Leena Mela

Interactions of La3+ and local anesthetic drugs with mitochondrial Ca++ and Mn++ uptake☆

Abstract Interactions of lanthanum and some local anesthetic drugs with divalent cation uptake have been studied in rat liver mitochondria. The oxidation-reduction changes of the respiratory chain components and the development of the membrane pH gradient were measured during ion uptake. The trivalent ion lanthanum was found to be a powerful inhibitor of divalent cation uptake with half-maximal effect observed at concentrations of approximately 0.1 mμmole La 3+ /mg protein. The neurotropic drugs butacaine, dibucaine, chlorpromazine, and phenergan enhanced remarkably the divalent cation-induced membrane pH gradient. The possible mechanism of the action of lanthanum and local anesthetics ou mitochondrial membranes is discussed.

Factors in oxygen delivery to tissue

An animal model for evaluating oxygen delivery to tissue is based upon the determination of the intracellular oxygen tension by surface fluorometry of NADH under conditions where the inspired oxygen is reduced through the critical value for the particular organ. This procedure allows the evaluation of the overall efficiency of the pulmonocardiac tissue in delivering oxygen to tissue, and can be useful in new, quantitative measurements of the effect of various factors occurring between the inspired air and the final tissue oxygen tension.

Control of mitochondrial respiration by the phosphate potential

Abstract The rate of respiration of suspensions of mitochondria in the presence of excess oxygen and substrate is shown to be dependent on the ratio of the concentration of adenosine triphosphate (ATP) to the product of the concentrations of adenosine diphosphate and orthophosphate. The mitochondrial respiratory chain is essentially in equilibrium with the reactions for ATP synthesis. The rate of mitochondrial respiration is controlled by the free energy requirement for ATP synthesis and this control is expressed on the rates of the reactions for reduction of the dehydrogenases by substrate and the oxidation of cytochrome a 3 by molecular oxygen.

Local anesthetic induced changes of a membrane-bound fluorochrome A link between ion uptake and membrane structure

Calcium uptake by mitochondria represents their most rapid and effective energy-linked function. Cytochrome b responds to calcium addition in 20 msec [ 11; calcium-stimulated respiration exceeds the rate of ADP-stimulated respiration and causes a maximum rate of external hydrogen ion ejection [2,3] and in the absence of a permeant anion, a maximal membrane alkalinization [4]. The affinity of mitochondria for calcium is remarkably high; external calcium concentrations in equilibrium with the endogenous calcium of rat liver mitochondria are of the order of lo-* M, as detected with the calcium indicator, aequorin [ 11. Thus, extremely high inside/outside calcium concentration ratios are possible. The response of the membrane bound pH indicator, bromothymol blue [4], to calcium accumulation in the absence of a permeant anion is greatly stimulated by relatively low concentrations of a local anesthetic, butacaine. The rate of calcium uptake, as reported by the indicator, murexide, has been shown to be enhanced several-fold by butacaine [5,1]. A striking effect of butacaine on the mitochondrial membrane is revealed by a membrane-bound fluorochrome, 8-aniline-1-naphthalene sulfonic acid, ANS, which is insensitive to pH and to oxidation-reduction changes, but shows a sensitive response to changes in the state of the membrane [6,7]. The properties of the membrane-bound fluorochrome and the effects of butaCaine and calcium upon the kinetics and stoichiometry of the ANS reaction are described. 2. Materials and methods

Mechanism and Physiological Significance of Calcium Transport Across Mammalian Mitochondrial Membranes

Publisher Summary This chapter describes the early findings of investigators, which led to the notion that intact isolated mitochondria can accumulate calcium (Ca 2+ ), Sr 2+ , and Mn 2+ in an energy-dependent process, whether permeant anions are present. Evidence is presented in support of a three-step mechanism of mitochondrial Ca 2+ accumulation, which is energy-independent binding, carrier-mediated accumulation into the membrane, and anion dependent release from the carrier into the mitochondrial matrix. The physiological significance of mitochondrial Ca 2+ accumulation system in terms of the regulation of cellular Ca 2+ free is also discussed in the chapter. Experimental evidence is presented, indicating physiological control of cellular free Ca 2+ by mitochondria in a variety of mammalian tissues. Examples of pathological alterations of the mitochondrial Ca 2+ transport system are presented for two disease states: tumor cells and ischemically injured cells.

Mitochondrial function in cerebral ischemia and hypoxia: comparison of inhibitory and adaptive responses.

In cerebral ischemia, brain oxygen supply is totally exhausted within seconds. This necessitates cessation of mitochondrial electron transfer and energy (ATP) production. After certain periods of ATP deficiency of from 5 to 90 min, irreversible damage of mitochondrial membranes occurs. This results in decreased mitochondrial function, characterized by inhibited State 3 respiratory rates, low respiratory control ratios, and inhibited Ca2+ transport activities. A 30-min recirculation period of the ischemic brain tissue induces total restitution of mitochondrial respiratory capacity after complete ischemia, but not after incomplete ischemia. Regional in situ measurements of brain pyridine nucleotide redox levels, tissue ATP, and lactate concentrations indicate variable metabolic responses of different brain regions to oligemia. Macroheterogeneity from region to region, as well as microheterogeneity within a region are demonstrated. Contrary to the effect of tissue ischemia involving reduced or zero cerebral blood flow and tissue oxygenation, sublethal hypoxia alone at normal or increased levels of blood flow induces adaptation of the mitochondrial enzyme system to a new level of respiratory capacity, without any indications of inhibited mitochondrial energy production. Acute hypoxia induces increased respiratory capacities within 30-60 min. Under chronic conditions, alterations of mitochondrial cytochrome concentrations accompany the increased respiratory capacities. Instead of the decreased efficiency of mitochondrial energy-producing mechanisms induced by ischemia, hypoxia induces increased efficiency of energy production.

Inactivation of alcohol dehydrogenase by 3-butyn-1-ol☆

Abstract Horse liver and yeast alcohol dehydrogenases are rapidly inactivated during their catalysis of the oxidation of 3-butyn-1-ol. In the case of the horse liver enzyme, the inactivation is secondary to covalent modification of the apoenzyme by an electrophilic product that accumulates in the reaction solution and that can also react with water, glutathione, and other enzymes. The modified protein exhibits enhanced ultraviolet absorbance, which is not bleached upon dialysis of the denatured enzyme at pH 7.4 for 24 h. The inactivation by 3-butyn-1-ol is more rapid than that which is afforded by the related alcohols 2-propyn-1-ol and 2-propen-1-ol under identical conditions and no inactivation is seen upon incubation with 3-hydroxypropanoic nitrile plus nicotinamide-adenine dinucleotide.

Inactivation of alanine aminotransferase by the neurotoxin β-cyano-L-alanine

Abstract β-Cyano-L-alanine inactivates pig heart alanine aminotransferase. The nitrile and enzyme form a freely dissociable Michaelis complex which rearranges to a form of inactive enzyme. The inactivated enzyme slowly recovers activity at 25° in 100 mM phosphate buffer, pH 7.4. The observations are consistent with a mechanism of inactivation similar to that thought to apply to the suicide inactivator propargylglycine except that the putative covalent modification of the apoenzyme is relatively labile in the case of the nitrile.

Correlation of mitochondrial cytochrome concentration and activity to oxygen availability in the newborn

Effect of in vivo oxygenation on mitochondrial respiratory chain capacity was studied in newborn puppies. Heart mitochondria were isolated from 0 to 7 days. As the arterial PO2 rises sharply during the first 3–4 days, State 3 respiratory capacity and Ca++ transport activity decrease linearly. Cytochrome c and a + a3 concentrations increase rapidly. Thus the enzymatic turnover of the respiratory chain decreases to one-fifth of the fetal term value in 4 days. These data are in agreement with earlier results indicating a strong controlling effect of in vivo oxygenation on mitochondrial respiratory capacity.

Early alterations in mitochondrial membrane transport during endotoxemia

Abstract Endotoxemia and changes in reaction medium pH and ionic composition were studied to determine their effect on mitochondrial membrane transport of calcium, potassium, and adenine nucleotides. Endotoxemia decreases calcium translocation from 373 ± 13 to 177 ± 19 nmoles calcium per minute per mg mitochondrial protein in normal and endotoxemic mitochondria respectively. Lowering the pH of the reaction medium to 6.5 decreases the calcium uptake in normal mitochondria to 101 ± 20 nmoles per minute per mg mitochondrial protein. Similarly, the calcium uptake is reduced in a medium designed to simulate the intracellular shock state. Endotoxemia also increases mitochondrial membrane permeability for potassium to unphysiologic levels, and this may be responsible for mitochondrial swelling noted in vivo . The translocation and phosphorylation of ADP is not significantly inhibited in early endotoxemia. Thus, some of the early endotoxemic changes are seen in the ion transport reactions of the mitochondrial membrane. The inhibition of ATP production occurs later when the mitochondrial membrane has become more extensively damaged.