Hayden W. Hyatt

A Ketogenic Diet in Rodents Elicits Improved Mitochondrial Adaptations in Response to Resistance Exercise Training Compared to an Isocaloric Western Diet

Purpose: Ketogenic diets (KD) can facilitate weight loss, but their effects on skeletal muscle remain equivocal. In this experiment we investigated the effects of two diets on skeletal muscle mitochondrial coupling, mitochondrial complex activity, markers of oxidative stress, and gene expression in sedentary and resistance exercised rats. Methods: Male Sprague-Dawley rats (9–10 weeks of age, 300–325 g) were fed isocaloric amounts of either a KD (17 g/day, 5.2 kcal/g, 20.2% protein, 10.3% CHO, 69.5% fat, n = 16) or a Western diet (WD) (20 g/day, 4.5 kcal/g, 15.2% protein, 42.7% CHO, 42.0% fat, n = 16) for 6 weeks. During these 6 weeks animals were either sedentary (SED, n = 8 per diet group) or voluntarily exercised using resistance-loaded running wheels (EXE, n = 8 per diet group). Gastrocnemius was excised and used for mitochondrial isolation and biochemical analyses. Results: In the presence of a complex II substrate, the respiratory control ratio (RCR) of isolated gastrocnemius mitochondria was higher (p < 0.05) in animals fed the KD compared to animals fed the WD. Complex I and IV enzyme activity was higher (p < 0.05) in EXE animals regardless of diet. SOD2 protein levels and GLUT4 and PGC1α mRNA expression were higher (p < 0.05) in EXE animals regardless of diet. Conclusion: Our data indicate that skeletal muscle mitochondrial coupling of complex II substrates is more efficient in chronically resistance trained rodents fed a KD. These findings may provide merit for further investigation, perhaps on humans.

Comparative adaptations in oxidative and glycolytic muscle fibers in a low voluntary wheel running rat model performing three levels of physical activity.

A unique polygenic model of rat physical activity has been recently developed where rats were selected for the trait of low voluntary wheel running. We utilized this model to identify differences in soleus and plantaris muscles of sedentary low voluntary wheel running rats and physically active low voluntary wheel running rats exposed to moderate amounts of treadmill training. Three groups of 28‐day‐old male Wistar rats were used: (1) rats without a running wheel (SEDENTARY, n = 7), (2) rats housed with a running wheel (WHEEL, n = 7), and (3) rats housed with a running wheel and exercised on the treadmill (5 days/week for 20 min/day at 15.0 m/min) (WHEEL + TREADMILL, n = 7). Animals were euthanized 5 weeks after the start of the experiment and the soleus and plantaris muscles were excised and used for analyses. Increases in skeletal muscle gene expression of peroxisome proliferator‐activated receptor gamma coactivator 1 alpha and fibronectin type III domain‐containing protein 5 in WHEEL + TREADMILL group were observed. Also, WHEEL + TREADMILL had higher protein levels of superoxide dismutase 2 and decreased levels of oxidative damage. Our data demonstrate that the addition of treadmill training induces beneficial muscular adaptations compared to animals with wheel access alone. Furthermore, our data expand our understanding of differential muscular adaptations in response to exercise in mitochondrial, antioxidant, and metabolic markers.

Graded hypoxia and blood oxidative stress during exercise recovery

Abstract Altitude exposure and exercise elicit oxidative stress in blood; however, exercise recovery at 5000 m attenuates oxidative stress. The purpose was to determine the altitude threshold at which blood oxidative stress is blunted during exercise recovery. Twelve males 18–28 years performed four-cycle ergometry bouts (60 min, 70% VO2max, at 975 m). In a randomised counterbalanced crossover design, participants recovered 6 h at 0, 1667, 3333 and 5000 m in a normobaric hypoxia chamber (recovery altitudes were simulated by using a computerised system in an environmental chamber by lowering the partial pressure of oxygen to match that of the respective altitude). Oxygen saturation was monitored throughout exercise recovery. Blood samples obtained pre-, post-, 1 h post- and 5 h post-exercise were assayed for ferric-reducing antioxidant plasma, Trolox equivalent antioxidant capacity, uric acid, lipid hydroperoxides and protein carbonyls. Muscle biopsies obtained pre and 6 h were analysed by real-time polymerase chain reaction to quantify expression of hemeoxgenase 1, superoxide dismutase 2 and nuclear factor (euthyroid-derived 2)-like factor. Pulse oximetry data were similar during exercise, but decreased for the three highest recovery elevations (0 m = 0%, 1667 m = −3%; 3333 m = −7%; 5000 m = −17%). A time-dependent oxidative stress occurred following exercise for all variables, but the two highest recovery altitudes partially attenuated the lipid hydroperoxide response (0 m = +135%, 1667 m = +251%, 3333 m = +99%; 5000 m = +108%). Data may indicate an altitude threshold between 1667 and 3333 m, above which the oxidative stress response is blunted during exercise recovery.

The 1-Week and 8-Month Effects of a Ketogenic Diet or Ketone Salt Supplementation on Multi-Organ Markers of Oxidative Stress and Mitochondrial Function in Rats

We determined the short- and long-term effects of a ketogenic diet (KD) or ketone salt (KS) supplementation on multi-organ oxidative stress and mitochondrial markers. For short-term feedings, 4 month-old male rats were provided isocaloric amounts of KD (n = 10), standard chow (SC) (n = 10) or SC + KS (~1.2 g/day, n = 10). For long-term feedings, 4 month-old male rats were provided KD (n = 8), SC (n = 7) or SC + KS (n = 7) for 8 months and rotarod tested every 2 months. Blood, brain (whole cortex), liver and gastrocnemius muscle were harvested from all rats for biochemical analyses. Additionally, mitochondria from the brain, muscle and liver tissue of long-term-fed rats were analyzed for mitochondrial quantity (maximal citrate synthase activity), quality (state 3 and 4 respiration) and reactive oxygen species (ROS) assays. Liver antioxidant capacity trended higher in short-term KD- and SC + KS-fed versus SC-fed rats, and short-term KD-fed rats exhibited significantly greater serum ketones compared to SC + KS-fed rats indicating that the diet (not KS supplementation) induced ketonemia. In long term-fed rats: (a) serum ketones were significantly greater in KD- versus SC- and SC + KS-fed rats; (b) liver antioxidant capacity and glutathione peroxidase protein was significantly greater in KD- versus SC-fed rats, respectively, while liver protein carbonyls were lowest in KD-fed rats; and (c) gastrocnemius mitochondrial ROS production was significantly greater in KD-fed rats versus other groups, and this paralleled lower mitochondrial glutathione levels. Additionally, the gastrocnemius pyruvate-malate mitochondrial respiratory control ratio was significantly impaired in long-term KD-fed rats, and gastrocnemius mitochondrial quantity was lowest in these animals. Rotarod performance was greatest in KD-fed rats versus all other groups at 2, 4 and 8 months, although there was a significant age-related decline in performance existed in KD-fed rats which was not evident in the other two groups. In conclusion, short- and long-term KD improves select markers of liver oxidative stress compared to SC feeding, although long-term KD feeding may negatively affect skeletal muscle mitochondrial physiology.

Lactation has persistent effects on a mother’s metabolism and mitochondrial function

Human epidemiological data show that breastfeeding reduces the prevalence of numerous diseases compared to mothers that give birth but do not participate in lactation. The goal of this study was to determine if differences in metabolism, mitochondrial function, and oxidative stress underlie the protective phenotype found in lactating women. Ten-week old female Sprague-Dawley rats were divided into three groups (n = 8 per group): 1) rats that did not reproduce (NR), 2) rats that were allowed to mate and become pregnant but did not suckle their pups after giving birth (NL), and 3) rats that were allowed to mate and become pregnant and suckled their pups for 21 days before weaning (L). All animals were sacrificed at approximately 7 months of age, a time corresponding to 15 weeks after the NL and L females gave birth. Liver mitochondrial respiration was higher in L rats when using NADH-linked substrates and these rats had lower serum glucose concentration. Additionally, the L group exhibited changes in liver, skeletal muscle, and white adipose tissue PPARδ protein levels that may, in part, explain the observed lower serum glucose concentration. These novel animal findings provide evidence of differences in metabolic processes that persist months after weaning.

Changes in Metabolism, Mitochondrial Function, and Oxidative Stress Between Female Rats Under Nonreproductive and 3 Reproductive Conditions

Women who do not lactate display increased incidence of obesity, type II diabetes, and cancer. Stuebe and Rich-Edwards proposed that these effects occur because physiological changes that ensue during pregnancy are not reversed without lactation. To empirically test this hypothesis, we compared markers of metabolism, mitochondrial function, and oxidative stress between 4 groups of Sprague-Dawley rats: (1) nonreproductive (NR) rats, (2) rats killed at day 20 of gestation, (3) rats that gave birth but were not allowed to suckle their pups (nonlactating), and (4) rats that suckled their young for 14 days. Nonlactating females displayed higher body fat compared to all other groups. Peroxisome proliferator-activated receptor δ (PPARδ) in skeletal muscle and white adipose tissue of nonlactating rats was lower than the other groups. The PPARδ is associated with lipid metabolism suggesting that the higher fat mass in nonlactating females was not associated with the retention of a physiological state that was set during pregnancy but instead an independent drop in PPARδ. Relative mitochondrial respiratory function and complex activity in the liver and skeletal muscle of nonlactating mice were not predictive of higher body mass, and measures of oxidative stress displayed minimal variation between groups.

Life History Trade-offs within the Context of Mitochondrial Hormesis

Evolutionary biologists have been interested in the negative interactions among life history traits for nearly a century, but the mechanisms that would create this negative interaction remain poorly understood. One variable that has emerged as a likely link between reproductive effort and longevity is oxidative stress. Specifically, it has been proposed that reproduction generates free radicals that cause oxidative stress and, in turn, oxidative stress damages cellular components and accelerates senescence. We propose that there is limited support for the hypothesis because reactive oxygen species (ROS), the free radicals implicated in oxidative damage, are not consistently harmful. With this review, we define the hormetic response of mitochondria to ROS, termed mitochondrial hormesis, and describe how to test for a mitohormetic response. We interpret existing data using our model and propose that experimental manipulations will further improve our knowledge of this response. Finally, we postulate how the mitohormetic response curve applies to variation in animal performance and longevity.

Change in the Lipid Transport Capacity of the Liver and Blood during Reproduction in Rats

To support the high energetic demands of reproduction, female mammals display plasticity in many physiological processes, such as the lipid transport system. Lipids support the energy demands of females during reproduction, and energy and structural demands of the developing offspring via the placenta in utero or milk during the suckling period. We hypothesized that key proteins supporting lipid transport in reproductive females will increase during pregnancy and lactation, but drop to non-reproductive levels shortly after reproduction has ended. We compared the relative protein levels of liver-type cytosolic fatty acid transporter (L-FABPc), plasma membrane fatty acid transporter (FABPpm), fatty acid translocase (FAT/CD36) in the liver, a key site of lipid storage and synthesis, and free fatty acid transporter albumin and triglyceride transporter [represented by apolipoprotein B (apoB)] levels in serum in reproductive Sprague-Dawley rats during late pregnancy, peak-lactation, and 1-week post-lactation as well as in non-reproductive rats. We found that all lipid transporter levels were greater in pregnant rats compared to non-reproductive rats. Lactating rats also showed higher levels of FAT/CD36 and FABPpm than non-reproductive rats. Moreover, all fat transporters also dropped back to non-reproductive levels during post-lactation except for FAT/CD36. These results indicate that fat uptake and transport capacities in liver cells are elevated during late gestation and lactation. Liver lipid secretion is up-regulated during gestation but not during lactation. These data supported the plasticity of lipid transport capacities in liver and blood during reproductive stages.