O2 Consumption Rate Research Articles

The effects of red LED light on pig sperm function rely upon mitochondrial electron chain activity rather than on a PKC-mediated mechanism.

While irradiation with red LED light has been reported to modulate sperm function in different mammalian species, the mechanisms underlying their response are poorly understood. This work sought to provide new insights into whether this effect relies on a direct action upon mitochondrial electron chain and/or on PKC-linked mechanisms such as those related to opsins. For this purpose, pig semen was light-stimulated for 1, 5 or 10 min in the presence/absence of antimycin A, an inhibitor of the mitochondrial electron chain, or PKC 20–28® (PKCi), a PKC inhibitor. Antimycin A completely blocked the effects of light at all the performed irradiation patterns. This effect was linked to a complete immobility of sperm, which was accompanied with a significant (p < 0.05) drop in several markers of mitochondrial activity, such as JC-1 staining and O2 consumption rate. Antimycin A, however, did not affect intracellular ATP levels, intramitochondrial calcium, total ROS, superoxides or cytochrome C oxidase (CCO) activity. In the case of PKCi, it did also counteract the effects of light on motility, O2 consumption rate and CCO activity, but not to the same extent than that observed for antimycin A. Finally, the effects observed when sperm were co-incubated with antimycin A and PKCi were similar to those observed with antimycin A alone. In conclusion, red LED light acts on sperm function via a direct effect on mitochondrial electron chain. Additionally, light-activated PKC pathways have a supplementary effect to that observed in the electron chain, thereby modulating sperm parameters such as motility and CCO activity.

Sex-specific effects of chronic hypoxia on routine cardiovascular function and metabolism in CD-1 mice.

Hypoxia can have significant impacts on cardiovascular physiology, but the effects of chronic exposure to moderate hypoxia and how they differ between sexes remain poorly understood. We used physiological telemetry to examine this issue in CD-1 mice. Adult mice were chronically exposed to normoxia or hypobaric hypoxia (12 kPa O2) for 6 wk and then subjected to telemetry measurements of routine physiology across the diel cycle. Heart rate (fH), mean arterial blood pressure (Pmean), body temperature (Tb), and activity were greater during the nighttime active phase than the daytime inactive phase. Chronic hypoxia had no effect on these traits at night but had sex-specific effects during the day, when chronic hypoxia reduced fH, Tb, and activity in males but not females. These differences existed without any effect of chronic hypoxia on α-adrenergic or nitric oxide tone on the vasculature (assessed as Pmean response to pharmacological blockade). Responses to acute hypoxia were then measured during stepwise reductions in inspired O2 from 21 to 8 kPa O2. O2 consumption rate, fH, Pmean, and Tb declined in severe hypoxia, but the O2 tension at which this began was lower in mice held in chronic hypoxia. However, the hypoxic ventilatory response was augmented by exposure to chronic hypoxia in females but not in males. Females also exhibited larger increases in lung mass and less right ventricle hypertrophy than males in chronic hypoxia. Our results support the growing evidence that there can be considerable sex differences in the cardiorespiratory responses to hypoxia.

Abstract 099: Role Of Nox-induced Oxidative Stress In Mitochondrial Bioenergetic Dysfunction In The Kidney Cortex And Outer Medulla Of Dahl Salt-sensitive Rats During The Development Of Hypertension

Background: Although renal mitochondrial dysfunction has been observed in various models of hypertension, the events leading to this dysfunction remain unclear. The present study focused on the role of salt (NaCl) intake upon renal mitochondrial bioenergetic function in Dahl salt-sensitive (SS) rats hypothesizing that a high salt (HS) diet would impair mitochondrial bioenergetics via stimulation of NOX-induced oxidative stress. Method: The HS induced progressive alteration of oxidative phosphorylation (OxPhos) and ATP synthesis in mitochondria obtained from renal cortex and outer medulla (OM) of male SS rats were compared to those of SS rats with dual knockout of NOX4 and p67 phox subunit of NOX2 (NOX DKO). Mitochondria were isolated from the cortex and OM of groups of age matched rats fed a 0.4% NaCl (LS) diet since weaning and at days 3, 7, 14 &amp; 21 of a 4.0% NaCl (HS) diet feeding. ADP-induced (state 3) O 2 consumption rates (OCR) were measured in mitochondria energized with pyruvate and malate as substrates using an Oroboros Oxygraph-2k Instrument. Results: In SS rats, a biphasic pattern in state 3 OCR was observed with progressive uncoupling of OxPhos occurring in both cortex and OM. In the early phase of hypertension (days 3 &amp; 7), state 3 OCR was increased (155 ± 8.2 to 190 ± 12.2 to 203.4 ± 12.1 nmol/min/mg in cortex and 101.4 ± 5.1 to 121.8 ± 7.3 to 112.8 ± 6.8 nmol/min/mg in OM; n = 5). In the established phase of hypertension (days 14 &amp; 21), this increase was not sustained as state 3 OCR was reduced to 153 ± 7.65 and 132 ± 6.6 nmol/min/mg in cortex and 105 ± 7.3 and 63.3 ± 6.5 nmol/min/mg in OM. In contrast, SS NOX DKO rats fed a HS diet showed no significant differences in state 3 OCR in both cortex and OM, as was also observed in a control group of age matched NOX DKO rats maintained on a LS diet throughout the study. Conclusions: In SS rats, a HS diet causes initial increase in the efficiency of OxPhos for ATP production followed by a progressive decrease in the efficiency of OxPhos for ATP production in the established phase of hypertension. This biphasic pattern with ultimate reduction in the efficiency of OxPhos for energy production was not observed in SS NOX DKO rats demonstrating the important causal role of NOX-induced oxidative stress in the renal mitochondrial bioenergetic dysfunction.

Prevalence and mechanisms of environmental hyperoxia-induced thermal tolerance in fishes.

Recent evidence has suggested environmental hyperoxia (O2 supersaturation) can boost cardiorespiratory performance in aquatic ectotherms, thereby increasing resilience to extreme heat waves associated with climate change. Here, using rainbow trout (Oncorhynchus mykiss) as a model species, we analysed whether improved cardiorespiratory performance can explain the increased thermal tolerance of fish in hyperoxia (200% air saturation). Moreover, we collated available literature data to assess the prevalence and magnitude of hyperoxia-induced thermal tolerance across fish species. During acute warming, O2 consumption rate was substantially elevated under hyperoxia relative to normoxia beyond 23°C. This was partly driven by higher cardiac output resulting from improved cardiac contractility. Notably, hyperoxia mitigated the rise in plasma lactate at temperatures approaching upper limits and elevated the critical thermal maximum (+0.87°C). Together, these findings show, at least in rainbow trout, that hyperoxia-induced thermal tolerance results from expanded tissue O2 supply capacity driven by enhanced cardiac performance. We show 50% of the fishes so far examined have increased critical thermal limits in hyperoxia (range: 0.4–1.8°C). This finding indicates environmental hyperoxia could improve the ability of a large number of fishes to cope with extreme acute warming, thereby increasing resilience to extreme heat wave events resulting from climate change.

Abstract P3041: DWORF Overexpression In The Heart Results In Enhanced Mitochondrial Function

Background: Calcium (Ca 2+ ) plays a pivotal role in regulating physiological processes in the heart including excitation-contraction coupling, Ca 2+ signaling, and mitochondrial function. It is well established that both Ca 2+ dysregulation and altered cardiac energy metabolism are universal features of heart failure (HF). Dwarf open reading frame (DWORF), a newly discovered microprotein, increases SERCA2a activity in part by relieving SERCA2a inhibition by phospholamban (PLN). DWORF overexpression via cardiac specific transgene or gene therapy is cardioprotective in genetic and experimental HF mouse models. In addition to activation of SERCA2a by PLN displacement, preliminary data indicates that enhanced mitochondrial function contributes to the cardioprotective effects of DWORF. Here we tested the effects of DWORF overexpression on cardiac mitochondrial function and Ca 2+ dynamics. Methods and Results: We isolated mitochondria from wildtype (WT) and DWORF transgenic (Tg) mouse hearts and measured O 2 consumption rates using a Seahorse mito stress test. We found that DWORF Tg mice have increased respiratory capacity compared to WT (DWORF Tg 1150.0 pmol/min, WT 799.4 pmol/min +/-146.2) and enhanced spare respiratory capacity (DWORF Tg 489.1 pmol/min, WT 229.4 pmol/min +/- 102.1). To measure mitochondrial Ca 2+ dynamics, we performed Ca 2+ retention capacity assays and found that DWORF Tg mice have increased Ca 2+ uptake rates compared to WT (DWORF Tg 32.32 s, WT 46.46 s +/- 4.39). Western blot analysis of the mitochondrial Ca 2+ uniporter (MCU) in mito extracts from WT and DWORF Tg hearts indicated elevated levels of MCU. However, electron transport chain (ETC) protein expression indicated no significant differences, indicating the enhanced mitochondrial activity observed in DWORF Tg mice was not due to increased ETC levels. Conclusions: DWORF Tg mouse heart mitochondria exhibit increased maximal respiration rates and enhanced spare respiratory capacity. However, ETC protein levels are unchanged, indicating altered enzyme activity rather than expression. Additionally, mitochondria from DWORF Tg hearts uptake Ca 2+ faster into the mitochondria through the increased levels of MCU, suggesting enhanced mitochondrial Ca 2+ dynamics.

Effect of sub-atmospheric pressure on the characteristics of concurrent/upward flame spread over a thin solid

The variation of ambient pressure is a potential tool for studying the driving parameters of fire dynamics and heat release in low-pressure environments such as high-altitude locations, aircraft, and spacecraft. The study of upward flame spread over a solid fuel has direct implications on material flammability and fire development, and low pressure environments have recently gained more attention for the possible comparison with the reduced gravity conditions encountered during space missions. In this work, we consider upward spreading flames over thin acrylic sheets in ambient pressures between 30 and 100 kPa. A forced flow velocity of 20 cm/s is added to the naturally-driven buoyant flow, creating a mixed flow field (natural and forced) that varies with pressure. Flame characteristics such as spread rate and standoff distance are measured from the video analysis of the experiments. It is observed that the former decreases with pressure while the latter increases. The larger flame stand-off distance at low pressures partially explains the decrease of the flame spread rate since the convective heat flux from the flame to the solid decreases. Additionally, volumetric concentrations of the combustion products are measured during the experiments. The results show lower O2 consumption and CO2 production rates at lower pressures. Based on these rates, we could calculate the heat release rate from upward spreading flames at low pressure, providing fundamental information for better understanding pressure-gravity correlations. According to the results, the volumetric heat release rate is proportional to pressure, which is consistent with previous studies on pressure modeling of fires. This suggests that chemical kinetics is not a constraint for the conditions tested in this study, which could help make future flammability tests comparable to low gravity ones.

Combustion and NOx formation characteristics from a 330 MWe retrofitted anthracite-fired utility boiler with swirl burner under deeply-staged-combustion

In order to achieve peak carbon dioxide emissions and carbon neutrality goals, a higher requirement is put forward for coal type adaptability medium, load stable combustion performance, and pollutant emission for wall-fired boiler with swirl burners. In this work, through industry experiments, the combustion characteristics of a retrofitted wall-firing mode 330 MWe power unit feed with anthracite were investigated. The combustion system was retrofitted employing the centrally fuel-rich (CFR) swirl burner and deeply-staged combustion technology. The experiment data were measured, including flue gas temperature and composition, furnace temperature, and the main operational parameter under high-load (300 MWe), medium-load (230 MWe), and low-load (150 MWe) conditions. The stable combustion of anthracite is achieved at various loads. With the excessive air coefficients of the main combustion region increase, the corresponding second air damper opening increase under high-load condition or over-fire-air damper opening decrease, both the flue gas temperature and heating rate of pulverized coal increase advance the ignition and promote the stable combustion and burn-out of anthracite coal. In the initial combustion stage, the O2 given into the chamber increase, both O2 consumption rate and CO concentration decrease, but NOx concentration increases. While in the later stage, both the O2 and CO concentration tend to become flat. Meanwhile, the O2 concentration is lower than 2.1%, the CO concentration maintains a relatively high level, and NOx production is inhibited effectively and becomes flat gradually. With the excess air coefficient increase, the temperature in the main combustion region increases, and the difference in furnace temperature between the burn-out region and the main combustion region diminishes. The unburned carbon content in fly ash and exhaust gas temperature decreases, while NOx emission increases. There is little difference for the flue gas temperature distribution in the side-wall region, and the measured O2 concentration is always higher than 3.5% with the different mass flux of secondary air and over-air-fire. With decreasing load, the NOx emission decreases from the highest 1392.66 to the lowest 958.38 mg/m3 (O2 6%), and the boiler efficiency improves. Compared with high-load and medium-load, the slagging tendency of the water wall is lower under low-load condition.

Use of a human patient simulator for apnea studies: a preliminary in vitro trial.

BackgroundModern human patient simulators (HPSs) could be used for researching critical scenarios such as apnea oxygenation. We aimed to study the use of a high-fidelity HPS to assess prolonged apnea using various oxygenation strategies with a simple high-flow nasal cannula (15 L/min).MethodsAn experimental simulation study using an HPS (CAE Healthcare™) was conducted after obtaining approval from the Institutional Review Board. The HPS responded according to real-time physiologically modeled responses to external gases, such as oxygen (O2). Apnea experiments were performed with different physiological settings, such as shunt fraction (5%) and O2 consumption (250, 500, and 750 ml/min). The following four apnea experiments were conducted: no oxygenation (NO), apnea oxygenation alone (AO), preoxygenation alone (PO), and para-oxygenation (PAO). The time to 92%, 75%, and 50% saturation was recorded. Alveolar and arterial gas levels were recorded till 50% saturation.ResultsAt 250 ml/min, PO (1121 s) and PAO (1274.5 s) had a significantly longer time to 50% saturation (400% increase) compared to NO (222.5 s) and AO (239 s). A similar trend was observed for the time to 92% and 75% saturation. At higher O2 consumption rates, a shorter time to desaturation was observed.ConclusionsApnea trends in the HPS correlated with similar prior human experiments. AO without preoxygenation was found to provide no additional benefit. Preoxygenation with high-flow O2 via nasal cannula prolonged the time to desaturation in the PAO more than PO scenario. Therefore, HPSs can be used in future studies where patient safety is a concern.

What does respirometric analysis tell us about Metarhizium robertsii

Metarhizium comprises species of importance as biological control agents whose conidia have been well produced in lab-scale packed column bioreactors. However, difficulties such as low oxygen supply, CO2 removal, and overheating limit bioreactor scale-up. To select suitable operating conditions in industrial-scale bioreactors, it is necessary to analyze the bioprocess in terms of O2 consumption and CO2 production. Metarhizium robertsii cultures reached maxima of O2 consumption and CO2 production rates of 0.23 and 0.35 mg/gidm h, respectively. The total O2 consumption (23.81 mg/gidm) and CO2 production (34.52 mg/gidm) were determined by experimental measurements and estimates by mathematical models with a coefficient of determination higher than 0.99. The respiratory quotient presented values from 1 to 1.2 (from 48 to 175 h). The respirometric analysis showed that the culture of is an aerobic process with low CO2 and low metabolic heat production, favoring the scale-up of the bioprocess

Ferroptosis: A Potential Therapeutic Target in Acute Kidney Injury.

Ferroptosis is a recently recognized form of nonapoptotic cell death that is triggered by reactive oxidative species (ROS) due to iron overload, lipid peroxidation accumulation, or the inhibition of phospholipid hydroperoxidase glutathione peroxidase 4 (GPX4). Recent studies have reported that ferroptosis plays a vital role in the pathophysiological process of multiple systems such as the nervous, renal, and pulmonary systems. In particular, the kidney has higher rates of O2 consumption in its mitochondria than other organs; therefore, it is susceptible to imbalances between ROS and antioxidants. In ischemia/reperfusion (I/R) injury, which is damage caused by the restoring blood flow to ischemic tissues, the release of ROS and reactive nitrogen species is accelerated and contributes to subsequent inflammation and cell death, such as ferroptosis, as well as apoptosis and necrosis being induced. At the same time, I/R injury is one of the major causes of acute kidney injury (AKI), causing significant morbidity and mortality. This review highlights the current knowledge on the involvement of ferroptosis in AKI via oxidative stress.

Bioactives From Coffee By-Products Stimulate Liver Metabolism and Mitochondrial Bioenergetics via AMPK/PGC-1α/Nrf2 and IRS/AKT/GLUT2 Pathways In Vitro

ObjectivesThis study aimed to investigate the effect of the main bioactive compounds from the coffee silverskin and coffee husk on hepatic lipid and glucose metabolism and mitochondrial bioenergetics using an in vitro model mimicking non-alcoholic fatty liver disease. MethodsHepG2 cells were treated with 50 μmol L–1 caffeine, chlorogenic, caffeic, protocatechuic, or gallic acids, or kaempferol, or the aqueous extracts from the coffee silverskin (CSE, 100 μg mL–1) or the coffee husk (CHE, 100 μg mL–1) in the presence of palmitic acid (500 μmol L–1). Cell metabolism biomarkers were assessed 24 h after the co-treatment in cell lysates and supernatants using chemical and immunochemical techniques. Differences among treatments were considered significant at p < 0.05. ResultsBioactive compounds in coffee by-products, CSE, and CHE decreased lipid accumulation (53–115%) by reducing fatty acid synthase expression (44–76%) and stimulated β-oxidation by triggering carnitine palmitoyltransferase I activity (1.6 to 2.6-fold) and up-phosphorylating AMP-activated protein kinase (AMPK, 1.5–2.4-fold). Glucose uptake was promoted via the increase in the insulin receptor substrate (IRS)-1 (1.5 to 2.1-fold) and protein kinase B (Akt1, 1.7–2.4-fold) phosphorylation, and glucose transporter (GLUT)-2 expression (2.1 to 3.7-fold). Mitochondrial function was enhanced (ATP production and O2 consumption rate, 1.7–2.5-fold and 1.8 to 2.5-fold, respectively), and oxidative phosphorylation complexes and peroxisome proliferator-activated receptor-γ coactivator (PGC)-1α expression restored (30–105%). Oxidative stress was diminished (56 – 85%) through augmented superoxide dismutase (1.3 to 1.7-fold) and catalase (1.5 to 2.0-fold) activities, and nuclear factor-erythroid 2-related factor 2 (Nrf2) expression (2.3 to 3.8-fold). ConclusionsThe major bioactive compounds in coffee by-products, primarily chlorogenic and protocatechuic acids, could regulate hepatic lipid and glucose metabolism and prevent oxidative stress and mitochondrial dysfunction by activating AMPK/PGC-1α/Nrf2 and IRS/AKT/GLUT2 signaling pathways. Altogether, our results suggest the use of coffee by-products as a sustainable source of natural bioactives to counteract non-alcoholic fatty liver disease. Funding SourcesUSDA-NIFA-HATCH and the Spanish Ministry of Science and Innovation.

The EP3 and EP4 Receptor Subtypes both Mediate the Fever-producing Effects of Prostaglandin E2 in the Rostral Ventromedial Preoptic Area of the Hypothalamus in Rats

This study aimed to re-examine the receptor subtype that mediates the fever-producing effects of prostaglandin E2 (PGE2) in the rostral ventromedial preoptic area (rvmPOA) of the hypothalamus. Among the four subtypes of PGE2 receptors (EP1, EP2, EP3, and EP4), EP3 receptor is crucially involved in the febrile effects of PGE2. However, it is possible for other subtypes of PGE2 receptor to contribute in the central mechanism of fever generation. Accordingly, effects of microinjection of PGE2 receptor subtype-specific agonists or antagonists were examined at the locus where a microinjection of a small amount (420 fmol) of PGE2 elicited prompt increases in the O2 consumption rate (VO2), heart rate, and colonic temperature (Tc) in the rvmPOA of urethane–chloralose-anesthetized rats. The EP3 agonist sulprostone mimicked, whereas its antagonist L-798,106 reduced, the febrile effects of PGE2 microinjected into the same site. Similarly, the EP4 agonist rivenprost mimicked, whereas its antagonist ONO-AE3-208 reduced, the effects of PGE2 microinjected into the same site. In contrast, microinjection of the EP1 agonist iloprost induced a very small increase in VO2 but did not have significant influences on the heart rate and Tc, whereas its antagonist, AH6809, did not affect the PGE2-induced responses. Microinjection of the EP2 agonist butaprost had no effects on the VO2, heart rate, and Tc. The results suggest that the EP3 and EP4 receptor subtypes are both involved in the fever generated by PGE2 in the rvmPOA.

Maximum critical temperature and oxygen consumption during thermoregulation in Macrobrachium americanum (Bate, 1868) adult prawns

Macrobrachium americanum is a large prawn living in rivers and coastal water bodies of tropical and subtropical ecosystems of the Pacific side of the Americas. Water temperature and salinity are the main physicochemical variables determining its distribution. Temperature also influences its metabolic rate, which determines growth rate and reproduction. Because of this, preferred temperature, critical thermal maximum (CTMax), and O2 consumption rates at four temperatures (20, 24, 28, and 32°C) were tested from wild prawns in laboratory assays. Those temperatures correspond to the annual temperature fluctuations at the sampling site. The final preferred temperature of this species is close to 26.4°C. The acclimatization temperature has an almost linear effect on the CTMax (R2 = 0.987, P &lt; 0.05), and the oxygen consumption routine rates increased as the acclimation temperature increased from 20 to 32°C.

Intermittent Hypoxia (IH) Impairs Hippocampal Oxygen Consumption and Neurophysiological Responses to Metabolic Challenge

IH is a consequence of several clinical conditions such as sleep apnea. Previous work has shown that IH impairs many neurophysiological functions such as hippocampal adult neurogenesis and NMDAr‐dependent synaptic plasticity. While IH produces hippocampal oxidative stress, IH‐dependent changes in hippocampal metabolism may also contribute to impaired neurophysiology. The objective of this study is to understand how IH impacts hippocampal physiology in response to metabolic challenge. We hypothesize that ten days of IH perturbs hippocampal synaptic transmission when metabolically challenged. In vitro recordings of field excitatory post‐synaptic potentials (fEPSPs) from hippocampal slices showed that glucose deprivation (i.e., 30mM Fructose aCSF with 95% O2) caused a complete loss of the fEPSP within 20min; whereas, the fEPSP in control tissue was reduced to 40% of baseline within the same period of time. This difference coincided with a reduction in both maximal and ATP‐linked O2 consumption rates (OCRs) in hippocampal tissue following IH. Despite the reductions in hippocampal OCRs, neither ATP/ADP ratio nor lactate concentrations were different between the two groups. These data indicate that while ATP/ADP levels following IH are sufficiently maintained under basal conditions, metabolic processes fail to support synaptic physiology during glucose deprivation. Our ongoing experiments will further delineate how IH impacts the contribution of glycolytic and oxidative metabolism in the hippocampus. These findings may better define how IH impacts hippocampal responses to metabolic stressors, which could contribute to impairing hippocampal physiology in conditions such as sleep apnea.

TNFα Reduces the Maximum Respiratory Capacity of Mitochondria in Human Airway Smooth Muscle Cells

Previously, we reported that airway inflammation mediated by pro‐inflammatory cytokines such as TNFα induces hypercontractility and increased ATP consumption in human airway smooth muscle (hASM). We also found that TNFα induces an increase in maximum O2 consumption rate as well as mitochondrial biogenesis and an increase in mitochondrial volume density. However, when normalized to mitochondrial volume, maximum O2 consumption rate per mitochondrion is reduced in hASM after TNFα exposure. Unfortunately, standard respirometry (e.g., Seahorse) measures only the averaged maximum O2 consumption rate of a large number of cells. In the present study, we used a quantitative histochemical technique to determine the maximum velocity of the succinate dehydrogenase reaction (SDHmax) together with mitochondrial volume density in individual hASM cells. SDH is a key enzyme in the tricarboxylic acid (TCA) cycle as well as complex II in the electron transport chain (ETC), and SDHmax reflects the maximum respiratory capacity of individual mitochondrion. We hypothesized that TNFα decreases SDHmax per mitochondrion in hASM cells. hASM cells were dissociated from bronchial biopsies obtained during thoracic surgeries in patients with no history of asthma or chronic obstructive pulmonary disease. The hASM cells were treated with TNFα (20 ng/ml) or vehicle (DMSO) for 24 h, and SDHmax was measured using a solution containing 80 mM succinate (maximum substrate concentration) and 1.5 mM nitro blue tetrazolium (NBT) as the reaction indicator. As the SDH reaction proceeded, the hASM cells were imaged in 3D (Z optical slice of 0.5 μm) using an oil‐immersion ×60/1.4 NA objective on a Nikon Eclipse A1 laser scanning confocal system. The change in optical density (OD), reflecting the accumulation of NBT diformazan reaction product in the hASM cell, was measured every 15 s over a 10 min period. Based on the Beer‐Lambert equation, the slope of the change in OD was used to determine SDHmax expressed as mM fumarate/L tissue/min. To determine mitochondrial volume density, the same hASM cells were also loaded with MitoTracker Green (500 nM) and imaged in 3D. We found that mitochondria were more fragmented after TNFα exposure although mitochondrial volume density increased. When SDHmax was normalized to mitochondrial volume, we found that SDHmax per mitochondrion decreased in hASM cells, consistent with the reduction in maximum O2 consumption per mitochondrion that we previously observed. Thus, the increased energetic demand induced by inflammation (TNFα) are met by mitochondrial fragmentation leading to biogenesis and an increase in mitochondrial volume density rather than an increase in mitochondrial respiratory capacity.

Single Cell Assessment of Mitochondrial Function

Cellular mitochondrial function can be assessed using high resolution respirometry that measures O2 consumption rate during various conditions that systematically alter the tricarboxylic acid (TCA) cycle or the electron transport chain (ETC). However, current high resolution respirometry does not measure O2 consumption rate at the single cell level, but actually measures average mitochondrial function across a number of cells (either isolated or in tissues). Thus, respirometry assumes physiological homogeneity across cells. However, in many tissues, mitochondrial function varies across cells and this heterogeneity is physiologically important. Therefore, a direct measurement of cellular mitochondrial function will provide valuable novel information and physiological insight. In the present study, we used a quantitative histochemical technique to measure the activity of succinate dehydrogenase (SDH), a key enzyme located in the inner mitochondrial membrane, and the only enzyme to participate in both the TCA cycle and the ETC as Complex II. SDH mediates the oxidation of succinate to fumarate in the TCA cycle, which is coupled to the reduction of ubiquinone to ubiquinol in the ETC. In this study we determined the maximum velocity of the SDH reaction (SDHmax) in isolated human airway smooth muscle (hASM) cells using 1‐methoxyphenazine methosulphate (mPMS), as an exogenous electron carrier, and azide to inhibit cytochrome oxidase. To measure SDHmax, the cells were exposed to a solution containing 80 mM succinate and 1.5 mM nitroblue tetrazolium (NBT) as the reaction indicator. hASM cells were imaged in 3D (Z optical slice of 0.5 μm) using a Nikon Eclipse A1 laser scanning confocal system with a ×60/1.4 NA oil‐immersion lens. In the quantitative histochemical procedure, changes in cell optical density (OD) due to the progressive reduction of NBT to its diformazan (peak absorbance wavelength of 570 nm) were measured every 15 s over a 10 min period. Linearity of the SDH reaction was confirmed across the 10 min period, and SDHmax was expressed as mM fumarate/liter of tissue/min. Validation of this technique included specific ETC inhibitors including oligomycin (ATP synthase inhibitor), FCCP (proton ionophore), antimycin A (Complex III inhibitor) and rotenone (Complex I inhibitor), similar to those used in high resolution respirometry. We observed that FCCP‐mediated disruption of the mitochondrial proton gradient does not affect SDHmax, while SDHmax is decreased by rotenone and antimycin A. In addition, we used MitoTracker Green to label and image mitochondria in hASM cells and determined mitochondrial volume density. The SDHmax was then normalized to mitochondrial content. Our results confirm that this quantitative technique is rigorous and reproducible, and that measurements of cellular SDHmax can serve as a reliable surrogate for the measurement of maximum mitochondrial respiration in single cells.

Application of a Second‐Order Control‐Systems Model to Investigate Effects of Impaired Capillary Signalling in Skeletal Muscle on Regulation of Capillary Blood Flow Velocity and Tissue Oxygenation

Microcirculation is the main region of the vasculature where oxygen (O2) is transferred directly from blood to metabolically active tissue. Tissue O2 partial pressure (PO2) in the microcirculation has been historically infeasible to measure, but critical as an indicator of capillary and surrounding tissue health. Thus, theoretical modelling of capillary‐tissue O2 transport is crucial as a complement to in vivo experiments in exploring structure‐function relationships in microvascular networks (Ellis et al, Microcirc 2012, 19:5). To properly complement experiments which stimulate O2 flow‐dependent regulation systems, modelling of microcirculatory O2 transport requires incorporation of knowledge on regulatory mechanisms.Capillary networks modulate their blood velocity to recruit sufficient O2 delivery in many local O2 conditions; this has led to the conclusion that endothelium signals upstream to arterioles in an O2‐dependent manner (Ghonaim et al, Microcirc 2021, e12699). Recently, a second‐order control systems model was developed to investigate the interaction between capillary‐tissue O2 transport and O2‐dependent blood flow regulation under an externally applied O2 stimulus. This utilized elements from previous arteriolar diameter alteration models in response to skeletal muscle O2 gradients, as well as elements from a recently developed continuous‐capillary O2 transport model (Afas et al, Math Biosci 2021, 333:108535).The control systems model aimed to predict modulations in blood velocity through endothelial signalling which homogenized outlet capillary PO2 in skeletal muscle. The model demonstrated that variations in O2 transport parameters such as capillary network density and tissue O2 consumption rate had varying effects on the averaged blood velocity in a small skeletal muscle segment perfused by a capillary network. In addition, the externally supplied PO2 had an effect on both blood velocity and the tissue‐capillary O2 balance. While the tissue‐capillary PO2 balance and velocity adaptation were investigated, the endothelial signalling parameters were not reported (Afas &amp; Goldman, Vasc. Bio. 2021, conference).The present study investigates implications of capillary network metric variations on the endothelial signal communicated upstream to modulate capillary blood flow in response to various muscle surface PO2 levels. Constraints imposed by pathogenic impairment of endothelial signalling will be investigated and interpreted in the context of capillary flow regulation.

Kingfish (Seriola lalandi) adjust to low salinity with only subtle effects to cardiorespiratory and growth performance

At reduced salinities nearing isosmotic conditions, marine fishes experience reduced osmotic gradients between body fluids and the surrounding water, which should lower energetic costs of osmoregulation. Such energy savings on osmoregulation can potentially be directed to other processes such as growth, suggesting the possibility that rearing euryhaline fishes under reduced salinity in aquaculture may be a beneficial practice. The yellowtail kingfish (Seriola lalandi) is one such aquaculture species that appears to benefit from low salinity, as juveniles display faster growth when reared at salinities approaching isosmotic conditions. Here, we tested the hypothesis that, due to reduced osmoregulatory costs, growth is improved in sub-adult kingfish reared at low salinity, and this is associated with a down-regulation of resting cardiorespiratory activity reflecting energetic savings. Furthermore, to gain a more complete picture of how low salinity impacts cardiorespiratory function in kingfish, we also assessed maximal cardiorespiratory responses to enforced exercise. Two experiments were carried out. The first assessed resting and maximum cardiac and respiratory performance of kingfish acclimated to seawater (35 ppt), and after ~20 h (acute) or > 4 weeks (chronic) exposure to brackish water (17 ppt). Acute 17 ppt exposure reduced standard metabolic rate (i.e. resting O2 consumption rate; 179.4 mg O2∙h−1∙kg−1) relative to fish maintained in seawater (198.3 mg O2∙h−1∙kg−1), but this difference was no longer apparent following chronic acclimation to 17 ppt (191.3 mg O2∙h−1∙kg−1). While maximum metabolic rate was not affected by salinity, kingfish both acutely transferred and acclimated to 17 ppt had a reduced excess post-exercise O2 consumption compared to fish in 35 ppt. Acclimation to 17 ppt was also associated with a reduced proportion of ventricle compact myocardium. In a second experiment, we analyzed growth rate and feed conversion efficiency of kingfish acclimated to seawater (35 ppt) or brackish water (24 and 12 ppt) for 8 weeks. Contrary to our initial hypothesis, growth rate and feed conversion efficiency were slightly deteriorated (by ~5 and ~ 6% for growth rate and feed conversion efficiency, respectively) after 4 weeks acclimation to 12 ppt; however, these differences were no longer apparent after 8 weeks acclimation. This study shows neither substantial benefits nor costs of brackish water exposure to cardiorespiratory and growth performance in sub-adult kingfish, suggesting this species can be reared at salinities as low as 12 ppt.

Study of ambient temperature oxidation in low metamorphic coal and the oxidation mechanism

In view of the problem where there is abnormal continuous CO exceedance in the tailgate corner without any sign of spontaneous combustion associated with the mining of low metamorphic coal seam, both desorption experiments and ambient temperature oxidation experiments were carried out, and the ambient temperature oxidation characteristics and its mechanism were studied. The results demonstrate that the original coal seam from which the experimental coal sample was taken, does not contain CO and CO2, and the coal sample can oxidize at ambient temperature. The O2 consumption rate, CO and CO2 production rate is not directly related to the particle size. Low O2 concentration can inhibit but not eliminate the ambient temperature oxidation. In addition to the active sites of coal itself, the ambient temperature oxidation reaction rate is also affected by the inhibitory substances produced by the reaction itself. R1(△CO/△O2), R2(△CO2/△O2) and R3(△CO/△CO2) indexes are mostly not affected by the changes of particle size, O2 concentration and oxidation rates, and the ambient temperature oxidation of coal is dominated by a single mechanism. The ambient temperature oxidation mechanism is also theoretically deduced. The research will have important significance for controlling the abnormal exceedance of CO.

Breathing versus feeding in the Pacific hagfish.

Hagfish represent the oldest extant connection to the ancestral vertebrates, but their physiology is not well understood. Using behavioural (video), physiological (respirometry, flow measurements), classical morphological (dissection, silicone injection) and modern imaging approaches (micro-MRI, DICE micro-CT), we examined the interface between feeding and the unique breathing mechanism (nostril opening, high-frequency velum contraction, low-frequency gill pouch contraction and pharyngo-cutaneous duct contraction) in the Pacific hagfish, Eptatretus stoutii. A video tour via micro-MRI is presented through the breathing and feeding passages. We have reconciled an earlier disagreement as to the position of the velum chamber, which powers inhalation through the nostril, placing it downstream of the merging point of the food and water passage, such that the oronasal septum terminates at the anterior end of the velum chamber. When feeding occurs by engulfment of large chunks by the dental plates, food movement through the chamber may transiently interfere with breathing. Swallowing is accelerated by peristaltic body undulation involving the ventral musculature, and is complete within 5 s. After a large meal (anchovy, 20% body mass), hagfish remain motionless, defaecating bones and scales at 1.7 days and an intestinal peritrophic membrane at 5 days. O2 consumption rate approximately doubles within 1 h of feeding, remaining elevated for 12-24 h. This is achieved by combinations of elevated O2 utilization and ventilatory flow, the latter caused by varying increases in velar contraction frequency and stroke volume. Additional imaging casts light on the reasons for the trend for greater O2 utilization by more posterior pouches and the pharyngo-cutaneous duct in fasted hagfish.