Visualization and Analysis of Blood Flow and Oxygen Consumption in Hepatic Microcirculation: Application to an Acute Hepatitis Model

There is a considerable discrepancy between oxygen supply and demand in the liver because hepatic oxygen consumption is relatively high but about 70% of the hepatic blood supply is poorly oxygenated portal vein blood derived from the gastrointestinal tract and spleen. Oxygen is delivered to hepatocytes by blood flowing from a terminal branch of the portal vein to a central venule via sinusoids, and this makes an oxygen gradient in hepatic lobules. The oxygen gradient is an important physical parameter that involves the expression of enzymes upstream and downstream in hepatic microcirculation, but the lack of techniques for measuring oxygen consumption in the hepatic microcirculation has delayed the elucidation of mechanisms relating to oxygen metabolism in liver. We therefore used FITC-labeled erythrocytes to visualize the hepatic microcirculation and used laser-assisted phosphorimetry to measure the partial pressure of oxygen in the microvessels there. Noncontact and continuous optical measurement can quantify blood flow velocities, vessel diameters, and oxygen gradients related to oxygen consumption in the liver. In an acute hepatitis model we made by administering acetaminophen to mice we observed increased oxygen pressure in both portal and central venules but a decreased oxygen gradient in the sinusoids, indicating that hepatocyte necrosis in the pericentral zone could shift the oxygen pressure up and affect enzyme expression in the periportal zone. In conclusion, our optical methods for measuring hepatic hemodynamics and oxygen consumption can reveal mechanisms related to hepatic disease.     

A multi-layer model of retinal oxygen supply and consumption helps explain the muted rise in inner retinal PO(2) during systemic hyperoxia

A multi-layer mathematical model of oxygen supply and consumption in the rat retina is described. The model takes advantage of the highly layered structure of the retina and the compartmentalisation of the available oxygen sources. The retina is divided into eight layers, each with a distinct oxygen consumption or supply rate. When applied to the available data from intraretinal oxygen measurements in the rat under normal physiological conditions, a close fit between the model and the data was achieved (r(2)=0.98+0.005, n=6). The model was then used to investigate recent evidence of oxygen regulating mechanisms in the rat retina during systemic hyperoxia. Fitting our model to the experimental data (r(2)=0.988+0.004, n=25) allowed the relative oxygen delivery or consumption of the key retinal layers to be determined. Two factors combine to produce the relative stability of inner retinal oxygen levels in hyperoxia. The retinal layer containing the outer plexiform layer/deep retinal capillaries, switches from a net source to a net consumer of oxygen, and the oxygen consumption of the outer region of the inner plexiform layer increases significantly. The model provides a useful tool for examining oxygen consumption and supply in all retinal layers, including for the first time, those layers within the normally perfused inner retina.     

Alteration in myocardial oxygen balance during exercise after beta-adrenergic blockade in dogs

The effects of beta-adrenergic blockade upon myocardial blood flow and oxygen balance during exercise were evaluated in eight conscious dogs, instrumented for chronic measurements of coronary blood flow, left ventricular pressure, aortic blood pressure, heart rate, and sampling of arterial and coronary sinus venous blood. The administration of propranolol (1.5 mg/kg iv) produced a decrease in heart rate, peak left ventricular (LV) dP/dt, LV (dP/dt/P, and an increase in LV end-diastolic pressure during exercise. Mean coronary blood flow and myocardial oxygen consumption were lower after propranolol than at the same exercise intensity in control conditions. The oxygen delivery-to-oxygen consumption ratio and the coronary sinus oxygen content were also significantly lower. It is concluded that the relationship between myocardial oxygen supply and demand is modified during exercise after propranolol, so that a given level of myocardial oxygen consumption is achieved with a proportionally lower myocardial blood flow and a higher oxygen extraction.     

Exchange of oxygen across the epicardial surface distorts estimates of myocardial oxygen consumption

The rate of oxygen consumption of isolated, Langendorff-circulated, saline-perfused hearts of guinea pigs, rats, and rabbits was measured using the classical Fick Principle method. The heart was suspended in a glass chamber the oxygen partial pressure, PO2, of which could be varied. The measured rate of oxygen consumption was found to vary inversely with the ambient (heart chamber) PO2. This result prevailed whether the chamber was filled with air, saline, or oil, and whether the pericardium was present or the heart was wrapped in Saran. The effect varied inversely with heart size both within and across species. It is concluded that the epicardial surface is permeable to oxygen which will diffuse either into or out of the heart as the PO2 gradient dictates. In either case the classically measured rate of oxygen consumption will be in error. The error can be large in studies of cardiac basal metabolism. A simple model is developed to describe the observed rate of oxygen consumption as classically measured. The measured rate is partitioned into two components: the true rate of oxygen consumption of the heart, and the rate of loss of oxygen by diffusive exchange across the epicardial surface. The latter component is proportional to the gradient of oxygen partial pressure from myocardium to environment and to the diffusive oxygen conductance of myocardial tissue. Application of the model allows the true rate of oxygen consumption of the heart to be recovered from measured values which may be considerably in error.     

Simultaneous measurement of cellular respiration and acidification with a single CMOS ISFET

In vivo, the pH value and oxygen partial pressure are the most important physico-chemical parameters in the microenvironment of human tissues. In vitro, the extracellular acidification rate of cell cultures is an indicator of global cellular metabolism, while the rate of oxygen consumption is a measure of mitochondrial activity. Earlier approaches had the disadvantage that these two values had to be measured with two separate sensors at different loci within the tissue or cell culture. Furthermore, conventional Clark-type oxygen sensors are not very compatible for miniaturisation, making it impossible to measure at small cell volumes or even at the single cell level. We have, therefore, developed an ISFET based sensor structure which is able to measure both pH and oxygen partial pressure. This sensor structure was tested in vitro for simultaneous records of cellular acidification and respiration rates at the same site within the cell culture. This sensor is manufactured by a CMOS-process.     

The effect of myoglobin-facilitated oxygen transport on the basal metabolism of papillary muscle.

A mathematical model of oxygen diffusion into cylindrical papillary muscles is presented. The model partitions total oxygen flux into its simple and myoglobin-facilitated components. The model includes variable sigmoidal, exponential, or hyperbolic functions relating oxygen partial pressure to both fractional myoglobin saturation and rate of oxygen consumption. The behavior of the model was explored for a variety of saturation- and consumption-concentration relations. Facilitation of oxygen transport by myoglobin was considerable as indexed both by the elevation of oxygen partial pressure on the longitudinal axis of the muscle and by the fraction of total oxygen flux at the muscle center contributed by oxymyoglobin. Despite its facilitation of oxygen flux at the muscle center, myoglobin made only a negligible contribution to the total oxygen consumption averaged over the muscle cross-section. Hence the presence of myoglobin fails to explain either the experimentally determined basal metabolism-muscle radius relation or the stretch effect observed in isolated papillary muscle.     

The mechanism of stimulation of respiration by fatty acids in isolated hepatocytes

Addition of fatty acids to isolated hepatocytes raised respiration rate by 92% and raised mitochondrial membrane potential (delta psi m) in situ from 155 to 162 mV suggesting that the increased fuel supply had a greater effect on respiration rate than any increases in processes that consumed mitochondrial protonmotive force (delta p). The relationship between delta psi m and respiration rate was changed by addition of fatty acids or lactate, showing that there was also stimulation of delta p-consuming reactions. In the presence of oligomycin the relationship between delta psi m and respiration rate was unaffected by substrate addition, showing that the kinetics of delta p consumption by the H+ leak across the mitochondrial inner membrane were unchanged. The stimulation of delta p consumers by fatty acids therefore must be in the pathways of ATP synthesis and turnover. Inhibition of several candidate ATP-consuming reactions had little effect on basal or fatty acid-stimulated respiration, and the nature of the ATP turnover reactions in hepatocytes remains speculative. We conclude that fatty acids (and other substrates) stimulate respiration in hepatocytes in two distinct ways. They provide substrate for the electron transport chain, raising delta p and increasing the non-ohmic proton leak across the mitochondrial inner membrane and the rate of oxygen consumption. They also directly stimulate an unidentified delta p-consuming reaction in the cytoplasm. They do not work by uncoupling or by stimulation of intramitochondrial ATP-turnover reactions.     

Modeling of oxygen transport and cellular energetics explains observations on in vivo cardiac energy metabolism

Observations on the relationship between cardiac work rate and the levels of energy metabolites adenosine triphosphate (ATP), adenosine diphosphate (ADP), and phosphocreatine (CrP) have not been satisfactorily explained by theoretical models of cardiac energy metabolism. Specifically, the in vivo stability of ATP, ADP, and CrP levels in response to changes in work and respiratory rate has eluded explanation. Here a previously developed model of mitochondrial oxidative phosphorylation, which was developed based on data obtained from isolated cardiac mitochondria, is integrated with a spatially distributed model of oxygen transport in the myocardium to analyze data obtained from several laboratories over the past two decades. The model includes the components of the respiratory chain, the F₀F₁-ATPase, adenine nucleotide translocase, and the mitochondrial phosphate transporter at the mitochondrial level; adenylate kinase, creatine kinase, and ATP consumption in the cytoplasm; and oxygen transport between capillaries, interstitial fluid, and cardiomyocytes. The integrated model is able to reproduce experimental observations on ATP, ADP, CrP, and inorganic phosphate levels in canine hearts over a range of workload and during coronary hypoperfusion and predicts that cytoplasmic inorganic phosphate level is a key regulator of the rate of mitochondrial respiration at workloads for which the rate of cardiac oxygen consumption is less than or equal to approximately 12 μmol per minute per gram of tissue. At work rates corresponding to oxygen consumption higher than 12 μmol min⁻¹ g⁻¹, model predictions deviate from the experimental data, indicating that at high work rates, additional regulatory mechanisms that are not currently incorporated into the model may be important. Nevertheless, the integrated model explains metabolite levels observed at low to moderate workloads and the changes in metabolite levels and tissue oxygenation observed during graded hypoperfusion. These findings suggest that the observed stability of energy metabolites emerges as a property of a properly constructed model of cardiac substrate transport and mitochondrial metabolism. In addition, the validated model provides quantitative predictions of changes in phosphate metabolites during cardiac ischemia.     

Aerobic performance: role of oxygen delivery and utilization, glycolytic flux

Oxygen delivery to muscle, its consumption and glycolytic flux, all of each affect and restrict aerobic performance, are discussed. Energy supply of intensive exercise till exhaustion lasting 3 to 4 min is provided mainly by oxidative metabolism, simultaneously glycolytic flux may be increased considerably. Other conditions being equal, capacity of oxygen delivery determines oxygen partial pressure in myoplasm of exercising/contracting muscle. With PO2 in myoplasm increasing from 0 to 1-2 mm Hg oxygen consumption (VO2) in mitochondria enhances dramatically, with further increase of PO2 its rise slows down. At the ascending part of VO2-PO2 relationship for mitochondria the increase of VO2 is noticeably restricted by oxygen delivery to contracting muscle. When PO2 approaches plateau of the VO2-PO2 relationship, an increase of VO2 is restricted by mitochondria capacity to accumulate oxygen and augmented oxygen delivery will not lead to a significant increase of muscle VO2. On the other hand considerable accumulation of glycolytic metabolites in contracting muscle causes a decrease of contractility which in its turn may restrict aerobic performance. Noteworthy no strict relationship between glycolytic flux and PO2 in myoplasm exists. That is why correct evaluation of factors limiting aerobic performance presupposes simultaneous evaluation of both glycolytic flux and oxygen consumption in muscle which in its turn depends on oxygen delivery to mitochondria and its utilization.     

Radius-dependent decline of performance in isolated cardiac muscle does not reflect inadequacy of diffusive oxygen supply

The study of cardiac energetics commonly involves the use of isolated muscle preparations (papillary muscles or trabeculae carneae). Their contractile performance has been observed to vary inversely with thickness. This inverse dependence has been attributed, almost without exception, to inadequate diffusion of oxygen into the centers of muscles of large diameter. It is thus commonly hypothesized that the radius-dependent diminution of performance reflects the development of an anoxic core. We tested this hypothesis theoretically by solving a modification of the diffusion equation, in which the rate of oxygen consumption is a sigmoidal function of the partial pressure of oxygen. The model demonstrates that sufficiently thick muscles, operating at sufficiently high rates of oxygen demand or sufficiently low ambient partial pressures of oxygen, will indeed show diminished energetic performance, whether indirectly indexed as stress (force per cross-sectional area) development or as the rate of heat production. However, such simulated behavior requires the adoption of extreme parameter values, often differing by an order of magnitude from their experimental equivalents. We thus conclude that the radius-dependent diminution of muscle performance in vitro cannot be attributed entirely to an insufficient supply of oxygen via diffusion.     

Monoamine oxidase, an intracellular probe of oxygen pressure in isolated cardiac myocytes

The activity of monamine oxidase, an enzyme located almost exclusively at the outer mitochondrial membrane, toward the substrate phenylethylamine is used to report the oxygen pressure at the outer mitochondrial membrane of intact cardiac myocytes isolated from hearts of adult rats. The rate of substrate oxidation, under the conditions used, follows the Michaelis-Menten relation, and accordingly can be used as a measure of the local chemical activity of dissolved oxygen. The oxygen pressure at the outer mitochondrial membrane of myocytes, at rest and after 2- to 3-fold stimulation of respiratory oxygen consumption, differs from the extracellular oxygen pressure by at most 2 torr. This implies that most of the large, about 20 torr, difference in oxygen pressure between capillary lumen and mitochondria of the working heart must be extracellular. At physiologically relevant concentrations of the substrates phenylethylamine and norepinephrine, monoamine oxidase activity is relatively insensitive to extracellular oxygen pressure in the range 155 to 8 torr, suggesting a limited role for regulation of biogenic amine oxidation by oxygen availability.     

A theoretical analysis of the effect of placental metabolism on fetal oxygenation under conditions of limited oxygen availability.

The purpose of this theoretical paper is to examine the effects of placental metabolism on fetal oxygenation under conditions of limited oxygen availability. Features of the mathematical model used here include: (1) ordinary non-linear differential equations defining the oxygen partial pressure profiles in the maternal and fetal streams for a concurrent flow pattern; (2) the presence of maternal and fetal blood flow shunts; (3) consumption of oxygen by a metabolically active placenta; and (4) modification of the fetal input to the placenta by changing the rate of fetal oxygen consumption in response to changes in the rate of oxygen delivered to the fetus via the umbilical vein. Model parameters were chosen to be well within the range of values cited in the literature. Based on these calculations, we conclude that: (1) under normal conditions, approximately one-half of the fetal uterine-umbilical venous oxygen partial pressure difference can be attributed to placental oxygen consumption; (2) utilization of fetal oxygen to help maintain the metabolic activities of the placenta does not significantly impair fetal oxygenation under normal conditions; (3) consumption of oxygen by the placenta will have a significant detrimental effect on the rate of oxygen delivered to the fetus if oxygen availability is compromised; and (4) for the same rate of maternal oxygen delivered to the placenta, maternal hypoxemia has a significantly greater adverse effect on fetal oxygenation than does maternal anemia.     

Influence of dicarboxylic phosphatidylcholines on the stability and phase transition of phosphatidylcholine liposomes

Utilizing reflectance spectrophotometry, hemoperfusion, rate of oxygen consumption and redox level of mitochondrial cytochrome c (+c1) in livers in situ of anesthetized rats were measured. The transition to the anoxic state was induced by raising the pressure on the liver surface to more than the hepatic blood pressure by pressing with the tip of the optical guide of the reflectance spectrophotometer. During this transition, the average oxygen saturation of hemoglobin in the liver in situ decreased linearly with time until it became 10--20% of the total. This was followed by reduction of mitochondrial cytochrome c (+c1), which reached completion in 10--20 s. The measured O2 consumption rate remained constant until the percentage of oxyhemoglobin in situ decreased to a critical level. There was then a decrease in the rate of O2 consumption which was accompanied by a progressive reduction of cytochrome c (+c1). It was shown that amounts of hemoglobin and mitochondrial respiratory chain cytochromes in the liver in situ could be measured non-invasively and could provide important signals for vital cellular functions. The changes in hemoperfusion and rate of O2 consumption of the liver in situ following ethanol ingestion were also shown in rats, and are briefly discussed with respect to possible application of this method to study the pathophysiology of tissues.     

Gas Exchange of Algae: II. Effects of Oxygen, Helium, and Argon on the Photosynthesis of Chlorella pyrenoidosa

Changes in the oxygen partial pressure of air over the range of 8 to 258 mm of Hg did not adversely affect the photosynthetic capacity of Chlorella pyrenoidosa. Gas exchange and growth measurements remained constant for 3-week periods and were similar to air controls (oxygen pressure of 160 mm of Hg). Oxygen partial pressures of 532 and 745 mm of Hg had an adverse effect on algal metabolism. Carbon dioxide consumption was 24% lower in the gas mixture containing oxygen at a pressure 532 mm of Hg than in the air control, and the growth rate was slightly reduced. Oxygen at a partial pressure of 745 mm of Hg decreased the photosynthetic rate 39% and the growth rate 37% over the corresponding rates in air. The lowered metabolic rates remained constant during 14 days of measurements, and the effect was reversible after this time. Substitution of helium or argon for the nitrogen in air had no effect on oxygen production, carbon dioxide consumption, or growth rate for 3-week periods. All measurements were made at a total pressure of 760 mm of Hg, and all gas mixtures were enriched with 2% carbon dioxide. Thus, the physiological functioning and reliability of a photosynthetic gas exchanger should not be adversely affected by: (i) oxygen partial pressures ranging from 8 to 258 mm of Hg; (ii) the use of pure oxygen at reduced total pressure (155 to 258 mm of Hg) unless pressure per se affects photosynthesis, or (iii) the inclusion of helium or argon in the gas environment (up to a partial pressure of 595 mm of Hg).     

Fatty liver and insulin resistance in obese Zucker rats: no role for mitochondrial dysfunction

The relationship between insulin resistance and mitochondrial function is of increasing interest. Studies looking for such interactions are usually made in muscle and only a few studies have been done in liver, which is known to be a crucial partner in whole body insulin action. Recent studies have revealed a similar mechanism to that of muscle for fat-induced insulin resistance in liver. However, the exact mechanism of lipid metabolites accumulation in liver leading to insulin resistance is far from being elucidated. One of the hypothetical mechanisms for liver steatosis development is an impairment of mitochondrial function. We examined mitochondrial function in fatty liver and insulin resistance state using isolated mitochondria from obese Zucker rats. We determined the relationship between ATP synthesis and oxygen consumption as well as the relationship between mitochondrial membrane potential and oxygen consumption. In order to evaluate the quantity of mitochondria and the oxidative capacity we measured citrate synthase and cytochrome c oxidase activities. Results showed that despite significant fatty liver and hyperinsulinemia, isolated liver mitochondria from obese Zucker rats display no difference in oxygen consumption, ATP synthesis, and membrane potential compared with lean Zucker rats. There was no difference in citrate synthase and cytochrome c oxidase activities between obese and lean Zucker rats in isolated mitochondria as well as in liver homogenate, indicating a similar relative amount of hepatic mitochondria and a similar oxidative capacity. Adiponectin, which is involved in bioenergetic homeostasis, was increased two-fold in obese Zucker rats despite insulin resistance. In conclusion, isolated liver mitochondria from lean and obese insulin-resistant Zucker rats showed strictly the same mitochondrial function. It remains to be elucidated whether adiponectin increase is involved in these results.     

Renal hypoxia and dysoxia after reperfusion of the ischemic kidney

Ischemia is the most common cause of acute renal failure. Ischemic-induced renal tissue hypoxia is thought to be a major component in the development of acute renal failure in promoting the initial tubular damage. Renal oxygenation originates from a balance between oxygen supply and consumption. Recent investigations have provided new insights into alterations in oxygenation pathways in the ischemic kidney. These findings have identified a central role of microvascular dysfunction related to an imbalance between vasoconstrictors and vasodilators, endothelial damage and endothelium-leukocyte interactions, leading to decreased renal oxygen supply. Reduced microcirculatory oxygen supply may be associated with altered cellular oxygen consumption (dysoxia), because of mitochondrial dysfunction and activity of alternative oxygen-consuming pathways. Alterations in oxygen utilization and/or supply might therefore contribute to the occurrence of organ dysfunction. This view places oxygen pathways' alterations as a potential central player in the pathogenesis of acute kidney injury. Both in regulation of oxygen supply and consumption, nitric oxide seems to play a pivotal role. Furthermore, recent studies suggest that, following acute ischemic renal injury, persistent tissue hypoxia contributes to the development of chronic renal dysfunction. Adaptative mechanisms to renal hypoxia may be ineffective in more severe cases and lead to the development of chronic renal failure following ischemia-reperfusion. This paper is aimed at reviewing the current insights into oxygen transport pathways, from oxygen supply to oxygen consumption in the kidney and from the adaptation mechanisms to renal hypoxia. Their role in the development of ischemia-induced renal damage and ischemic acute renal failure are discussed.     

O2 transport in cerebral microregions (mathematical simulation)

The effects of the circulation rate in capillaries, the intensity of O2 consumption by nerve cells and the capillary network density on the O2 tension distribution in the cerebral cortex have been studied, utilizing a mathematical model simulating actual neuron-capillary relationships. The model has been written as a system of equations in partial derivatives, its solution obtained by the net-point method. Regulatory variations of the capillary circulation rate in certain cerebral microregions have been shown to ensure similar changes in oxygen supply throughout the region. A drop of the pO2 level in a cerebral microregion with a rising O2 consumption by nerve cells is shown to be due, by 75 percent, to the increase of O2 consumption and by 25 percent, to the lower pO2 in the capillaries. Conversely, an increase in pO2 in microregions resulting from a lower O2 consumption by neurons is due by 75 percent, to a pO2 rise in capillaries and by 25 percent, at the expense of an O2 consumption decrease. In cerebral regions differing in capillary network density by 20 percent, changes in the conditions for oxygen supply to tissue are due by 1/3 to pO2 variations in the capillaries and by 2/3 to alterations in the diffusion distances.     

The potential uses of ethephon in apple growing

A reduced partial oxygen pressure retards the onset and the rate of ripening of tomato fruits, irrespective of the total atmospheric pressure and the absorption of endogenously evolved ethylene. The reduction of the partial oxygen supply is therefore a main factor in retarding senescence at low pressure storage.