Hierarchies of ATP-consuming processes: direct compared with indirect measurements, and comparative aspects

2001 ◽  
Vol 355 (2) ◽  
pp. 389-395 ◽  
Author(s):  
Wolfgang WIESER ◽  
Gerhard KRUMSCHNABEL
Keyword(s):  
Protein Synthesis ◽  
Oxygen Consumption ◽  
Atpase Activity ◽  
Fish Species ◽  
The Other ◽  
Leucine Incorporation ◽  
Hepatoma Cell Line ◽  
Total Oxygen ◽  
Other Hand ◽  
Energy Limitation

The original aim of the present study was to deal with two problems that had emerged from a study on hierarchies of ATP-consuming processes in cells [Buttgereit and Brand (1995) Biochem. J. 312, 163-167]. Firstly, we wanted to find out whether the results of that study had been influenced by the method used for the determination of process activity and, secondly, we wondered whether and to what extent the structure of the hierarchy established for cell suspensions under energy-limiting conditions might depend on the type of cell or on the lifestyle, ecology and phylogenetic status of the species from which the cells were derived. We confined our study to the two most prominent ATP consumers of cells: protein synthesis and the Na+/K+-ATPase, measuring their activity directly by [3H]leucine incorporation and Rb+-flux respectively. We found large differences in the sensitivity of protein synthesis to energy limitation between hepatocytes from an anoxia-tolerant fish species and an anoxia-sensitive fish species (goldfish and rainbow trout respectively). On the other hand, Na+/K+-ATPase activity was hardly affected by energy limitation in the hepatocytes from both fish species. We also studied the response of a human hepatoma cell line, HepG2, to energy limitation and found both protein synthesis and Na+/K+-ATPase activity to be equally sensitive to energy limitation, but more sensitive than the Na+/K+-ATPase of the two fish species. A comparison of the indirect and direct methods for measuring protein synthesis revealed the rate of oxygen consumption to be functionally related to the concentration of cycloheximide, the inhibitor used. It was found that at 15mM cycloheximide [three orders of magnitude higher than the concentration at which the incorporation of free amino acids (FAA) into protein is inhibited] total oxygen consumption was suppressed by 71-75%, whereas the measured rate of [3H]leucine incorporation into protein suggested that the cycloheximide-sensitive fraction should have amounted to not more than approx. 10% of the total oxygen consumption. On the other hand, the amount of oxygen consumption suppressed with the high concentration of cycloheximide corresponded almost exactly to the increase in oxygen consumption of cells incubated in an FAA-enriched medium compared with cells incubated in a standard, FAA-free medium. Our major conclusions are, firstly, that high concentrations of cycloheximide disrupt cellular metabolism, bringing to a standstill all those processes that can be stimulated by incubating starved cells in an FAA-enriched medium, secondly, that the attempt to estimate the metabolic cost of protein synthesis by inhibiting oxygen consumption with cycloheximide leads to spurious results, and, thirdly, that the structure of a ‘hierarchy’ of ATP-consumers may reflect the lifestyle and physiology of the species studied.

EFFECT OF ACTINOMYCIN D ON TSH-INDUCED STIMULATION OF THYROIDAL PROTEIN SYNTHESIS IN THE RAT

1974 ◽  
Vol 77 (1) ◽  
pp. 64-70 ◽  
Author(s):  
Gustav Wägar
Keyword(s):  
Protein Synthesis ◽  
Actinomycin D ◽  
The Other ◽  
Leucine Incorporation ◽  
Short Term ◽  
Other Hand ◽  
Thyroid Glands ◽  
Translational Level ◽  
Stimulation Of

ABSTRACT Whether the short-term regulation of thyroidal protein synthesis by TSH occurs at the transcriptional or the translational level was tested by measuring the effect of actinomycin D (act D) on the TSH-induced stimulation of L-14C-leucine incorporation into the thyroidal proteins of rats. TSH was injected 6 h before the rats were killed. The thyroid glands were then removed and incubated in vitro in the presence of L-14C-leucine for 2 h. The pronounced stimulation of leucine incorporation in the TSH-treated animals was depressed as compared with controls but still significant even when the animals had been pre-treated with 100 μg act D 24 and 7 h before sacrifice. On the other hand, act D strongly decreased incorporation of 3H-uridine into RNA. Short-term regulation of thyroidal protein synthesis by TSH appears to be partly but not wholly dependent on neosynthesis of RNA. Hence regulation may partly occur at the translation level of protein synthesis.

Hypothermia in iron deficiency due to altered triiodothyronine metabolism

1980 ◽  
Vol 239 (5) ◽  
pp. R377-R381 ◽  
Author(s):  
E. Dillman ◽  
C. Gale ◽  
W. Green ◽  
D. G. Johnson ◽  
B. Mackler ◽  
...  
Keyword(s):  
Oxygen Consumption ◽  
Iron Deficiency ◽  
Ambient Temperature ◽  
The Other ◽  
Other Hand ◽  
Iron Deficient ◽  
Catecholamine Response ◽  
Cutaneous Vasoconstriction

Iron-deficient rats become hypothermic and have an excessive catecholamine response when exposed to an ambient temperature of 4 degrees C. This is not due to changes in body insulation, since thickness is unaltered, since differences persist after removal of hair, and since cutaneous vasoconstriction is intact. On the other hand, oxygen consumption of iron-deficient animals at 4 degrees C is reduced, 39 +/- 3 ml . kg-1 . min-1 compared to 63 +/- 2 in control animals. Thyroxine (T4) values at 4 degrees C were 4.34 +/- 0.20 microgram/dl sera as compared to control values of 3.6 +/- 0.32. Triiodothyronine (T3) values of iron-deficient animals in the cold were 48 +/- 6.8 ng/dl as compared to 72 +/- 5.6 in control animals. Treatment of iron-deficient animals with iron was shown to normalize the plasma T3 response at 4 degrees C within 6 days. Thyroidectomized iron-deficient animals injected with T3 did not show hypothermia at 4 degrees C, whereas thyroidectomized iron-deficient animals injected with T4 showed hypothermia, increased catecholamines, and decreased T3 levels as compared to non-iron-deficient animals similarly treated. It is proposed that iron deficiency impairs conversion of T4 to T3 and that this is primarily responsible for the hypothermia observed.

Action of adenosine on energetics, protein synthesis and K(+) homeostasis in teleost hepatocytes

2000 ◽  
Vol 203 (17) ◽  
pp. 2657-2665 ◽  
Author(s):  
G. Krumschnabel ◽  
C. Biasi ◽  
W. Wieser
Keyword(s):  
Protein Synthesis ◽  
Oxygen Consumption ◽  
Atpase Activity ◽  
Oncorhynchus Mykiss ◽  
Carassius Auratus ◽  
Goldfish Carassius Auratus ◽  
Inhibition Of Protein Synthesis ◽  
Rate Of Oxygen Consumption ◽  
A1 Receptor ◽  
Lactate Formation

In a comparative study, we analysed the effects of adenosine on the energetics, protein synthesis and K(+)homeostasis of hepatocytes from the anoxia-tolerant goldfish Carassius auratus and the anoxia-intolerant trout Oncorhynchus mykiss. The rate of oxygen consumption did not respond immediately to the addition of adenosine to the cells from either species, but showed a significant decrease in trout hepatocytes after 30 min. The anaerobic rate of lactate formation was not significantly affected by adenosine in goldfish hepatocytes, but was increased in trout cells. We also studied the effects of adenosine on the two most prominent ATP consumers in these cells, protein synthesis and Na(+)/K(+)-ATPase activity. Under aerobic conditions, adenosine inhibited protein synthesis of hepatocytes from goldfish by 51% and of hepatocytes from trout by 32%. During anoxia, the rate of protein synthesis decreased by approximately 50% in goldfish hepatocytes and by 90% in trout hepatocytes, and this decrease was not altered by the presence of adenosine. Adenosine inhibited normoxic Na(+)/K(+)-ATPase activity and K(+)efflux by 20–35% in the cells of both species. An investigation into the mechanism underlying the inhibition of protein synthesis by adenosine indicated that, in the goldfish cells, adenosine acts via a membrane receptor-mediated pathway, i.e. the effect of adenosine was abolished by applying the A1 receptor antagonist 8-phenyltheophylline. In the trout, however, the uptake of adenosine into hepatocytes seems to be required for an effect on protein synthesis. [Ca(2+)](i) does not seem to be involved in the inhibition of protein synthesis by adenosine.

Polysomal organization in growing and resting cultures and its relation to protein synthesis in Tetrahymena

1983 ◽  
Vol 59 (1) ◽  
pp. 121-131
Author(s):  
P. Isberner ◽  
G. Cleffmann
Keyword(s):  
Protein Synthesis ◽  
Amino Acid ◽  
Growth Conditions ◽  
The Other ◽  
Protein Accumulation ◽  
Resting Cells ◽  
Amino Acid Incorporation ◽  
Total Rna ◽  
Acid Incorporation ◽  
Other Hand

Cytosol from Tetrahymena cells growing at different rates was isolated and separated by centrifugation into polysomal and non-polysomal fractions. The RNAs of either fraction were separated chromatographically into poly(A)+ RNA and poly(A)-RNA. It was found that in resting cultures the total RNA per cell is only about half of that of rapidly growing cultures. All fractions of RNA were reduced proportionally. Thus, the percentage of polysomally bound total RNA (70% of cytosol RNA) and polysomally bound poly(A)+ RNA (72% of cytosol poly(A)+ RNA) is the same in growing and resting cultures. Differences, however, were found in the polysomal structure. Polysomes from resting cultures contained significantly fewer ribosomes. The amounts of RNA bound to polysomes were related to the rate of protein synthesis under different growth conditions. The decrease in cellular RNA corresponded well with the reduction in amino acid incorporation in resting cells. The rate of protein accumulation in resting cells, on the other hand, was considerably less, suggesting that polypeptides in resting cultures are less stable.

Trypanosoma cruzi Epimastigotes Express the Ouabain-and Vanadate-Sensitive (Na++K+)ATPase Activity

1998 ◽  
Vol 53 (11-12) ◽  
pp. 1049-1054 ◽  
Author(s):  
Celso Caruso-Neves ◽  
Marcelo Einicker-Lamas ◽  
Carlos Chagas ◽  
Mecia Maria Oliveira ◽  
Adalberto Vieyra ◽  
...  
Keyword(s):  
Trypanosoma Cruzi ◽  
Atpase Activity ◽  
The Other ◽  
Other Hand ◽  
P Type

Abstract The presence of (Na++K+)ATPase activity in CL14 clone and NIH NTY strain of Trypano­soma cruzi epimastigotes is demonstrated. A Na+ plus K+ stimulated ATPase activity is found in both strains. The optimal Na+/K+ ratio is 5:1 and 9:1 in CL14 clone and NIH NTY strain, respectively. In both strains, vanadate completely inhibits the ouabain-sensitive ATPase activ­ity indicating that it belongs to the P-type (E 1/E2) family of ion-transporting ATPases. The I50 for vanadate is 0.66 ± 0.04 and 0.04 ± 0.02 μᴍ in CL14 clone and NIH NTY strain, respectively. These data indicate that both strains of T. cruzi epimastigotes express the oua­ bain-and vanadate-sensitive (Na++K+)ATPase activity. On the other hand, the discrepancy between the parameters analyzed for the inhibitors suggests that they express different iso­ forms of this enzyme.

Effect of different oxygen concentrations in the gas phase on biochemical activities of goat mammary tissue in organ culture

1989 ◽  
Vol 56 (1) ◽  
pp. 1-5 ◽  
Author(s):  
Josef Škarda ◽  
Eva Urbanová
Keyword(s):  
Protein Synthesis ◽  
Organ Culture ◽  
Gas Phase ◽  
The Other ◽  
Mammary Tissue ◽  
Relative Response ◽  
Other Hand ◽  
Rna And Protein Synthesis ◽  
Casein Synthesis ◽  
Oxygen Concentrations

SummaryNon-secretory mammary expiants from virgin goats showed higher RNA and protein synthesis in a low O2 gas phase (air) than in high O2 (95% O2). Lipid and casein synthesis was not affected significantly by the concentration of O2 in the atmosphere during culture. on the other hand, the more developed mammary tissue from primigravid goats showed higher lipid, casein and protein synthesis in 95% O2. The relative response of mammary tissue to hormones was not substantially different when cultured in the presence of a low or high O2 gas phase. As Hepes-buffered medium was found not to need a supply of CO2 to maintain the correct pH and as Hepes did not interfere with biochemical activities of cells, it is recommended to use it for cultures in a low O2 gas phase.

Leucine and sport

2007 ◽  
Vol 277 ◽  
pp. 1-3 ◽  
Keyword(s):  
Amino Acids ◽  
Protein Synthesis ◽  
Enhanced Recovery ◽  
The Other ◽  
Sports Performance ◽  
Other Hand ◽  
Improved Function ◽  
Endurance Activity

In a nutshellExercise increases oxidation of amino acids such as leucine.On the other hand, leucine is a powerful mediator of protein synthesis in muscle, promoting improved function, reduced fatigue and enhanced recovery from exertion. There is RCT evidence that leucine improves sports performance, particularly for endurance activity, but more evidence is still needed.

Assessment of the Effect of C-Peptide Level on Na-K ATPase Activity In Individuals with Type II Diabetic Peripheral Neuropathy

2021 ◽  
Vol 53 (03) ◽  
pp. 213-219
Author(s):  
Eman H Al-Rikabi ◽  
Keyword(s):  
Enzyme Activity ◽  
Diabetic Neuropathy ◽  
Atpase Activity ◽  
Negative Correlation ◽  
Clinical Symptoms ◽  
The Other ◽  
C Peptide ◽  
Other Hand ◽  
Peptide Level

Background: Both Type 1 and Type 2 diabetes can cause neuropathy, which is a frequent and severe consequence. C-peptide depletion may be partly related to in the occurrence of certain diabetic complications. It has been demonstrated that even a little amount of residual C-peptide has a considerable metabolic advantage. Objective: The study’s objective was to predict the relation of plasma C-peptide levels in patients with diabetic neuropathy, and its effect on Na-K ATPase activity. Design and Methods: In this case-control study, 150 individuals have been included: 80 patients with diabetic neuropathy, 40 diabetics without neuropathy and 30 non-diabetic subjects as a control. Patients in the first group were carefully chosen based on their clinical symptoms and nerve conduction studies results. The assessment of plasma C-peptide was done by ELISA, Na-K ATPase enzyme activity by spectrophotometer, and HbA1C by HPLC. Results: Mean plasma C-peptide level and Erythrocyte Na-K ATPase activity were substantially lower in neuropathy type 2 DM patients compared to diabetes without neuropathy and control (p= 0.002, 0.000 respectively). The negative correlation between C-peptide with HbA1c, and diabetes period were all negligible (p= 0.447,0.098), Even though there was a notable negative correlation with age (p= 0.03). On the other hand, the relationship linking C-peptide and Na-K ATPase enzyme activity was shown to be insignificant (p=0.69). Conclusions: Diabetic neuropathy is related to a low C-peptide level. The association between C-peptide and Na-K ATPase enzyme activity, on the other hand, was shown to be insignificant. C-peptide HbA1c, and duration of diabetes all had minor negative associations.

CORTISOLWIRKUNGEN AUF DEN PROTEINUND ENERGIESTOFFWECHSEL IN DER PERIPHERIE

1968 ◽  
Vol 59 (2) ◽  
pp. 193-202
Author(s):  
H. Kröner ◽  
W. Staib
Keyword(s):  
Skeletal Muscle ◽  
Protein Synthesis ◽  
Energy Consumption ◽  
Connective Tissue ◽  
Oxidative Metabolism ◽  
The Other ◽  
Other Hand ◽  
Inhibition Of Protein Synthesis ◽  
Short Time ◽  
Rna And Dna

ABSTRACT By measuring protein, RNA and DNA in skeletal muscle and spleen we were unable to demonstrate an enhanced catabolism due to cortisol. On the other hand we could demonstrate a reduced anabolism by diminished incorporation of 14C labelled leucin in protein of muscle, spleen and skin. Simultaneously an increase of ATP and an equivalent decrease of ADP were found in connective tissue, which were regarded as a consequence of reduced energy consumption. In skeletal muscle a decrease of energy consumption due to cortisol was only indirect demonstrable by reduced glycolysis when oxidative metabolism was eliminated for a short time. It seems obvious, that inhibition of protein synthesis causes the diminished energy consumption.

scholarly journals The fuel of the spleen. Studies using a new method for perfusing the rat spleen with whole blood

1989 ◽  
Vol 263 (2) ◽  
pp. 325-332 ◽  
Author(s):  
M A Mindham ◽  
P A Mayes
Keyword(s):  
Oxygen Consumption ◽  
Fatty Acid ◽  
Glucose Uptake ◽  
Whole Blood ◽  
Glucose Oxidation ◽  
The Other ◽  
Cell Populations ◽  
Total Oxygen ◽  
Total Glucose ◽  
Oxidation Of Glucose

1. An improved rat spleen perfusion is described incorporating a method of defibrination which avoids the use of heparin and enables the spleen to be perfused with rat blood for several hours at a haematocrit of 40% and for 12 h or more at a haematocrit of 20%. 2. Glucose oxidation accounted for 11.6% of the total oxygen consumption but this represented only 8% of total glucose uptake, which was largely converted to lactate and released into the perfusate. However, significant amounts of lactate were oxidized. These results can be explained by the presence of at least two cell populations, one emphasizing the anaerobic oxidation of glucose and the other aerobic metabolism, particularly of lactate. 3. Non-esterified fatty acid and 3-hydroxybutyrate, when available at physiological concentrations, were shown to be major oxidative fuels of the spleen. 4. Chylomicron triacylglycerol was hydrolysed readily and taken up, but not oxidized extensively.

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