Use of spin probe in the determination of oxygen consumption in human neutrophils

A quantitative method of the oxygen consumption rate measurement by human blood neutrophils upon their activation is described. This method is based on the spin exchange determination between 3-carbamoyl-2,2,5,5-tetramethyl-3-pyrroline-1-yloxyl spin probe and oxygen molecules from the ESR spectra. Method allows the determination of about 1 micromolar concentrations of oxygen.     

Kinetics of oxygen consumption after a single isometric tetanus of frog sartorius muscle at 20 degrees C

The time-course of the rate of oxygen consumption (QO2) has been measured in the excised frog sartorius muscle after single isometric tetani of 0.1-1.0 s at 20 degrees C. To measure deltaQO2(t), the change in QO2 from its basal level, a novel method was devised, based on the validity in this tissue of the one-dimensional diffusion equation for oxygen, established in the preceding paper. After a tetanus, deltaQO2 reached a peak within 45-90 s, then declined exponentially, and could be well fit by deltaQO2(t) = QO + Q1(epsilon -k1t - epsilon-k2t). tau2 (= 1/k2), which characterized the rise of deltaQO2, was a decreasing function of tetanus duration (range: from 1.1 +/- 0.28 min [nu = 5] for a 0.1-s tetanus, to 0.34 +/- 0.05 min [nu = 8] for a 1.0-sec tetanus). tau1 (= 1/k1), which characterized the decline of deltaQO2, was not dependent on tetanus duration, with mean 3.68 +/- -.24 min (nu = 46). A forthcoming paper in this series shows that these kinetics of deltaQO2 are the responses to impulse-like changes in the rate of ATP hydrolysis. The variation of tau2 with tetanus duration thus indicates the involvement of a nonlinear process in the coupling of O2 consumption to ATP hydrolysis. However, the monoexponential decline of deltaQO2(t), with time constant independent of tetanus duration, suggests that during this phase, the coupling is rate-limited by a single reaction with apparent first order kinetics.     

Method for the determination of oxygen consumption rates and diffusion coefficients in multicellular spheroids

A method has been developed for the quantitative evaluation of oxygen tension (PO2) distributions in multicellular spheroids measured with O2-sensitive microelectrodes. The experimental data showed that multicellular tumor spheroids in stirred growth media were characterized by a diffusion-depleted zone surrounding the spheroids. This zone was elicited by an unstirred layer of medium next to the spheroid leading to a continuous decrease in the PO2 values from the bulk medium towards the spheroid surface. Theoretical considerations demonstrate that the volume-related O2 consumption rate, Q, in the spheroids can be assessed by measuring the PO2 gradient in the diffusion-depleted zone outside the spheroids. Accordingly, Krogh's diffusion constant, KS, in the spheroids can be determined through measuring the PO2 gradient within the spheroids. The results obtained suggest that multicellular spheroids represent useful in vitro tumor models for the experimental and theoretical analysis of the interrelationship among O2 supply to tumor cells, O2 metabolism in tumors tissue, and the responsiveness of cancer cells to treatment.     

Kinetic studies of biodegradation of insoluble compounds by continuous determination of oxygen consumption

Continuous determination of oxygen consumption by electrolytic respirometry has been experimented as a means to study the biodegradation kinetics of scarcely soluble environmental pollutants. The substrates used were the polycyclic aromatic hydrocarbons (PAH), naphthalene, phenanthrene, anthracene, fluoranthene and pyrene. The definition of an appropriate mode of PAH supply, either as crystals or more generally as a solution in a water non-miscible solvent, was found essential for yielding reproducible biodegradation kinetics. In these conditions, for all compounds tested, oxygen determination was found suitable for quantitative evaluation of PAH biodegradation and formation of biomass and soluble metabolites. The study of biodegradation kinetics with this methodology showed that a first phase of exponential growth could be characterized in most cases, followed by a phase of limited growth. Possible mechanisms involved in insoluble substrate uptake are discussed. During exponential growth, the bacteria utilized (although not necessarily exclusively) the PAH solubilized in the aqueous medium.     

Arachidonate-dependent oxygen consumption in Dictyostelium discoideum

A central feature of the processes of aggregation and differentiation in the cellular slime mould Dictyostelium discoideum is the periodic excitatory cycle. Originally thought to involve primarily fluctuations in cyclic AMP levels, this excitatory cycle has since been shown to involve changes in several other second messengers including cyclic GMP, calcium and inositol trisphosphate. Previous work from this laboratory using specific inhibitors strongly suggested a role for eicosanoids in this stimulus-response process. Production of eicosanoids from fatty acid precursors is an oxygen-consuming process. In this paper, we report on oxygen consumption measurements in intact D. discoideum cells and in cell extracts. We demonstrate the existence of an azide-insensitive component of oxygen consumption which can be stimulated by the addition of arachidonate and other polyunsaturated fatty acids, and at least partially inhibited by meclofenamate and eicosatetraynoic acid, both of which block eicosanoid biosynthesis in higher organisms. These observations provide further evidence for the existence of an eicosanoid-metabolizing system in D. discoideum.     

Below-halocline oxygen consumption in the Kattegat

Sediment and seston oxygen consumption rates below the sharp halocline in the south-eastern part of the shallow Kattegat were measured and compared to calculated rates of carbon addition through the halocline. The mean rate of decrease in deep-water oxygen concentrations between March and September 1988 was 1.0 ml O₂ M^–3 h^–1. Measurements of benthic oxygen uptake using laboratory-incubated sediment cores from depths 30 m gave a mean value of 7.8 ml O₂ m^–2 h^–1. Below-halocline water (from 20 m, 30 m and 1 m above bottom) incubated in bottles showed oxygen consumption rates varying from 0.5 ml O₂ m ^–3 h^–1 in March to 2.8 ml O₂ M^–3 h^-1 in late August. The sum of benthic and deep-water oxygen consumption was equivalent to a mean oxygen decrease rate of 1.7 ml O₂ m^–3 h^–1 below the halocline. Of the total oxygen consumption below the halocline 65% was due to oxygen up-take in the water and 35% was due to benthic oxygen consumption. The sum of oxygen consumption measured in sediment cores and in bottles corresponds to a carbon utilisation of 80.1 g C m^–2 (respiratory quotient (RQ), assumed 1.0 and 1.4 for water and sediment, respectively), while the decrease in deep-water oxygen concentration was equivalent to 43.0 g C m^–2 (RQ assumed = 1.0). Using published values for the external N loading (including deep-water supply), ¹⁵NO₃-uptake, ¹⁴CO₂-uptake in combination with % ¹⁵NO₃-uptake of total ¹⁵N-uptake (nitrate, ammonia and urea) and a Redfield C/N ratio of 6.6, rates of carbon addition (new or export production) through the halocline were calculated to 31.9, 46.7 and 36.3 g C m^–2, respectively, with a mean value of 38.3 g C m^–2 for the 8 month period March–September. This is somewhat less than the value (50.5 g C m^–2) calculated from a published empirical relationship between total and export production. The fact that the calculated carbon addition through the halocline was appreciably less than the carbon equivalent of the measured below-halocline respiration may be an effect of sediment focusing (horizontal transport of sedimenting material to deeper areas), since the bottom area below the halocline is much smaller than the total area of the Kattegat. A lower observed decrease in the oxygen concentration below the halocline compared to the sum of measured sediment and deep-water oxygen consumption on the other hand indicates oxygen supply to below-halocline waters through advection and/or vertical entrainment.     

Matching coronary blood flow to myocardial oxygen consumption.

At rest the myocardium extracts approximately 75% of the oxygen delivered by coronary blood flow. Thus there is little extraction reserve when myocardial oxygen consumption is augmented severalfold during exercise. There are local metabolic feedback and sympathetic feedforward control mechanisms that match coronary blood flow to myocardial oxygen consumption. Despite intensive research the local feedback control mechanism remains unknown. Physiological local metabolic control is not due to adenosine, ATP-dependent K(+) channels, nitric oxide, prostaglandins, or inhibition of endothelin. Adenosine and ATP-dependent K(+) channels are involved in pathophysiological ischemic or hypoxic coronary dilation and myocardial protection during ischemia. Sympathetic beta-adrenoceptor-mediated feedforward arteriolar vasodilation contributes approximately 25% of the increase in coronary blood flow during exercise. Sympathetic alpha-adrenoceptor-mediated vasoconstriction in medium and large coronary arteries during exercise helps maintain blood flow to the vulnerable subendocardium when cardiac contractility, heart rate, and myocardial oxygen consumption are high. In conclusion, several potential mediators of local metabolic control of the coronary circulation have been evaluated without success. More research is needed.     

The validation of backward extrapolation of submaximal oxygen consumption from the oxygen recovery curve.

The purpose of this study was to determine the validity and practicality of exponential vs linear backward extrapolation of the O2 recovery curve for prediction of exercise oxygen consumption (VO2). Eight men and women, age 20.1, 0.9 years, body mass 66.0, 2.5 kg (mean, SEM), completed seven bouts of cycle ergometer exercise at submaximal power outputs ranging from 50 to 175 W. Respiratory gases were collected from each subject during exercise and recovery. The monoexponential extrapolation of five recovery samples (r2 = 0.85) and linear extrapolation of one recovery sample taken during the first 20-s of recovery (r2 = 0.83) accounted for similar amounts of variance in predicting exercise VO2. The linear regression equation was the most practical predictor, as only one recovery gas sample was necessary and it did not require the complicated mathematical techniques used in exponential regression.     

Cell surface oxygen consumption: a major contributor to cellular oxygen consumption in glycolytic cancer cell lines

Oxygen consumption for bioenergetic purposes has long been thought to be the prerogative of mitochondria. Nevertheless, mitochondrial gene knockout (rho(0)) cells that are defective in mitochondrial respiration require oxygen for growth and consume oxygen at the cell surface via trans-plasma membrane electron transport (tPMET). This raises the possibility that cell surface oxygen consumption may support glycolytic energy metabolism by reoxidising cytosolic NADH to facilitate continued glycolysis. In this paper we determined the extent of cell surface oxygen consumption in a panel of 19 cancer cell lines. Non-mitochondrial (myxothiazol-resistant) oxygen consumption was demonstrated to consist of at least two components, cell surface oxygen consumption (inhibited by extracellular NADH) and basal oxygen consumption (insensitive to both myxothiazol and NADH). The extent of cell surface oxygen consumption varied considerably between parental cell lines from 1% to 80% of total oxygen consumption rates. In addition, cell surface oxygen consumption was found to be associated with low levels of superoxide production and to contribute significantly (up to 25%) to extracellular acidification in HL60rho(0) cells. In summary, cell surface oxygen consumption contributes significantly to total cellular oxygen consumption, not only in rho(0) cells but also in mitochondrially competent tumour cell lines with glycolytic metabolism.