AIN-93M (Semi-purified diet, according to the American Institute of Nutrition, AIN-93M; [12]) The diet was composed of 70% carbohydrates, 14% protein, and 4% fat at 3,802.7 kcal/g. The remainder of the ingredients were comprised of minerals, fibre, and vitamins. Adaptation to water Before undergoing

the PF-04929113 ic50 lactate minimum protocol, all the animals were adapted only one time to water. The adaptation occurred over a total period of five continuous days, by placing the animals in shallow water in the tank where the tests occurred. The water temperature was maintained at 31 ± 1°C [19]. The purpose of the adaptation was to reduce the stress of the animals, without promoting physiological adaptations that result from physical training. Evaluation of aerobic and anaerobic capacity To determine acutely aerobic and anaerobic capacity, we used the

lactate minimum test, which enabled us to determine both parameters MK-4827 in a single protocol [20, 21]. This test consists of an induction phase to hyperlactatemia (anaerobic exercise) followed MK-1775 in vivo by progressive exercise. The induction phase consisted of two efforts with a load equivalent to 13% of the animals’ body weight. The first effort lasted 30 s, followed by a 30-s passive recovery period. After the recovery period, the animals performed a maximum effort to obtain the time to exhaustion, considered as the parameter of anaerobic fitness. Nine minutes after the exhaustion period, we collected 25 μl of blood via a cut at the distal end of the tail to determine lactate concentrations. After collecting the blood, the animals began a progressive phase with an initial intensity of 4.0% of body weight, which was increased by increments of 0.5% of body weight over 5 min intervals. At the end of each stage, 25 μl of blood was collected to determine lactate concentrations. The anaerobic threshold, considered as the parameter

for aerobic capacity, was equivalent to the Bacterial neuraminidase zero derivative of a second-order polynomial fit that was obtained from the relationship between lactate concentrations and the exercise intensity. Consequently, we determined lactate concentrations based on the anaerobic threshold. During all the efforts, the animals were placed individually in tanks (100 × 80 × 80 cm) containing water at 31 ± 1°C. Blood samples were collected using calibrated capillary tubes and heparinised, and blood lactate was determined using an enzymatic method [22]. Evaluations conducted during the intervention and before euthanasia Throughout the experimental period, the body weights (all groups) and feed intakes (ad libitum group) were recorded daily using an analytical balance. The results were analysed based on the weight change of the animals (weight change = initial weight – final weight). Parameters obtained following euthanasia At the end of the experiment, animals were anesthetised in a CO2 chamber, 48 h after measuring the lactate minimum test.