A computational model of mitochondrial metabolism and electrophysiology is introduced and applied to analysis of data from isolated cardiac mitochondria and data on phosphate metabolites in striated muscle in vivo. This model is constructed based on detailed kinetics and thermodynamically balanced reaction mechanisms and a strict accounting of rapidly equilibrating biochemical species. Since building such a model requires introducing a large number of adjustable kinetic parameters, a correspondingly large amount of independent data from isolated mitochondria respiring on different substrates and subject to a variety of protocols is used to parameterize the model and ensure that it is challenged by a wide range of data corresponding to diverse conditions. The developed model is further validated by both in vitro data on isolated cardiac mitochondria and in vivo experimental measurements on human skeletal muscle. The validated model is used to predict the roles of NAD and ADP in regulating the tricarboxylic acid cycle dehydrogenase fluxes, demonstrating that NAD is the more important regulator. Further model predictions reveal that a decrease of cytosolic pH value results in decreases in mitochondrial membrane potential and a corresponding drop in the ability of the mitochondria to synthesize ATP at the hydrolysis potential required for cellular function.Mitochondrial energy metabolism centers on the tricarboxylic acid cycle reactions, oxidative phosphorylation, and associated transport reactions. It is a system in which biochemical reactions are coupled to membrane electrophysiology, nearly every intermediate acts as an allosteric regulator of several enzymes in the system, and nearly all intermediates are transported into and out of the mitochondria via a host of electroneutral and electrogenic exchangers and cotransporters. Thus, it is a system with a level of complexity that begs for computational modeling to aid in analysis of experimental data and development and testing of quantitative hypotheses. In addition, as is demonstrated in this work, computer modeling of mitochondrial function facilitates the translation of observations made in one experimental regime (isolated ex vivo mitochondria) to another (in vivo cellular energy metabolism).In addition, the developed model provides the basis for examining how mitochondrial energetics is controlled in vivo. The model is used to predict the roles of NAD and ADP on regulating tricarboxylic acid cycle dehydrogenase fluxes, demonstrating that NAD is the more important regulator. The mitochondrial redox state in turn is affected by cytoplasmic P i concentration, since inorganic phosphate is a co-factor for transport of tricarboxylic acid cycle substrates a substrate for the tricarboxylic acid cycle. Since ADP and P i are the biochemical substrates for oxidative phosphorylation, the primary mechanism of control of mitochondrial energy metabolism (tricarboxylic acid cycle and oxidative phosphorylation) in striated muscle is feedback of the products of ATP hydrolysis....
To understand how cardiac ATP and CrP remain stable with changes in work rate -a phenomenon that has eluded mechanistic explanation for decades -data from 31 phosphate-magnetic resonance spectroscopy ( 31 P-MRS) are analysed to estimate cytoplasmic and mitochondrial phosphate metabolite concentrations in the normal state, during high cardiac workstates, during acute ischaemia and reactive hyperaemic recovery. Analysis is based on simulating distributed heterogeneous oxygen transport in the myocardium integrated with a detailed model of cardiac energy metabolism. The model predicts that baseline myocardial free inorganic phosphate (P i ) concentration in the canine myocyte cytoplasm -a variable not accessible to direct non-invasive measurement -is approximately 0.29 mm and increases to 2.3 mm near maximal cardiac oxygen consumption. During acute ischaemia (from ligation of the left anterior descending artery) P i increases to approximately 3.1 mm and ATP consumption in the ischaemic tissue is reduced quickly to less than half its baseline value before the creatine phosphate (CrP) pool is 18% depleted. It is determined from these experiments that the maximal rate of oxygen consumption of the heart is an emergent property and is limited not simply by the maximal rate of ATP synthesis, but by the maximal rate at which ATP can be synthesized at a potential at which it can be utilized. The critical free energy of ATP hydrolysis for cardiac contraction that is consistent with these findings is approximately −63.5 kJ mol −1 . Based on theoretical findings, we hypothesize that inorganic phosphate is both the primary feedback signal for stimulating oxidative phosphorylation in vivo and also the most significant product of ATP hydrolysis in limiting the capacity of the heart to hydrolyse ATP in vivo. Due to the lack of precise quantification of P i in vivo, these hypotheses and associated model predictions remain to be carefully tested experimentally.
Data from 31 P-nuclear magnetic resonance spectroscopy of human forearm flexor muscle were analyzed based on a previously developed model of mitochondrial oxidative phosphorylation (PLoS Comp Bio 1: e36, 2005) to test the hypothesis that substrate level (concentrations of ADP and inorganic phosphate) represents the primary signal governing the rate of mitochondrial ATP synthesis and maintaining the cellular ATP hydrolysis potential in skeletal muscle. Model-based predictions of cytoplasmic concentrations of phosphate metabolites (ATP, ADP, and Pi) matched data obtained from 20 healthy volunteers and indicated that as work rate is varied from rest to submaximal exercise commensurate increases in the rate of mitochondrial ATP synthesis are effected by changes in concentrations of available ADP and Pi. Additional data from patients with a defect of complex I of the respiratory chain and a patient with a deficiency in the mitochondrial adenine nucleotide translocase were also predicted the by the model by making the appropriate adjustments to the activities of the affected proteins associates with the defects, providing both further validation of the biophysical model of the control of oxidative phosphorylation and insight into the impact of these diseases on the ability of the cell to maintain its energetic state. computational model; mitochondria; cellular energetics; oxidative phosphorylation; 31 P-NMR spectroscopy MITOCHONDRIAL OXIDATIVE ADP phosphorylation is the primary source of ATP in skeletal muscle during aerobic exercise. Thus, to maintain the free energy state of the cytoplasmic phosphoenergetic compounds ATP, ADP, and P i , oxidative phosphorylation is modulated to match the rate of ATP utilization during exercise. It has recently been shown through computational model-based analysis of data obtained from 31 P-NMR spectroscopy of working in vivo dog hearts that the primary control mechanism operating in cardiomyocytes is feedback of substrate concentrations for ATP synthesis (5). In other words, changes in the concentrations of the products generated by the utilization of ATP in the cell, ADP and P i , effect changes in the rate at which mitochondria utilize those products to resynthesize ATP (5).Here the question of whether this same mechanism can explain the observed data on the control of oxidative metabolism in skeletal muscle is investigated. Previous analyses of 31 P-NMR spectroscopy ( 31 P-MRS) data on energy balance in exercising skeletal muscle have mainly focused on testing ADP feedback control of mitochondrial ATP synthesis using black box descriptions of the mitochondrial ATP synthetic pathway (8, 14 -16, 28), P i acceptor control (7), and thermodynamic control involving quasi-linear relations between cytoplasmic Gibbs free energy of ATP hydrolysis and mitochondrial ATP synthesis flux (13,18, 31). Yet, to date, these 31 P-MRS data have not been adequately explained based on a detailed mechanistic model of oxidative phosphorylation and cellular energetics.To analyze and interpret data from s...
The failing heart is hypothesized to suffer from energy supply inadequate for supporting normal cardiac function. We analyzed data from a canine left ventricular hypertrophy model to determine how the energy state evolves because of changes in key metabolic pools. Our findings-confirmed by in vivo 31 P-magnetic resonance spectroscopy-indicate that the transition between the clinically observed early compensatory phase and heart failure and the critical point at which the transition occurs are emergent properties of cardiac energy metabolism. Specifically, analysis reveals a phenomenon in which low and moderate reductions in metabolite pools have no major negative impact on oxidative capacity, whereas reductions beyond a critical tipping point lead to a severely compromised energy state. The transition point corresponds to reductions in the total adenine nucleotide pool (TAN) of Ϸ30%, corresponding to the reduction observed in humans in heart failure [Ingwall JS, Weiss RG (2004) Is the failing heart energy starved? On using chemical energy to support cardiac function. Circ Res 95(2):135-145]. At given values of TAN and the total exchangeable phosphate pool during hypertrophic remodeling, the creatine pool attains a value that is associated with optimal ATP hydrolysis potential. Thus, both increases and decreases to the creatine pool are predicted to result in diminished energetic state unless accompanied by appropriate simultaneous changes in the other pools.computational biology ͉ hypertrophy ͉ mitochondria ͉ energy metabolism T he mechanistic relationship between energy metabolism and cardiac contractility during the development of heart failure has long eluded understanding. Observations on failing hearts show that basal ATP and phosphocreatine (CrP), as well as the total creatine pool (CR tot ), decrease steadily as cardiac hypertrophic remodeling progresses. These decreases are associated with abnormal levels of other energy metabolites in both animals and humans with left ventricular hypertrophy (LVH) and congestive heart failure (CHF), implying abnormal regulation of oxidative phosphorylation in LVH and CHF. The central hypothesis that has emerged from these observations is that the evolution from the normal to failing heart involves ''energy starvation,'' in which the ability of mitochondria to synthesize ATP at the rate and free energy required for normal cardiac function is impaired.To understand the observed phenomena, we applied a validated computer model that simulates cellular oxygen consumption and cardiac energy metabolism. The predictions of this model challenged the widely accepted hypothesis (1) that biochemical feedback cannot explain how phosphate metabolite concentrations vary with cardiac work rate in the heart and show that the in vivo data are consistent with a critical role of inorganic phosphate (Pi) concentration as a feedback signal (2). Here we use the same model to analyze data on the evolution of LVH and CHF in a canine model. The model is adapted to LVH by varying 3 biochemical po...
Previous studies have demonstrated that the metabolic syndrome is associated with impaired skeletal muscle arteriolar function, although integrating observations into a conceptual framework for impaired perfusion in peripheral vascular disease (PVD) has been limited. This study builds on previous work to evaluate in situ arteriolar hemodynamics in cremaster muscle of obese Zucker rats (OZR) to integrate existing knowledge into a greater understanding of impaired skeletal muscle perfusion. In OZR cremaster muscle, perfusion distribution at microvascular bifurcations (γ) was consistently more heterogeneous than in controls. However, while consistent, the underlying mechanistic contributors were spatially divergent as altered adrenergic constriction was the major contributor to altered γ at proximal microvascular bifurcations, with a steady decay with distance, while endothelial dysfunction was a stronger contributor in distal bifurcations with no discernible role proximally. Using measured values of γ, we found that simulations predict that successive alterations to γ in OZR caused more heterogeneous perfusion distribution in distal arterioles than in controls, an effect that could only be rectified by combined adrenoreceptor blockade and improvements to endothelial dysfunction. Intravascular (125)I-labeled albumin tracer washout from in situ gastrocnemius muscle of OZR provided independent support for these observations, indicating increased perfusion heterogeneity that was corrected only by combined adrenoreceptor blockade and improved endothelial function. These results suggest that a defining element of PVD in the metabolic syndrome may be an altered γ at microvascular bifurcations, that its contributors are heterogeneous and spatially distinct, and that interventions to rectify this negative outcome must take a new conceptual framework into account.
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