Similar mitochondrial activation kinetics in wild-type and creatine kinase-deficient fast-twitch muscle indicate significant Pi control of respiration

Abstract

Past simulations of oxidative ATP metabolism in skeletal muscle have predicted that elimination of the creatine kinase (CK) reaction should result in dramatically faster oxygen consumption dynamics during transitions in ATP turnover rate. This hypothesis was investigated. Oxygen consumption of fast-twitch (FT) muscle isolated from wild-type (WT) and transgenic mice deficient in the myoplasmic (M) and mitochondrial (Mi) CK isoforms (MiM CK(-/-)) were measured at 20°C at rest and during electrical stimulation. MiM CK(-/-) muscle oxygen consumption activation kinetics during a step change in contraction rate were 30% faster than WT (time constant 53 ± 3 vs. 69 ± 4 s, respectively; mean ± SE, n = 8 and 6, respectively). MiM CK(-/-) muscle oxygen consumption deactivation kinetics were 380% faster than WT (time constant 74 ± 4 s vs. 264 ± 4 s, respectively). Next, the experiments were simulated using a computational model of the oxidative ATP metabolic network in FT muscle featuring ADP and Pi feedback control of mitochondrial respiration (J. A. L. Jeneson, J. P. Schmitz, N. A. van den Broek, N. A. van Riel, P. A. Hilbers, K. Nicolay, J. J. Prompers. Am J Physiol Endocrinol Metab 297: E774-E784, 2009) that was reparameterized for 20°C. Elimination of Pi control via clamping of the mitochondrial Pi concentration at 10 mM reproduced past simulation results of dramatically faster kinetics in CK(-/-) muscle, while inclusion of Pi control qualitatively explained the experimental observations. On this basis, it was concluded that previous studies of the CK-deficient FT muscle phenotype underestimated the contribution of Pi to mitochondrial respiratory control.

Fig. 1.

Typical results for the respiratory flux response during a rest-stimulation-recovery experiment for wild-type (WT) (top) and mitochondrial (Mi) creatine kinase isoforms (MiM-CK^−/−) (bottom) extensor digitorum longus (EDL) muscle. The stimulation frequency was 2 Hz. Monoexponential best fits of the stimulation frequency transients from rest-to-active (0 to 2 Hz) and active-to-rest (2 to 0 Hz) are indicated as thickened line parts.

Fig. 2.

Steady-state oxygen consumption (in nmol·min⁻¹·g⁻¹) of WT (n = 8) vs. MiM-CK^−/− (n = 6) EDL muscle at rest and during serial stimulation at 0.5, 1, and 2 Hz, respectively. Values are expressed as means ± SE. *P < 0.05. The solid lines represent the fits of a monoexponential function to the data. Regression equations: WT: JO₂ = 63.1 + 202.7 [1 − exp (−1.6/x)]; MiM-CK^−/−: JO₂ = 131.1 + 249.9 [1 − exp (−1.3/x)].

Fig. 3.

Computed timecourses of oxygen consumption (A–C), the cytoplasmic ADP (D–F), and Pi (G, H) concentrations, and the free energy of ATP hydrolysis (ΔG_p; I, J) during simulated rest-stimulation-recovery experiments at four incremental ATP turnover rates in the network (0.02, 0.03, 0.04, and 0.05 mM/s, respectively) for three model configurations: 1) WT muscle (Fig. 3, A and D, G, and I); 2) CK^−/− muscle (Fig. 3, B, E, H, and J); 3) CK^−/− muscle with a constant intramitochondrial Pi concentration ([Pi_x]) of 10 mM (Fig. 3, C and F).

Fig. 4.

Computed oxygen consumption activation time constant and 95% activation time (A) and 95% recovery time (B) as a function of the ATP turnover rate in the network for the three model configurations: 1) WT, 2) CK^−/−, and 3) CK^−/− with [Pi_x] clamped at 10 mM. The kinetic parameters were derived from the computed time course of the oxygen consumption rate for ATP turnover rates between 0.005 and 0.05 mM for each model phenotype as described in methods.