The value of the diffusion coefficient for oxygen in muscle is uncertain. The diffusion coefficient is important because it is a determinant of the extracellular oxygen tension at which the core of muscle fibers becomes anoxic (Po_2crit). Anoxic cores in muscle fibers impair muscular function and may limit adaptation of muscle cells to increased load and/or activity. We used Hill’s diffusion equations to determine Krogh’s diffusion coefficient (Dα) for oxygen in single skeletal muscle fibers from Xenopus laevis at 20°C (n = 6) and in myocardial trabeculae from the rat at 37°C (n = 9). The trabeculae were dissected from the right ventricular myocardium of control (n = 4) and monocrotaline-treated, pulmonary hypertensive rats (n = 5). The cross-sectional area of the preparations, the maximum rate of oxygen consumption (V̇o_{2 max}), and Po_2crit were determined. Dα increased in the following order: Xenopus muscle fibers Dα = 1.23 nM·mm²·mmHg⁻¹·s⁻¹ (SD 0.12), control rat trabeculae Dα = 2.29 nM·mm²·mmHg⁻¹·s⁻¹ (SD 0.24) (P = 0.0012 vs. Xenopus), and hypertrophied rat trabeculae Dα = 6.0 nM·mm²·mmHg⁻¹·s⁻¹ (SD 2.8) (P = 0.039 vs. control rat trabeculae). Dα increased with extracellular space in the preparation (Spearman’s rank correlation coefficient = 0.92, P < 0.001). The values for Dα indicate that Xenopus muscle fibers cannot reach V̇o_{2 max} in vivo because Po_2crit can be higher than arterial Po₂ and that hypertrophied rat cardiomyocytes can become hypoxic at the maximum heart rate.

The leading cause of mortality due to pulmonary arterial hypertension (PAH) is failure of the cardiac right ventricle. It has long been hypothesized that during the development of chronic cardiac failure the heart becomes energy deprived, possibly due to shortage of oxygen at the level of cardiomyocyte mitochondria. However, direct evaluation of oxygen tension levels within the in vivo right ventricle during PAH is currently lacking. Here we directly evaluated this hypothesis by using a recently reported technique of oxygen-dependent quenching of delayed fluorescence of mitochondrial protoprophyrin IX, to determine the distribution of mitochondrial oxygen tension (mitoPO2) within the right ventricle (RV) subjected to progressive PAH. PAH was induced through a single injection of monocrotaline (MCT). Control (saline-injected), compensated RV hypertrophy (30 mg/kg MCT; MCT30), and RV failure (60 mg/kg MCT; MCT60) rats were compared 4 wk after treatment. The distribution of mitoPO2 within the RV was determined in mechanically-ventilated, anaesthetized animals, applying different inspired oxygen (FiO2) levels and two increment dosages of dobutamine. MCT60 resulted in RV failure (increased mortality, weight loss, increased lung weight), MCT30 resulted in compensated RV hypertrophy. At 30% or 40% FiO2, necessary to obtain physiological arterial PO2 in the diseased animals, RV failure rats had significantly less mitochondria (15% of total mitochondria) in the 0-20 mmHg mitoPO2 range than hypertrophied RV rats (48%) or control rats (54%). Only when oxygen supply was reduced to 21% FiO2, resulting in low arterial PO2 for the MCT60 animals, or when oxygen demand increased with high dose dobutamine, the number of failing RV mitochondria with low oxygen became similar to control RV. In addition, metabolic enzyme analysis revealed similar mitochondrial mass, increased glycolytic hexokinase activity following MCT, with increased lactate dehydrogenase activity only in compensated hypertrophied RV. Our novel observation of increased mitochondrial oxygenation suggests down-regulation of in vivo mitochondrial oxygen consumption, in the absence of hypoxia, with transition towards right ventricular failure induced by pulmonary arterial hypertension.

The purpose of this study was to determine the myoglobin concentration in skeletal muscle fibers of chronic heart failure (CHF) patients and to calculate the effect of myoglobin on oxygen buffering and facilitated diffusion. Myoglobin concentration, succinate dehydrogenase (SDH) activity, and cross-sectional area of individual muscle fibers from the vastus lateralis of five control and nine CHF patients were determined using calibrated histochemistry. CHF patients compared with control subjects were similar with respect to myoglobin concentration: type I fibers 0.69 ± 0.11 mM (mean ± SD), type II fibers 0.52 ± 0.07 mM in CHF vs. type I fibers 0.70 ± 0.09 mM, type II fibers 0.49 ± 0.07 mM in control, whereas SDH activity was significantly lower in CHF in both fiber types ( P < 0.01). The myoglobin concentration in type I fibers was higher than in type II fibers ( P < 0.01). Consequently, the oxygen buffering capacity, calculated from myoglobin concentration/SDH activity was increased in CHF: type I fibers 11.4 ± 2.1 s, type II fibers 13.6 ± 3.9 s in CHF vs. type I fibers 7.8 ± 0.9 s, type II fibers 7.5 ± 1.0 s in control, all P < 0.01). The calculated extracellular oxygen tension required to prevent core anoxia (Po_2crit) in muscle fibers was similar when controls were compared with patients in type I fibers 10.3 ± 0.9 Torr in CHF and 11.5 ± 3.3 Torr in control, but was lower in type II fibers of patients 6.1 ± 2.8 Torr in CHF and 14.7 ± 6.2 Torr in control, P < 0.01. The lower Po_2crit of type II fibers may facilitate oxygen extraction from capillaries. Reduced exercise tolerance in CHF is not due to myoglobin deficiency.

Cytochrome c release from the intermembrane space of mitochondria is one of the triggers of apoptosis. There is no histochemical method available to demonstrate cytochrome c in cryostat sections, possibly because small cytosolic proteins diffuse readily into aqueous fixation media. This report shows that it is possible to demonstrate cytochrome c release in cardiomyocytes in failing myocardium using vapor fixation of cryostat sections and immunohistochemistry. The method is calibrated using sections from gelatin blocks containing known concentrations of cytochrome c. The method is applied to the hypertrophied right ventricular wall of rats in which pulmonary hypertension was induced by monocrotaline. Cytochrome c release is found in a fraction of the cardiomyocytes, leading to a mosaic-staining pattern. Cytochrome c release was found in myocytes over the full range of cross-sectional area (from 1 to 3.9 times control) in the hypertrophied myocardium. Cytosolic cytochrome c concentrations up to 0.4–0.5 mM occur frequently.