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Problem statement: There are limited data on age related changes in the physical, hematological and biochemical parameters of endurance horses. Therefore, the purpose of this study was to investigate any differences in these parameters associated to aging. Approach: This study was conducted in Malaysia to determine the post-race hematological, blood electrolyte, biochemical and physical parameters of endurance horses based on age for the eliminated horses and those that completed the races with good performance. Whole blood, plasma and serum samples were collected after each race. Results: After physical examination, n=12(6-10) years, n=33(11-15) years, n=12(16-20) years, n=9(21-25) years, and n=6(26-30) years were eliminated from the race while the good performance horses that completed the race successfully are n=3(6-10) years and n=6(11-15) years. The mean heart rate of the good performance horses in the 11-15 years category was 54±7.9 beats per minute and was lower than that of the eliminated horses in all age brackets. The blood lactate concentration of good performance horses was 9.6±2.3 mmol L-1 in the 11-15 years category, which was significantly higher than the eliminated horses in the entire age category. The blood glucose concentration was lower in the good performance endurance horses within the 6-10 years bracket with 1.3±0.8mmol L-1 than the eliminated endurance horses in the entire age category. Conclusion: The study showed that eliminated horses within the 16-20 years bracket exhibited poorer glucose utilization than the good performance horses within the 6-10 years bracket, these may have resulted in poor lactate production. Thus the blood lactate and glucose concentrations of horses during training may be used to predict performance based on age in endurance races.
Key words: Hematological, biochemical parameters, physical parameters, glucose, endurance horses, plasma electrolyte, Lactate Dehydrogenase (LDH), Ethyl Diaminotetra-Acetic Acid (EDTA), and age.
Recent studies have showed that up to 15% of the horse population are over 20 years of age with many of these horses are able to perform in athletic competitions (Robert, et al., 2004). Several horses are in their late teens and are in the peak of their performance careers, competing in endurance rides, dressage, jumping, three-day eventing, and other athletic competitions. Age seems to affect cardiopulmonary function in the horse (McKeever and Malinowski, 1997; McKeever et al., 1998). It has also been indicated that older horses may be at higher risk of developing hyperthermia due to changes in fluid and electrolyte status giving way to the derangement of cardiovascular function and heat loss mechanisms (McKeever et al., 2000). While recent studies have elaborated on age-related changes in central factors affecting exercise ability in the horse, other studies have examined peripheral changes in relation to aging (Robert, et al., 2004).
Adaptation of muscular fibers to training begins at an early age and this potentiates their citrate synthetase activity in the muscles, as well as type IIA and type IIB fiber proportions. The adaptational changes, such as the increased type IIA and type IIB muscles fiber ratios and increase oxidative enzymes capacity of the muscles are, however, to a great extent are influenced by training (Robert, et al., 2004).
The intermediate age group (10-16)years is an age where many horses are still in their peak athletically and are considered to be physiologically analogous to 40 years and above in humans (Robert, et al., 2004).
The distribution of muscle fiber type is one set of measures of muscle function that may be changed by aging. Other measures that may have a more vital effect on the ability to perform exercise are body composition and, more importantly, total fat-free mass (FFM). The larger component of a horse's FFM is muscle mass and a recent study has documented the strong correlation between FFM and performance in elite horses (Robert, 2004).
As one advances in age, the oxidative capability of human skeletal muscle also declines (Rooyackers et al., 1996; Conley et al., 2000). Evidence of the decreased oxidative capability is indicated in both the rate of mitochondrial protein biosynthesis and activity of oxidative enzymes such as cytochrome c oxidase and citrate synthase (Rooyackers et al., 1996; Houmard et al., 1998).
The reduction in the oxidative capability seems to be associated to the age-related decrease in aerobic capacity and muscle performance. There is also evidence of reduced glycolytic capacity in aged skeletal muscle (Welle et al., 2000). These age-associated adaptations are due to physiological changes and exacerbated by physical inactivity during the ageing process (Kim, et al., 2005).
The major physiological adaptations that can directly influence exercise capacity and stamina of endurance horses include the ability and efficiency of gas exchange, oxygen uptake and delivery to the exercising muscles. The working muscle of endurance horses depends on aerobic metabolism of its glycogen stores, blood fatty acids and volatile fatty acids from hindgut fermentation, heart size and capacity to deliver large volumes of blood to the tissue (Lawan, et al., 2010).
Many factors can affect the hematological and biochemical parameters including the breed, age and type of exercise. Also, the effect of age on several parameters has been observed in different breeds and age groups (McKeever et al., 1998).
Measurement of the fitness or exercise tolerance of a horse is by assessment, through physical examination of heart rates and respiratory rates (Cottin et al., 2006; Bashir and Rasedee, 2009). Hematological and biochemical alterations can also be analyzed by obtaining the post-ride blood samples (Valberg, 2009). Post-ride blood lactate concentrations are sometimes used to show fitness of the horse. As fitness increases, post-ride blood lactate concentrations of the horse should become low. In fact, maximal blood lactate steady state concentration and anaerobic threshold have been indicated to predict training and performances (Gondim et al., 2007). In endurance competitions, stress and fatigue are vividly shown by changes in circulating erythrocytes and leucocyte numbers and in creatinine concentration of the horses. Tissue remodeling can also occur in endurance competitions and this is indicated by changes in plasma fibrinogen, urea, proteins and creatine kinase (Benamou-Smith et al., 2006). Endurance horses require calcium for muscle contractions and low plasma levels of calcium during strenuous endurance rides can lead to metabolic problems and failures, including synchronous diaphragmatic flutter. However, high blood calcium concentration is needless because it may increase the frequency of thumps during endurance competitions (Lewis, 1995). In tying-up, horses' exhibit increased muscle enzymes, Creatine Kinase (CK), aspartate aminotransferase (AST) and Lactate Dehydrogenase (LDH) (Hodgson and Rose, 1994).
It is known that factors affecting lung health can have added effect in the horse. Over a long time older horses can be exposed to more pathogens and allergens that can lead to small airway diseases. Pathologies like hyperreactive airway disease and chronic obstructive pulmonary disease, exercise-induced pulmonary hemorrhage, tend to be more prevalent in older animals (McKeever, 2003).
The ageing process typically is responsible for physical derangements such as a reduction in muscle force generating capacity and impaired mobility in mammals. This phenomenon appears to be primarily due to reduction in the ability of muscles to generate and sustain power output in association with changes in muscular structure and function during ageing (Kim, et al, 2005).
MATERIALS AND METHODS
Subjects: Eighty-one endurance horses that participated in three endurance competitions each consisting of, n=12(6-10) years, n=33(11-15) years, n=12(16-20) years, n=9(21-25) years, and n=6(26-30) years were eliminated from the race while the good performance horses that completed the race successfully are n=3(6-10) years and n=6(11-15) years were sampled. Among these, 72 horses were eliminated and 9 horses completed the race successfully.
Veterinary inspection: Veterinary inspection was conducted after each leg of the race on all competing horses and physical parameters recorded. The physical parameters evaluated were the resting heart rate (44-64 = normal, 65-70 = high, 71-90 = very high); mucous membrane (1 = normal, 2 = moderately congested, 3 = severe congestion); capillary refill time (1 sec = normal, 2 sec = moderate, 3 sec = severe); skin recoil (1 = normal skin, 2 = moderate dehydration, 3 = severely dehydrated); gut motility or sound (1 = normal, 2 = moderate, 3 = no motility or sound) and gait (1 = normal, 2 = moderate limp, 3 = severe limp). The horses were also observed for soreness or injuries on the back, withers, girth area, body or distal extremities.
Sampling: Blood samples were obtained from all horses, anticoagulated with Ethyl Diaminotetra-Acetic Acid (EDTA) for hematological analysis and lithium heparin for biochemical analysis. The hematological parameters determined were erythrocyte, leucocyte thrombocyte counts and hemoglobin concentration (Cell DYN 3700, Abbot) and hematocrit (PCV) (Hettich-Hematocrit 210 and Hawksley microhematocrit reader) and differential leucocyte count, The plasma electrolyte and biochemical, sodium, potassium, chloride, calcium, urea, creatinine, bilirubin,
Aspartate Transaminase (AST), Creatine Kinase (CK), glucose, lactate, total protein, albumin and globulin concentrations were determined by chemistry analyzer (Hitachi 920) using standard diagnostic kits (Roche).
Seventy-two horses were eliminated from the endurance competition. Only nine horses managed to complete the race without metabolic signs and all were from the n=3(6-10) years and n=6(11-15) years category. All horses from the n=33(11-15) years, n=12(16-20) years, n=9(21-25) years, and n=6(26-30) years categories were eliminated because of metabolic or physical derangements. Horses that completed the races with good performance between the age of 11-15 years bracket have showed a lower mean heart rate (Table 5). The hematology, plasma/serum electrolyte and biochemical parameters are presented in Table 1-4. The mean segmented neutrophils is higher in the eliminated endurance horses within the 26-30 years bracket than other age groups among the eliminated horses and also compared to the good performance horses in the 6-10 and 11-15 years bracket. The most significant change in hematology and plasma biochemistry parameters were lactate concentrations which were significantly higher in good performance horses within the 11-15 years bracket than those eliminated from the endurance race in the same age bracket and other age brackets. The plasma glucose concentrations were lower in the good performance within the 6-10 years bracket than the eliminated horses in the same age bracket and the other age brackets. There were no tremendous changes in the other parameters.
Recent studies have showed that up to 20% of the horse population in Malaysia are over 20 years of age with many of these horses being eliminated from athletic competitions. Several horses are in their late teens and are in the peak of their performance careers, competing in endurance rides. Owing to the small number of these horses they are continuously being circulated monthly for endurance races subsequently leading to inappropriately conditioning.
This study was conducted on horses between the ages of 6-30 years participating in endurance competitions to determine the effect of age on the physical, hematology and blood biochemistry parameters. The result showed that only approximately 10% of these horses managed to complete the races in good condition. The rest of the horses were eliminated from the endurance race.
Age seems to affect cardiopulmonary function and tissue oxygenation in the endurance horses, subjecting the older endurance horses to have higher risk of developing hyperthermia due to changes in fluid and electrolyte status giving way to a large percentage of these horses eventually being eliminated from the endurance race because of various metabolic and physical disorders. These are also dependent on the oxygen carriage capacity of blood, which is dependent on erythrocyte number and hemoglobin concentrations.
Endurances horses that have metabolic disorders in races could not produce adequate muscle performance either because of insufficient oxidation or poor glucose utilization to produce enough energy for muscular actions during endurance rides. During rides the blood lactate concentrations are expected to be high in these horses. The value of blood lactate in the horse is used as an indicator of performance and fatigue (Lawan, et al., 2010).
It has been indicated, that during endurance rides blood lactate concentrations increase, but the levels seldom exceed 4 mmol L-1 even in sick horses. In fact the speed of the horse when the blood lactate concentration attains 4 mmol L-1 (VLA4) is used as an indicator of fitness of horse during conditioning (Fielding et al., 2009; Lindner et al., 2009; 2010). In our study, however, the mean blood lactate concentrations for all horses were higher than 6 mmol L-1. This value was attained after endurance races irrespective of age. Horses that completed the races with good performance had even higher blood lactate concentrations reaching mean values of higher than 9 mmol L-1, which were significantly higher than those in the eliminated horses.
The endurance horses that were eliminated in our study had lower circulating blood lactate and higher blood glucose concentrations than good performance horses. It is not vividly clear why the eliminated horses produced less lactate from tissue metabolism during endurance rides. From the high blood glucose concentration there is a possible indication that decreases in lactate production could be related to poor glucose uptake by the metabolizing tissue. In acute exercise, the glucose transporters in the skeletal muscle, which mediate insulin-responsive utilization of glucose, increases (Hayashi, et al., 1997; Tomás, et al., 2002; Hirshman, et al., 1988; Prenen, et al, 2005; Alhusseini, et al., 2010). Hence, the exercising muscles produce lactate from glucose through the anaerobic mechanisms. Thus from the result of this study, it seems that the insufficient levels of the glucose transporters in the skeletal muscle that leads to the poor performance of the susceptible horses (Hayashi et al., 1997), thus resulting in the increased levels of blood glucose concentrations and low blood lactate in the eliminated horses.
In conclusion, the study indicated that eliminated horses had low blood lactate and increased blood glucose concentrations than the good performance horses. This blood biochemical manifestation in eliminated horses may be related to insufficient muscle tissue metabolism. The parameters may be used as indicators of performance in horses during the conditioning policies.
The researchers really appreciate the effort and assistance offered Mr. Mohamed Halmi Othman, Mr. Abdullah Misron, the staffs of Veterinary Teaching Hospital, University Putra Malaysia especially Mr. Salehuddin and Drs Mohammad Fairuz Jamaluddin, Muhammad Munsiff Kamarudin and Mimi Armiladiana Mohamad for their assistance, advice and encouragement.