Hypoxia refines plasticity of mitochondrial respiration to repeated muscle work

Abstract

Purpose: We explored whether altered expression of factors tuning mitochondrial metabolism contributes to muscular adaptations with endurance training in the condition of lowered ambient oxygen concentration (hypoxia) and whether these adaptations relate to oxygen transfer as reflected by subsarcolemmal mitochondria and oxygen metabolism in muscle.

Methods: Male volunteers completed 30 bicycle exercise sessions in normoxia or normobaric hypoxia (4,000 m above sea level) at 65% of the respective peak aerobic power output. Myoglobin content, basal oxygen consumption, and re-oxygenation rates upon reperfusion after 8 min of arterial occlusion were measured in vastus muscles by magnetic resonance spectroscopy. Biopsies from vastus lateralis muscle, collected pre and post a single exercise bout, and training, were assessed for levels of transcripts and proteins being associated with mitochondrial metabolism.

Results: Hypoxia specifically lowered the training-induced expression of markers of respiratory complex II and IV (i.e. SDHA and isoform 1 of COX-4; COX4I1) and preserved fibre cross-sectional area. Concomitantly, trends (p < 0.10) were found for a hypoxia-specific reduction in the basal oxygen consumption rate, and improvements in oxygen repletion, and aerobic performance in hypoxia. Repeated exercise in hypoxia promoted the biogenesis of subsarcolemmal mitochondria and this was co-related to expression of isoform 2 of COX-4 with higher oxygen affinity after single exercise, de-oxygenation time and myoglobin content (r ≥ 0.75). Conversely, expression in COX4I1 with training correlated negatively with changes of subsarcolemmal mitochondria (r < -0.82).

Conclusion: Hypoxia-modulated adjustments of aerobic performance with repeated muscle work are reflected by expressional adaptations within the respiratory chain and modified muscle oxygen metabolism.

Fig. 1

Experimental design and assessed parameters. a Scheme depicting the outline of the experiments. For details see the “Methods” section. Subjects experienced an entry test during which anthropometry and oxygen metabolism in m. vastus lateralis at rest, and aerobic performance in hypoxia or normoxia were assessed, and a resting muscle biopsy was collected from vastus lateralis. A week later subjects carried out an endurance exercise test at the respective training condition (i.e. warm-up followed by 30-min bicycle exercise at 65 % of the respective P _max) during which biopsies were collected 24 h after the exercise bout. Subjects then entered an endurance-training protocol with 30 exercise sessions of 30-min on a stationary bicycle at 65 % P _max in normoxia or normobaric hypoxia equivalent to 4,000 m above sea level. This was followed by a post-training biopsy and an exit test essentially repeating the entry test. b Scheme depicting the localization of assessed factors of mitochondrial metabolism in muscle fibres. Continuous arrows indicate the flow of metabolic processes. Boxes highlighted by black filling with text in white font reflect the two respiratory complexes where hypoxia-dependent regulation is identified. Protein and transcript species being assessed are indicated with underlined font and in italics, respectively. AcCoA acetyl coenzyme A, B-OX beta oxidation, CO I–CO V complex I to complex V of the mitochondrial respiration chain. Further names are defined in the list of abbreviations. c dMb signals from 2 independent receive coils as determined by 1H MRS in function of time and as consequence of applying a blood pressure cuff placed on the thigh in a one subject before the start of exercise. Circles and the connecting grey line represent data points filtered as a moving average of 3 original fitted points. The black line is the result of fitting this signal with a model for a delayed linear increasing signal, followed by a plateau and an exponential recovery upon release of the air cuff

Fig. 2

Hypoxia-specific transcript expression after the exercise bout. Bar graph visualizing the mean fold changes + standard error (SE) of 13 selected gene transcripts in vastus lateralis muscle of the untrained men 24 h into recovery from the bout of endurance exercise in normoxia or hypoxia, respectively, vs. pre-exercise levels. ^# p < 0.05 for the interaction effect between the repeated factor pre/post-exercise × co-stimulus (normoxia/hypoxia, repeated ANOVA, n = 5–6). *p < 0.05 and ⁺0.05 ≤ p < 0.10 for post vs. pre exercise differences (post hoc test of Fisher)

Fig. 3

Hypoxia-modulated adaptations in muscle protein and oxygen metabolism to endurance training. a Examples of immunoblots for mitochondrial proteins in vastus lateralis muscle of a subject pre- and post-endurance training in normoxia and hypoxia, respectively. The abbreviated name and molecular weight of the detected proteins are indicated. The Ponceau S stained membrane before immunoblotting is shown below each blot to visualize protein loading based on the stained band corresponding to actin. b Bar graph visualizing the mean fold changes + SE of protein (activity) levels in m. vastus lateralis after 6 weeks of endurance training in normoxia (n = 6) or hypoxia (n = 5) training. ^# p < 0.05 and ^$0.05 ≤ p < 0.10, respectively, for the interaction between the ‘pre/post-exercise’ ×  ‘co-stimulus’ (repeated ANOVA). * and ⁺ denote p < 0.05 for values post vs. pre training (post hoc test of Fisher)

Fig. 4

Interrelationships for hypoxia-specific adjustments with exercise. Matrix visualising the correlations between factors demonstrating hypoxia-specific regulation with single endurance exercise and training. Fold changes which significance exceeded the threshold of |r| > 0.7 and p < 0.05 are shown in colour coding (red up, blue down). White colour refers to values below the threshold for significance (|r| < 0.70). Grey symbolises suppressed values. Correlations that are deemed of interest are circled. Double represented relationships are once circled in grey (colour figure online)