The present study evaluated whether intracellular partial pressure of O2 ([Formula: see text]) modulates the muscle O2 uptake (V˙o 2) as exercise intensity increased. Indirect calorimetry followedV˙o 2, whereas nuclear magnetic resonance (NMR) monitored the high-energy phosphate levels, intracellular pH, and intracellular[Formula: see text] in the gastrocnemius muscle of four untrained subjects at rest, during plantar flexion exercise with a constant load at a repetition rate of 0.75, 0.92, and 1.17 Hz, and during postexercise recovery.V˙o 2 increased linearly with exercise intensity and peaked at 1.17 Hz (15.1 ± 0.37 watts), when the subjects could maintain the exercise for only 3 min.V˙o 2 reached a peak value of 13.0 ± 1.59 ml O2 ⋅ min−1 ⋅ 100 ml leg volume−1. The31P spectra indicated that phosphocreatine decreased to 32% of its resting value, whereas intracellular pH decreased linearly with power output, reaching 6.86. Muscle ATP concentration, however, remained constant throughout the exercise protocol. The 1H NMR deoxymyoglobin signal, reflecting the cellular[Formula: see text], decreased in proportion to increments in power output andV˙o 2. At the highest exercise intensity and peakV˙o 2, myoglobin was ∼50% desaturated. These findings, taken together, suggest that the O2 gradient from hemoglobin to the mitochondria can modulate the O2flux to meet the increasedV˙o 2 in exercising muscle, but declining cellular [Formula: see text]during enhanced mitochondrial respiration suggests that O2 availability is not limitingV˙o 2 during exercise.

1H NMR has detected both the deoxygenated proximal histidyl NδH signals of myoglobin (deoxyMb) and deoxygenated Hb (deoxyHb) from human gastrocnemius muscle. Exercising the muscle or pressure cuffing the leg to reduce blood flow elicits the appearance of the deoxyMb signal, which increases in intensity as cellular[Formula: see text] decreases. The deoxyMb signal is detected with a 45-s time resolution and reaches a steady-state level within 5 min of pressure cuffing. Its desaturation kinetics match those observed in the near-infrared spectroscopy (NIRS) experiments, implying that the NIRS signals are actually monitoring Mb desaturation. That interpretation is consistent with the signal intensity and desaturation of the deoxyHb proximal histidyl NδH signal from the β-subunit at 73 parts per million. The experimental results establish the feasibility and methodology to observe the deoxyMb and Hb signals in skeletal muscle, help clarify the origin of the NIRS signal, and set a stage for continuing study of O2regulation in skeletal muscle.

The detection of the 1H NMR signal of myoglobin (Mb) in tissue opens an opportunity to examine its cellular diffusion property, which is central to its purported role in facilitating oxygen transport. In perfused myocardium the field-dependent transverse relaxation analysis of the deoxy Mb proximal histidyl NdeltaH indicates that the Mb rotational correlation time in the cell is only approximately 1.4 times longer than it is in solution. Such a mobility is consistent with the theory that Mb facilitates oxygen diffusion from the sarcoplasm to the mitochondria. The microviscosities of the erythrocyte and myocyte environment are different. The hemoglobin (Hb) rotational correlation time is 2.2 longer in the cell than in solution. Because both the overlapping Hb and Mb signals are visible in vivo, a relaxation-based NMR strategy has been developed to discriminate between them.

(1)H-NMR experiments have determined intracellular O(2) consumption (Vo(2)) with oxymyoglobin (MbO(2)) desaturation kinetics in human calf muscle during plantar flexion exercise at 0.75, 0.92, and 1.17 Hz with a constant load. At the onset of muscle contraction, myoglobin (Mb) desaturates rapidly. The desaturation rate constant of approximately 30 s reflects the intracellular Vo(2). Although Mb desaturates quickly with a similar time constant at all workload levels, its final steady-state level differs. As work increases, the final steady-state cellular Po(2) decreases progressively. After Mb desaturation has reached a steady state, however, Vo(2) continues to rise. On the basis of current respiratory control models, the analysis in the present report reveals two distinct Vo(2) phases: an ADP-independent phase at the onset of contraction and an ADP-dependent phase after Mb has reached a steady state. In contrast to the accepted view, the initial intracellular Vo(2) shows that oxidative phosphorylation can support up to 36% of the energy cost, a significantly higher fraction than expected. Partitioning of the energy flux shows that a 31% nonoxidative component exists and responds to the dynamic energy utilization-restoration cycle (which lasts for only milliseconds) as postulated in the glycogen shunt theory. The present study offers perspectives on the regulation of respiration, bioenergetics, and Mb function during muscle contraction.

The production of glycolytic end products, such as lactate, usually evokes a cellular shift from aerobic to anaerobic ATP generation and O 2 insufficiency. In the classical view, muscle lactate must be exported to the liver for clearance. However, lactate also forms under well-oxygenated conditions, and this has led investigators to postulate lactate shuttling from non-oxidative to oxidative muscle fiber, where it can serve as a precursor. Indeed, the intracellular lactate shuttle and the glycogen shunt hypotheses expand the vision to include a dynamic mobilization and utilization of lactate during a muscle contraction cycle. Testing the tenability of these provocative ideas during a rapid contraction cycle has posed a technical challenge. The present study reports the use of hyperpolarized [1- 13 C]lactate increases sharply and acetyl-L-carnitine, acetoacetate and glutamate levels also rise. Such a quick mobilization of pyruvate and lactate toward oxidative metabolism supports the postulated role of lactate in the glycogen shunt and the intracellular lactate shuttle models. The study thus introduces an innovative DNP approach to measure metabolite transients, which will help delineate the cellular and physiological role of lactate and glycolytic end products.