scholarly journals Deterioration of rat liver mitochondria under conditions of metabolite deprivation

1980 ◽  
Vol 188 (3) ◽  
pp. 817-822 ◽  
Author(s):  
J W Parce ◽  
P I Spach ◽  
C C Cunningham
Keyword(s):  
Atpase Activity ◽  
Sucrose Solution ◽  
Adenine Nucleotides ◽  
Respiratory Control ◽  
Liver Mitochondria ◽  
Phospholipid Metabolism ◽  
Rat Liver Mitochondria ◽  
Second Phase ◽  
Mitochondrial Atpase ◽  
Sequence Of Events

In a previous study [Parce, Cunningham & Waite (1978) Biochemistry 17, 1634-1639] changes in mitochondrial phospholipid metabolism and energy-linked functions were monitored as coupled mitochondria were aged in iso-osmotic sucrose solution at 18 degrees C. The sequence of events that occur in mitochondrial deterioration under the above conditions have been established more completely. Total adenine nucleotides are depleted early in the aging process, and their loss parallels the decline in respiratory control. Related to the loss of total adenine nucleotides is a dramatic decrease in ADP and ATP translocation (uptake). The decline of respiratory control is due primarily to a decrease in State-3 respiration; loss of this respiratory activity can be related to the decline in ADP translocation. Mitochondrial ATPase activity does not increase significantly until State-4 respiration has increased appreciably. At the time of loss of respiratory control the ATPase activity increases to equal the uncoupler-stimulated activity. The H+/O ratio and P/O ratios do not decrease appreciably until respiratory control is lost. Similarly, permeability of the membrane to the passive diffusion of protons increases only after respiratory control is lost. There observations reinforce our earlier conclusion that there are two main phases in mitochondrial aging. The first phase is characterized by loss of the ability to translocate adenine nucleotides. The second phase is characterized by a decline in the ability of the mitochondrion to conserve energy (i.e. maintain a respiration-driven proton gradient) and to synthesize ATP.

scholarly journals The pathway of inorganic-phosphate efflux from isolated liver mitochondria during adenosine triphosphate hydrolysis

1980 ◽  
Vol 192 (3) ◽  
pp. 821-828 ◽  
Author(s):  
David D. Tyler
Keyword(s):  
Atpase Activity ◽  
Large Amplitude ◽  
Atp Hydrolysis ◽  
Adenine Nucleotide ◽  
Sucrose Solution ◽  
Adenine Nucleotides ◽  
Liver Mitochondria ◽  
Rat Liver Mitochondria ◽  
Nucleotide Exchange ◽  
Major Activity

1. The distribution of Pi between mitochondria and suspending medium during uncoupler-stimulated hydrolysis of ATP by rat liver mitochondria [Tyler (1969) Biochem. J.111, 665–678] has been reinvestigated, by using either mersalyl or N-ethylmaleimide as inhibitors of Pi transport and either buffered sucrose/EDTA or LiCl/EGTA solutions as suspending medium. More than 75% of the total Pi liberated was retained in mitochondria treated with either inhibitor at all ATP concentrations tested (0.2–2.5mm). With low ATP concentrations and mersalyl-treated mitochondria incubated in sucrose/EDTA, virtually all the Pi liberated was retained in the mitochondria. 2. Larger amounts of Pi appeared in the suspending medium during ATPase activity, despite the presence of N-ethylmaleimide, when LiCl/EGTA was used as suspending medium compared with sucrose/EDTA. Two sources of this Pi were identified: (a) a slow efflux of Pi from mitochondria to suspending medium despite the presence of N-ethylmaleimide; (b) a slow ATPase activity insensitive to carboxyatractyloside, which was stimulated by added Mg2+, partially inhibited by oligomycin or efrapeptin and strongly inhibited by EDTA. 3. It is concluded that liver mitochondria preparations contain two distinct forms of ATPase activity. The major activity is associated with coupled mitochondria of controlled permeability to adenine nucleotides and Pi and is stimulated strongly by uncoupling agents. The minor activity is associated with mitochondria freely permeable to adenine nucleotides and Pi, is unaffected by uncoupling agents and is activated by endogenous or added Mg2+. 4. When mitochondria treated with mersalyl were incubated in buffered sucrose solution, almost all the Pi liberated was recovered in the suspending medium, unless inhibitors of Pi-induced large-amplitude swelling such as EDTA, EGTA, antimycin, rotenone, nupercaine or Mg2+ were added. Thus the loss of the specific permeability properties of the mitochondrial inner membrane associated with large-amplitude swelling also influences the extent of Pi retention during ATPase activity. 5. The results confirm the previous conclusion (Tyler, 1969) that the Pi transporter provides the sole pathway for Pi efflux during uncoupler-stimulated ATP hydrolysis by mitochondria. It is concluded that more recent hypotheses concerning the influence of Mg2+ on mersalyl inhibition of the Pi transporter [Siliprandi, Toninello, Zoccaroto & Bindoli (1975) FEBS Lett. 51, 15–17] and a postulated role of the adenine nucleotide exchange carrier in Pi efflux [Reynafarje & Lehninger (1978) Proc. Natl. Acad. Sci. U.S.A.75, 4788–4792] are erroneous and should be discarded.

scholarly journals Nutritional effects on mitochondrial bioenergetics. Alterations in oxidative phosphorylation by rat liver mitochondria

1984 ◽  
Vol 218 (1) ◽  
pp. 61-67 ◽  
Author(s):  
J Ferreira ◽  
L Gil
Keyword(s):  
Oxidative Phosphorylation ◽  
Respiratory Control ◽  
Atp Synthesis ◽  
Liver Mitochondria ◽  
Rat Liver Mitochondria ◽  
Mitochondrial Atpase ◽  
Protein Free Diet ◽  
Energy Dependent ◽  
Hydrolysis Of ◽  
Rate Controlling Step

Rats malnourished since birth and fed on a protein-free diet for 2 weeks showed a 23-27% decrease in the State-3 oxidation of glutamate, succinate and ascorbate + NNN′ N′-tetramethyl-p-phenylenediamine by liver mitochondria compared with control fed animals. ATP synthesis and the respiratory control index were diminished at the three coupling sites, but significant alterations were not observed in ADP/O ratios. Vmax. for NADH oxidation in electron-transport particles was 40% lower. Mitochondrial cytochromes b and c1 remained unchanged, but cytochrome c was increased by 26%. Cytochromes a + a3 were diminished by 22%. Vmax. for mitochondrial ATPase was 23% lower. These results suggest that the lower content of cytochrome a + a3 at the rate-controlling step of oxidative phosphorylation in malnourished rats might be mainly responsible for the decrease in substrate oxidations as well as ATP synthesis at the three coupling sites. The decreased synthesis and hydrolysis of ATP suggests that other energy-dependent mitochondrial processes could be decreased during malnutrition.

scholarly journals The effect of butacaine on adenine nucleotide binding and translocation in rat liver mitochondria

1975 ◽  
Vol 148 (3) ◽  
pp. 527-531 ◽  
Author(s):  
D R Fayle ◽  
G J Barritt ◽  
F L Bygrave
Keyword(s):  
Rat Liver ◽  
Local Anaesthetic ◽  
Binding Sites ◽  
Adenine Nucleotide ◽  
Adenine Nucleotides ◽  
Nucleotide Binding ◽  
Local Anaesthetics ◽  
Liver Mitochondria ◽  
Rat Liver Mitochondria ◽  
Mitochondrial Inner Membrane

The effect of the local anaesthetic, butacaine, on adenine nucleotide binding and translocation in rat liver mitochondria partially depleted of their adenine nucleotide content was investigated. The range of butacaine concentrations that inhibit adenine nucleotide translocation and the extent of the inhibition are similar to the values obtained for native mitochondria. Butacaine does not alter either the total number of atractyloside-sensitive binding sites of depleted mitochondria, or the affinity of these sites for ADP or ATP under conditions where a partial inhibition of the rate of adenine nucleotide translocation is observed. The data are consistent with an effect of butacaine on the process by which adenine nucleotides are transported across the mitochondrial inner membrane rather than on the binding of adenine nucleotides to sites on the adenine nucleotide carrier. The results are briefly discussed in relation to the use of local anaesthetics in investigations of the mechanism of adenine nucleotide translocation.

The polyphasic nature of the respiratory process at the mitochondrial level

1990 ◽  
Vol 258 (3) ◽  
pp. C504-C511 ◽  
Author(s):  
B. D. Reynafarje ◽  
P. W. Davies
Keyword(s):  
Oxygen Consumption ◽  
Energy Demand ◽  
Initial Conditions ◽  
Electron Flow ◽  
Liver Mitochondria ◽  
Rat Liver Mitochondria ◽  
Second Phase ◽  
Third Phase ◽  
Metabolic Conditions ◽  
Kinetics Of

The kinetics of oxygen consumption by rat liver mitochondria, respiring under a variety of metabolic conditions, have been studied. Respiration was initiated by injecting oxygen into anaerobic suspensions of mitochondria. It was found that, irrespective of the metabolic state of the mitochondria and the nature of the respiratory substrate, the rates of electron flow and oxygen consumption follow the pattern of a polyphasic reaction. The rates of oxygen uptake during the first phase are extremely fast and depend on oxygen concentration. The second phase represents a transition in which net oxidation of cytochrome-c oxidase stops and the rates of oxygen consumption suddenly decrease. The third phase is characterized by its changeability. Depending on initial conditions the rates may increase, decrease, or remain constant, although the reaction is not one of zero order. During the last phase, the rates decrease and the oxidase becomes increasingly reduced. It is postulated that the mitochondrial respiratory process is basically a cyclic event in which the redox state of the membrane and the rates of oxygen consumption oscillate with amplitudes and frequencies conditioned by the energy demand and energy-yielding capacity of the cell.

scholarly journals STRUCTURAL CHANGES IN ISOLATED LIVER MITOCHONDRIA OF RATS DURING ESSENTIAL FATTY ACID DEFICIENCY

1964 ◽  
Vol 21 (1) ◽  
pp. 15-26 ◽  
Author(s):  
Janet A. Smith ◽  
Hector F. DeLuca
Keyword(s):  
Atpase Activity ◽  
Structural Damage ◽  
Structural Changes ◽  
Essential Fatty Acids ◽  
Saturated Fat ◽  
Respiratory Control ◽  
Liver Mitochondria ◽  
Structural Defects ◽  
Thin Sections ◽  
Fatty Acid Deficiency

Liver mitochondria isolated in 0.44 M sucrose from rats deficient in essential fatty acids (EFA) oxidized citrate, succinate, α-ketoglutarate, glutamate, and pyruvate at a faster rate than did mitochondria isolated from normal rats; however, the oxidation of malate, caprylate, and ß-hydroxybutyrate was not significantly increased. The mitochondria from deficient rats exhibited an increased ATPase activity and extensive structural damage as revealed by electron microscope examination of thin sections. An increase in citrate oxidation and ATPase activity, together with some structural damage, could be demonstrated as early as the 4th week in rats on a fat-free diet. Saturated fat in the diet did not prevent the change in mitochondrial structure but accelerated its appearance. Both the biochemical and structural defects could be reversed within three weeks after feeding deficient rats a source of EFA. In the presence of a phosphate acceptor the effect of EFA deficiency on substrate oxidation was largely eliminated. A trend toward a reduced efficiency of oxidative phosphorylation was noted in mitochondria from EFA-deficient rats, but significant uncoupling was found only in the case of citrate, ß-hydroxybutyrate, and glutamate in the presence of malonate. Together with the increased ATPase activity, the uncoupling of phosphorylation could account for the poor respiratory control found with the deficient preparation. However, EFA deficiency was without effect on the respiration of liver slices, which supports the belief that the observed changes in oxidation and phosphorylation are an artifact resulting from damage sustained by the deficient mitochondria during their isolation.

scholarly journals The control of isocitrate oxidation by rat liver mitochondria

1969 ◽  
Vol 114 (2) ◽  
pp. 215-225 ◽  
Author(s):  
D. G. Nicholls ◽  
P. B. Garland
Keyword(s):  
Respiratory Chain ◽  
Rat Liver ◽  
Isocitrate Dehydrogenase ◽  
Adenine Nucleotides ◽  
Kinetic Studies ◽  
Liver Mitochondria ◽  
Rat Liver Mitochondria ◽  
Kinetic Properties ◽  
Dependent Inhibition ◽  
Nad 4

1. The factors capable of affecting the rate of isocitrate oxidation in intact mitochondria include the rate of isocitrate penetration, the activity of the NAD-specific and NADP-specific isocitrate dehydrogenases, the activity of the transhydrogenase acting from NADPH to NAD+, the rate of NADPH oxidation by the reductive synthesis of glutamate and the activity of the respiratory chain. A quantitative assessment of these factors was made in intact mitochondria. 2. The kinetic properties of the NAD-specific and NADP-specific isocitrate dehydrogenases extracted from rat liver mitochondria were examined. 3. The rate of isocitrate oxidation through the respiratory chain in mitochondria with coupled phosphorylation is approximately equal to the maximal of the NAD-specific isocitrate dehydrogenase but at least ten times as great as the transhydrogenase activity from NADPH to NAD+. 4. It is concluded that the energy-dependent inhibition of isocitrate oxidation by palmitoylcarnitine oxidation is due to an inhibition of the NAD-specific isocitrate dehydrogenase. 5. Kinetic studies of NAD-specific isocitrate dehydrogenase demonstrated that its activity could be inhibited by one or more of the following: an increased reduction of mitochondrial NAD, an increased phosphorylation of mitochondrial adenine nucleotides or a fall in the mitochondrial isocitrate concentration. 6. Uncoupling agents stimulate isocitrate oxidation by an extent equal to the associated stimulation of transhydrogenation from NADPH to NAD+. 7. A technique is described for continuously measuring with a carbon dioxide electrode the synthesis of glutamate from isocitrate and ammonia.

scholarly journals Studies on the nature of the high-affinity trialkyltin binding site of rat liver mitochondria

1982 ◽  
Vol 202 (1) ◽  
pp. 163-169 ◽  
Author(s):  
A P Dawson ◽  
B G Farrow ◽  
M J Selwyn
Keyword(s):  
Binding Site ◽  
Rat Liver ◽  
Liver Mitochondria ◽  
Rat Liver Mitochondria ◽  
Affinity Binding ◽  
Mitochondrial Atpase ◽  
High Affinity Binding ◽  
High Affinity ◽  
High Affinity Binding Site ◽  
Triphenyltin Chloride

1. The proteolipid fraction isolated from rat liver mitochondria pretreated with [3H]triphenyltin chloride is enriched in triphenyltin compared with the original mitochondria. 2. Part of this [3H]triphenyltin is eluted with a protein of Mr 5000-6000 on Sephadex LH20 chromatography. 2. Mössbauer spectra of the proteolipid fraction treated with 119Sn-enriched triethyltin chloride show a doublet which corresponds closely with that assigned previously [Farrow & Dawson (1978) Eur. J. Biochem. 86. 85-95] to the absorption of triethyltin bound to the high-affinity binding site of the mitochondrial ATPase.

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