Atsushi Hiraga

Assessment of autonomic nervous function by power spectral analysis of heart rate variability in the horse

We studied power spectral analysis of heart rate (HR) variability in the horse, with the hypothesis that the quantitative information provided by the spectral analysis of HR variability reflects the interaction between sympathetic and parasympathetic regulatory activities. For this purpose, electrocardiogram, blood pressure (BP) and respiratory (Resp) waveform were simultaneously recorded from Thoroughbred horses (3-5 years old) and analyzed by power spectrum. There were two major spectral components at low-frequency (LF) and high-frequency (HF) bands for HR variability. The peak of Resp variability clearly occurred at the HF range. In contrast to Resp variability, the power spectra of BP variability occurred at lower frequencies. The maximum coherence between HR and Resp variabilities and HR and BP variabilities occurred at approximately 0.15 and approximately 0.03 Hz, respectively. These relationships were similar to the ensemble spectra. On the basis of these data, we have defined two frequency bands of interest: LF (0.01-0.07 Hz) and HF (0.07-0.6 Hz). Therefore, we believe that power spectral analysis of HR variability provides a very powerful technique for assessing autonomic nervous activity in the horse.

Effects of Novelty Stress on Neuroendocrine Activities and Running Performance in Thoroughbred Horses

This study investigated the effects of novelty stress on neuroendocrine activities and running performance in Thoroughbred horses. First, to examine the neuroendocrine responses to novelty stress, we exposed horses to two types of novel environmental stimuli (audiovisual or novel field stimuli). After the stimuli, plasma concentrations of vasopressin, catecholamines and adrenocorticotropin (ACTH), as well as heart rates, were significantly increased in each experiment. Second, we investigated neuroendocrine activities during incremental exercise. Plasma concentrations of vasopressin, catecholamines, ACTH and blood lactate increased as the exercise load increased. Finally, we investigated the effects of novelty stimuli on neuroendocrine activities and running performance during supra‐maximal exercise (110% VHRmax). When the novelty stimuli were presented to horses, the increases in plasma vasopressin and catecholamines due to exercise load were significantly smaller than those in the control experiments. Blood lactate during supra‐maximal exercise was also significantly lower and total run time until exhaustion was prolonged in the novel environmental stimuli compared to the control. These results suggest that novelty stimuli facilitate vasopressin release from the posterior pituitary in addition to activating the sympatho‐adrenomedullary and the hypothalamic‐pituitary‐adrenocortical axes in thoroughbred horses, and increase exercise capacity, resulting in improvement of running performance during supra‐maximal exercise.

Effect of training and detraining on monocarboxylate transporter (MCT) 1 and MCT4 in Thoroughbred horses

The aim of this study was to investigate the effects of training and detraining on the monocarboxylate transporter (MCT) 1 and MCT4 levels in the gluteus medius muscle of Thoroughbred horses. Twelve Thoroughbred horses were used for the analysis. For 18 weeks, all the horses underwent high‐intensity training (HIT), with running at 90–110% maximal oxygen consumption ( ) for 3 min, 5 days week−1. Thereafter, the horses either underwent detraining for 6 weeks by either 3 min of moderate‐intensity training (MIT) at 70% , 5 days week−1 (HIT‐MIT group) or stall rest (HIT‐SR group). The horses underwent an incremental exercise test, was measured and resting muscle samples were obtained from the middle gluteus muscle at 0, 18 and 24 weeks. The content of MCT1 and MCT4 proteins increased after 18 weeks of HIT. At the end of this period, an increase was noted in the citrate synthase activity, while phosphofructokinase activity remained unchanged. After 6 weeks of detraining, all these indexes returned to the pretraining levels in the HIT‐SR group. However, in the HIT‐MIT group, the increase in the MCT1 protein content and citrate synthase activity was maintained after 6 weeks of MIT, while the MCT4 protein content decreased to the pretraining value. These results suggest that the content of MCT1 and MCT4 proteins increases after HIT in Thoroughbred horses. In addition, the increase in the MCT1 protein content and oxidative capacity induced by HIT can be maintained by MIT of 70% , but the increase in the MCT4 protein content cannot be maintained by MIT.

Effects of altered FiO2 on maximum V̇O2 in the horse

Although the horse is considered an elite athlete with a specific VO2max some 2-4 times higher than man, maximal O2 transport is compromised both by moderately severe arterial desaturation and by failure to extract all O2 from blood perfusing exercising muscle. This prompted the present study to ascertain whether correction of arterial desaturation would proportionally augment VO2max and, if so, would O2 extraction behave in a manner predicted by diffusional transport limitation. Six two year old thoroughbreds were exercised to VO2max on a treadmill each on three separate occasions breathing gases of FIO2 = 0.15, 0.21 and 0.35, each used once in balanced order. VO2, ventilation, arterial and pulmonary arterial blood gases, pressures and lactate levels were measured both submaximally and maximally at each FIO2 and cardiac output was computed by mass balance for O2. At FIO2 = 0.21, VO2max = 143.9 +/- 4.8 ml kg-1 min-1, arterial saturation (SaO2) was 81.6 +/- 3.3% while venous PO2 (PvO2) was 15.3 +/- 1.4 Torr. At FIO2 = 0.35, VO2max was 172.6 +/- 8.2 ml kg-1 min-1, SaO2 reached 97.4 +/- 0.4% and PvO2 was 23.4 +/- 0.7 Torr. VO2max at FIO2 = 0.15 was 109.8 +/- 4.1 ml kg-1 min-1, SaO2 fell to 68.1 +/- 2.5% and PvO2 was 10.6 +/- 1.0 Torr, all changes being significant, p < 0.01. As FIO2 was varied, VO2max changed proportionally to calculated mean capillary Po2 as well as to total O2 delivery. These data confirm substantial O2 supply dependence of VO2max in the horse, and in such a manner as to be consistent with the hypothesis of combined diffusive and convective transport limitation within muscle.

Muscle fiber population and biochemical properties of whole body muscles in thoroughbred horses.

We examine the muscle fiber population and metabolic properties of skeletal muscles from the whole body in Thoroughbred horses. Postmortem samples were taken from 46 sites in six Thoroughbred horses aged between 3 and 6 years. Fiber type population was determined on muscle fibers stained with monoclonal antibody to each myosin heavy chain isoform and metabolic enzyme activities were determined spectrophotometrically. Histochemical analysis demonstrated that most of the muscles had a high percentage of Type IIa fibers. In terms of the muscle characteristic in several parts of the horse body, the forelimb muscles had a higher percentage of Type IIa fiber and a significantly lower percentage of Type IIx fiber than the hindlimb muscles. The muscle fiber type populations in the thoracic and trunk portion were similar to those in the hindlimb portion. Biochemical analysis indicated high succinate dehydrogenase activity in respiratory‐related muscle and high phosphofructokinase activity in hindlimbs. We suggested that the higher percentage of Type IIa fibers in Thoroughbred racehorses is attributed to training effects. To consider further the physiological significance of each part of the body, data for the recruitment pattern of each muscle fiber type during exercise are needed. The muscle fiber properties in this study combined with the recruitment data would provide fundamental information for physiological and pathological studies in Thoroughbred horses. Anat Rec, 2009.

Hypoxic helium breathing does not reduce alveolar-arterial PO2 difference in the horse

In a previous study we evaluated the mechanism of alveolar-arterial PO2 (AaPO2) reduction when nitrogen is replaced with helium in normoxia (FIO2 = 0.21). The reduction in AaPO2 was not due to changes in VA/Q inequality, pulmonary O2 diffusing capacity, or cardiac output, but to more complete diffusion equilibration as a consequence of the higher ventilation and thus PAO2 (which reduced the average slope of the hemoglobin O2 dissociation curve (ODC), and thus enhanced diffusive equilibration). We hypothesized that hypoxic He/O2 breathing in contrast would not reduce the AaPO2 because PAO2 and PaO2, although higher with He than N2, would remain constrained to the linear region of the ODC. Breathing hypoxic gas mixtures did constrain the PAO2 to the linear region of the ODC, even when PAO2 was increased by He/O2 breathing. Thus, the average slope of the ODC did not change when He replaced N2 and this explains the lack of change in AaPO2, as hypothesized.

Muscle satellite cells are activated after exercise to exhaustion in Thoroughbred horses.

REASONS FOR PERFORMING STUDY Although satellite cells are well known as muscle stem cells capable of adding myonuclei during muscle repair and hypertrophy, the response of satellite cells in horse muscles to a run to exhaustion is still unknown. OBJECTIVES To investigate the time course of satellite cell activation in Thoroughbred horse muscle after running to exhaustion. We hypothesised that this type of intense exercise would induce satellite cell activation in skeletal muscle similar to a resistance exercise. METHODS Nine de-trained Thoroughbred horses (6 geldings and 3 mares) aged 3-6 years were studied. Biopsy samples were taken from the gluteus medius muscle of the horses before and 1 min, 3 h, 1 day, 3 days, 1 week and 2 weeks after a treadmill run to exhaustion. The numbers of satellite cells for each fibre type were determined by using immunofluorescence staining. Total RNA was extracted from these samples, and the expressions of interleukin (IL)-6, paired box transcriptional factor (Pax) 7, myogenic differentiation 1 (MyoD), myogenin, proliferating cell nuclear antigen (PCNA), insulin-like growth factor (IGF)-I and hepatocyte growth factor (HGF) mRNA were analysed using real-time reverse transcription-PCR. RESULTS The numbers of satellite cells were significantly increased in type I and IIa fibres at 1 week and in type IIa/x fibre at 2 weeks post exercise. The expression of IL-6 mRNA increased significantly by 3 h post exercise. The expression of PCNA mRNA also increased by 1 day after running, indicating that running can initiate satellite cell proliferation. The expression of Pax7, MyoD, myogenin, IGF-I and HGF mRNA peaked at 1 week post exercise. CONCLUSION Satellite cell activation and proliferation could be enhanced after a run to exhaustion without detectable injury as assessed by the histochemical analysis. Understanding the response of satellite cell activation to running exercise provides fundamental information about the skeletal muscle adaptation in Thoroughbred horses.

Changes in heart rate and heart rate variability during transportation of horses by road and air

OBJECTIVE To determine the influence of transportation by road and air on heart rate (HR) and HR variability (HRV) in horses. Animals-6 healthy horses. PROCEDURES ECG recordings were obtained from horses before (quarantine with stall rest [Q]; 24 hours) and during a journey that included transportation by road (RT; 4.5 hours), waiting on the ground in an air stall (W; 5.5 hours), and transportation by air (AT; 11 hours); HR was determined, and HRV indices of autonomic nervous activity (low-frequency [LF; 0.01 to 0.07 Hz] and high-frequency [HF; 0.07 to 0.6 Hz] power) were calculated. RESULTS Mean ± SD HRs during Q, RT, W, and AT were 38.9 ± 1.5 beats/min, 41.7 ± 5.6 beats/min, 41.5 ± 4.3 beats/min, and 48.8 ± 5.6 beats/min, respectively; HR during AT was significantly higher than HR during Q. The LF power was significantly higher during Q (3,454 ± 1,087 milliseconds(2)) and AT (3,101 ± 567 milliseconds(2)) than it was during RT (1,824 ± 432 milliseconds(2)) and W (2,072 ± 616 milliseconds(2)). During Q, RT, W, and AT, neither HF powers (range, 509 to 927 milliseconds(2)) nor LF:HF ratios (range, 4.1 to 6.2) differed significantly. The HR during RT was highly correlated with LF power (R(2) = 0.979), and HR during AT was moderately correlated with the LF:HF ratio (R(2) = 0.477). CONCLUSIONS AND CLINICAL RELEVANCE In horses, HR and HRV indices during RT and AT differed, suggesting that exposure to different stressors results in different autonomic nervous influences on HR.

Effect of acute exercise on monocarboxylate transporters 1 and 4 in untrained and trained Thoroughbreds

OBJECTIVE To evaluate the effects of a single incremental exercise test (IET) on mRNA expression and protein content of monocarboxylate transporter (MCT) 1 and MCT4 in the gluteus medius muscle of Thoroughbreds. ANIMALS 12 Thoroughbreds (6 males and 6 females; age, 3 to 4 years). PROCEDURES Horses underwent an IET before and after 18 weeks of high-intensity exercise training (HIT). Horses were exercised at 90% of maximal oxygen consumption for 3 minutes during the initial 10 weeks of HIT and 110% of maximal oxygen consumption for 3 minutes during the last 8 weeks of HIT. Gluteus medius muscle biopsy specimens were obtained from horses before (baseline), immediately after, and at 3, 6, and 24 hours after the IET. RESULTS Expression of MCT1 and MCT4 mRNA was upregulated at 3 and 6 hours after the IET in muscle specimens obtained from horses prior to HIT (untrained horses) and at 6 hours after the IET in muscle specimens obtained from horses after HIT (trained horses). For both untrained and trained horses, MCT1 and MCT4 protein contents were increased at 6 hours after the IET and did not differ at 24 hours after the IET, compared with those at baseline. CONCLUSIONS AND CLINICAL RELEVANCE Results indicated that a single IET resulted in transient increases in MCT1 and MCT4 mRNA expression and protein content in untrained and trained horses. These results may be important for the elucidation of exercise-induced alterations in lactate metabolism.

Effects of three warm‐up regimens of equal distance on V̇o2 kinetics during supramaximal exercise in Thoroughbred horses

REASONS FOR PERFORMING STUDY Several studies have indicated that even low-intensity warm-up increases O(2) transport kinetics and that high-intensity warm-up may not be needed in horses. However, conventional warm-up exercise for Thoroughbred races is more intense than those utilised in previous studies of equine warm-up responses. OBJECTIVES To test the hypothesis that warm-up exercise at different intensities alters the kinetics and total contribution of aerobic power to total metabolic power in subsequent supramaximal (sprint) exercise in Thoroughbred horses. METHODS Nine well-trained Thoroughbreds ran until fatigue at 115% of maximal oxygen consumption (VO2max) 10 min after warming-up under each of 3 protocols of equal running distance: 400 s at 30% VO2max (LoWU), 200 s at 60% VO2max (MoWU) and 120 s at 100% VO2max (HiWU). Variables measured during exercise were rates of O(2) and CO(2) consumption/production (VO2,VO2), respiratory exchange ratio (RER), heart rate, blood lactate concentration and accumulation rate and blood gas variables. RESULTS VO2 was significantly higher in HiWU than in LoWU at the onset of the sprint exercise and HR was significantly higher in HiWU than in LoWU throughout the sprint. Accumulation of blood lactate, RER, P(a)CO(2) and PvCO2 in the first 60 s were significantly lower in HiWU than in LoWU and MoWU. There were no significant differences in stroke volume, run time or arterial-mixed venous O(2) concentration. CONCLUSIONS These results suggest HiWU accelerates kinetics and reduces reliance on net anaerobic power compared with LoWU at the onset of the subsequent sprint.