Diffusion barriers for ADP in the cardiac cell.

The regulation of mitochondrial respiration in the intact heart may differ from that of isolated mitochondria if intracellular diffusion is restricted. Here we consider which factors may hinder diffusion in vivo and, based on computational analysis, design a reverse engineering approach to estimate the role of diffusional resistance in mitochondrial regulation from an experiment on the intact heart. Computational analysis of respiration measurements on skinned heart fibers shows that the outer mitochondrial membrane does not hinder diffusion enough to cause ADP gradients of tens of micromolars. A diffusion model further shows that the mesoscale structure of the myofibrillar space also does not hinder diffusion appreciably. However, ADP gradients are suggested by the measured activation time of oxidative phosphorylation and may be caused by diffusion restriction of other intracellular structures or the in vivo microstructure of networks of physically interacting proteins. Based on computational modeling we propose an experiment on the intact heart that allows to estimate the effective diffusion restriction between ATP producing and consuming sites in the cardiac cell.

Role of the creatine/phosphocreatine system in the regulation of mitochondrial respiration.

The mechanism of metabolic regulation of mitochondrial respiration in cardiac muscle cells was studied experimentally in the permeabilized heart fibres of mice and by computer modelling in silico. The experiments showed that the rate of mitochondrial respiration could be controlled by local production of ADP by mitochondrial creatine kinase in the intermembrane space of mitochondria. The spatially inhomogenous reaction-diffusion model of compartmentalized energy transfer was used to analyse which metabolite level in cytoplasm may be important for regulation of respiration. At low and moderate workloads, up to VO2 equal to 70 micromol min-1 g-1 dry weight, the only factor to which respiration responded was inorganic phosphate. At the values of VO2 higher than 70 micromol min-1 g-1 dry weight, the respiration rate responded mostly to changes in creatine, phosphocreatine and then time-averaged (over the contractile cycle) ADP concentrations in the cytoplasm. These results are taken to show that under conditions of moderate workloads, creatine kinase activity at given physiological creatine and phosphocreatine concentrations (apparent maximal activity achievable under these conditions) is in excess to oxidative phosphorylation rate, which is controlled by Pi concentration changes starting from very low values of the latter. At higher workloads mi-CK should be upregulated by increasing creatine and decreasing phosphocreatine concentrations, and only at very high workloads the ADP diffusion flux should be increased to upregulate oxidative phosphorylation. Thus, on the basis of the study in silico of compartmentalized energy transfer by phophocreatine/creatine system, the authors conclude that there exist multiple parallel regulatory factors controlling the rate of oxygen consumption in dependence of the workload. If creatine kinase is inhibited (and there is no myokinase activity), respiration requires high diffusive flux of ADP back into mitochondria, which is the sole regulator of respiration. This needs, however, increased ADP concentrations in the cytoplasm, which in turn result in inhibition of contraction.

Regulation of mitochondrial respiration in heart cells analyzed by reaction-diffusion model of energy transfer.

The purpose of this study is to investigate theoretically which intracellular factors may be important for regulation of mitochondrial respiration in working heart cells in vivo. We have developed a model that describes quantitatively the published experimental data on dependence of the rate of oxygen consumption and metabolic state of working isolated perfused rat heart on workload over its physiological range (Williamson JR, Ford G, Illingworth J, Safer B. Circ Res 38, Suppl I, I39-I51, 1976). Analysis of this model shows that for phosphocreatine, creatine, and ATP the equilibrium assumption is an acceptable approximation with respect to their diffusion in the intracellular bulk water phase. However, the ADP concentration changes in the contraction cycle in a nonequilibrium workload-dependent manner, showing the existence of the intracellular concentration gradients. The model shows that workload-dependent alteration of ADP concentration in the compartmentalized creatine kinase system may be taken, together with the changes in P(i) concentration, to be among the major components of the metabolic feedback signal for regulation of respiration in muscle cells.