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Exercise training can increase both mitochondrial content and mitochondrial respiration. Despite its popularity, high-intensity exercise can be accompanied by mild acidosis (also present in certain pathological states), which may limit exercise-induced adaptations to skeletal muscle mitochondria. The aim of this study was to determine if administration of ammonium chloride (0.05 g/kg) to Wistar rats before each individual exercise session (5 high-intensity exercise sessions per week for eight weeks) reduced training-induced increases in mitochondrial content (measured by citrate synthase activity and protein content of electron transport system complexes) and respiration (measured in permeabilised muscle fibres). In the soleus muscle, the exercise-training-induced increase in mitochondrial respiration was reduced in rats administered ammonium chloride compared to control animals, but mitochondrial content was not altered. These effects were not present in the white gastrocnemius muscle. In conclusion, ammonium chloride administration before each exercise session over eight weeks reduced improvements in mitochondrial respiration in the soleus muscle but did not alter mitochondrial content. This suggests that mild acidosis may impact training-induced improvements in the respiration of mitochondria in some muscles.

The aim of this study was to investigate the relationship between mitochondrial content and respiratory function and whole-body insulin resistance in high-fat diet (HFD) fed rats. Male Wistar rats were given either a chow diet or an HFD for 12 weeks. After 4 weeks of the dietary intervention, half of the rats in each group began 8 weeks of interval training. In vivo glucose and insulin tolerance were assessed. Mitochondrial respiratory function was assessed in permeabilised soleus and white gastrocnemius (WG) muscles. Mitochondrial content was determined by the measurement of citrate synthase (CS) activity and protein expression of components of the electron transport system (ETS). We found HFD rats had impaired glucose and insulin tolerance but increased mitochondrial respiratory function and increased protein expression of components of the ETS. This was accompanied by an increase in CS activity in WG. Exercise training improved glucose and insulin tolerance in the HFD rats. Mitochondrial respiratory function was increased with exercise training in the chow-fed animals in soleus muscle. This exercise effect was absent in the HFD animals. In conclusion, exercise training improved insulin resistance in HFD rats but without changes in mitochondrial respiratory function and content. The lack of an association between mitochondrial characteristics and whole-body insulin resistance was reinforced by the absence of strong correlations between these measures. Our results suggest that improvements in mitochondrial respiratory function and content are not responsible for improvements in whole-body insulin resistance in HFD rats.

There are epidemiological associations between obesity and type 2 diabetes, cardiovascular disease and Alzheimer’s disease. While some common aetiological mechanisms are known, the role of amyloid beta 42 (Aβ42) in these diverse chronic diseases is obscure. Here we show that adipose tissue releases Aβ42, which is increased from adipose tissue of obese mice and is associated with higher plasma Aβ42. Increasing circulating Aβ42 levels in non-obese mice had no effect on systemic glucose homeostasis but had obesity-like effects on the heart, including reduced cardiac glucose clearance and impaired cardiac function. These effects on cardiac function were not observed when circulating levels of the closely related Aβ40 isoform were increased. Administration of an Aβ neutralising antibody prevented obesity-induced cardiac dysfunction and hypertrophy. Furthermore, Aβ neutralising antibody administration in established obesity prevented further deterioration of cardiac function. Multi-contrast transcriptomic analyses revealed that Aβ42 impacted pathways of mitochondrial metabolism and exposure of cardiomyocytes to Aβ42 inhibited mitochondrial function. These data reveal a role for systemic Aβ42 in the development of cardiac disease in obesity and suggest that therapeutics designed for Alzheimer’s disease could be effective in combating obesity-induced heart failure.

Autophagy is a key intracellular mechanism by which cells degrade old or dysfunctional proteins and organelles. In skeletal muscle, evidence suggests that exercise increases autophagosome content and autophagy flux. However, the exercise-induced response seems to differ between rodents and humans, and little is known about how different exercise prescription parameters may affect these results. The present study utilised skeletal muscle samples obtained from four different experimental studies using rats and humans. Here, we show that, following exercise, in the soleus muscle of Wistar rats, there is an increase in LC3B-I protein levels immediately after exercise (+109%), and a subsequent increase in LC3B-II protein levels 3 h into the recovery (+97%), despite no change in Map1lc3b mRNA levels. Conversely, in human skeletal muscle, there is an immediate exercise-induced decrease in LC3B-II protein levels (−24%), independent of whether exercise is performed below or above the maximal lactate steady state, which returns to baseline 3.5 h following recovery, while no change in LC3B-I protein levels or MAP1LC3B mRNA levels is observed. SQSTM1/p62 protein and mRNA levels did not change in either rats or humans following exercise. By employing an ex vivo autophagy flux assay previously used in rodents we demonstrate that the exercise-induced decrease in LC3B-II protein levels in humans does not reflect a decreased autophagy flux. Instead, effect size analyses suggest a modest-to-large increase in autophagy flux following exercise that lasts up to 24 h. Our findings suggest that exercise-induced changes in autophagosome content markers differ between rodents and humans, and that exercise-induced decreases in LC3B-II protein levels do not reflect autophagy flux level.

Assessments of mitochondrial respiration and mitochondrial content are common in skeletal muscle research and exercise science. However, many sources of technical and biological variation render these analyses susceptible to error. This study aimed to better quantify the reliability of different experimental designs and/or techniques so as to assist researchers to obtain more reliable data.We examined the repeatability of maximal mitochondrial oxidative phosphorylation in permeabilized muscle fibres via high-resolution respirometry, and citrate synthase activity (a biomarker for mitochondrial content) in a microplate with spectrophotometery.For mitochondrial respiration using permeabilized skeletal muscle fibres, the variability was reduced using three chambers and removing outliers compared to two chambers (CV reduced from 12.7% to 11.0%), and the minimal change that can be detected with 10 participants reduced from 17% to 13% according to modelling. For citrate synthase activity, the within-plate CV (3.5%) increased when the assay was repeated after 4 hours (CV = 10.2%) and 4 weeks (CV = 30.5%). The readings were correlated, but significantly different after 4 hours and 4 weeks.This research provides evidence for important technical considerations when measuring mitochondrial respiration and content using citrate synthase activity as a biomarker. When assessing mitochondrial respiration in human skeletal muscle, the technical variability of high-resolution respirometry can be reduced by increasing technical repeats and excluding outliers, practices which are not currently common. When analysing citrate synthase activity, our results highlight the importance of analysing all samples from the same study at the same time.