Voluntary Wheel Running In Mice Research Articles

Voluntary wheel running and lithium supplementation promotes fatigue resistance, fat oxidation, and improves insulin tolerance in D2 mdx mice

BACKGROUND: Duchenne muscular dystrophy (DMD) is an X-linked disorder characterized by progressive muscle wasting and premature death. Many DMD patients will also exhibit signs of metabolic dysfunction such as insulin resistance that predisposes them to type 2 diabetes and worsened cardiovascular disease. Most treatments for DMD aim to lower muscle inflammation and improve strength, however, none have addressed the complication of insulin resistance. The current standard of treatment is corticosteroids, which prolongs ambulation, however, chronic use increases susceptibility to obesity, insulin resistance, and type 2 diabetes. It is well-established that regular aerobic exercise enhances insulin sensitivity. Further, we have recently found that inhibiting glycogen synthase kinase 3 with lithium (Li), a known insulin mimetic, improves force production and fatigue resistance in wildtype (WT) mice. Here, we tested whether voluntary wheel running (VWR) and Li supplementation, together, could improve whole-body fatigue, metabolism, and insulin sensitivity in the DBA/2J (D2) mdx mouse model of DMD. METHODS: 5-week old male D2 WT and mdx mice were separated into four groups: WT, mdx sedentary (SED), mdx VWR, and mdx Li+VWR (n=11/group). The mdx VWR and mdx Li+VWR mice had unlimited access to a cagewheel, and mdx Li+VWR mice were given a low dose of Li (50 mg/kg/day) via their drinking water throughout the study. A treadmill time-to-fatigue test was performed to assess whole-body fatigue. Mice were housed for 48-hours in metabolic cages to measure energy expenditure. Glucose handling was measured using glucose and insulin tolerance tests. RESULTS: As expected, WT mice had ~2-fold greater total cage activity compared to all mdx groups. When examining cagewheel distance, we found that mdx Li+VWR mice ran half the total distance in kilometres than mdx VWR mice (39.1 ± 9.6 km vs. 74.8 ± 13.6 km). Despite this, VWR with and without Li improved whole-body fatigue as only mdx SED mice (22.8 ± 0.9 min) had a shorter time to exhaustion than WT mice (32.6 ± 0.9 min). All mdx groups had higher energy expenditure than WT mice, suggestive of a hypermetabolic phenotype. When examining the respiratory exchange ratio (RER) among mdx mice, mdxLi+VWR (0.87) mice had a lower RER than mdx SED mice (0.91), indicative of enhanced fat oxidation. Glucose tolerance was equally impaired among mdx groups compared to WT mice. However, insulin tolerance was notably improved in mdx VWR and mdx Li+VWR mice, with a greater improvement observed in the mdx Li+VWR mice. CONCLUSION: Despite running less than mdx VWR mice, mdx Li+VWR mice had enhanced fatigue resistance, fat utilization, and insulin sensitivity, suggesting Li may be a viable treatment to improve muscle and metabolic function in mdx mice. CIHR CRC (Tier II) to VAF This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Resolution of cisplatin-induced fatigue does not require endogenous interleukin-10 in male mice

Based on previous results showing a pivotal role of endogenous interleukin-10 (IL-10) in the recovery from cisplatin-induced peripheral neuropathy, the present experiments were carried out to determine whether this cytokine plays any role in the recovery from cisplatin-induced fatigue in male mice. Fatigue was measured by decreased voluntary wheel running in mice trained to run in a wheel in response to cisplatin. Mice were treated with a monoclonal neutralizing antibody (IL-10na) administered intranasally during the recovery period to neutralize endogenous IL-10. In the first experiment, mice were treated with cisplatin (2.83 mg/kg/day) for five days and IL-10na (12 μg/day for three days) five days later. In the second experiment, they were treated with cisplatin (2.3 mg/kg/day for 5 days twice at a five-day interval) and IL10na (12 μg/day for three days) immediately after the last injection of cisplatin. In both experiments, cisplatin decreased body weight and reduced voluntary wheel running. However, IL-10na did not impair recovery from these effects. These results show that the recovery from the cisplatin-induced decrease in wheel running does not require endogenous IL-10 in contrast to the recovery from cisplatin-induced peripheral neuropathy

The Effects of Thermoneutral Housing on GSK3 activation and PGC‐1alpha Muscle Content After Voluntary Wheel Running in Mice

Background Glycogen synthase kinase-3 (GSK-3) is a serine/threonine protein kinase constitutively active at rest. GSK-3 is known to regulate muscle glucose homeostasis and PGC-1α expression. Studies have shown that acute and chronic muscle activation can inhibit GSK-3 by promoting inhibitory serine9 phosphorylation; and this inhibition may mediate some of the beneficial effects of regular exercise including enhanced fatigue resistance. In light of recent evidence demonstrating the importance of housing mice at thermoneutrality to better mimic human metabolism and physiology, we investigated the protein levels of phosphorylated/total GSK3 and PGC-1a in exercised and sedentary mice housed within their thermoneutral zone (TN, 29°C) or at room temperature (RT, 22°C). Methods Tricep muscles were obtained as part of another study conducted at the University of Guelph. Briefly, eight-week old male mice (Charles River, QC, Canada) were purchased and acclimated to RT or TN for two weeks. Mice were then weight-matched into sedentary (SED) or voluntary wheel running (VWR) groups, where mice had free access to a running wheel for up to 6 weeks. Following the intervention, mice were euthanized and tricep muscles were collected. Western blot analyses were done to examine protein levels of phosphorylated (Ser9) GSK3, total GSK3, and PGC-1a. Pearson's correlational analyses were conducted to examine the associations between total GSK3 and PGC-1a and phosphorylated:total GSK3 and PGC-1a. Results Our results indicate that at RT GSK3 content was significantly reduced (-38%) with VWR, which led to a 1.5-fold increase in the Ser9 phosphorylated:total GSK3 ratio. This suggests that when housed at RT, VWR led to lower GSK3 activation. However, when housed at TN, these effects were not observed. When examining, PGC-1α content, we found a significant elevation in both RT (2.3-fold) and TN (3.5-fold) VWR groups. Correlational analyses detected a negative association between total GSK3 content and PGC-1α (r = -0.45, p = 0.03). Conversely, we found a positive association between phosphorylated:total GSK3 and PGC-1α (r = 0.45, p = 0.04) suggesting that mice with greater phosphorylated GSK3, and thus lower activation, had greater levels of PGC-1α. Conclusions Together our data suggests that VWR inhibits GSK3 in muscle when mice are housed at RT but not at TN. Though PGC-1α was elevated in both VWR conditions, our correlational analyses reinforce the link between muscle GSK3 content/activation and PGC-1α. In the future, it would be of interest to determine the impact of combined VWR and GSK3 inhibition in mice.

Endurance Exercise Training Modes to Improve Physical Function in Older Mice: HIIT vs. VWR

With age, we experience a progressive loss of physical function. Exercise is a promising therapy to preserve muscle health and functional aptitude. Animal models are needed to examine the underlying molecular mechanisms at the intersection of aging, exercise, and functional decline. In this study, we compare the efficacy of two types of individualized endurance exercise training in older C57BL/6 mice (26-months old at completion): HIIT (high intensity interval training on a treadmill, n=10) and VWR (voluntary wheel running, n=8). We hypothesized that while both exercises would improve function, HIIT would promote more extensive adaptation. For four months the VWR mice ran 4 days/week and the HIIT group ran 3x/week. We determined function pre/post-training by utilizing our composite scoring system, the Comprehensive Functional Assessment Battery (CFAB). CFAB consists of the following: treadmill running (endurance), rotarod (overall motor function), wheel running (volitional exercise rate/activity), grip strength (fore-limb strength), and inverted cling (overall strength/endurance). EchoMRI determined body composition. After training, we found significant CFAB improvement (repeated measures t-test, p<0.05) in both exercise groups, specifically including: rotarod (+37%, HIIT and VWR); treadmill (+61% VWR; +58% HIIT), grip strength (+20% VWR), body mass (-17% VWR, -10% HIIT), and fat percentage (-44% VWR, -20% HIIT). Contrary to our hypothesis, HIIT did not improve function more than VWR, though we suspect increasing training intensity would improve response. Thus, future studies will need to address defining HIIT dose response and optimal training volume for older mice. We conclude that our models will be useful for future mechanistic investigations.

Recruitment of parvalbumin and somatostatin interneuron inputs to adult born dentate granule neurons

GABA is a key regulator of adult-born dentate granule cell (abDGC) maturation so mapping the functional connectivity between abDGCs and local interneurons is required to understand their development and integration into the hippocampal circuit. We recorded from birthdated abDGCs in mice and photoactivated parvalbumin (PV) and somatostatin (SST) interneurons to map the timing and strength of inputs to abDGCs during the first 4 weeks after differentiation. abDGCs received input from PV interneurons in the first week, but SST inputs were not detected until the second week. Analysis of desynchronized quantal events established that the number of GABAergic synapses onto abDGCs increased with maturation, whereas individual synaptic strength was constant. Voluntary wheel running in mice scaled the GABAergic input to abDGCs by increasing the number of synaptic contacts from both interneuron types. This demonstrates that GABAergic innervation to abDGCs develops during a prolonged post-mitotic period and running scales both SST and PV synaptic afferents.

Cardiac up-regulation of NBCe1 emerges as a beneficial consequence of voluntary wheel running in mice

Physical training stimulates the development of physiologic cardiac hypertrophy (CH), being a key event in this process the inhibition of the Na+/H+ exchanger. However, the role of the sodium bicarbonate cotransporter (NBC) has not been explored yet under this circumstance. C57/Bl6 mice were allowed to voluntary exercise (wheel running) for five weeks. Cardiac mass was evaluated by echocardiography and histomorphometry detecting that training promoted the development of physiological CH (heart weight/tibia length ratio, mg/mm: 6.54 ± 0.20 vs 8.81 ± 0.24; interstitial collagen content, %: 3.14 ± 0.63 vs. 1.57 ± 0.27; and cross-sectional area of cardiomyocytes, μm2: 200.6 ± 8.92 vs. 281.9 ± 24.05; sedentary (Sed) and exercised (Ex) mice, respectively). The activity of the electrogenic isoform of the cardiac NBC (NBCe1) was estimated by recording intracellular pH under high potassium concentration and by measuring action potential duration (APD). NBCe1 activity was significantly increased in isolated cardiomyocytes of trained mice. Additionally, the APD was shorter and the alkalization due to high extracellular potassium-induced depolarization was greater in this group, indicating that the NBCe1 was hyperactive. These results are online with the observed myocardial up-regulation of the NBCe1 (Western Blot, %: 100 ± 13.86 vs. 202 ± 29.98; Sed vs. Ex, n = 6 each group). In addition, we detected a reduction in H2O2 production in the myocardium of trained mice.These results support that voluntary training induces the development of physiologic CH with up-regulation of the cardiac NBCe1 in mice. Furthermore, the improvement in the antioxidant capacity contributes to the beneficial cardiovascular consequences of physical training.

Chronic Environmental or Genetic Elevation of Galanin in Noradrenergic Neurons Confers Stress Resilience in Mice

The neuropeptide galanin has been implicated in stress-related neuropsychiatric disorders in humans and rodent models. While pharmacological treatments for these disorders are ineffective for many individuals, physical activity is beneficial for stress-related symptoms. Galanin is highly expressed in the noradrenergic system, particularly the locus coeruleus (LC), which is dysregulated in stress-related disorders and activated by exercise. Galanin expression is elevated in the LC by chronic exercise, and blockade of galanin transmission attenuates exercise-induced stress resilience. However, most research on this topic has been done in rats, so it is unclear whether the relationship between exercise and galanin is species specific. Moreover, use of intracerebroventricular (ICV) galanin receptor antagonists in prior studies precluded defining a causal role for LC-derived galanin specifically. Therefore, the goals of this study were twofold. First, we investigated whether physical activity (chronic wheel running) increases stress resilience and galanin expression in the LC of male and female mice. Next, we used transgenic mice that overexpress galanin in noradrenergic neurons (Gal OX) to determine how chronically elevated noradrenergic-derived galanin, alone, alters anxiogenic-like responses to stress. We found that three weeks of ad libitum access to a running wheel in their home cage increased galanin mRNA in the LC of mice, which was correlated with and conferred resilience to stress. The effects of exercise were phenocopied by galanin overexpression in noradrenergic neurons, and Gal OX mice were resistant to the anxiogenic effect of optogenetic LC activation. These findings support a role for chronically increased noradrenergic galanin in mediating resilience to stress.SIGNIFICANCE STATEMENT Understanding the neurobiological mechanisms underlying behavioral responses to stress is necessary to improve treatments for stress-related neuropsychiatric disorders. Increased physical activity is associated with stress resilience in humans, but the neurobiological mechanisms underlying this effect are not clear. Here, we investigate a potential causal mechanism of this effect driven by the neuropeptide galanin from the main noradrenergic nucleus, the locus coeruleus (LC). We show that chronic voluntary wheel running in mice increases stress resilience and increases galanin expression in the LC. Furthermore, we show that genetic overexpression of galanin in noradrenergic neurons causes resilience to a stressor and the anxiogenic effects of optogenetic LC activation. These findings support a role for chronically increased noradrenergic galanin in mediating resilience to stress.

Late-life voluntary wheel running reverses age-related aortic stiffness in mice: a translational model for studying mechanisms of exercise-mediated arterial de-stiffening.

Aortic stiffening, assessed as pulse-wave velocity (PWV), increases with age and is an important antecedent to, and independent predictor of, cardiovascular diseases (CVD) and other clinical disorders of aging. Aerobic exercise promotes lower levels of aortic stiffness in older adults, but the underlying mechanisms are incompletely understood, largely due to inherent challenges of mechanistic studies of large elastic arteries in humans. Voluntary wheel running (VWR) is distinct among experimental animal exercise paradigms in that it allows investigation of the physiologic effects of aerobic training without potential confounding influences of aversive molecular signaling related to forced exercise. In this study, we investigated whether VWR in mice may be a suitable model for mechanistic studies (i.e., "reverse translation") of the beneficial effects of exercise on arterial stiffness in humans. We found that 10weeks of VWR in old mice (~ 28months) reversed age-related elevations in aortic PWV assessed in vivo (Old VWR: 369 ± 19 vs. old sedentary: 439 ± 20cm/s, P < 0.05). The de-stiffening effects of VWR were accompanied by normalization of age-related increases in ex vivo mechanical stiffness of aortic segments and aortic accumulation of collagen-I and advanced glycation end products, as well as lower levels of aortic superoxide and nitrotyrosine. Our results suggest that late-life VWR in mice recapitulates the aortic de-stiffening effects of exercise in humans and indicates important mechanistic roles for decreased oxidative stress and extracellular matrix remodeling. Therefore, VWR is a suitable model for further study of the mechanisms underlying beneficial effects of exercise on arterial stiffness.

Sex-specific alterations in whole body energetics and voluntary activity in heterozygous R163C malignant hyperthermia-susceptible mice.

Malignant hyperthermia (MH) is characterized by induction of skeletal muscle hyperthermia in response to a dysregulated increase in myoplasmic calcium. Although altered energetics play a central role in MH, MH‐susceptible humans and mouse models are often described as having no phenotype until exposure to a triggering agent. The purpose of this study was to determine the influence of the R163C ryanodine receptor 1 mutation, a common MH mutation in humans, on energy expenditure, and voluntary wheel running in mice. Energy expenditure was measured by indirect respiration calorimetry in wild‐type (WT) and heterozygous R163C (HET) mice over a range of ambient temperatures. Energy expenditure adjusted for body weight or lean mass was increased (P < .05) in male, but not female, HET mice housed at 22°C or when housed at 28°C with a running wheel. In female mice, voluntary wheel running was decreased (P < .05) in the HET vs WT animals when analyzed across ambient temperatures. The thermoneutral zone was also widened in both male and female HET mice. The results of the study show that the R163C mutations alters energetics even at temperatures that do not typically induce MH.

Neural mechanisms underlying regulation of motivation for voluntary wheel running in mice

Neural mechanisms underlying regulation of motivation for voluntary wheel running in mice

Blood Pressure Normalization-Independent Cardioprotective Effects of Endogenous, Physical Activity-Induced αCGRP (α Calcitonin Gene-Related Peptide) in Chronically Hypertensive Mice.

αCGRP (α calcitonin gene-related peptide), one of the strongest vasodilators, is cardioprotective in hypertension by reducing the elevated blood pressure. However, we hypothesize that endogenous, physical activity-induced αCGRP has blood pressure-independent cardioprotective effects in chronic hypertension. Chronically hypertensive (one-kidney-one-clip surgery) wild-type and αCGRP-/- sedentary or voluntary wheel running mice were treated with vehicle, αCGRP, or the αCGRP receptor antagonist CGRP8-37. Cardiac function and myocardial phenotype were evaluated echocardiographically and by molecular, cellular, and histological analysis, respectively. Blood pressure was similar among all hypertensive experimental groups. Endogenous αCGRP limited pathological remodeling and heart failure in sedentary, chronically hypertensive wild-type mice. In these mice, voluntary wheel running significantly improved myocardial phenotype and function, which was abolished by CGRP8-37 treatment. In αCGRP-/- mice, αCGRP treatment, in contrast to voluntary wheel running, improved myocardial phenotype and function. Specific inhibition of proliferation and myofibroblast differentiation of primary, murine cardiac fibroblasts by αCGRP suggests involvement of these cells in αCGRP-dependent blunting of pathological cardiac remodeling. Endogenous, physical activity-induced αCGRP has blood pressure-independent cardioprotective effects and is crucial for maintaining cardiac function in chronic hypertension. Consequently, inhibiting endogenous αCGRP signaling, as currently approved for migraine prophylaxis, could endanger patients with hypertension.

Comparative molecular analysis of endurance exercise in vivo with electrically stimulated in vitro myotube contraction.

Exercise has positive effects on health and improves a variety of disease conditions. An in vitro model of exercise has been developed to better understand its molecular mechanisms. While various conditions have been used to mimic in vivo exercise, no specific conditions have matched a specific type of in vivo exercise. Here, we screened various electrical pulse stimulation (EPS) conditions and compared the molecular events under each condition in myotube culture with that obtained under voluntary wheel running (VWR), a mild endurance exercise, in mice. Both EPS and VWR upregulated the mRNA levels of genes involved in the slow-type twitch (Myh7 and Myh2) and myogenesis (Myod and Myog) and increased the protein expression of peroxisome proliferator-activated receptor-γ coactivator-1α, which is involved in mitochondrial biogenesis. These changes were accompanied by activation of p38 and AMPK. However, neither condition induced the expression of muscle-specific E3 ligases such as MAFbx and MuRF1. Both EPS and VWR consistently induced antioxidant genes such as Sod3 and Gpx4 but did not cause similar changes in the expression levels of the calcium channel/pump-related genes Ryr and Serca. Furthermore, both EPS and VWR reduced glycogen levels but not lactate levels as assessed in post-EPS culture medium and post-VWR serum, respectively. Thus we identified an in vitro EPS condition that effectively mimics VWR in mice, which can facilitate further studies of the detailed molecular mechanisms of endurance exercise in the absence of interference from multiple tissues and organs.NEW & NOTEWORTHY This study establishes an optimal condition for electrical pulse stimulation (EPS) in myotubes that shows a similar molecular signature as voluntary wheel running. The specific EPS condition 1) upregulates the mRNA of slow-twitch muscle components and myogenic transcription factors, 2) induces antioxidant genes without any muscle damage, and 3) promotes peroxisome proliferator-activated receptor-γ coactivator-1α and its upstream regulators involved in mitochondrial biogenesis.

Voluntary exercise in mice fed an obesogenic diet alters the hepatic immune phenotype and improves metabolic parameters \u2013 an animal model of life style intervention in NAFLD

Reproducible animal models to recapitulate the pathophysiology of non-alcoholic fatty liver disease (NAFLD) are urgently required to improve the understanding of the mechanisms of liver injury and to explore novel therapeutic options. Current guidelines recommend life-style interventions as first-line therapy for NAFLD and these types of intervention are considered standard-of-care. The current study establishes a reproducible mouse model of a life-style intervention in NAFLD using voluntary wheel running (VWR). Male C57BL/6J mice were fed a high-fat, high-carbohydrate diet (HFD) to induce NAFLD or a corresponding control diet for 12 weeks. Starting at week 9 of the obesogenic NAFLD diet, mice were randomized to either free access to a running wheel or being single caged resembling a sedentary (SED) life-style. VWR induced a transient weight reduction in HFD-fed mice up until week 10. In contrast to the SED mice, VWR mice exhibited normal ALT at the end of the intervention, while the metabolic alterations including elevated fasting glucose, insulin, triglyceride, and total cholesterol levels remained almost unchanged. Additionally, VWR prevented HFD-induced hepatic steatosis by alterations in key liver metabolic processes including the induction of fatty acid β-oxidation and lipogenesis inhibition following increased AMP-activated protein kinase (AMPK)-α activity. Phosphorylation of the serine kinase Akt in hepatic tissue was enhanced following VWR. Furthermore, VWR mice were protected from HFD-induced expression of pro-inflammatory cytokines, chemokines and liver macrophage infiltration. The SED/HFD group exhibited increasing activity of hepatic nuclear factor (NF)-κB p65, which was absent following exercise in the VWR/HFD group. In summary, in an obesogenic mouse model of NAFLD physical exercise improves fatty acid and glucose homeostasis and protects from macrophage-associated hepatic inflammation.

Voluntary wheel running: patterns and physiological effects in mice.

Exercise can prevent and improve the pathophysiology of diseases and promote healthy aging. Thus, understanding the mechanisms that regulate the beneficial effects of exercise may lead to the development of new strategies to enhance quality of life and to counteract chronic diseases. Voluntary wheel running is an interesting model to study the effects of exercise in mice. Compared to forced treadmill exercise, voluntary wheel running presents several advantages such as: 1) running pattern is similar to natural running behavior of mice; 2) it is performed under non-stressed conditions, according to the rhythmicity of the animal; 3) it does not require direct interference from the researcher, and can be easily applied in long-term studies. Mice run spontaneously when given access to running wheels, for a total distance of ∼4 to 20 km per day and a total activity time of ∼3 to 7 hours a day. Hence, voluntary wheel running can result in robust endurance-like adaptation in skeletal and cardiac muscles and protect from sarcopenia. However, due to the lack of control over exercise parameters in voluntary exercise models, it is important for the researcher to understand the patterns and variability of wheel running in mice, as well as the factors that can affect voluntary running activity. Overall, voluntary wheel running in mice is a very interesting approach to study the chronic adaptation to exercise, analyze the effects of exercise, and test exercise capacity in different experimental models.

Medial habenula maturational deficits associate with low motivation for voluntary physical activity

The habenula is a small, diencephalic structure comprised of distinct subnuclei which receives inputs from the limbic forebrain and sends projections to various regions in the midbrain, making this region well positioned to influence reward and motivation. Genetic ablation of the dorsal medial habenula is known to decrease voluntary wheel-running in mice. However, the extent to which the medial habenula (MHb) mediates wheel-running motivation in the context of high or low motivation for voluntary physical activity remains to be determined. In so, we utilized 5-week-old female rats selectively bred to voluntarily run high (HVR) or low (LVR) distances in order to determine if inherent differences in medial habenula maturation accompany inherent differences in wheel-running motivation. We report a significantly higher expression of genes associated with MHb development (Brn3a, Nurr1, Tac1, and Kcnip) in HVR versus LVR rats. Furthermore, there was a positive correlation between Brn3a and Nurr1 expression and run distance in HVR, but not LVR rats. Similarly, NeuN and Synapsin 1, markers of neuronal maturation, were higher in HVR compared to LVR rats. Lastly, dendritic density was determined to be higher in the MHb of HVR versus LVR rats, while LVR rats showed a higher percentage of thin spines, suggesting a higher prevalence of immature dendrites in LVR rats. Taken together, the above findings highlight the involvement of MHb in driving the motivation to be physically active. Given pandemic levels of global physical inactivity, the role of the MHb offers a novel potential to improve our global health.

Sexual-Dimorphism in Exercise-Induced Beiging and Altered Mitochondrial Function in Inguinal White Adipose Tissue (iWAT) from the Mouse

Regular physical exercise has numerous benefits on metabolic health, likely involving adaptations to many tissues, including adipose tissue. There are conflicting reports on the effects of exercise training on adaptations to iWAT and we hypothesized that these discrepancies may stem in part from sex-specific differences. Male and female 10 week old C57BL6/J mice were housed in cages with (trained) or without (sedentary) running wheels for 11 days (n=6-9/sex/experiment). Males performed less voluntary exercise vs. females (56±7 vs. 78±5 km; p&amp;lt;0.05). In males, training decreased iWAT fat pad weight (19.6%) and adipocyte size (19%) (both p&amp;lt;0.05), whereas training did not have these effects on iWAT in females, most likely due to increased food intake. Training did not alter perigonadal WAT weight or adipocyte size in either sex. In iWAT, training in males increased mRNA of the beiging genes Ucp1 (304%; p&amp;lt;0.01); Pgc1α (228%; p&amp;lt;0.05); Cidea (206%; p&amp;lt;0.05); and Dio2 (197%; p&amp;lt;0.01), as well as Ucp1 protein (124%; p&amp;lt;0.01), but none of these markers were increased in females. Exercise training also increased mitochondrial parameters in iWAT only from males, including basal respiration (170%; p&amp;lt;0.01), glycolytic capacity (227%; p&amp;lt;0.001) and the mitochondrial fission genes Opa1 (235%; p&amp;lt;0.05), Mfn1 (231%; p&amp;lt;0.001), and Mfn2 (229%; p&amp;lt;0.01). Since exercise may increase testosterone in male mice, we tested the effects of testosterone on iWAT beiging and mitochondria. iWAT of male and female mice were treated with testosterone (10μM) for 24 hours. Interestingly, testosterone increased Ucp1 protein (175%; p&amp;lt;0.05) and mRNA (386%; p&amp;lt;0.05) in iWAT from male, but not in females. We conclude that voluntary wheel running in mice results in beiging and mitochondrial adaptations to iWAT exclusively in males. Sex-specific differences in WAT function will be important to consider in all research investigating adipose tissue beiging and mitochondrial function. Disclosure P. Nigro: None. R. Middelbeek: None. C.R. Alves: None. B.G. Albertson: None. L.A. Rowland: None. A.B. Wagner: None. N.B. Prince: None. M.F. Hirshman: None. L.J. Goodyear: None.

Voluntary Wheel Running Does Not Alter Mortality to or Immunogenicity of Vaccinia Virus in Mice: A Pilot Study.

Exercise has been shown to improve immune responses to viral infections and vaccines in several mouse models. However, previous pathogen studies have primarily used infections limited to the respiratory tract. Additionally, previous studies have utilized forced treadmill exercise paradigms, and voluntary wheel running (VWR) has been shown to have differential effects on the immune system in non-infection models. We examined whether VWR could improve morbidity and mortality to a 50% lethal dose of vaccinia virus (VACV), a systemic pathogen commonly used to examine immune responses. Additionally, we examined whether VWR could improve antibody response to a replication-deficient strain of VACV, mimicking a vaccination. Male C57Bl/6J mice underwent 8 weeks of VWR or remained sedentary, then were infected intranasally with 105 PFU VACV strain WR and followed 14 days for weight loss. Mice in the vaccination study ran or were sedentary for 8 weeks, then were given 106 PFU of replication-deficient VACV strain MVA intraperitoneally. Blood was collected at 1, 2, and 4 weeks post-inoculation, and anti-VACV IgG titer was determined by ELISA. VWR did not improve mortality due to VACV infection (p = 0.26), although fewer VWR mice (4/10) died compared to sedentary (SED, 6/10). VWR did not prevent body weight loss due to infection compared to SED (p = 0.20), although VWR mice loss slightly less weight compared to SED through the first 6 days post-infection. Food intake was significantly reduced in SED post-infection compared to VWR (p = 0.05). VWR mice developed a greater IgG antibody response, although this was not significant (p = 0.22). In summary, VWR did not protect against mortality to VACV or prevent infection-induced weight loss, and VWR did not enhance antibody responses. However, there were non-significant trends toward VWR-related improvements in these outcomes, and post-infection food intake was improved by VWR.

The Importance Of Accurate Measurements In Voluntary Wheel Running In Mice

The Importance Of Accurate Measurements In Voluntary Wheel Running In Mice

Voluntary Wheel Running in Mice Is Safe During Vaccinia Virus Infection and Does not Impair Vaccine Responses

Vaccinia virus (VACV) is a double‐stranded DNA virus of the poxvirus family. Since the eradication of smallpox, VACV is most commonly used as an experimental infection model, but VACV‐based vaccines (either live or attenuated) have received increased attention in previous years as vehicles for inoculation against other agents. Exercise has previously been shown to impair immune responses to viral infections. Although this has primarily been described in intense exercise, to our knowledge no study has examined voluntary wheel running (VWR) in mice, which impacts immune responses differently compared to forced exercise. Therefore, we examined the impact of VWR on infection and antibody responses to several strains of VACV. Male C57Bl/6J mice ran for 8 weeks on wheels or remained sedentary, then were infected with 10^5 PFU of pathogenic VACV strain Western Reserve (WR) intranasally or inoculated with 10^6 PFU of replication‐deficient VACV strain Modified Vaccinia Ankara (MVA) intraperitoneally. Mice receiving WR were followed for morbidity and mortality for 14 days, while mice receiving MVA had blood collected prior to inoculation and at week 1, 2, and 4 post‐inoculation for examination of antibody responses. VWR in mice did not alter mortality or body weight loss due to WR infection, although food intake was significantly reduced in sedentary mice (p=0.05), indicating a potential protective effect of VWR against morbidity. VWR also did not alter anti‐VACV IgG response, although mice with wheel access had non‐significantly increased IgG (p=0.22). In conclusion, VWR is safe prior to and during VACV infection and does not impair vaccine responses to the attenuated MVA strain. VWR may increase immune responses and reduce morbidity, although the present study was underpowered to determine this.Support or Funding InformationFunded by a grant from the American College of Sports Medicine to BDP.

Caffeine stimulates voluntary wheel running in mice without increasing aerobic capacity

The “energy drink” Red Bull and the “sports drink” Gatorade are often marketed to athletes, with claims that they cause performance gains. However, both are high in sugars, and also consumed by non-athletes. Few studies have addressed the effects of these drinks or their biologically active components in rodent exercise models. We used three experiments to test effects on both voluntary exercise behavior and maximal aerobic capacity in lines of mice known to differ in “athletic” traits. Mice from four replicate High Runner (HR) lines have been selectively bred for voluntary running on wheels, and run approximately three times as many revolutions per day as do mice from four non-selected Control (C) lines. HR mice also have higher endurance and maximal oxygen consumption (VO2max) during forced treadmill exercise. In Experiment 1, we tested the hypothesis that Gatorade or Red Bull might cause or allow mice to increase their voluntary wheel running. On days 5 and 6 of 6days of wheel access, as is used to select breeders, HR mice ran 3.3-fold more than C, and females ran 1.2-fold more than males, with no linetype by sex interaction. On day 7, mice were administered Gatorade, Red Bull or tap water. During the subsequent 19-hour period, Gatorade had no statistical effect on running, but Red Bull significantly increased distance run by both sexes and in both HR and C lines. The increase in distance run caused by Red Bull was attributable to time spent running, not an increase in mean (or maximum) speed. As previous studies have found that sucrose alone does not generally increase wheel running, we tested two other active ingredients in Red Bull, caffeine and taurine, in Experiment 2. With a similar testing protocol, caffeine alone and caffeine+taurine increased running by about half the magnitude of Red Bull. In Experiment 3, we tested the hypothesis that Red Bull or caffeine alone can increase physiological performance ability during aerobic exercise, measured as VO2max. In a repeated-measures design spanning 6days, females were housed with water bottles containing Red Bull, caffeine or water in a randomized order, and tested for VO2max twice while receiving each fluid (6 total trials). Neither Red Bull nor caffeine significantly affected either VO2max or a measure of trial cooperativity (rated on a scale of 1–5), but both treatments significantly reduced tiredness (rated on a scale of 1–3) scored at the end of trials for both HR and C lines. Taken together, our results suggest that caffeine increases voluntary exercise levels of mice by delaying fatigue, rather than increasing aerobic capacity.