Evaluation of a Rat Model of Exercise-Induced Fatigue Using Treadmill Running with Progressively Increasing Load
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Anais da Academia Brasileira de Ciências (2019) 91(4): e20180957 (Annals of the Brazilian Academy of Sciences) Printed version ISSN 0001-3765 / Online version ISSN 1678-2690 http://dx.doi.org/10.1590/0001-3765201920180957 www.scielo.br/aabc | www.fb.com/aabcjournal Evaluation of a rat model of exercise-induced fatigue using treadmill running with progressively increasing load LIJUAN HOU1, KE LI1, YANRU HU2, YIJUN BIAN3, WEI JI1, KAIXUAN SHI1, YITING LI1, MENGJIAO CHEN1, JIAXIN LI1, XIAOLI LIU1 and DECAI QIAO1 1Beijing Normal University, No. 19, XinJieKouWai St., HaiDian District, Beijing, 100875, P.R. China 2Central South University, No. 932 South Lushan Road, Changsha, Hunan, 410083, P.R. China 3University of Science and Technology of China, No. 96, JinZhai Road Baohe District, Hefei, Anhui, 230027, P.R. China Manuscript received on September 17, 2018; accepted for publication on January 23, 2019 How to cite: HOU L, LI K, HU Y, BIAN Y, JI W, SHI K, LI Y, CHEN M, LI J, LIU X AND QIAO D. 2019. Evaluation of a rat model of exercise-induced fatigue using treadmill running with progressively increasing load. An Acad Bras Cienc 91: e20180957. DOI 10.1590/0001-3765201920180957. Abstract: The mechanism behind exercise-induced fatigue is a significant topic in the field of sports physiology. Therefore, establishing and evaluating an acute exercise-induced fatigue animal model that explores the limits of the motor system may provide greater insight into these mechanisms. Heart rate is an important quantitative parameter that accurately reflects the immediate change in physical function due to exercise load. And there is likely to be an important correlation between heart rate and behavioral performance. In this study, changes in heart rate and behavioral indexes during exercise-induced fatigue were quantitatively analyzed in rats using heart rate telemetry and video methods respectively. The behavioral indexes were used as independent variables and the degree of fatigue was used as the forecast value. Ternary quadratic function curve fitting was used to deduce a formula to calculate a fatigue score: 2 2 Y = 15.2548+0.4346∙xa-0.1154∙xb+0.6826∙xc+0.0044∙xa∙xb-0.0021∙xb∙xc-0.0013∙xc∙xa-0.0023∙xa -0.0016∙xb (r2=0.906). It identified a linear relationship between heart rate and exercise intensity, with a plateau in heart rate occurring during difference periods. It will serve as an effective reference for the modeling of exercise-induced fatigue. In addition, it also provides a theoretical method for analyzing the correlation between peripheral and central parameters. Key words: exercise-induced fatigue, animal model, treadmill running, heart rate. INTRODUCTION or the individual cannot maintain a predetermined exercise intensity. Professor Mosso pioneered the Exercise-induced fatigue is a common phenomenon research on exercise-induced fatigue in 1891 and in athletics and a key factor that restricts an athlete’s performance level (Knuttgen 1983). The subsequently introduced multiple hypotheses, term “exercise-induced fatigue” refers to a state including that “muscle fatigue is a peripheral in which the physiological systems of the body phenomenon independent of the nervous system,” cannot continue to function at the required level, “carbon dioxide and lactic acid metabolites are the main factors that reduce muscle contractility,” Correspondence to: Decai Qiao E-mail: [email protected] and “fatigue is an alert signal for the body’s self- ORCid: http://orcid.org/0000-0002-2724-3139 protection.” At the Fifth International Exercise BIOLOGICAL SCIENCES An Acad Bras Cienc (2019) 91(4) LIJUAN HOU et al. EXERCISE-INDUCED FATIGUE ANIMAL MODEL Biochemistry Conference held in 1982, the term In this study, changes in heart rate and was finally defined as “the body’s physiological behavioral indexes during exercise-induced fatigue processes that cannot maintain its functions were quantitatively analyzed in rats using heart at a particular level and/or cannot maintain rate telemetry and video methods, respectively. predetermined exercise intensity,” as mentioned The behavioral indexes were used as independent above. variables and the degree of fatigue was used as In recent years, with the rapid theoretical the forecast value. Ternary quadratic function and technological developments in the field of curve fitting was used to deduce a formula, and neuroscience, the central nervous system (CNS) evaluation of the degree of exercise-induced fatigue has been known to play an important role in was performed to provide a reference for future moderating exercise-induced fatigue. For example, exploration of physiological, biochemical, and once exercise-induced fatigue appears, nerve neurobiological mechanisms of exercise-induced conduction velocity decreases, the concentration of fatigue. 5-hydroxy tryptamine (5-HT) neurotransmitter in MATERIALS AND METHODS the CNS increases, and the excitability of neurons in the cerebral cortex decreases (Tanaka et al. 2003, ANIMALS USED IN THIS STUDY Tian and Gao 1995, Li et al. 2008). In addition, Twelve male Wistar rats (Beijing Vital River it is proposed that exercise-induced fatigue is the Laboratory Animal Technological Company, result of cerebral protective inhibition, and that Beijing, China) at 8-10 weeks of age (weighing the associated reduction in muscle contractility 280 ± 20 g) were used in this study. All rats were is the result of functional failure of the CNS. It is randomly divided into two groups with three rats to presumed that exercise-induced fatigue occurs due a cage. The animals were maintained on a 12-h light/ to changes in the electrical activities of neurons dark cycle and given food and water ad libitum. in the cerebral cortex and basal ganglia. However, The rats were maintained in an environment with a owing to the lack of quantitative methods to relative humidity of 40–60% and room temperature evaluate exercise-induced fatigue in animals, it is of 20–25°C. All experimental procedures were difficult to obtain conclusive experimental results, approved by the Ethics Committee of Beijing and further research is required to deduce the Normal University (LUNLI20130901). mechanisms behind exercise-induced fatigue. In the present study, the degree of exercise-induced FATIGUE MODEL ESTABLISHMENT AND ETHOLOGICAL INDICES OBSERVATION fatigue was evaluated by observing the running distance and posture of rats, which are not fully After being fed in the laboratory for 3 days, the quantitative parameters. In exercise physiology, rats were subjected to 5-days of adaptive treadmill heart rate is an important quantitative parameter training. A modified Bedford treadmill exercise that accurately reflects the immediate change in with a progressively increasing load was used physical function due to exercise load. Behavioral for the exercise-induced fatigue model (Liu et indexes, such as running distance and posture, al. 2012). There were three load levels. Level I: may reflect the overall condition of experimental 8.2 m/min, lasting for 15 min; level II: 15 m/min, animals (Kruk 1997). Therefore, there is likely to lasting for 15 min; level III: 20 m/min, lasting until be an important correlation between heart rate and exhaustion. The criteria used to indicate exercise- behavioral performance. induced fatigue were as follows: the running An Acad Bras Cienc (2019) 91(4) e20180957 2 | 12 LIJUAN HOU et al. EXERCISE-INDUCED FATIGUE ANIMAL MODEL posture shifted from thrusting against the ground to fur above the left side of the chest and right side of prostration, the rat stayed at the end of the treadmill the neck was removed (approximately 2 cm × 1 cm), without the ability to continue running, and the rat and a heart rate telemetry electrode vest was affixed was unable to be driven to continue running even to each rat. A 0.8 cm incision was made in the skin, with sound or electrical stimulation. and the electrode was implanted subcutaneously A transparent polymethylmethacrylate with the positive electrode near the heart, in the enclosure was placed over the treadmill, and a scale left side of the chest, and the negative electrode was marked below the two sides of the enclosure. implanted in the right side of the neck, at the same The midpoint of each rat’s body and the root of level as the positive electrode. The wound was then the tail were marked in red. Two cameras were sutured. Before the treadmill exercise, a battery- placed on each side of the treadmill to record the operated transmitter was attached to the vest, and data which was to be analyzed later. If the marked the electrode was connected to the transmitter. point of the root of the tail was ≥ 34 cm away While the rats were performing the Bedford from the board in front of the treadmill, sound treadmill exercise, their heart rate was measured by stimulation was applied; if the marks were ≥ 39 the wireless animal heart rate telemetry system and cm away from the board, electrical stimulation was the behavioral changes were observed at the same applied. Meanwhile, if the midpoint of the rat was time, by video (Figure 1). ≥ 39 cm away from the board for > 0.5 s during a 1 DATA COLLECTION AND PRESENTATION s period, the rat was considered to have slipped off the treadmill belt. Five behavioral indexes were selected based on previous preliminary experiments: the relative ELECTRODE IMPLANTATION AND HEART RATE COLLECTION DURING EXERCISE velocity of the rat, relative position on the treadmill, sound stimulation time (as a percentage of the total The electrode for heart rate telemetry was implanted time) electrical stimulation time (as a percentage of the day after the adaptive treadmill training finished. the total time), and slippage time (as a percentage For the electrophysiological recording, the rats of the total time). were anesthetized with 10% chloral hydrate (0.35 The method by which relative velocity was mL/100 g) and placed on a stereotaxic frame.