Biochemical Basis of Heart Function. X. Reduction in the Na+–K+-Stimulated ATPase Activity in Failing Rat Heart Due to Hypoxia

1973 ◽  
Vol 51 (7) ◽  
pp. 504-510 ◽  
Author(s):  
V. Balasubramanian ◽  
D. B. McNamara ◽  
J. N. Singh ◽  
N. S. Dhalla
Keyword(s):  
Atpase Activity ◽  
Specific Activity ◽  
Heart Function ◽  
Perfusion Medium ◽  
Biochemical Basis ◽  
Exogenous Glucose ◽  
Hypoxic Heart ◽  
Rat Hearts ◽  
Membrane Bound ◽  
Assay Conditions

The activities of the membrane bound enzyme Na+–K+-stimulated ATPase were studied in control and hypoxic isolated rat hearts. The control hearts were perfused with medium gassed with 95% O2 and 5% CO2, whereas hypoxia was induced by perfusing the hearts with medium gassed with 95% N2 and 5% CO2. The specific activity of the Na+–K+ ATPase was lower in the hypoxic hearts than in the control preparations. The enzyme activity of the hypoxic hearts was also lower when determined under different assay conditions obtained by changing the concentration of Na+, K+, H+, and MgATP in the incubation medium. The ouabain-induced inhibition of Na+–K+ ATPase activity in the hypoxic hearts was similar to that in the control. The depression in Na+–K+ ATPase activity due to hypoxia was reversible on perfusing the hypoxic heart with aerobic medium. The decreased Na+–K+ ATPase activity in the hypoxic heart preparations supports the concept of membrane abnormality associated with heart failure due to lack of oxygen. Since the depression in the activity of Na+–K+ ATPase in the hypoxic heart was markedly pronounced on omitting the substrate from the perfusion medium, it is likely that exogenous glucose plays an important role in maintaining the integrity of hypoxic heart membranes.

Local ATP regeneration is important for sarcoplasmic reticulum Ca2+ pump function

1994 ◽  
Vol 267 (2) ◽  
pp. C357-C366 ◽  
Author(s):  
P. Korge ◽  
K. B. Campbell
Keyword(s):  
Sarcoplasmic Reticulum ◽  
Atpase Activity ◽  
Muscle Relaxation ◽  
Creatine Phosphate ◽  
Coupling Ratio ◽  
Pump Function ◽  
Membrane Bound ◽  
Low Efficiency ◽  
Atp Regeneration ◽  
Assay Conditions

Ca2+ pump function of skeletal muscle sarcoplasmic reticulum (SR) vesicles was measured by monitoring Ca2+ uptake and efflux with a Ca(2+)-sensitive minielectrode and adenosinetriphosphatase (ATPase) activity of the same preparation under the same conditions. The efficiency of Ca2+ transport into SR vesicles, defined by the amount of Ca2+ transported per ATP hydrolyzed (coupling ratio), varied significantly depending on assay conditions. Coupling ratio increased in parallel with increase in precipitating anion concentration, which is supposed to decrease accumulation of free Ca2+ inside vesicles and its subsequent efflux. Membrane-bound creatine kinase-creatine phosphate (CK-CP) system, acting as a ADP sensor and local ATP regenerator, significantly improved Ca2+ pump function when the pump worked with low efficiency (coupling ratio < 1). The effect of CK-CP system on Ca2+ pump function was also dependent on extravesicular Ca2+ concentration ([Ca2+]o), the effect being most significant at high initial [Ca2+]o. Under conditions in which SR vesicles were allowed to decrease [Ca2+]o, as occurs also during muscle relaxation, plateau values of Ca(2+)-ATPase activity were reached at significantly higher [Ca2+]o (54 +/- 5.7, n = 6), compared with leaky vesicles or the condition in which [Ca2+]o was maintained. By preventing local accumulation of ADP, generated in ATPase reactions, CK-CP system also inhibited Ca2+ efflux under conditions in which this efflux was stimulated by the increase of free Ca2+ inside vesicles. This effect was at least partially responsible for the CK-CP-supported increase in Ca2+ uptake and coupling ratios that were more expressed at low precipitating anion concentration. We hypothesize that local ATP regeneration by CK-CP system is one mechanism the cell can use to improve Ca2+ uptake by SR in emergency conditions, where excessive increase in cytoplasmic [Ca2+] may have deleterious effects.

Characterization studies on the membrane-bound adenosine triphosphatase (ATPase) of Azotobacter vinelandii

1975 ◽  
Vol 21 (11) ◽  
pp. 1807-1814 ◽  
Author(s):  
Peter Jurtshuk ◽  
John E. McEntire
Keyword(s):  
Electron Transport ◽  
Atpase Activity ◽  
Azotobacter Vinelandii ◽  
Specific Activity ◽  
Maximal Activity ◽  
Molar Ratio ◽  
Supernatant Fraction ◽  
Time Interval ◽  
Time Intervals ◽  
Membrane Bound

The adenosinetriphosphatase (ATPase) (EC 3.6.1.3) activity in Azotobacter vinelandii concentrates in the membranous R3 fraction that is directly associated with Azotobacter electron transport function. Sonically disrupted Azotobacter cells were examined for distribution of ATPase activity and the highest specific activity (and activity units) was consistently found in the particulate R3 membranous fraction which sediments on ultracentrifugation at 144 000 × g for 2 h. When the sonication time interval was increased, the membrane-bound ATPase activity could neither be solubilized nor released into the supernatant fraction. Optimal ATPase activity occurred at pH 8.0; Mg2+ ion when added to the assay was stimulatory. Maximal activity always occurred when the Mg2+:ATP stoichiometry was 1:1 on a molar ratio at the 5 mM concentration level. Sodium and potassium ions had no stimulatory effect. The reaction kinetics were linear for the time intervals studied (0–60 min). The membrane-bound ATPase in the R3 fraction was stimulated 12-fold by treatment with trypsin, and fractionation studies showed that trypsin treatment did not solubilize ATPase activity off the membranous R3 electron transport fraction. The ATPase was not cold labile and the temperature during the preparation of the R3 fraction had no effect on activity; overnight refrigeration at 4 °C, however, resulted in a 25% loss of activity as compared with a 14% loss when the R3 fraction was stored overnight at 25 °C. A marked inactivation (although variable, usually about 60%) did occur by overnight freezing (−20 °C), and subsequent sonication failed to restore ATPase activity. This indicates that membrane reaggregation (by freezing) was not responsible for ATPase inactivation. The addition of azide, ouabain, 2,4-dinitrophenol, or oligomycin to the assay system resulted in neither inhibition nor stimulation of the ATPase activity. The property of trypsin activation and that ATPase activity is highest in the R3 electron transport fraction suggests that its probable functional role is in coupling of electron transport to oxidative phosphorylation.

Biochemical Basis of Heart Function. VI. Influence of Cations on Norepinephrine Transport, Storage, and Synthesis in the Isolated Perfused Rat Heart

1972 ◽  
Vol 50 (5) ◽  
pp. 423-431 ◽  
Author(s):  
V. Balasubramanian ◽  
Naranjan S. Dhalla
Keyword(s):  
Rat Heart ◽  
Heart Function ◽  
Adrenergic Nerve ◽  
Spontaneous Release ◽  
Perfusion Medium ◽  
Experimental Conditions ◽  
Biochemical Basis ◽  
Perfused Rat Heart ◽  
Isolated Perfused Rat Heart ◽  
Adrenergic Nerve Endings

The effects of Ca2+ (0–5 mM), Mg2+ (0–16 mM), K+ (0–20 mM), and Na+ (0–145 mM) on the uptake, subcellular distribution, and release of exogenously given 3H-norepinephrine were investigated in the isolated perfused rat heart. The uptake of 14C-tyrosine, the synthesis of 14C-catecholamines, and the endogenous levels of norepinephrine were also determined under these experimental conditions. The uptake of 3H-norepinephrine was markedly decreased on perfusing the hearts with media containing low concentrations of Na+. Addition of Ca2+ to the perfusion medium containing Na+ increased the uptake of 3H-norepinephrine while Mg2+ or K+ had no effect. Increasing the concentration of Na+ from 0 to 145 mM in the perfusion medium increased the 3H-label in the granular fraction and decreased it in the soluble fraction whereas Ca2+, Mg2+, or K+ did not show any action. The spontaneous release of 3H-label was not influenced by the presence or absence of Ca2+, Mg2+, or K+ in the medium. On the other hand, the absence of Na+ was found to accelerate the spontaneous release of 3H-label and this effect was further enhanced when Ca2+ was also omitted from the perfusion medium. Increasing the concentrations of Ca2+ or Na+ in the perfusion medium decreased the rate of synthesis of 14C-catecholamincs from 14C-tyrosine whereas Mg2+ and K+ showed no effect. Absence of Na+, but not of K+, Ca2+, or Mg2+, resulted in a decrease in the endogenous level of norepinephrine. The amount of newly synthesized 14C-catecholamines in the perfusate was more on perfusing the hearts with Ca2+-free or Na+-free medium. These results are consistent with the view that Na+ is required for storage as well as uptake of norepinephrine in the adrenergic nerve terminals in the heart. Ca2+ appears necessary for the optimal uptake of norepinephrine due to Na+ and has been shown to inhibit the spontaneous release of norepinephrine due to Na+ lack. In addition, Na+ and Ca2+ reduce the rate of synthesis of catecholamines in the adrenergic nerve endings in the heart.

A High Resolution Cytochemical Study of Nuclear ATPase in Human Spermatozoa

1975 ◽  
Vol 33 ◽  
pp. 594-595
Author(s):  
A. Sosa ◽  
L. Calzada
Keyword(s):  
High Resolution ◽  
Atpase Activity ◽  
Nuclear Membrane ◽  
Human Sperm ◽  
Sperm Nucleus ◽  
Human Spermatozoa ◽  
Cytochemical Study ◽  
Membrane Bound ◽  
Sperm Nuclei ◽  
Interchromatin Space

The dependence of nuclear metabolism on the function of the nuclear membrane is not well understood. Whether or not the function of the nuclear membrane is partial or totally responsible of the repressed template activity of human sperm nucleus has not at present been elucidated. One of the membrane-bound enzymatic activities which is concerned with the mechanisms whereby substances are thought to cross cell membranes is adenosintriphosphatase (ATPase). This prompted its characterization and distribution by high resolution photogrammetry on isolated human sperm nuclei. Isolated human spermatozoa nuclei were obtained as previously described. ATPase activity was demonstrated by the method of Wachstein and Meisel modified by Marchesi and Palade. ATPase activity was identified as dense and irregularly distributed granules confined to the internal leaflet of the nuclear membrane. Within the nucleus the appearance of the reaction product occurs as homogenous and dense precipitates in the interchromatin space.

scholarly journals Hepatic endosome fractions contain an ATP-driven proton pump

1985 ◽  
Vol 225 (1) ◽  
pp. 51-58 ◽  
Author(s):  
T Saermark ◽  
N Flint ◽  
W H Evans
Keyword(s):  
Endoplasmic Reticulum ◽  
Golgi Apparatus ◽  
Atpase Activity ◽  
Proton Pump ◽  
Subcellular Distribution ◽  
Plasma Membranes ◽  
Specific Activity ◽  
Ph Gradients ◽  
Subcellular Organelle ◽  
Liver Homogenates

Endosome fractions were isolated from rat liver homogenates on the basis of the subcellular distribution of circulating ligands, e.g. 125I-asialotransferrin internalized by hepatocytes by a receptor-mediated process. The distribution of endocytosed 125I-asialotransferrin 1-2 min and 15 min after uptake by liver and a monensin-activated Mg2+-dependent ATPase activity coincided on linear gradients of sucrose and Nycodenz. The monensin-activated Mg2+-ATPase was enriched relative to the liver homogenates up to 60-fold in specific activity in the endosome fractions. Contamination of the endosome fractions by lysosomes, endoplasmic reticulum, mitochondria, plasma membranes and Golgi-apparatus components was low. By use of 9-aminoacridine, a probe for pH gradients, the endosome vesicles were shown to acidify on addition of ATP. Acidification was reversed by addition of monensin. The results indicate that endosome fractions contain an ATP-driven proton pump. The ionophore-activated Mg2+-ATPase in combination with the presence of undegraded ligands in the endosome fractions emerge as linked markers for this new subcellular organelle.

Biochemical Basis of Heart Function. IV. Energy Metabolism and Calcium Transport in Hearts of Vitamin E Deficient Rats

1971 ◽  
Vol 49 (10) ◽  
pp. 909-918 ◽  
Author(s):  
Margaret Fedelesova ◽  
Prakash V. Sulakhe ◽  
John C. Yates ◽  
Naranjan S. Dhalla
Keyword(s):  
Vitamin E ◽  
Sarcoplasmic Reticulum ◽  
Heart Function ◽  
Adenine Nucleotides ◽  
Creatine Phosphate ◽  
Normal Diet ◽  
Vitamin E Deficiency ◽  
Deficient Diet ◽  
Cardiac Sarcoplasmic Reticulum ◽  
Biochemical Basis

Feeding a vitamin E deficient diet to rats for 10 weeks was found to decrease myocardial creatine phosphate, ATP, ATP/ADP ratio, NAD+, NADP+, and NADPH, whereas the level of ADP was increased without any changes in the levels of AMP, total adenine nucleotides, NADH, and ATP/AMP ratio. The levels of ATP and pyridine nucleotides were restored fully, whereas creatine phosphate was restored partially on feeding a normal diet for 4 weeks to animals previously on the vitamin E deficient diet for 10 weeks. Vitamin E deficiency was found to increase cardiac lactate, pyruvate, and lactate/pyruvate ratio and decrease the activities of lactate dehydrogenase and malate dehydrogenase. The activity of Na+–K+-stimulated, ouabain-sensitive ATPase was markedly elevated in the hearts of animals on the vitamin E deficient diet. The ATP-dependent calcium accumulation by the sarcoplasmic reticular fraction in the absence and presence of P1 or oxalate was greater in the vitamin E deficient heart. Vitamin E deficiency also increased the Ca2+-stimulated ATPase activity of the cardiac sarcoplasmic reticulum. Although myocardial contractility of the hearts from vitamin E deficient rats was depressed, no damage to the ultrastructures of mitochondria and sarcoplasmic reticulum was apparent. These results indicate marked alterations in myocardial metabolism due to vitamin E deficiency and it is suggested that such changes are due to abnormalities in the processes of both energy production and utilization.

THE INCORPORATION OF PALMITATE-1-14C BY RAT LUNG IN VITRO

1967 ◽  
Vol 45 (4) ◽  
pp. 597-607 ◽  
Author(s):  
A. Naimark ◽  
D. Klass
Keyword(s):  
Specific Activity ◽  
Specific Radioactivity ◽  
Phosphatidyl Inositol ◽  
Phosphatidyl Serine ◽  
Rat Lung ◽  
Exchange Reactions ◽  
Ethanolamine Phosphatidyl ◽  
Exogenous Glucose ◽  
Lipid Fractions

The incorporation of palmitate-1-14C into various lipid fractions was studied in rat lung in vitro. Most of the radioactivity was found in phospholipid, although in terms of decreasing specific activity the lipids were ranked: free fatty acid (FFA), glycerides, phospholipid. In addition to the usual glycerophosphatides, rat lung contained a substance with some of the chromatographic characteristics of phosphatidyl dimethylethanolamine. In terms of decreasing specific activities the phospholipids were ranked: phosphatidyl choline, phosphatidyl ethanolamine, phosphatidyl dimethylethanolamine, phosphatidyl serine plus phosphatidyl inositol, sphingomyelin plus lysophosphatidyl choline. Inhibition of oxidative energy production by hypoxia, cyanide, or low temperature markedly depressed the esterification of palmitate-1-14C. Less marked depression was observed in the absence of exogenous glucose. In all cases the decreased incorporation was associated with an increase in the total and specific radioactivity in tissue FFA. It is concluded that energy-independent exchange reactions are probably of little importance in the incorporation of FFA into esterified lipids of lung tissue. Under conditions of metabolic inhibition the penetration of labelled FFA into the tissue FFA pool does not appear to limit esterification.

scholarly journals Characterization of a Mutant ofLactococcus lactiswith Reduced Membrane-bound ATPase Activity under Acidic Conditions

1998 ◽  
Vol 62 (8) ◽  
pp. 1574-1580 ◽  
Author(s):  
Seigo AMACHI ◽  
Kohei ISHIKAWA ◽  
Shuji TOYODA ◽  
Yasuo KAGAWA ◽  
Atsushi YOKOTA ◽  
...  
Keyword(s):  
Atpase Activity ◽  
Membrane Bound ◽  
Acidic Conditions

scholarly journals Dynamics of myocardial adaptation to low-flow ischemia and hypoxemia

1996 ◽  
Vol 271 (6) ◽  
pp. H2300-H2305
Author(s):  
G. Merati ◽  
S. Allibardi ◽  
L. D. Monti ◽  
J. W. de Jong ◽  
M. Samaja
Keyword(s):  
Low Flow ◽  
Control Performance ◽  
Perfusion Medium ◽  
Atp Production ◽  
Lactate Release ◽  
Rat Hearts ◽  
O2 Supply ◽  
Developed Pressure ◽  
Myocardial Adaptation ◽  
Isolated Rat

We investigated whether one or more factors control performance in O2-limited hearts. For this purpose, we measured the dynamics of myocardial adaptation to reduced O2 supply with a specially designed setup, analyzing early changes after reduction in either flow of the perfusion medium or its PO2. For 10 min, 38 isolated rat hearts underwent low-flow ischemia or hypoxemia, matched for O2 supply. Early during ischemia, developed pressure declined at a rate of 311 +/- 25 mmHg/s; lactate release increased and then leveled off to 3.4 +/- 0.7 mumol/min within 2 min. During hypoxemia, pressure dropped initially, as observed during ischemia. However, it then increased before slowly decreasing. Lactate release during hypoxemia peaked at 13.0 +/- 2.3 mumol/min after 2 min, leveling off to 3.5 +/- 1.3 mumol/min. Glycogen decreased by 52 and 81% in ischemic and hypoxemic hearts, respectively (P < 0.05). Reexposure to ischemia or hypoxemia induced comparable changes in both groups. We conclude that, at the beginning of ischemia, a single factor does limit myocardial performance. This variable, which remains undisturbed for 10 min, is presumably O2 availability. In contrast, approximately 20 s after induction of hypoxemia, glycolytic ATP production can partially override low O2 availability by providing most of the energy needed. During repeated restriction of O2 supply, O2 availability alone limits performance during both ischemia and hypoxemia.

Sign in / Sign up

Export Citation Format

Share Document