Changes In Oxygen Consumption Rate Research Articles

Metabolic Profile of Pancreatic Acinar and Islet Tissue in Culture

BackgroundThe amount and condition of exocrine impurities may affect the quality of islet preparations, especially during culture. In this study, the objective was to determine the oxygen demand and viability of islet and acinar tissue post-isolation and whether they change disproportionately while in culture. MethodWe compared the oxygen consumption rate (OCR) normalized to DNA (OCR/DNA, a measure of fractional viability in units of nmol/min/mg DNA), and the percent change in OCR and DNA recoveries between adult porcine islet and acinar tissue from the same preparation (paired) over 6–9 days of standard culture. Paired comparisons were done to quantify differences in OCR/DNA between islet and acinar tissue from the same preparation, at specified time points during culture. ResultsThe mean (±SE) OCR/DNA was 74.0 ± 11.7 units higher for acinar (vs islet) tissue on the day of isolation (n = 16, P < .0001), but 25.7 ± 9.4 units lower after 1 day (n = 8, P = .03), 56.6 ± 11.5 units lower after 2 days (n = 12, P = .0004), and 65.9 ± 28.7 units lower after 8 days (n = 4, P = .2) in culture. DNA and OCR recoveries decreased at different rates for acinar versus islet tissue over 6–9 days in culture (n = 6). DNA recovery decreased to 24 ± 7% for acinar and 75 ± 8% for islets (P = .002). Similarly, OCR recovery decreased to 16 ± 3% for acinar and remained virtually constant for islets (P = .005). ConclusionDifferences in the metabolic profile of acinar and islet tissue should be considered when culturing impure islet preparations. OCR-based measurements may help optimize pre–islet transplantation culture protocols.

Bioenergetic characterization of macrophage inflammatory responses attenuated by anthocyanins (383.2)

Anthocyanins present strong anti‐inflammatory properties, thus leading to the potential use as functional food ingredients for the prevention and treatment of chronic inflammatory diseases and increased intestinal permeability to endotoxins, notably LPS. However, the differences between individual anthocyanins and their effects on cellular signaling and metabolism need to be further elucidated. In this study, we evaluated the protection afforded by anthocyanins against LPS‐induced inflammatory response in RAW 264.7 macrophages and determined changes in oxygen consumption rates (OCR) and extracellular acidification rates (ECAR) during the macrophage activation. Basal OCR and ECAR increased as the macrophages underwent LPS stimulation from 283.3±38.4 to 361.7±47.9 pmol/min, and from 9.7±2.4 to 27.6±3.8 mpH/min, respectively, suggesting that energy required for activation is rapidly provided by changes in glycolytic pathways. When tested in physiological concentrations (0.1‐3 uM), malvidin and, to a certain degree, delphinidin, were more effective than cyaniding or pelargonidin in the reduction of mRNA and protein biomarkers of inflammation. Different effects exerted by individual anthocyanins suggested that the presence of the methylated hydroxyls on the B ring may be critical for their greater biological activity. Anthocyanin‐treated macrophages developed a distinct metabolic profile with respect to glycolysis versus oxidative phosphorylation, in part by increasing spare respiratory capacity and therefore shifting to mitochondria as their main source of ATP.Grant Funding Source: Supported by NCSU faculty start up funds to SK.

Systematic Study of Mitochondrial Toxicity of Environmental Chemicals Using Quantitative High Throughput Screening

A goal of the Tox21 program is to transit toxicity testing from traditional in vivo models to in vitro assays that assess how chemicals affect cellular responses and toxicity pathways. A critical contribution of the NIH Chemical Genomics center (NCGC) to the Tox21 program is the implementation of a quantitative high throughput screening (qHTS) approach, using cell- and biochemical-based assays to generate toxicological profiles for thousands of environmental compounds. Here, we evaluated the effect of chemical compounds on mitochondrial membrane potential in HepG2 cells by screening a library of 1,408 compounds provided by the National Toxicology Program (NTP) in a qHTS platform. Compounds were screened over 14 concentrations, and results showed that 91 and 88 compounds disrupted mitochondrial membrane potential after treatment for 1 or 5 h, respectively. Seventy-six compounds active at both time points were clustered by structural similarity, producing 11 clusters and 23 singletons. Thirty-eight compounds covering most of the active chemical space were more extensively evaluated. Thirty-six of the 38 compounds were confirmed to disrupt mitochondrial membrane potential using a fluorescence plate reader, and 35 were confirmed using a high content imaging approach. Among the 38 compounds, 4 and 6 induced LDH release, a measure of cytotoxicity, at 1 or 5 h, respectively. Compounds were further assessed for mechanism of action (MOA) by measuring changes in oxygen consumption rate, which enabled the identification of 20 compounds as uncouplers. This comprehensive approach allows for the evaluation of thousands of environmental chemicals for mitochondrial toxicity and identification of possible MOAs.

Bioenergetic characterization of adipogenesis impaired by inflammation

Adipocytes have been fundamental in identifying basic cellular mechanisms associated with diabetes and obesity, but their bioenergetics has not been fully understood. We measured changes in oxygen consumption rates (OCR) and extracellular acidification rates (ECAR) during differentiation of naïve and TNF‐α exposed 3T3‐L1 preadipocytes using Seahorse XF24 Extracellular Flux Analyzer, and correlated them to mRNA and protein markers of adipogenesis. Basal OCR and ECAR increased as the 3T3‐L1 cells underwent adipogenic differentiation from 110.7±26.5 to 901.1±86.9 pmol/min, and from 7.7±1.4 to 21.6±1.8 mpH/min, respectively. The complex V inhibitor oligomycin reduced OCR of the mature adipocytes to 63% of baseline rates, suggesting that 37% of cellular oxygen consumption was coupled to ATP synthesis. The combination of complex I and II inhibitors rotenone and antimycin reduced OCR to 16% of baseline, indicating that mitochondrial fraction accounted for 84% of total cellular respiration. Bioenergetic parameters of naïve and TNF‐α exposed preadipocytes and adipocytes responded differently to certain substrates (glucose, pyruvate, lactate, and palmitate), PPARα/γ activators, and β‐adrenergic agonists, highlighting intrinsic differences in maintaining energy homeostasis during adipocyte differentiation and possible molecular targets for nutritional preemption of metabolic disorders.

Accumulation of Copper in Different Tissues and Changes in Oxygen Consumption Rate in Indian Flying Barb , Esomus Danricus ( Hamilton-Buchanan ) Exposed to Sub-Lethal Concentrations of Copper

In the present work, the accumulation pattern of sub-lethal doses (0.005, 0.0025 and 0.001 µgl -1 ) of copper (Cu) in different tissues of Indian flying barb, Esomus danricus and its influence on the rate of oxygen consumption of this fish at the end of 28 days of exposure were studied. It was found that the pattern of accumulation changed with concentration. Cu concentration increased in all the tissues except bone and brain and the rate of uptake of Cu increased with time in the gill, liver and flesh but remained same in kidney. Treatment of 0.005 and 0.0025 µgl -1 of Cu produced significant decline in the rates of oxygen consumption of the fish at all exposure duration, while treatment of 0.001 µgl -1 produced similar decline only after 14 days of exposure when compared to control. Higher doses of exposure had more severe effects.

Loss of coupling between calcium influx, energy consumption and insulin secretion associated with development of hyperglycaemia in the UCD-T2DM rat model of type 2 diabetes.

Previous studies on isolated islets have demonstrated tight coupling between calcium (Ca(2+)) influx and oxygen consumption rate (OCR) that is correlated with insulin secretion rate (ISR). To explain these observations, we have proposed a mechanism whereby the activation of a highly energetic process (Ca(2+)/metabolic coupling process [CMCP]) by Ca(2+) mediates the stimulation of ISR. The aim of the study was to test whether impairment of the CMCP could play a role in the development of type 2 diabetes. Glucose- and Ca(2+)-mediated changes in OCR and ISR in isolated islets were compared with the time course of changes of plasma insulin concentrations observed during the progression to hyperglycaemia in a rat model of type-2 diabetes (the University of California at Davis type 2 diabetes mellitus [UCD-T2DM] rat). Islets were isolated from UCD-T2DM rats before, 1week, and 3weeks after the onset of hyperglycaemia. Glucose stimulation of cytosolic Ca(2+) and OCR was similar for islets harvested before and 1week after the onset of hyperglycaemia. In contrast, a loss of decrement in islet OCR and ISR in response to Ca(2+) channel blockade coincided with decreased fasting plasma insulin concentrations observed in rats 3weeks after the onset of hyperglycaemia. These results suggest that phenotypic impairment of diabetic islets in the UCD-T2DM rat is downstream of Ca(2+) influx and involves unregulated stimulation of the CMCP. The continuously elevated levels of CMCP induced by chronic hyperglycaemia in these islets may mediate the loss of islet function.

Oxygen consumption rate and Na+/K+-ATPase activity in early developmental stages of the sea urchin Paracentrotus lividus Lam.

Changes in oxygen consumption rate and Na+/K+-ATPase activity during early development were studied in the sea urchin Paracentrotus lividus Lam. The oxygen consumption rate increased from 0.12 μmol O2 mg protein−1 h−1 in unfertilized eggs to 0.38 μmol O2 mg protein−1 h−1 25 min after fertilization. Specific activity of the Na+/K+-ATPase was significantly stimulated after fertilization, ranging up to 1.07 μmol Pi h−1 mg protein−1 in the late blastula stage and slightly lower values in the early and late pluteus stages.

Changes in Oxygen Consumption Rate during Development of Larval Japanese Whiting, Sillago japonica

Changes in Oxygen Consumption Rate during Development of Larval Japanese Whiting, Sillago japonica

Complex I Is Rate-limiting for Oxygen Consumption in the Nerve Terminal

Metabolic control analysis was used to determine the spread of control exerted by the electron transport chain complexes over oxygen consumption rates in the nerve terminal. Oxygen consumption rates and electron transport chain complex activities were titrated with appropriate inhibitors to determine the flux control coefficients and the inhibition thresholds in rat brain synaptosomes. The flux control coefficients for complex I, complex II/III, complex III, and complex IV were found to be 0.30 +/- 0.07, 0.20 +/- 0.03, 0.20 +/- 0.05, and 0.08 +/- 0.05, respectively. Inhibition thresholds for complex I, complex II/III, complex III, and complex IV activities were determined to be approximately 10, approximately 30, approximately 35, and 50-65%, respectively, before major changes in oxygen consumption rates were observed. These results indicate that, of the electron transport chain components, complex I exerts a high level of control over synaptosomal bioenergetics, suggesting that complex I deficiencies that are present in neurodegenerative disorders, such as Parkinson disease, are sufficient to compromise oxygen consumption in the synaptosomal model of the nerve terminal.

Islet Oxygen Consumption and Insulin Secretion Tightly Coupled to Calcium Derived from L-type Calcium Channels but Not from the Endoplasmic Reticulum

The aim of the study was to test whether the source of intracellular calcium (Ca2+) is a determinant of beta cell function. We hypothesized that elevations in cytosolic Ca2+ caused by the release of Ca2+ from the endoplasmic reticulum (ER) have little physiologic impact on oxygen consumption and insulin secretion. Ca2+ release from the ER was induced in isolated rat islets by acetylcholine and response of oxygen consumption rate (OCR), NAD(P)H, cytosolic Ca2+, and insulin secretory rate (ISR) were measured. Glucose increased all four parameters, and thereafter acetylcholine further increased cytosolic Ca2+, OCR, and ISR. To assess the contribution of Ca2+ release from the ER in mediating the effects of acetylcholine, ER Ca2+ stores were first emptied by inhibiting the sarcoendoplasmic reticulum Ca2+-ATPase, which subsequently reduced the effect of acetylcholine on cytosolic Ca2+ but not its effects on OCR or ISR. As predicted, OCR and ISR were acutely sensitive to changes in L-type Ca2+ channel activity; nimodipine completely inhibited glucose-stimulated ISR and suppressed OCR by 36%, despite only inhibiting cytosolic Ca2+ by 46%. Moreover, in the presence of nimodipine and high glucose, acetylcholine still elevated cytosolic Ca2+ levels above those observed in the presence of high glucose alone but did not significantly stimulate ISR. In conclusion, Ca2+ flux through L-type Ca2+ channels was tightly coupled to changes in OCR and ISR. In contrast, the results obtained support the notion that Ca2+ release from the ER has little or no access to the intracellular machinery that regulates OCR and ISR.

Starvation resistance and metabolic response to food deprivation and recovery feeding in Fenneropenaeus chinensis juveniles

Starvation resistance in the juveniles of Chinese shrimp, Fenneropenaeus chinensis Osbeck, was studied by point-of-no-return (PNR) and point-of-reserve-saturation (PRS) experiments. Changes in oxygen consumption rate, ammonia-N excretion rate, and oxygen consumed to nitrogen excreted (O:N) ratio were investigated immediately before and after the period of recovery feeding for each regime to assess metabolic response during starvation and refeeding. There was a significant change in survival rate and wet weight of test shrimp throughout the PNR and PRS experiments (P < 0.05). The estimated PNR and PRS for F. chinensis juveniles could be directly calculated from the selected equations as PNR50 = 7.86 days, PNR100 = 21.42 days, PRS0 = 1.16 days, and PRS50 = 11.75 days. A decrease in metabolic rate during starvation as a means of conserving energy was found in F. chinensis juveniles. Furthermore, although F. chinensis juveniles only utilized protein as an energy source during starvation, they showed a shift from pure protein during starvation to an equal utilization of protein and lipid or a lipid-carbohydrate mixture during later satiate feeding. The results in the present study indicate that F. chinensis juveniles have the ability to withstand and recover from relatively prolonged starvation after longer initial feeding periods. The knowledge derived from the understanding of starvation resistance and the corresponding metabolic response of F. chinensis juveniles will be useful in the design of feeding regimes in this species.

No acute toxicity to Uca pugnax, the mud fiddler crab, following a 96‐h exposure to sediment‐bound permethrin

In coastal areas, the application of pyrethroid insecticides and the resulting sediment residues pose a potential threat to marine benthic ecosystems. Pyrethroids cause acute toxicity and exhibit a wide range of sublethal effects on fish and crustaceans when exposure is aqueous. Fiddler crabs that inhabit salt marsh sediment are sensitive to sediment-associated pollutants and serve as a sentinel species for xenobiotic exposure. We exposed adult U. pugnax to salt marsh sediment spiked with different 60% trans/40% cis permethrin concentrations for 96 h, and evaluated changes in oxygen consumption rate, hemolymph osmolarity, and glutathione S-transferase activity (GST) following exposure. Marsh sediment was not lethal to U. pugnax at permethrin concentrations of 100-10,000 microg/kg. Sediment-bound permethrin had no significant effect on respiration and osmoregulation. Exposure caused an induction of hepatopancreas GST in a dose-dependent manner. Gill and midgut tissues showed induction at permethrin concentrations at 10,000 microg/kg. We conclude that short term exposure to permethrin-contaminated sediment does not pose a significant threat to this species or impact respiration and osmoregulation. Furthermore, increased GST activity allows us to evaluate this enzyme's induction as a generalist biomarker for sediment-bound pyrethroid exposures.

Thyroid Status is a Key Modulator of Tumor Oxygenation: Implication for Radiation Therapy

In normal tissues, thyroid hormones play a major role in the metabolic activity and oxygen consumption of cells. Because the rate of oxygen consumption is a key factor in the response of tumors to radiation, we hypothesized that thyroid hormones may affect the metabolic activity of tumor cells and hence modulate the response to cytotoxic treatments. We measured the influence of thyroid status on the tumor microenvironment in experimental tumors. Hypothyroidism and hyperthyroidism were generated in mice by chronic treatment with propyl thiouracil and l-thyroxine. Thyroid status significantly modified tumor pO(2) as measured with EPR oximetry. Mechanistically, this was the result of the profound changes in oxygen consumption rates. Thyroid status was associated with a significant change in tumor radiosensitivity since the regrowth delay was increased in hypothyroid mice compared to euthyroid mice, an effect that was abolished when temporarily clamped tumors were irradiated. This study provides unique insights into the impact of modulating tumor oxygen consumption and could have implications in the management of cancer patients with thyroid disorders.

Increased oxygen consumption rates in response to high glucose detected by a novel oxygen biosensor system in non-human primate and human islets

In this study, we investigated the use of a novel oxygen biosensor system to detect changes in oxygen consumption rates (OCRs) by islets in response to glucose. Islets from non-human primate and human pancreata were seeded into an oxygen biosensor system microplate and exposed to basal (2.8 or 5.6 mM) or high (16.7 or 33.3 mM) glucose over either a long-term or a short-term culture. Our data clearly demonstrated that non-human primate islets cultured in high glucose conditions exhibited significant increases in OCRs over a 168 h extended culture period (P<0.05), which indicates an accelerated rate of beta-cell metabolism triggered by glucose over time. Significant increases in OCRs (P<0.01) were also attained in both non-human primate and human islets exposed to high glucose conditions in a 120 min short-term incubation period. OCRs exhibited by human islets exposed to different glucose concentrations correlated with insulin secretion (r(2)=0.7681, P<0.01). Moreover, the OCR stimulation index (i.e. OCR at high glucose/OCR at basal glucose) was significantly greater in human islets displaying high viabilities as opposed to islets exhibiting low viabilities (P<0.05). Together these data demonstrate that this novel oxygen biosensor system documents significant increases in islet oxygen consumption upon acute and chronic exposure to high glucose concentrations. Importantly, this methodology rapidly and robustly detects changes in OCRs by islets in response to high glucose stimulation that correlate well with the metabolic activities and functional viability of islets and clearly delineates significant differences in OCR stimulation index between high and low viability human islets, and therefore may prove to be an effective approach for quickly assessing the functional viability of islets prior to transplantation.

Growth and oxygen consumption in embryonic and early postembryonic development of European pond turtle Emys orbicularis (Reptilia: Emydidae)

Experiments on developing eggs of European pond turtle (Emys orbicularis) demonstrated S-shaped changes in the rate of oxygen consumption and body weight during embryonic development. The rate of oxygen consumption and weight progressively increased within 70 days after hatching. During embryogenesis, the rate of oxygen consumption decreased. After hatching, it increased but then decreased to a certain level, which remained constant to the end of the studied period. We observed unidirectional changes in oxygen consumption rate during embryonic period and this pattern was maintained after hatching as well. The coefficients of the allometric relationship between oxygen consumption and body weight were a = 0.33 and k = 0.52 during the embryonic period and a = 0.17 and k = 0.89 during the postembryonic period.

Metabolic recovery in Atlantic salmon fry and parr following forced activity

Atlantic salmon Salmo salar fry and parr were subjected to 5 min of forced activity and the subsequent changes in oxygen consumption and ammonia excretion rates were evaluated over a 24 h period. In a second experiment, individual Atlantic salmon fry and parr were freeze‐clamped in liquid nitrogen, before, immediately following a 5 min activity period, or after periods of recovery up to 2 h. Samples were analysed for whole body phosphocreatine (PCr), ATP and lactate. Five minutes of forced activity resulted in significant increases in both oxygen consumption and ammonia excretion rates. Changes in the oxygen consumption rates were greater in the parr compared with the fry. In contrast, the post‐exercise ammonia excretion rates were nearly twice as high for the fry compared with the parr. Exercise also caused a marked decrease in PCr levels (c. 47 and 65% in fry and parr, respectively), no change in ATP levels and a significant increase in lactate levels in Atlantic salmon fry and parr. Recovery of PCr occurred quickly (between 15 and 30 min) in fry and parr. Although the post‐activity levels of lactate were lower in fry (c. 3 μmol g−1) compared with parr (c. 14 μmol g−1), lactate levels returned to control levels within 60 min in fry, but it took &gt;2 h for this metabolite to recover in parr. Compared with parr, these findings show that Atlantic salmon fry possess a reduced anaerobic capacity, and these results are consistent with the theoretical and experimental evidence that smaller fish support burst swimming through aerobic processes.

Effects of exposure to toxic and non-toxic dinoflagellates on oxygen consumption and locomotion in stage 1 larvae of the crabs Cancer oregonensis and C. magister

Sublethal effects on larval crabs upon exposure to toxic dinoflagellates were examined in the laboratory in early 1999. Specifically, oxygen consumption rates and geotaxis responses were determined for stage 1 larvae of the crabs Canceroregonensis (Dana) and C. magister Dana that were exposed to non-toxic (Alexandriumtamarense, strain 115) or toxic (A. fundyense, strain 1719) dinoflagellates or to freshly hatched nauplii of the brine shrimp Artemia sp. In C. oregonensis, larvae exposed to the toxic dinoflagellate showed reduced rates of oxygen consumption compared to those exposed to non-toxic dinoflagellates or brine shrimp nauplii. Larvae exposed to a filtrate of the non-toxic dinoflagellate showed no change in oxygen consumption, but a reduced rate when exposed to filtrate from the toxic alga at densities >5×102 cells ml–1. In C. magister, larvae exposed to the non-toxic A. tamarense or the toxic A. fundyense had reduced oxygen consumption rates. Larvae exposed to filtrates of non-toxic and toxic dinoflagellates had no change in oxygen consumption. In geotaxis tests, C. oregonensis larvae exposed for 1 day to the toxic A. fundyense reduced their level of locomotion compared to those exposed to non-toxic A. tamarense or to brine shrimp nauplii. C. magister larvae showed no change in activity after a 1-day exposure to the toxic A. fundyense. After a 4-day exposure to A. fundyense, C. magister larvae had much reduced locomotion. Reduced locomotory activity in larvae exposed to toxic algae is consistent with the changes in oxygen consumption rates. Responding to exposure to toxic algae by reducing locomotion may affect vertical migration in these negatively buoyant crab larvae, resulting in sinking below a toxic alga bloom, at least temporarily.

17O relaxation time and NMR sensitivity of cerebral water and their field dependence.

17O spin relaxation times and sensitivity of detection were measured for natural abundance H(2)(17)O in the rat brain at 4.7 and 9.4 Tesla. The relaxation times were found to be magnetic field independent (T(2) = 3.03 +/- 0.08 ms, T(*)(2) = 1.79 +/- 0.04 ms, and T(1) = 4.47 +/- 0.14 ms at 4.7T (N = 5); T(2) = 3.03 +/- 0.09 ms, T(*)(2) = 1.80 +/- 0.06 ms, and T(1) = 4.84 +/- 0.18 ms at 9.4T (N = 5)), consistent with the concept that the dominant relaxation mechanism is the quadrupolar interaction for this nucleus. The (17)O NMR sensitivity was more than fourfold higher at 9.4T than at 4.7T, for both the rat brain and a sodium chloride solution. With this sensitivity gain, it was possible to obtain localized (17)O spectra with an excellent signal-to-noise ratio (SNR) within 15 s of data acquisition despite the relatively low gyromagnetic ratio of this nucleus. Such a 15-s 2D (17)O-MRS imaging data set obtained for natural abundance H(2)(17)O in the rat brain yielded an SNR greater than 40:1 for a approximately 16 microl voxel. This approach was employed to measure cerebral blood flow using a bolus injection of H(2)(17)O via one internal carotid artery. These results demonstrate the ability of (17)O-MRS imaging to reliably map the H(2)(17)O dynamics in the brain tissue, and its potential for determining tissue blood flow and oxygen consumption rate changes in vivo. Magn Reson Med 45:543-549, 2001.

Reduced superoxide dismutase activity in Palaemonetes argentinus (Decapoda, Palemonidae) infected by Probopyrus ringueleti (Isopoda, Bopyridae).

Cellular oxidative stress may promote damage or death in biological systems and may be caused by production of pro-oxidant molecules known as reactive oxygen species (ROS). The aim of this work was to analyze the activity of antioxidant enzymes (catalase [CAT], superoxide dismutase [SOD] and glutathione peroxidase [GPx]) in the shrimp Palaemonetes argentinus Nobili, 1901 infected by Probopyrus ringueleti (Verdi & Schuldt, 1987), a gill chamber parasite known for its capacity to cause host metabolic changes, including changes in oxygen consumption rates. Infested and non-infested shrimp were collected in the Patos Lagoon estuary (southern Brasil), where the prevalence of the parasite may be as high as 70%. No significant differences were observed for either CAT or GPx activities. However, SOD activity was significantly reduced in infected shrimp, suggesting that bopyrid isopod respiratory impairment resulted in reduced SOD enzyme activity.

Seasonal changes in oxygen consumption rate and redox property of floating peat in a pond in central Japan

Seasonal changes and temperature dependence of the oxygen consumption rate were measured in the surface peat of a floating mat. A simple model succeeded in reproducing the seasonal pattern in Eh changes at the peat surface. This included two terms: oxygen supply rate as a function of water-table depth and oxygen consumption rate as a function of temperature and microbial activity. The model explained well the minimum and maximum Eh in July and October, respectively. The activity of aerobic microbes in the peat was important in determining redox properties in the floating mat.