velocity (Vmax) were not affected. A concentration-dependent negative inotropic effect was also seen. All drug effects were reversible after washout. In atria, 6.24 mmol/l Na-salicylate induced a slight shortening of the AP duration, a decrease of the AP amplitude and Vmax, but no decrease of the contractile force. The effects of the uncoupling agent, 2,4-DNP (10 mumol/l), were similar to those of the largest concentration of Na-salicylate in papillary muscles and in atria. Na-benzoate and Na-2,6-dihydroxybenzoate had no significant influence on AP duration, AP amplitude, resting potential, Vmax, refractory period or force of contraction of either papillary muscles or atria. These results suggest that Na-salicylate exerts its effects on isolated guinea-pig myocardium by uncoupling the oxidative phosphorylation, whereas two other possible mechanisms of action, namely an increase of membrane surface charge and an inhibition of prostaglandin synthesis, seem to be of minor importance.

2,4-Dinitrophenol↗

PubMed1967

Respiration-driven proton translocation in rat liver mitochondria.

1. Pulses of acidity of the outer aqueous phase of rat liver mitochondrial suspensions induced by pulses of respiration are due to the translocation of H(+) (or OH(-)) ions across the osmotic barrier (M phase) of the cristae membrane and cannot be attributed to the formation (with acid production) of a chemical intermediate that subsequently decomposes. 2. The effective quantity of protons translocated per bivalent reducing equivalent passing through the succinate-oxidizing and beta-hydroxybutyrate-oxidizing spans of the respiratory chain are very close to 4 and 6 respectively. These quotients are constant between pH5.5 and 8.5 and are independent of changes in the ionic composition of the mitochondrial suspension medium provided that the conditions permit the accurate experimental measurement of the proton translocation. 3. Apparent changes in the -->H(+)/O quotients may be induced by conditions preventing the occurrence of the usual backlash; these apparent changes of -->H(+)/O are attributable to a very fast electrically driven component of the decay of the acid pulses that is not included in the experimental extrapolations. 4. Apparent changes in the -->H(+)/O quotients may also be induced by the presence of anions, such as succinate, malonate and phosphate, or by cations such as Na(+). These apparent changes of -->H(+)/O are due to an increase in the rate of the pH-driven decay of the acid pulses. 5. The uncoupling agents, 2,4-dinitrophenol, carbonyl cyanide p-trifluoromethoxyphenylhydrazone and gramicidin increase the effective proton conductance of the M phase and thus increase the rate of decay of the respiration-driven acid pulses, but do not change the initial -->H(+)/O quotients. The increase in effective proton conductance of the M phase caused by these uncouplers accounts quantitatively for their uncoupling action; and the fact that the initial -->H(+)/O quotients are unchanged shows that uncoupler-sensitive chemical intermediates do not exist between the respiratory-chain system and the effective proton-translocating mechanism. 6. Stoicheiometric acid-base changes associated with the activity of the regions of the respiratory chain on the oxygen side of the rotenone- and antimycin A-sensitive sites gives experimental support for a suggested configuration of loop 3.

Journal Article↗

PubMed2003

Uncoupling of horizontal cells alters the receptive fields of retinal bipolar cells.

Effects of uncoupling of horizontal cells by 1-octanol, a non-specific gap junction uncoupling agent, on the receptive field organization of cone-dominant bipolar cells were investigated in isolated, superfused carp retina, using intracellular recording techniques. At 1 mM, 1-octanol increased responses of cone driven horizontal cells to light spots, but decreased those to light annuli, indicating a reduction of the receptive field size of these cells by uncoupling. Furthermore, 1-octanol eliminated the surround response of OFF type bipolar cells and increased their center response. Similar effects of 1-octanol on the center and surround responses were observed in ON type bipolar cells. These results suggest that uncoupling of horizontal cells can significantly alter the receptive field organization of retinal bipolar cells.

1-Octanol↗

PubMed1997

The immunosuppressant drug FK506 ameliorates secondary mitochondrial dysfunction following transient focal cerebral ischemia in the rat.

Recirculation following 2 h of focal ischemia due to transient middle cerebral artery (MCA) occlusion has previously been found to be accompanied by an initial, partial recovery of the cellular bioenergetic state and of mitochondrial respiratory functions, with secondary deterioration during the first 2-4 h of reflow. Both the free radical spin trap alpha-phenyl-N-tert-butyl nitrone (PBN) and the immunosuppressant drug FK506 ameliorate the damage incurred by the 2-h period of focal ischemia, even when given 1-3 h after the start of the recirculation. The primary objective of this study was to find out if FK506, like PBN, prevents the secondary deterioration of mitochondrial function, as this can be studied in vitro. Since this proved to be the case, we addressed the question of whether the secondary mitochondrial dysfunction and bioenergetic failure were related to a secondary compromise of microcirculation and cellular oxygen delivery. Six groups of male Wistar rats were studied for measurement of mitochondrial respiratory activity (total, n = 36). One group was used as control (n = 6). In the other groups of animals, MCA occlusion of 2 h duration was induced by an intraluminal filament technique, Neocortical focal and perifocal ("penumbra") tissues were sampled after 2 h of ischemia (n = 6) and after 1 h (n = 6), 2 h (n = 6 with vehicle), and 4 h (n = 6 with vehicle; n = 6 with FK506) of recirculation. The vehicle or 1.0 mg.kg-1 of FK506 was injected intravenously after 1 h of recirculation. Homogenates were prepared, and stimulated (+ADP), nonstimulated (-ADP), and uncoupled respiratory rates were measured polarographically. The uncoupling agent used was carbonyl cyanide m-chlorophenylhydrazone. Local CBF and tissue oxygen tension were evaluated by laser-Doppler flowmetry and PO2 microelectrodes, respectively, throughout the whole periods of 2 h of ischemia and 4 h of recirculation, using a remote MCA occlusion technique. After 2 h of ischemia, the penumbra showed a moderate decrease and the focus a marked decrease in ADP-stimulated and uncoupled respiratory rates, with a marked fall in the respiratory control ratio, defined as ADP-stimulated divided by nonstimulated respiration. Recirculation (1 h) brought about partial recovery, but continued reflow (2 and 4 h) was associated with a secondary deterioration of respiratory functions. The secondary deterioration was prevented by FK506. The results thus confirm previous findings showing that secondary mitochondrial dysfunction occurs following transient focal cerebral ischemia and demonstrate that FK506, like PBN, improves the in vitro performance of mitochondria in focal and penumbral areas. Following MCA occlusion, local CBF in a penumbral area and tissue PO2 in a focal area decreased to about 30 and 5% of control, respectively. However, recirculation brought about rapid recovery of blood flow and oxygen delivery. During the whole 4-h period of recirculation, local CBF and tissue PO2 were maintained close to 100% and at about 160% of the preischemic level, respectively. The results make it highly unlikely that the secondary bioenergetic failure during recirculation is due to a compromised microcirculation. It follows that oxygen delivery is not rate-limiting for recovery events. Very likely, FK506 (and PBN) acts at the cellular level to improve mitochondrial energy functions.

Adenosine Diphosphate↗

PubMed1994

Effect of methyl methacrylate on mitochondrial function and structure.

1. Treatment of isolated rat liver mitochondria with methyl methacrylate (MM) produced membrane disruption as evidenced by the release of citrate synthase, and changes in the ultrastructure of mitochondria. 2. At concentration 0.1%, MM uncoupled oxidative phosphorylation as evidenced by stimulation of state 4 respiration supported either by pyruvate plus malate or succinate (+rotenone) and ATP-ase activity in intact mitochondria. 3. At concentration 1% MM stimulated ATP-ase activity in intact mitochondria and succinate (+ rotenone) oxidation at state 4 and was without effect on this substrate oxidation at state 3. 4. MM inhibited pyruvate plus malate oxidation either at state 3 or in the presence of uncoupling agents. 5. MM inhibited the NADH oxidase of electron transport particles at a concentration which failed to inhibit either succinic oxidase or the NADH-ferricyanide reductase activity. 6. The data presented suggest that in the isolated mitochondria MM inhibits NADH oxidation in the vicinity of the rotenone sensitive site of complex I. 7. The general conclusion is that MM may block an electron transport and to uncouple oxidative phosphorylation in rat liver mitochondria. The overall in vitro effect would be to prevent ATP synthesis which could result in cell death under in vivo conditions.

Adenosine Triphosphatases↗

PubMed2004

Higher respiratory activity decreases mitochondrial reactive oxygen release and increases life span in Saccharomyces cerevisiae.

Increased replicative longevity in Saccharomyces cerevisiae because of calorie restriction has been linked to enhanced mitochondrial respiratory activity. Here we have further investigated how mitochondrial respiration affects yeast life span. We found that calorie restriction by growth in low glucose increased respiration but decreased mitochondrial reactive oxygen species production relative to oxygen consumption. Calorie restriction also enhanced chronological life span. The beneficial effects of calorie restriction on mitochondrial respiration, reactive oxygen species release, and replicative and chronological life span could be mimicked by uncoupling agents such as dinitrophenol. Conversely, chronological life span decreased in cells treated with antimycin (which strongly increases mitochondrial reactive oxygen species generation) or in yeast mutants null for mitochondrial superoxide dismutase (which removes superoxide radicals) and for RTG2 (which participates in retrograde feedback signaling between mitochondria and the nucleus). These results suggest that yeast aging is linked to changes in mitochondrial metabolism and oxidative stress and that mild mitochondrial uncoupling can increase both chronological and replicative life span.

Mitochondria↗

PubMed2001

Efficacy of antipsychotic agents at human 5-HT(1A) receptors determined by [3H]WAY100,635 binding affinity ratios: relationship to efficacy for G-protein activation.

5-HT(1A) receptors are implicated in the aetiology of schizophrenia. Herein, the influence of 15 antipsychotics on the binding of the selective 'neutral' antagonist, [3H]WAY100,635 ([3H]N-[2-[4-(2-methoxyphenyl)-1-piperazinyl]ethyl]-N-(2-pyridinyl)-cyclo-hexanecarboxamide), was examined at human 5-HT(1A) receptors expressed in Chinese Hamster Ovary cells. In competition binding experiments, 5-HT displayed biphasic isotherms which were shifted to the right in the presence of the G-protein uncoupling agent, GTPgammaS (100 microM). In analogy, the isotherms of ziprasidone, quetiapine and S16924 (((R-2-[1-[2-(2,3-dihydro-benzo[1,4]dioxin-5-yloxy)-ethyl]-pyrrolidin-3yl]-1-(4-fluoro-phenyl)-ethanone), were displaced to the right by GTPgammaS, consistent with agonist actions. Binding of several other antipsychotics, such as ocaperidone, olanzapine and risperidone, was little influenced by GTPgammaS. Isotherms of the neuroleptics, haloperidol, chlorpromazine and thioridazine were shifted to the left in the presence of GTPgammaS, suggesting inverse agonist properties. For most ligands, the magnitude of affinity changes induced by GTPgammaS (alteration in pK(i) values) correlated well with their previously determined efficacies in [35S]GTPgammaS binding studies [Eur. J. Pharmacol. 355 (1998) 245]. In contrast, the affinity of the 'atypical' antipsychotic agent, clozapine, which is a known partial agonist at 5-HT(1A) receptors, was less influenced by GTPgammaS. When the ratio of high-/low-affinity values was plotted against efficacy, hyperbolic isotherms were obtained, consistent with a modified ternary complex model which assumes that receptors can adopt active conformations in the absence of agonist. In conclusion, modulation of [3H]-WAY100,635 binding by GTPgammaS differentiated agonist vs. inverse agonist properties of antipsychotics at 5-HT(1A) receptors. These may contribute to differing profiles of antipsychotic activity.

Animals↗

PubMed1975

Reconstitution of ion transport and respiratory control in vesicles formed from reduced coenzyme Q-cytochrome c reductase and phospholipids.

Reduced coenzyme Q-cytochrome c reductase from bovine heart mitochondria (complex III) was incorporated into phospholipid vesicles by the cholate dialysis procedure. Soybean phospholipids or mixtures of purified phosphatidylcholine, phosphatidylethanolamine, and cardiolipin could be used. Oxidation of reduced coenzyme Q2 by the reconstituted vesicles with cytochrome c as oxidant showed the following energy-coupling phenomena. 1. Protons were translocated outward with a coupling ratio, H+/2e, of 1.9 +/- 0.2. Measurements with mitochondria under similar conditions showed an H+/2e ratio of 1.8. Proton translocation was not seen in the presence of uncoupling agents and was in addition to the net acidification of the medium from the over-all oxidation reaction. 2. Potassium ions were taken up by the reconstituted vesicles in the presence of valinomycin in a reaction coupled to electron transfer. The coupling ratio for K+ uptake, K+/2e, was 2.0 in the vesicles and approximately 1.5 in mitochondria. 3. The rate of oxidation of reduced coenzyme Q2 by the reconstituted vesicles was stimulated up to 10-fold by uncouplers or by valinomycin plus nigericin and K+ ions. Addition of valinomycin alone in a K+ medium caused a transient stimulation of electron transfer. The results indicate that energy coupling can be observed with isolated reduced coenzyme Q-cytochrome c reductase if the enzyme complex is properly incorporated into a phospholipid vesicle.

Animals↗

PubMed1998

Ineffectiveness of oxytocin on intercellular communication between term pregnant human myometrial cells before labor.

OBJECTIVE: The effects of the uterotonic agent oxytocin on intercellular communication between term pregnant human myometrial cells before labor were studied to investigate its contribution to the synchronicity of uterine contractions. STUDY DESIGN: The membrane potential and the input resistance of myometrial cells were measured with intracellular micropipettes, and dye-coupling assays were concomitantly performed while the tissues were perfused with three concentrations of oxytocin (10(-8) mol/L, 10(-7) mol/L, and 10(-6) mol/L). The results were compared with those obtained from tissues perfused with gap junctional uncoupling agent octanol and those with Tyrode's solution (control). RESULTS: Octanol increased input resistance and inhibited dye coupling but did not affect membrane potential compared with controls. Oxytocin depolarized membrane potential at all studied concentrations. The mean input resistance values and detectable dye propagation with oxytocin were similar to controls. CONCLUSION: Oxytocin does not contribute to gap junctional intercellular communication in term pregnant human myometrium before labor but may affect contractions by depolarizing membrane potential.

Cell Communication↗

PubMed1984

Influence of fatty acids on energy metabolism. 2. Kinetics of changes in metabolic rates and changes in subcellular adenine nucleotide contents and pH gradients following addition of octanoate and oleate in perfused rat liver.

The uncoupling-like effect of fatty acids [ Scholz , R., Schwabe , U., and Soboll , S. (1984) Eur. J. Biochem. 141, 223-230] was further substantiated in experiments with perfused rat livers by two ways: firstly the kinetics of changes in metabolic rates (oxygen consumption, ketogenesis, fatty acid oxidation) were analysed; secondly subcellular contents of adenine nucleotides and pH gradients across the mitochondrial membrane were determined following fractionation of freeze-fixed and dried tissues in non-aqueous solvents. The following results were obtained. The relaxation kinetics of the increase in oxygen consumption following fatty acid infusion revealed two components, a rapid one with a half-time around 10 s and a slow one with a half-time of more than 100 s. The rapid component was similar to the kinetics of fatty acid oxidation (ketogenesis and 14CO2 production from labelled fatty acids) whereas the half-time of the slow component was in the range of half-times observed with the increase in oxygen consumption following addition of carbonylcyanide p-trifluoromethoxyphenylhydrazone. In the presence of fatty acids, the cytosolic ATP concentrations and ATP/ADP ratios decreased, whereas the corresponding parameters for the mitochondrial space were either increased (oleate) or decreased (octanoate). The effects of oleate were dependent on the albumin concentrations in the perfusate. The normally large difference between cytosolic and mitochondrial ATP/ADP ratios became smaller. Similar observations were obtained with uncoupling agents. The pH gradient across the mitochondrial membrane as calculated from the subcellular distribution of 5,5 dimethyl[2-14C]oxazolidine-2,4-dione was inversed following the addition of both carbonylcyanide p-trifluoromethoxyphenylhydrazone and fatty acids, i.e. the mitochondrial matrix became more acidic than the cytosol. The pH gradient was not affected when oleate was added in the presence of high albumin concentrations. The data support the hypothesis that the increase in hepatic oxygen consumption due to octanoate or oleate is, in part, caused by a mechanism similar to uncoupling of oxidative phosphorylation. This mechanism seems not to be an artifact of isolated systems; it may be of physiological importance for processes in which reducing equivalents are removed independently of the ATP demand of the hepatocyte.

Adenine Nucleotides↗