Mitochondrial energetics, pH regulation, and ion dynamics: a computational-experimental approach.

We developed a computational model of mitochondrial energetics that includes Ca(2+), proton, Na(+), and phosphate dynamics. The model accounts for distinct respiratory fluxes from substrates of complex I and complex II, pH effects on equilibrium constants and enzyme kinetics, and the acid-base equilibrium distributions of energy intermediaries. We experimentally determined NADH and ΔΨ(m) in guinea pig mitochondria during transitions from de-energized to energized, or during state 2/4 to state 3 respiration, or into hypoxia and uncoupling, and compared the results with those obtained in model simulations. The model quantitatively reproduces the experimentally observed magnitude of ΔΨ(m), the range of NADH levels, respiratory fluxes, and respiratory control ratio upon transitions elicited by sequential additions of substrate and ADP. Simulation results are also able to mimic the change in ΔΨ(m) upon addition of phosphate to state 4 mitochondria, leading to matrix acidification and ΔΨ(m) polarization. The steady-state behavior of the integrated mitochondrial model qualitatively simulates the dependence of respiration on the proton motive force, and the expected flux-force relationships existing between respiratory and ATP synthesis fluxes versus redox and phosphorylation potentials. This upgraded mitochondrial model provides what we believe are new opportunities for simulating mitochondrial physiological behavior during dysfunctional states involving changes in pH and ion dynamics.