Effects of transport and racing on ionic changes in thoroughbred race horses.

1. Packed cell volume (PCV), blood glucose, total plasma proteins (TPP) and plasma electrolytes, osmolality, cortisol and aldosterone alterations produced by transport and racing, were investigated in race horses. 2. Plasma cortisol, sodium and blood glucose, found after transport, were higher, while aldosterone was lower than control levels. 3. After racing, PCV, blood glucose, TPP and plasma cortisol, sodium and osmolality were higher than control, while chloride diminished and aldosterone returned to control values. 4. These results demonstrate that transport and racing are different kinds of stressors, suggesting that the sympathetic system and hypophysis-suprarenal cortex axis have a dissimilar contribution to the physiological response.     

Transcriptional expression changes of glucose metabolism genes after exercise in thoroughbred horses

Physical exercise induces gene expression changes that trigger glucose metabolism pathways in organisms. In the present study, we monitored the expression levels of LDHA (lactate dehydrogenase) and GYS1 (glycogen synthase 1) in the blood, to confirm the roles of these genes in exercise physiology. LDHA and GYS1 are related to glucose metabolism and fatigue recovery, and these processes could elicit economically important traits in racehorses. We collected blood samples from three retired thoroughbred racehorses, pre-exercise and immediately after 30 min of exercise. We extracted total RNA and small RNA (≤ 200 nucleotide-long) from the blood, and assessed the expression levels of LDHA, GYS1, and microRNAs (miRNAs), by using qRT-PCR. We showed that LDHA and GYS1 were down-regulated, whereas eca-miR-33a and miR-17 were up-regulated, after exercise. We used sequences from the 3′ UTR of LDHA and GYS1, containing eca-miR-33a and miR-17 binding sites, to observe the down-regulation activity of each gene expression. We observed that the two miRNAs, namely, eca-miR-33a and miR-17, inhibited LDHA and GYS1 expression via binding to the 3′ UTR sequences of each gene. Our results indicate that eca-miR-33a and miR-17 play important roles in the glucose metabolism pathway. In addition, our findings provide a basis for further investigation of the exercise metabolism of racehorses.     

Effects of racetrack exercise on third metacarpal and carpal bone of New Zealand thoroughbred horses

The response of equine bone to training has not been quantified in racetrack trained horses, only in treadmill exercised horses. Seven two-year-old thoroughbred fillies were trained on sand and grass at a racetrack, in a typical New Zealand flatrace training regime. The horses were exercised 6 days per week for up to 13 weeks. During the day the horses were confined in 4 x 4m sand yards, and were stalled at night. Another 7 fillies of the same age were allowed free exercise in grass yards. The bones of the animals were available after the 13 week experimental period, and were examined using a Siemens Somatom AR CT scanner. To quantify the response of epiphyseal bone, 3mm thick sagittal plane images of the carpus (through the middle of the medial condyle of distal radius) and the distal third metacarpal bone (Mc3) (immediately lateral and medial to the junction of the condyle and the median sagittal ridge) were studied. Appropriate areas of interest were chosen, and the mean tissue density equivalent (Houndsfield Units) was determined. In the carpus, there was a significant effect of exercise in the dorso-distal aspect of the radius (p<0.01), dorsal aspect of radial and third carpal bones (p<0.01 and p<0.001 respectively). In palmaro-distal subchondral bone of Mc3, there was a significant effect on the medial/lateral site (p<0.01), which differed between right and left legs, probably due to the effect of the horses having been trained in one direction around the training track. The mean tissue density of the Mc3 epiphysis of the exercised group was 36.8% greater than that of the non-exercised group (p<0.001). The study demonstrates that bone response is both rapid and substantial, which should prompt the use of non-invasive diagnostic aids to determine the stage of training in which tissue density changes occur.     

Role of the antioxidant ascorbate in hibernation and warming from hibernation

Ground squirrels tolerate up to 90% reductions in cerebral blood flow during hibernation as well as rapid reperfusion upon periodic arousal from torpor without apparent neurological damage. Thus, hibernation is studied as a model of tolerance to cerebral ischemia and other types of brain injury. Metabolic suppression likely plays a primary adaptive role that allows hibernating species to tolerate dramatic fluctuations in blood flow. Several other aspects of hibernation physiology are also consistent with tolerance to ischemia and reperfusion suggesting that multiple neuroprotective adaptations may work in concert during hibernation. The purpose of the present work is to review evidence for enhanced antioxidant defense systems during hibernation, with a focus on ascorbate, and discuss potential roles of these antioxidants during hibernation. In concert with dramatic decreases in blood flow, nutrient and oxygen delivery, plasma concentrations of the antioxidant ascorbate [(Asc)p] increase 3-5-fold during hibernation. In contrast, during re-warming, [Asc]p declines at a relatively rapid rate that peaks at the time of maximal O(2) consumption. This peak in O(2) consumption also coincides with a brief rise in plasma urate concentration consistent with a surge in reactive oxygen species production. Overall, data suggest that elevated concentration of plasma ascorbate is poised for distribution to metabolically active tissues during the surge in oxidative metabolism that accompanies re-warming during hibernation. This pool of ascorbate, as well as increased expression of other antioxidant defense systems, may protect vulnerable tissues from oxidative stress during hibernation and re-warming from hibernation. Better understanding of the role of ascorbate in hibernation may guide use of ascorbate and other antioxidants in treatment of stroke, head trauma and neurodegenerative disease.     

Effect of tryptophan and of glucose on exercise capacity of horses

We hypothesized that central fatigue may have a role in limitingthe endurance capacity of horses. Therefore, we tested the effect ofinfusing tryptophan and/or glucose on endurance time and plasmaconcentrations of free tryptophan and other substrates thought toaffect tryptophan uptake into the brain of seven mares (3-4 yr ofage, 353-435 kg) that ran on a treadmill at 50% of maximalO₂ consumption to fatigue. Withuse of a counterbalanced crossover design, the horses were infused withtryptophan (100 mg/kg in saline solution) or a similar volume of salinesolution (placebo) before exercise. During exercise, horses receivedinfusions of glucose (2 g/min, 50% wt/vol) or a similar volume ofsaline. Thus the treatments were 1)tryptophan and glucose (T & G), 2) tryptophan and placebo (T & P), 3)placebo and glucose (P & G), and 4)placebo and placebo (P & P). Mean heart rate, hematocrit, andconcentration of plasma total solids before and during exercise weresimilar for all trials. Mean time to exhaustion was reduced (P < 0.05) for T & P and T & Gcompared with P & P [86.1 ± 6.9 and 87.1 ± 6.8 vs. 102.3 ± 10.3 (SE) min], whereas endurance for P & G(122.4 ± 11.9 min) was greater than for all other trials (P < 0.05). Compared withnontryptophan trials, during the tryptophan trials plasma prolactinincreased (P < 0.05) nearlythreefold before exercise and almost twofold early in exercise. Muscleglycogen concentrations were reduced(P < 0.05) below preexercise values in the P & G and P & P trials only. However, glucose infusions (P & G)did not affect (P > 0.05)concentrations of plasma free fatty acids or ratios of branched-chainamino acids to free tryptophan. In conclusion, tryptophan infusionreduced endurance time, which was consistent with the central fatiguehypothesis. The failure of glucose infusion to alleviate the effects oftryptophan and the absence of significant muscle glycogen reduction inthe tryptophan trials suggest that the early onset of fatigue in thetryptophan trials is not due to a lack of readily available substrate.

     

Amino acids and plasma antioxidant capacity

Summary Amino acids antioxidant capacity has been investigated and compared with the chain-breaking antioxidant activity of known compounds as ascorbic acid and Trolox. Basic and acidic amino acids and most of neutral ones showed no antioxidant capacity. On the contrary, tryptophan, tyrosine, cysteine and homocysteine showed antioxidant ability at concentrations which are within the usually reported physiological ranges.These findings are discussed in connection with the antioxidant capacity ascribed to plasma proteins, as human serum albumin.     

Running and shipping elevate plasma levels of beta-endorphin-like substance (B-END-LI) in thoroughbred horses

A specific RIA for beta-endorphin (B-END) was developed to measure horse plasma levels of B-END-like material (B-END-LI) during exercises and shipping. Three exercise speeds and durations were: trot at 260-300 m/min for 10 min; slow gallop at 390-420 m/min for 5 min and fast gallop at 700-800 m/min for 2 min. Blood samples were taken from 4 horses before, immediately after, 30 and 60 min after exercise. Trotting increased plasma B-END-LI from a basal level of 109 +/- 7 pg/ml to 172 +/- 22 at the end of exercise and returned to 127 +/- 17 and 107 +/- 10 pg/ml at 30 and 60 min after exercise. Similar results were obtained in slow gallop (121 +/- 6 to 210 +/- 17 then 155 +/- 8 and 131 +/- 11 pg/ml). However, fast gallop caused the greatest increase (352%) in B-END-LI to concentrations of 544 +/- 93 pg/ml and 276 +/- 74 pg/ml at 5 and 30 min after exercise. Plasma B-END-LI returned to 199 +/- 46 pg/ml in 1 hr. Sequential exercises of trot, slow and fast gallop were conducted in 6 horses. Plasma B-END-LI were 116 +/- 19 pg/ml (pre-exercise), 198 +/- 21 (trot), 361 +/- 51 (slow gallop), 500 +/- 57 (fast gallop) and 248 +/- 29, 171 +/- 24, 143 +/- 20 and 139 +/- 21 pg/ml at 0.5, 1, 2 and 3 hr, respectively, following exercises. Transportation in horse trailer also significantly increased plasma levels of B-END-LI from a basal level of 138 +/- 12 to 196 +/- 24 pg/ml within 30 min and this levels were maintained at 45 min (177 +/- 3 pg/ml). Plasma levels of B-END-LI began to decline at 60 min of shipping. These results showed that plasma B-END-LI was increased in all speeds of exercise and by shipping and returned to pre-exercise and pre-shipping level in 30 min except fast gallop which returned to pre-exercise level in 1 hr.     

Role of selenium on antioxidant capacity in methomyl-treated mice

Methomyl carbamate is a pesticide widely used in the control of insects. The present work aims at studying the effect of selenium on the antioxidant system of methomyl-treated mice. Swiss albino mice were intraperitoneally administered a single dose of methomyl (7 mg/Kg body weight). Mice of another group were injected with sodium selenite (5 pmole/Kg b.wt.) 7 days before methomyl intoxication. After 24 hours, methomyl exposure resulted in significant increase in lactic dehydrogenase activity (LDH). The antioxidant capacity of hepatic cells in terms of the activities of superoxide dismutase (SOD), catalase (CAT), glutathione reductase (GR), glutathione-S-transferase (GST) and glutathione (GSH) content was diminished. It appears that methomyl exerts its toxic effect via peroxidative damage to hepatic, renal and splenic cell membranes. Also, methomyl induced DNA damage in these organs as detected by alkaline filter elution technique. The distribution of methomyl in different organs of mice was detected by HPLC. Selenium administration prior to methomyl injection produced pronounced protective action against methomyl effects. It is observed that selenium enhances the endogenous antioxidant capacity of the cells by increasing the activities of SOD, CAT, GR and GST as well as increasing GSH content. The activity of LDH was decreased in blood and the damage of DNA was suppressed comparable to controls. In conclusion, the adverse effects of methomyl in mice could be ameliorated by selenium.     

Effect of gender differences and voluntary exercise on antioxidant capacity in rats

The effects of gender difference and voluntary exercise on antioxidant capacity in rats were evaluated. The subjects were divided into two groups, physically active and sedentary. In the sedentary group, the level of hydroxyl radical in the liver was higher (P<0.001) in male rats than in female rats, however, in the physically active group, the level in male rats was lower (P<0.05) than in female rats. The levels of reduced glutathione (GSH) in physically active males and females were higher compared to those in the sedentary group. The physically active group also showed an increase in antioxidant enzymes, such as glutathione peroxidase (GPx), glutathione reductase (GR) and superoxide dismutase activities. The level of liver GSH was higher in physically active females than in physically active males. For both groups, GPx and GR activities in females were significantly higher than in males. These results indicate that female rats have an intrinsically higher antioxidant capacity, which resulted in increased levels of GSH via the glutathione redox cycle and gamma-glutamyl cycle enzymes. The adaptation to altered antioxidant capacity, induced by physical activity, appeared to be affected by gender differences.     

Beer increases plasma antioxidant capacity in humans

The positive association of a moderate intake of alcoholic beverages with a low risk for cardiovascular disease, in addition to ethanol itself, may be linked to their polyphenol content. This article describes the effect of acute ingestion of beer, dealcoholized beer, and ethanol (4.5% v/v) on the total plasma antioxidant status of subjects, and the change in the high performance liquid chromatography profile of some selected phenolic acids (caffeic, sinapic, syringic, and vanillic acids) in 14 healthy humans. Plasma was collected at various times: before (T0), 1 hour after (T1), and 2 hours after (T2) drinking. The study is part of a larger research planned to identify both the impact of brewing on minor components potentially present in beer and their metabolic fate in humans. Beer was able to induce a significant (P < 0.05) increase in plasma antioxidant capacity at T1 (mean +/- SD: T0 1,353 +/- 320 microM; T1 1,578 +/- 282 microM), returning close to basal values at T2. All phenolic acids measured in plasma tended to increase after beer intake (20% at T1, 40% at T2). Syringic and sinapic acid reached statistical significance (P < 0.05 by one-way analysis of variance-Fisher's test) at T1 and T2, respectively. Plasma metabolic parameters (glucose, total cholesterol, triglycerides, and uric acid) and plasma antioxidants (alpha-tocopherol and glutathione) remained unchanged. Ethanol removal impaired the absorption of phenolic acids, which did not change over the time of the experiment, accounting for the low (and not statistically significant) increase in plasma antioxidant capacity after dealcoholized beer drinking. Ethanol alone did not affect plasma antioxidant capacity or any of the antioxidant and metabolic parameters measured.     

Selenium deficiency, endurance exercise capacity, and antioxidant status in rats

Increased O2 metabolism imposed by physical exercise is likely to augment the production of active O2 species that have been shown to react with lipids, proteins, and DNA. Antioxidants and antioxidant enzymes, such as the selenium enzyme glutathione peroxidase, minimize or prevent such potentially toxic reactions. This study shows that selenium deficiency decreases glutathione peroxidase activity in liver and muscle (less than 80%, P less than 0.001), increases total glutathione in liver, muscle, and plasma (P less than 0.05) and increases muscle cytochrome oxidase activity, and ubiquinone content (P less than 0.05) but has no effect on endurance capacity. Exercise to exhaustion resulted in a significant (P less than 0.001) elevation of total and oxidized glutathione (GSSG) and a significant (P less than 0.05) decrease of vitamin E in plasma of control and selenium-deficient rats. Acute exercise also increased tissue GSSG levels in both control and selenium-deficient groups of rats. Hence, despite a large depletion of selenium-deficient glutathione peroxidase, pronounced oxidation of glutathione to GSSG can be produced by the increased oxidative metabolism during physical exercise. The results suggest that the residual glutathione peroxidase activity is sufficient to detoxify hydroperoxides in exercising selenium-deficient animals and to prevent the impairment of endurance capacity.     

Skeletal muscle oxidative capacity, antioxidant enzymes, and exercise training

The purposes of this study were to determine whether exercise training induces increases in skeletal muscle antioxidant enzymes and to further characterize the relationship between oxidative capacity and antioxidant enzyme levels in skeletal muscle. Male Sprague-Dawley rats were exercise trained (ET) on a treadmill 2 h/day at 32 m/min (8% incline) 5 days/wk or were cage confined (sedentary control, S) for 12 wk. In both S and ET rats, catalase (CAT), superoxide dismutase (SOD), and glutathione peroxidase (GPX) activities were directly correlated with the percentages of oxidative fibers in the six skeletal muscle samples studied. Muscles of ET rats had increased oxidative capacity and increased GPX activity compared with the same muscles of S rats. However, SOD activities were not different between ET and S rats, but CAT activities were lower in skeletal muscles of ET rats than in S rats. Exposure to 60 min of ischemia and 60 min of reperfusion (I/R) resulted in decreased GPX and increased CAT activities but had little or no effect on SOD activities in muscles from both S and ET rats. The I/R-induced increase in CAT activity was greater in muscles of ET than in muscles of S rats. Xanthine oxidase (XO), xanthine dehydrogenase (XD), and XO + XD activities after I/R were not related to muscle oxidative capacity and were similar in muscles of ET and S rats. It is concluded that although antioxidant enzyme activities are related to skeletal muscle oxidative capacity, the effects of exercise training on antioxidant enzymes in skeletal muscle cannot be predicted by measured changes in oxidative capacity.     

Measuring the antioxidant capacity of blood plasma using potentiometry

The use of potentiometry to measure plasma antioxidant capacity to contribute to oxidative stress evaluation is presented. In this assay, plasma (n = 60) diluted (0.3 to 1 ml) in phosphate buffer, pH 7.4, NaCl 9%, was submitted to potentiometry. A platinum wire was the working electrode and saturated calomel the reference. The results are presented as the difference between sample and buffer potential (ΔE). ΔE presented a good inverse correlation with added increasing concentrations of ascorbate (2.5−75 μmol/L; R = −0.99), urate (9.0−150 μmol/L; R = −0.99), and bilirubin (0.78−13 μmol/L; R = −0.99). Increase in the antioxidant capacity decreased ΔE. Depletion of the antioxidant capacity by tert-butylhydroperoxide (6.5−50 μmol/L) presented a direct correlation (0.97) with ΔE. Furthermore, ΔE presented an inverse correlation (R = −0.99) with increased antioxidant capacity of plasma (FRAP) induced by the addition of ascorbate (2.5−75 μmol/L). The response of the potentiometric method proved be adequate for measuring the plasma antioxidant depletion induced by acute exhaustive exercise in rats (control, n = 15; exercised, n = 15). This exercise decreased the concentration of urate (p < 0.05), decreased FRAP (p < 0.5), increased TBARS (p < 0.5), and decreased the potentiometer sensor response (p = 6.5 × 10⁻³). These results demonstrate the adequacy of potentiometry for evaluating the antioxidant capacity of blood plasma samples.     

Differential effects of acetaminophen on enzymatic and non-enzymatic antioxidant factors and plasma total antioxidant capacity in developing and adult rats

Recently we reported that ferric reducing ability of plasma (FRAP) assay, as an index of total antioxidant activity, increases in growing rats in response to high dose of vitamin K. In this study, it was found that acetaminophen (APAP) can cause elevation in FRAP in suckling and adult rats. This study was initiated to assess the contribution of individual antioxidant factors on elevation in FRAP. A surge in FRAP, 1 h after high dose APAP (250 or 450 mg/kg BW) administration was recorded in both young as well as adults. Whereas, low dose drug (25 mg/kg) failed to alter FRAP in both the age groups. Time-course studies show that drug-dependent elevation in FRAP begin rapidly, reaching a maximum at 1 h (> 500%). Increased FRAP was associated with a marked increase (∼14-fold) in plasma bilirubin, 6 h after drug administration at 450 mg/kg only in suckling rats. Similarly, APAP-related increase in superoxide dismutase activity in erythrocytes was limited to young rats of both the age groups. Other factors measured during this period viz., plasma uric acid, bilirubin and total protein together with catalase activity of erythrocytes remained unchanged in treated rats. Under these circumstances, APAP-related depletion in liver glutathione was almost similar in both the age groups. During a 12 h study, the concentration of lipid peroxidation products, in liver of treated groups remained within the levels of respective controls. The endpoint hepatotoxic effects of APAP was almost similar in both the age groups, suggesting that like adults, immature rats can cope with toxic effects of APAP owing to their drug-dependent induction in certain antioxidant factors.     

The effect of one year's swimming exercise on oxidant stress and antioxidant capacity in aged rats

The effect of exercise on oxidant stress and on alterations in antioxidant defense in elderly has been investigated extensively. However, the impact of regularly performed long-term physical activity starting from adulthood and prolonged up to the old age is not yet clear. We have investigated the changes in the activities of antioxidant enzymes - superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPx) - and lipid peroxidation in various tissues of rats which had performed (old-trained) or had not performed (old-control) regular swimming exercise for one year. These animals were compared with young-sedentary rats. Increased lipid peroxidation was observed with ageing in all tissues (heart, liver, kidney, striated muscle) and swimming had no additional effect on this elevation of lipid peroxidation. Heart and striated muscle SOD activites, and striated muscle CAT activity increased as a consequence of ageing, whereas kidney and liver CAT activities, as well as GPx activities in kidney, liver, lung and heart were significantly decreased compared to young controls. Lung and heart SOD, liver CAT activities as well as GPx activities in liver, lung and heart were increased significantly in rats which performed exercise during ageing, compared to the old-control group. These findings suggest that lifelong exercise can improve the antioxidant defense in many tissues without constituting any additional oxidant stress.