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- Author or Editor: Ellen de Graaf-Roelfsema x
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Objective—To determine the influence of intensified training and subsequent reduced training on glucose metabolism rate and peripheral insulin sensitivity in horses and identify potential markers indicative of early overtraining.
Animals—12 Standardbred geldings.
Procedures—Horses underwent 4 phases of treadmill-based training. In phase 1, horses were habituated to the treadmill. In phase 2, endurance training was alternated with high-intensity exercise training. In phase 3, horses were divided into control and intensified training groups. In the intensified training group, training intensity, duration, and frequency were further increased via a protocol to induce overtraining; in the control group, these factors remained unaltered. In phase 4, training intensity was reduced. Standardized exercise tests were performed after each phase and hyperinsulinemic euglycemic clamp (HEC) tests were performed after phases 2, 3, and 4.
Results—10 of 12 horses completed the study. Dissociation between mean glucose metabolism rate and mean glucose metabolism rate-to-plasma insulin concentration ratio (M:I) was evident in the intensified training group during steady state of HEC testing after phases 3 and 4. After phase 4, mean glucose metabolism rate was significantly decreased (from 31.1 ± 6.8 μmol/kg/min to 18.1 ± 3.4 μmol/kg/min), as was M:I (from 1.05 ± 0.31 to 0.62 ± 0.17) during steady state in the intensified training group, compared with phase 3 values for the same horses.
Conclusions and Clinical Relevance—Dissociation between the glucose metabolism rate and M:I in horses that underwent intensified training may reflect non-insulin–dependent increases in glucose metabolism.
Objectives—To determine whether increased glucose metabolism is the potential cause of the decreased plasma glucose curve determined after oral glucose tolerance testing in horses with lower motor neuron degeneration.
Animals—3 horses with signs suggestive of lower motor neuron degeneration, 1 horse with malignant melanoma with multiple metastases, and an obese but otherwise healthy horse.
Procedures—Glucose metabolism was assessed by use of the hyperglycemic clamp and euglycemic hyperinsulinemic clamp techniques.
Results—Mean rate of glucose metabolism of horses with lower motor neuron degeneration was significantly greater (mean, 3.7 times greater than control horses; range, 2.1 to 4.8 times greater) than that reported in 5 healthy control horses (41 ± 13 µmol/kg/min vs 11 ± 4.5 µmol/kg/min, respectively). In addition, one of the affected horses, an 8-year-old warmblood gelding, had a 5.6- times increased sensitivity to exogenously administered insulin, compared with that reported in 5 healthy control horses. Pancreatic insulin secretion was not insufficient in horses with lower motor neuron degeneration. Findings in the 2 diseased control horses were unremarkable.
Conclusions and Clinical Relevance—Increased glucose metabolism in horses with lower motor neuron degeneration may be the cause of the decreased plasma glucose curve detected after oral glucose tolerance testing. This finding could aid in developing supportive treatments with respect to adequate glucose and vitamin E supplementation. (Am J Vet Res 2005;66:271–276)
Objective—To investigate the effects of exercise on activation of mitogen-activated protein kinase (MAPK) signaling proteins in horses.
Animals—6 young trained Standardbred geldings.
Procedure—Horses performed a 20-minute bout of exercise on a treadmill at 80% of maximal heart rate. Muscle biopsy specimens were obtained from the vastus lateralis and pectoralis descendens muscles before and after exercise. Amount of expression and intracellular location of phosphospecific MAPK pathway intermediates were determined by use of western blotting and immunofluorescence staining.
Results—Exercise resulted in a significant increase in phosphorylation of p38 pathway intermediates, c-Jun NH2 terminal kinase (JNK), and heat shock protein 27 (HSP27) in the vastus lateralis muscle, whereas no significant changes were found in phosphorylation of extracellular regulated kinase. In the pectoralis descendens muscle, phosphorylation of p38 and HSP27 was significantly increased after exercise. Immunohistochemical analysis revealed fiber-type– specific locations of phosphorylated JNK in type 2a/b intermediate and 2b fibers and phosphorylated p38 in type 1 fibers. Phosphorylated HSP27 was strongly increased after exercise in type 1 and 2a fibers.
Conclusions and Clinical Relevance—The p38 pathway and JNK are activated in the vastus lateralis muscle after a single 20-minute bout of submaximal exercise in trained horses. Phosphorylation of HSP27 as detected in the study reported here is most likely induced through the p38 signaling pathway.
Objective—To investigate the expression and localization of glucose transporter 4 (GLUT4) and fatty acid translocase (FAT/CD36) in equine skeletal muscle.
Sample Population—Muscle biopsy specimens obtained from 5 healthy Dutch Warmblood horses.
Procedures—Percutaneous biopsy specimens were obtained from the vastus lateralis, pectoralis descendens, and triceps brachii muscles. Cryosections were stained with combinations of GLUT4 and myosin heavy chain (MHC) specific antibodies or FAT/CD36 and MHC antibodies to assess the fiber specific expression of GLUT4 and FAT/CD36 in equine skeletal muscle via indirect immunofluorescent microscopy.
Results—Immunofluorescent staining revealed that GLUT4 was predominantly expressed in the cytosol of fast type 2B fibers of equine skeletal muscle, although several type 1 fibers in the vastus lateralis muscle were positive for GLUT4. In all muscle fibers examined microscopically, FAT/CD36 was strongly expressed in the sarcolemma and capillaries. Type 1 muscle fibers also expressed small intracellular amounts of FAT/CD36, but no intracellular FAT/CD36 expression was detected in type 2 fibers.
Conclusions and Clinical Relevance—In equine skeletal muscle, GLUT4 and FAT/CD36 are expressed in a fiber type selective manner. ( Am J Vet Res 2004;65:951–956)
Objective—To investigate whether protein kinase C (PKC) isoforms are expressed in equine skeletal muscle and determine their distribution in various types of fibers by use of immunofluorescence microscopy.
Animals—5 healthy adult Dutch Warmblood horses.
Procedure—In each horse, 2 biopsy specimens were obtained from the vastus lateralis muscle. Cryosections of equine muscle were stained with PKC isoform (α, β1, β2, δ, ξ, or ζ)-specific polyclonal antibodies and examined by use of a fluorescence microscope. Homogenized muscle samples were evaluated via western blot analysis.
Results—The PKC α, β1, β2, δ, ξ, and ζ isoforms were localized within the fibers of equine skeletal muscle. In addition, PKC α and β2 were detected near or in the plasma membrane of muscle cells. For some PKC isoforms, distribution was specific for fiber type. Staining of cell membranes for PKC α was observed predominantly in fibers that reacted positively with myosin heavy chain (MHC)-IIa; PKC δ and ξ staining were more pronounced in MHC-I-positive fibers. In contrast, MHC-I negative fibers contained more PKC ζ than MHC-I-positive fibers. Distribution of PKC β1 was equal among the different fiber types.
Conclusions and Clinical Relevance—Results indicated that PKC isoforms are expressed in equine skeletal muscle in a fiber type-specific manner. Therefore, the involvement of PKC isoforms in signal transduction in equine skeletal muscle might be dependent on fiber type. ( Am J Vet Res 2004; 65:69–73)
Objective—To investigate the effects of acute exercise and long-term training on Na+,K+-ATPase content, mRNA isoforms, and protein concentration in equine muscle.
Procedures—Horses performed a bout of exercise on a treadmill before and after 18 weeks of combined interval and endurance training. Muscle biopsy specimens were obtained from vastus lateralis muscle (VLM) and pectoralis descendens muscle (PDM) before and after exercise. The Na+,K+-ATPase content, mRNA isoforms, and protein concentrations were determined by use of [3H]ouabain binding, real-time PCR assay, and western blotting, respectively.
Results—6 Na+,K+-ATPase mRNA isoforms were present in equine muscle, but only A2 and B1 proteins were detected. Exercise before training resulted in increases of mRNA isoforms A1, A2, A3, and B2 in VLM and A1 and B3 in PDM. Training increased resting values for mRNA isoforms A3 and B1 in VLM and B3 in PDM. The Na+,K+-ATPase, [3H]ouabain binding, and proteins of mRNA A2 and B1 increased in VLM, whereas in PDM, only A2 protein increased as a result of training. After training, effects of strenuous exercise on mRNA expression were no longer detectable.
Conclusions and Clinical Relevance—Equine muscle contained all Na+,K+-ATPase mRNA isoforms, but only A2 and B1 proteins could be detected. Expression of these isoforms changed as a result of strenuous exercise and long-term training, representing an adaptive response. Determination of Na+,K+-ATPase gene expression may be relevant for understanding alterations in excitability during neuromuscular diseases.
Objective—To determine the effects of short-term IV administration of hydrocortisone or equine growth hormone (eGH) or long-term IM administration of eGH to horses on tissue sensitivity to exogenous insulin.
Animals—5 Standardbreds and 4 Dutch Warmblood horses.
Procedure—The euglycemic-hyperinsulinemic clamp technique was used to examine sensitivity of peripheral tissues to exogenous insulin 24 hours after administration of a single dose of hydrocortisone (0.06 mg/kg), eGH (20 µg/kg), or saline (0.9% NaCl) solution and after long-term administration (11 to 15 days) of eGH to horses. The amounts of metabolized glucose (M) and plasma insulin concentration (I) were determined.
Results—Values for M and the M-to-I ratio were significantly higher 24 hours after administration of a single dose of hydrocortisone than after single-dose administration of eGH or saline solution. After long-term administration of eGH, basal I concentration was increased and the mean M-to-I ratio was 22% lower, compared with values for horses treated with saline solution.
Conclusions and Clinical Relevance—Increases in M and the M-to-I ratio after a single dose of hydrocortisone imply that short-term hydrocortisone treatment increases glucose use by, and insulin sensitivity of, peripheral tissues. Assuming a single dose of hydrocortisone improves sensitivity of peripheral tissues to insulin, it may be an interesting candidate for use in reducing insulin resistance in peripheral tissues of horses with several disease states. In contrast, long-term administration of eGH decreased tissue sensitivity to exogenous insulin associated with hyperinsulinemia. Therefore, increased concentrations of growth hormone may contribute to insulin resistance in horses with various disease states. (Am J Vet Res 2005;66:1907–1913)
Objective—To evaluate alterations in skeletal muscle carnitine metabolism during exercise and training by measuring changes in plasma acylcarnitine concentrations in Standardbreds.
Animals—10 Standardbred geldings with a mean ± SD age of 20 ± 2 months and weight of 384 ± 42 kg.
Procedures—In a 32-week longitudinal study, training on a treadmill was divided into 4 phases as follows: phase 1, acclimatization for 4 weeks; phase 2, 18 weeks with alternating endurance and high-intensity exercise training; phase 3, increased training volume and intensity for another 6 weeks; and phase 4, deconditioning for 4 weeks. In phase 3, horses were randomly assigned to 2 groups as follows: control horses (which continued training at the same level as in phase 2) and high-intensity exercise trained horses. At the end of each phase, a standardized exercise test (SET) was performed. Plasma acylcarnitine, fatty acids, and lactic acid and serum β-hydroxybutyric acid (BHBA) concentrations were assessed before and at different time points after each SET.
Results—Plasma lactic acid, total nonesterified fatty acids, 3-hydroxyisobutyric acid, and acetylcarnitine (C2-carnitine) concentrations significantly increased during SETs, whereas serum BHBA, plasma propionylcarnitine (C3-carnitine), and plasma butyryl- and isobutyrylcarnitine (C4-carnitine) concentrations decreased significantly, compared with those before SETs.
Conclusions and Clinical Relevance—Our findings indicated that the plasma acylcarnitine profile in horses likely reflects skeletal muscle carnitine metabolism following exercise, thereby providing a possible practical method to investigate potential disorders in carnitine metabolism in horses with myopathy.