Effect of trimecaine, ajmaline, stenopril and chloracizine on fluctuations in K+ currents in rat liver mitochondria

Antianginal hypotensive preparations chloracyzin and stenopryl, antiarrhythmic aimalin as well as local anesthetic and antiarrhythmic trimekaine hydrochloride were studied for their effect on the K+ inflow and outflow rate in the rat liver mitochondria during Sr2+-induced vibrations. In spite of differences in the chemical structure and pharmacological effect, all these substances are shown to uniformly suppress ion flow vibrations in mitochondria, inhibiting K+ outflow. It is found that the inhibition of the K+ outflow rate depends on the concentration of the preparations. Activity of the studied substances as to inhibition of K+ outflow from mitochondria correlates with their pharmacologic activity.     

Calcium ion cycling in rat liver mitochondria

1. Addition of N-ethylmaleimide to rat liver mitochondria respiring with succinate as substrate decreases both the initial rate of Ca(2+) transport and the ability of mitochondria to retain Ca(2+). As a result, Ca(2+) begins to leave the mitochondria soon after it has entered. Half-maximal effects occur at an N-ethylmaleimide concentration of about 100nmol/mg of protein. 2. The efflux of Ca(2+) induced by N-ethylmaleimide is not prevented by Mg(2+) or by Ruthenium Red at concentrations known to prevent Ca(2+) efflux when exogenous phosphate also is present. Swelling of mitochondria does not accompany N-ethylmaleimide-induced Ca(2+) efflux. 3. Addition of Ca(2+) to rat liver mitochondria in the presence of N-ethylmaleimide produces an immediate decrease in DeltaE (membrane potential), which decreases further to only a slight extent over the next 8min. Concomitant with this is an immediate increase and then levelling off of the -59DeltapH (transmembrane pH gradient). 4. Preincubation of rat liver mitochondria with p-chloromercuribenzenesulphonate, which by contrast with N-ethylmaleimide is unable to penetrate the inner mitochondrial membrane, also prevents Ca(2+) retention. The DeltaE and -59DeltapH respond to Ca(2+) addition in a manner similar to that which occurs when N-ethylmaleimide is present. Subsequent addition of mercaptoethanol produces an immediate increase in both DeltaE and -59DeltapH. At the same time Ca(2+) is rapidly accumulated by the organelles. 5. The above data are interpreted as indicating that under the conditions of Ca(2+) efflux seen here, the mitochondria retain their functional integrity. This contrasts with the uncoupling effect of Ca(2+) seen in the presence of P(i), which generally leads to a loss of mitochondrial integrity. We suggest that a unique mechanism of Ca(2+) cycling is able to take place when mitochondria have been treated with N-ethylmaleimide.     

Effect of inhibitors on the sigmoidicity of the calcium ion transport kinetics in rat liver mitochondria

The kinetic plot (initial rate of Ca²⁺ transport versus concentration) of mitochondrial Ca²⁺ transport is hyperbolic in a sucrose medium. The plot becomes sigmoidal in the presence of competitive inhibitors of Ca²⁺ binding to low affinity sites of the membrane surface such as Mg²⁺ and K⁺. The plot also becomes sigmoidal in the presence of Ba²⁺. Ba²⁺ is a competitive inhibitor of both Ca²⁺ transport and Ca²⁺ binding to the low affinity sites. The K_i for the inhibition of Ca²⁺ transport by Ba²⁺ increases in the presence of K⁺ and Mg²⁺, which suggests a competition for the low affinity sites between the cations. The plot is still hyperbolic in the presence of La³⁺, which inhibits Ca²⁺ transport competitively. Ruthenium red which is a pure non-competitive inhibitor of mitochondrial Ca²⁺ transport, does not affect the shape of the kinetic plot. These results indicate that the surface potential, which depends on the ions bound to the low affinity sites, determines whether the kinetics of Ca²⁺ uptake in mitochondria is sigmoidal or hyperbolic.     

Effect of micromolar concentrations of manganese ions on calcium-ion cycling in rat liver mitochondria.

The effects of micromolar concentrations of Mn2+ on the rat liver mitochondrial Ca2+ cycle were investigated. It was found that the addition of Mn2+ to mitochondria which were cycling 45Ca2+ led to a rapid dose dependent decrease in the concentration of extramitochondrial 45Ca2+ of about 1 nmol/mg of protein. The effect was complete within 30 s, was half maximal with 10 microM Mn2+ and was observed in the presence of 3 mM Mg2+ and 1 mM ATP. It occurred over a broad range of incubation temperatures, pH and mitochondrial Ca2+ loads. It was not observed when either Mg2+ or phosphate was absent from the incubation medium, or in the presence of Ruthenium Red. These findings indicate that micromolar concentrations of Mn2+ stimulate the uptake of Ca2+ by rat liver mitochondria, and provide evidence for an interaction between Mg2+ and Mn2+ in the control of mitochondrial Ca2+ cycling.     

Effect of chelators of bivalent metal ions on the reaction of myometrium actomoysin superprecipitation]

Kinetic regularities of the reaction of superprecipitation of myometrium actomyosin, as well as the effect of different concentrations of EGTA, EDTA and diphosphonic acids on this process have been studied. Results obtained are of interest from the viewpoint of possible practical use of diphosphonates as factors modifying interaction of the contractile proteins of the uterus smooth muscles under the pathology of contractile response.     

Effect of bedaquiline on the functions of rat liver mitochondria

The paper considers the effects of bedaquiline (BDQ), an antituberculous preparation of the new generation, on rat liver mitochondria. It was shown that 50?μM BDQ inhibited mitochondrial respiration measured with substrates of complexes I and II (glutamate/malate and succinate/rotenone systems respectively) in the states V₃ and V_DNP. At the same time, at concentrations below 50?μM, BDQ slightly stimulated respiration with substrates of complex I in the state V₂. BDQ was also found to suppress, in a dose-dependent manner, the activity of complex II and the total activity of complexes II?+?III of the mitochondrial transport chain. It was discovered that at concentrations up to 10?μM, BDQ inhibited H₂O₂ production in mitochondria. BDQ (10–50?μM) suppressed the opening of Ca²⁺-dependent CsA-sensitive mitochondrial permeability transition pore. The latter was revealed experimentally as the inhibition of Ca²⁺/P_i-dependent swelling of mitochondria, suppression of cytochrome c release, and an increase in the Ca²⁺ capacity of the organelles. BDQ also decreased the rate of mitochondrial energy-dependent K⁺ transport, which was evaluated by the energy-dependent swelling of mitochondria in a K⁺ buffer and DNP-induced K⁺ efflux from the organelles. The possible mechanisms of BDQ effect of rat liver mitochondria are discussed.     

Effect of inflammatory mediators on respiration in rat liver mitochondria

Some inflammatory mediators have been studied for their influence on the energy reactions of the liver mitochondria. Mediators were injected intraperitoneally to rats 15 min before decapitation in the following doses (per 100 g of the body) weight: histamine--0.5 mg, serotonin--0.5 mg, bradykinin--0.2 mg, andekalin--0.5 units. Histamine action in the body is connected with modification of the respiratory mitochondria chain and, like the oligomycin action, is directed to attended oxidation and phosphorylation points. Serotonin increases the mitochondria sensitivity to separating agents in succinate oxidation. It is supposed that serotonin-induced inhibition of oxidation of NAD-dependent substances is connected with NADH2 dehydrogenase inhibition or transhydrogenase reaction activation. Bradykinin has activated NAD-dependent substance oxidation and increased respiratory chain sensitivity on the SoQ link to 2,4-dinitrophenol action. Andekalin exerts an analogous effect intensifying ADP-, DNP- and Ca-stimulated respiration of mitochondria during succinate oxidation. Mechanism of the inflammatory mediators influence on the energy metabolism is discussed.     

Modification by calcium ions of adenine nucleotide translocation in rat liver mitochondria

1. Added Ca(2+) stimulates the translocation of ATP by isolated rat liver mitochondria. 2. The apparent K(m) for added Ca(2+) in stimulating the translocation of 200mum-ATP is approx. 160mum (75mum ;free' Ca(2+)). 3. The greatest stimulation of ATP translocation by Ca(2+) occurs at the lower concentrations of ATP. 4. Sr(2+) (and to a lesser extent Ba(2+)) can replace Ca(2+) whereas Mg(2+) and Mn(2+) have only little ability to stimulate ATP translocation. 5. Translocation of dATP is also stimulated by Ca(2+) whereas that of ADP is stimulated to only a relatively small degree. 6. Studies with metabolic inhibitors and uncouplers provide evidence that stimulation by Ca(2+) and by uncouplers is additive and that the mechanism of Ca(2+) stimulation does not seem to involve the high-energy intermediate of oxidative phosphorylation. 7. In the presence of Ca(2+), ATP is able to effectively compete with ADP for translocation. 8. Added K(+) further enhances the ability of Ca(2+) to stimulate ATP translocation. 9. These findings are discussed in relation to the potential involvement of Ca(2+) in modifying enzymic reactions involved in the regulation of cell metabolism.     

Ba2+ ions inhibit the release of Ca2+ ions from rat liver mitochondria

The release of Ca2+ from respiring rat liver mitochondria following the addition of either ruthenium red or an uncoupler was measured by a Ca2+-selective electrode or by 45Ca2+ technique. Ba2+ ions are asymmetric inhibitors of both Ca2+ release processes. Ba2+ ions in a concentration of 75 microM inhibited the ruthenium red and the uncoupler induced Ca2+ release by 80% and 50%, respectively. For the inhibition, it was necessary that Ba2+ ions entered the matrix space: Ba2+ ions did not cause any inhibition of Ca2+ release if addition of either ruthenium red or the uncoupler preceded that of Ba2+. The time required for the development of the inhibition of the Ca2+ release and the time course of 140Ba2+ uptake ran in parallel. Ba2+ accumulation is mediated through the Ca2+ uniporter as 140Ba2+ uptake was competitively inhibited by extramitochondrial Ca2+ and prevented by ruthenium red. Due to the inhibition of the ruthenium red insensitive Ca2+ release, Ba2+ shifted the steady-state extramitochondrial Ca2+ concentration to a lower value. Ba2+ is potentially a useful tool to study mitochondrial Ca2+ transport.     

Effect of metal ions and calcium on purified PON1 and PON3 from rat liver

The effect of several metal ions and calcium on purified paraoxonases (PON1 and PON3) from rat liver was studied. PON1 and PON3 were also inhibited by EDTA and both enzyme activities were restored by the addition of free calcium. The reactivation by calcium was a time-dependent effect for PON1; however, this was not the case for PON3. We also studied the response of PON1 and PON3 to several inhibitors: Co, Cu, Mn, Hg and p-hydroxymercurybenzoate (pOHMB), and determined the type of inhibition and the inhibition constants. Among all the compounds tested, mercurials (Hg and pOHMB) were the most potent inhibitors of PON1. For PON3 mercurials and copper showed the highest inhibitory potency. Purified PON3 also showed different inhibition patterns as compared to PON1. A comparison of PON1 and PON3 shows qualitative and quantitative differences in the sensitivity against the inhibitors tested, showing major differences in the case of cobalt, copper and pOHMB, which may be related to structural differences of both PONs. These results increase our knowledge of the biochemical properties of PON1 and PON3 and may help in the understanding of their physiological role as a potential detoxification mechanism against environmental metal ions.     

Effect of cadmium ions on the respiration, cation transport and morphology of the liver mitochondria in the rat and the lamprey

The influence of Cd2+ on the function and structure of liver mitochondria of rats and lamprey (Lampetra fluviatilis L.) has been studied in vitro. It is shown that Cd2+ can penetrate into the mitochondrial matrix due to Ca2+-transport mechanism. Being stored in the mitochondria, Cd2+ inhibits respiration and an energy dependent transport of penetrating cations (Cs+-valinomycin), and disturbs passive permeability of the inner mitochondrial membrane for monovalent cations and H+. The effect of Cd2+ on the lamprey liver mitochondria is more pronounced than in the case of rats.     

The titration curve of insulin in the presence of various bivalent metal ions

1. Titration curves of insulin in the presence and absence of various metal ions are reported. 2. The difference in base consumption with and without the metal ions is compared with calculated curves. 3. These experiments suggest that in dilute solutions Zn(2+) and Cu(2+) ions are bound to alpha-amino groups.