Yutaka Matsunaga

Postexercise whole body heat stress additively enhances endurance training-induced mitochondrial adaptations in mouse skeletal muscle

A recent study demonstrated that heat stress induces mitochondrial biogenesis in C2C12 myotubes, thereby implying that heat stress may be an effective treatment to enhance endurance training-induced mitochondrial adaptations in skeletal muscle. However, whether heat stress actually induces mitochondrial adaptations in skeletal muscle in vivo is unclear. In the present study, we report the novel findings that 1) whole body heat stress produced by exposure of ICR mice to a hot environment (40°C, 30 min/day, 5 days/wk, 3 wk) induced mitochondrial adaptations such as increased mitochondrial enzyme activity (citrate synthase and 3-hydroxyacyl CoA dehydrogenase) and respiratory chain protein content (complexes I-V) in skeletal muscle in vivo and 2) postexercise whole body heat stress additively enhanced endurance training-induced mitochondrial adaptations (treadmill running, 25 m/min, 30 min/day, 5 days/wk, 3 wk). Moreover, to determine the candidate mechanisms underlying mitochondrial adaptations, we investigated the acute effects of postexercise whole body heat stress on the phosphorylation status of cellular signaling cascades that subsequently induce mitochondrial gene transcription. We found that whole body heat stress boosted the endurance exercise-induced phosphorylation of p38 MAPK, increased the phosphorylation status of p70S6K, a biomarker of mammalian target of rapamycin complex 1 activity, and unexpectedly dephosphorylated AMP-activated protein kinase and its downstream target acetyl-CoA carboxylase in skeletal muscle. Our present observations suggest that heat stress can act as an effective postexercise treatment. Heat stress treatment appeared to be clinically beneficial for people who have difficulty participating in sufficient exercise training, such as the elderly, injured athletes, and patients.

Daily heat stress treatment rescues denervation-activated mitochondrial clearance and atrophy in skeletal muscle

Traumatic nerve injury or nerve disease leads to denervation and severe muscle atrophy. Recent evidence shows that mitochondrial loss could be a key mediator of skeletal muscle atrophy. Here, we show that daily heat stress treatment rescues denervation‐induced loss of mitochondria and concomitant muscle atrophy. We also found that denervation‐activated autophagy‐dependent mitochondrial clearance (mitophagy) was suppressed by daily heat stress treatment. The molecular basis of this observation is explained by our results showing that heat stress treatment attenuates the increase of key proteins that regulate the tagging step for mitochondrial clearance and the intermediate step of autophagosome formation in denervated muscle. These findings contribute to the better understanding of mitochondrial quality control in denervated muscle from a translational perspective and provide a mechanism behind the attenuation of muscle wasting by heat stress.

Effects of decreased lactate accumulation after dichloroacetate administration on exercise training–induced mitochondrial adaptations in mouse skeletal muscle

Recent studies suggested that lactate accumulation can be a signal for mitochondrial biogenesis in skeletal muscle. We investigated whether reductions in lactate concentrations in response to dichloroacetate (DCA), an activator of pyruvate dehydrogenase, attenuate mitochondrial adaptations after exercise training in mice. We first confirmed that DCA administration (200 mg/kg BW by i.p. injection) 10 min before exercise decreased muscle and blood lactate concentrations after high‐intensity interval exercise (10 bouts of 1 min treadmill running at 40 m/min with a 1 min rest). At the same time, exercise‐induced signal cascades did not change by pre‐exercise DCA administration. These results suggested that DCA administration affected only lactate concentrations after exercise. We next examined the effects of acute DCA administration on mRNA expressions involved with mitochondrial biogenesis after same high‐intensity interval exercise and the effects of chronic DCA administration on mitochondrial adaptations after high‐intensity interval training (increasing intensity from 38 to 43 m/min by the end of training period). Acute DCA administration did not change most of the exercise‐induced mRNA upregulation. These data suggest that lactate reductions by DCA administration did not affect transcriptional activation after high‐intensity interval exercise. However, chronic DCA administration attenuated, in part, mitochondrial adaptations such as training‐induced increasing rates of citrate synthase (P = 0.06), β‐hydroxyacyl CoA dehydrogenase activity (P < 0.05), cytochrome c oxidase IV (P < 0.05) and a fatty acid transporter, fatty acid translocase/CD36 (P < 0.05), proteins after exercise training. These results suggest that lactate accumulation during high‐intensity interval exercise may be associated with mitochondrial adaptations after chronic exercise training.

Effects of Heat Stress Treatment on Age-dependent Unfolded Protein Response in Different Types of Skeletal Muscle

Mitochondrial and endoplasmic reticulum (ER) stress, and subsequently activated responses (mitochondrial/ER unfolded protein responses; UPRmt/UPRER), are involved in the pathogenesis of sarcopenia. To extend both basic and translational knowledge, we examined (i) whether age-induced mitochondrial and ER stress depend on skeletal muscle type in mice and (ii) whether heat stress treatment, a suggested strategy for sarcopenia, improves age-induced mitochondrial and ER stress. Aged (21-month-old) mice showed more severe mitochondrial stress and UPRmt than young (12-week-old) mice, based on increased oxidative stress, mitochondrial proteases, and mitochondrial E3 ubiquitin ligase. The aged mice also showed ER stress and UPRER, based on decreased ER enzymes and increased ER stress-related cell death. These changes were much more evident in soleus muscle than in gastrocnemius and plantaris muscles. After daily heat stress treatment (40 °C chamber for 30 minutes per day) for 4 weeks, mice showed remarkable improvements in age-related changes in soleus muscle. Heat stress had only minor effects in gastrocnemius and plantaris muscles. Based on these findings, age-associated mitochondrial stress, ER stress, and UPRmt/ER vary qualitatively with skeletal muscle type. Our results suggest a molecular basis for the beneficial effects of heat stress on muscle atrophy with age in soleus muscle.

Comparison between pre–exercise casein peptide and intact casein supplementation on glucose tolerance in high–fat diet–fed mice

We hypothesized that along with exercise, casein peptide supplementation would have a higher impact on improving glucose tolerance than intact casein. Male 6-week-old ICR mice were provided a high-fat diet to induce obesity and glucose intolerance. The mice were randomly divided into 4 treatment groups: control (Con), endurance training (Tr), endurance training with intact casein supplementation (Cas+Tr), and endurance training with casein peptide supplementation (CP+Tr). The mice in each group were orally administrated water, intact casein, or casein peptide (1.0 mg/g body weight, every day), and then subjected to endurance training (15-25 m/min, 60 min, 5 times/week for 4 weeks) on a motor-driven treadmill 30 min after ingestion. Our results revealed that total intra-abdominal fat was significantly lower in CP+Tr than in Con (p < 0.05). Following an oral glucose tolerance test, the blood glucose area under the curve (AUC) was found to be significantly smaller for CP+Tr than for Con (p < 0.05). Moreover, in the soleus muscle, glucose transporter 4 (GLUT4) protein levels were significantly higher in CP+Tr than in Con (p < 0.01). However, intra-abdominal fat, blood glucose AUC, and GLUT4 protein content in the soleus muscle did not alter in Tr and Cas+Tr when compared with Con. These observations suggest that pre-exercise casein peptide supplementation has a higher effect on improving glucose tolerance than intact casein does in mice fed a high-fat diet.

Effects of Glucose with Casein Peptide Supplementation on Post-Exercise Muscle Glycogen Resynthesis in C57BL/6J Mice

Numerous studies have reported that post-exercise ingestion of carbohydrates with protein supplementation can enhance glycogen recovery. However, few reports have focused on the degrees of degradation of the ingested proteins due to post-exercise glycogen resynthesis. Accordingly, the aim of this study was to clarify the effects of differences in protein degradation on muscle glycogen recovery. Male seven-week-old C57BL/6J mice performed a single bout of 60-min treadmill running exercise and were then orally administered glucose (Glu; 1.5 mg/g body weight (BW)), glucose with casein peptide (Glu + Pep; 1.5 + 0.5 mg/g BW) or its constituent amino acid mixture (Glu + AA; 1.5 + 0.5 mg/g BW). At 120 min after supplementation, the soleus muscle glycogen content in the Glu and Glu + AA groups was significantly higher than that immediately after exercise; however, no such difference was observed in the Glu + Pep group. Blood substrate concentration and insulin signaling did not differ among the three groups. Furthermore, energy expenditure during the recovery period in the Glu + Pep group was significantly higher than that in the Glu and Glu + AA groups. These findings suggest that post-exercise co-ingestion of glucose and casein peptide might delay glycogen resynthesis, at least in part through increased energy expenditure caused by casein peptide ingestion.

Pre-Exercise High-Fat Diet for 3 Days Affects Post-Exercise Skeletal Muscle Glycogen Repletion

Previous studies have shown that the short-term intake of a high-fat diet (HFD) impairs glucose metabolism. In this study, we investigated the influences of pre-exercise HFD intake for 3 d on post-exercise glycogen repletion in skeletal muscle in ICR mice. Mice received either an HFD (57% kcal from fat, 23% kcal from carbohydrate; HFD group) or standard laboratory chow (13% kcal from fat, 60% kcal from carbohydrate; Con group) for 3 d before exercise. Mice performed treadmill running at 25 m/min for 60 min and were orally administered a glucose (2 mg/g body weight) solution immediately after and at 60 min after exercise. A negative main effect of pre-exercise HFD intake was observed for skeletal muscle glycogen concentration from the pre-exercise phase to 120 min of post-exercise recovery (p<0.01). Blood glucose concentration in the HFD group was significantly higher than in the Con group at 120 min after exercise (p<0.01). No significant difference was observed in plasma insulin concentration. There were no significant between-group differences in the phosphorylation state of Akt Thr308, AMPK Thr172, AS160 Thr642, or glycogen synthase Ser641 or in glucose transporter 4 protein levels during post-exercise recovery. Our results suggest that the intake of a pre-exercise HFD for 3 d affects post-exercise glycogen repletion in skeletal muscle without impairing the insulin signaling cascade.