Effects of Standardized Exercise Tests on Plasma Thyroid Hormones' Kinetics in Standardbred Racehorses.

This study examined how a standard exercise test (SET) affected (1) thyroid hormones (THs) of horses and (2) the relationship between the VLa4 of horses and TH responses to the exercise in trained Standardbred racehorses (VLa4 is the velocity run at defined conditions at which a blood lactate concentration of 4 mmol/L is determined). Twelve trained Standardbred racehorses (six stallions and six mares) performed SETs until the horses' blood lactate concentration was at or above 4 mmol/L. The horses were divided into three age groups (2, 3, and 4 years old); each group consists of 4 horses respectively (two male and two female), to evaluate the effects of age and sex on hormonal responses to SET. During each SET, blood samples were taken at rest and after each interval and at the end of SET. Blood was analysed for total and free triiodothyronine (T3, fT3) as well as total and free thyroxine (T4, fT4). The statistical model included three fixed factors (SET, sex and age) and their main interactions. ANOVA analysis revealed that T3 and fT3 were significantly influenced by SETs. Plasma T3 and fT3 concentrations were higher in 4-year-old horses compared to the other age groups. All plasma THs concentrations were higher in mares than in stallions. Correlations revealed that a higher VLa4 was negatively related to all THs responses in 2-year-old Standardbred mares only. The SET used to determine VLa4 increased selected THs (T3, fT3); these increases were inversely related to VLa4 and affected by age and sex of the horses. The correlation of VLa4 with thyroid exercise' response might provide some additional information for performance evaluation of Standardbred racehorses, especially for evaluating training adaptation, according to sex, and age. Further studies are necessary to provide support on the value of measuring THs in Standardbred racehorses of different sex and age.

Interspecific two-dimensional visual discrimination of faces in horses (Equus caballus).

In social animals, recognizing conspecifics and distinguishing them from other animal species is certainly important. We hypothesize, as demonstrated in other species of ungulates, that horses are able to discriminate between the faces of conspecifics and the faces of other domestic species (cattle, sheep, donkeys and pigs). Our hypothesis was tested by studying inter-and intra-specific visual discrimination abilities in horses through a two-way instrumental conditioning task (discrimination and reversal learning), using two-dimensional images of faces as discriminative stimuli and food as a positive reward. Our results indicate that 8 out of 10 horses were able to distinguish between two-dimensional images of the faces of horses and images showing the faces of other species. A similar performance was obtained in the reversal task. The horses' ability to learn by discrimination is therefore comparable to other ungulates. Horses also showed the ability to learn a reversal task. However, these results were obtained regardless of the images the tested horses were exposed to. We therefore conclude that horses can discriminate between two dimensional images of conspecifics and two dimensional images of different species, however in our study, they were not able to make further subcategories within each of the two categories. Despite the fact that two dimensional images of animals could be treated differently from two dimensional images of non-social stimuli, our results beg the question as to whether a two-dimensional image can replace the real animal in cognitive tests.

The different hormonal system during exercise stress coping in horses.

The review discusses the hormonal changes during exercise stress. The exercise generally produces a rise of adrenaline (A), noradrenaline (NA), adrenocorticotropic hormone (ACTH), cortisol, glucagon, growth hormone, arginine vasopressine, etc., and a drop of insulin. The hormonal events during reestablishment of homeostasis due to exercise stress can be divided into a catabolic phase, with decreased tolerance of effort, and reversible biochemical, hormonal and immunological changes, and an anabolic phase, with a higher adaptive capacity, and enhanced performance. The two main hormonal axes activated in the catabolic phase are sympathetic-adrenal-medullary system and hypothalamic-pituitary-adrenal (HPA) axis, while in the anabolic phase, growth hormone-insulin-like factor I axis, and gonadal axes. The hormonal responses during exercise and recovery can be regarded as regulatory and integrated endocrine responses. The increase of catecholamines and ACTH is dependent on the intensity of exercise; a marked increase in plasma A occurs during exercises with high emotional content. The response of cortisol is correlated with the duration of exercise, while the effect of exercise duration on b-endorphin changes is highly dependent on the type of exercise performed. Cortisol and b-endorphin changes usually occur in phase, but not during exercises with high emotional content. Glucocorticoids and iodothyronines are involved in meeting immediate energy demands, and a model of functional interactions between HPA axis and hypothalamic-pituitary-thyroid axis during exercise stress is proposed. A modulation of coping responses to different energy demanding physical activities required for sport activities could be hypothesized. This review supports the proposed regulation of hypophysiotropic TRHergic neurons as metabolic integrators during exercise stress. Many hormonal systems (ghrelin, leptin, glucose, insulin, and cortisol) are activated to control substrate mobilizations and utilization. The cardiovascular homeostasis, the fluid and electrolyte balance during exercise are highly dependent on vasoactive hormones (antidiuretic hormone, atrial natriuretic peptide, renin-angiotensin-aldosterone, and prostaglandins) control.

Hematochemical Patterns in Follicular Fluid and Blood Stream in Cycling Mares: A Comparative Note.

The aim of this study was to verify the existence of possible cross-talk between biochemical contents of follicular fluid (FF) and systemic concentrations according to the follicular development of the metabolites: glucose (GLU), lactate (LACT), cholesterol (CHOL), triglycerides (TG), total bilirubin (T BIL), blood urea nitrogen (BUN), and creatinine (CREAT); enzymatic activities: gamma-glutamyl transferase (GGT), lactate dehydrogenase (LDH), alkaline phosphatase (ALP), and aspartate aminotransferase (AST); electrolytes: calcium (Ca), phosphorus (P), sodium (Na), chloride (Cl), potassium (K), magnesium (Mg), and iron (Fe); total proteins (TP) and their fractions: albumin (ALB), α1-, α2-, ß-, and γ-globulins (GLOB) of FF and blood strain and their correlations with follicular size in cycling mares. Systemic concentrations of GLU, T BIL, BUN, Fe, TP, ALB, α-1, and α-2 and δ-GLOB and of ALP, GGT, and AST activities were higher than in the FF (P < .05); LACT, CHOL, and TG were higher in FF than systemic ones (P < .05). Glucose, CHOL, TG, LACT, and T BIL were higher in large follicles than in medium and small follicles (P < .05); however, BUN, Fe, ALP, and AST were lower in large follicles than in medium or small follicles (P < .05). Alkaline phosphatase, GGT, and AST activities decreased in medium and large follicles compared with small follicles (P < .05). These results suggest that the metabolic, enzymatic, electrolytic, and protein composition of FF of growing follicles could occur according to the bloodstream changes; hence, it is possible to presume that the nutritional environment of oocyte and follicular cells could improve the clinical diagnoses of infertility in the mare.

Intrafollicular and systemic serotonin, oestradiol and progesterone concentrations in cycling mares.

The hypothesis that a local serotonergic network might also exist in the follicle of mares remains poorly documented, with exception for humans and laboratory species. For this reason, the aim of the present study was to clarify this possibility, investigating intrafollicular serotonin concentrations of the cycling mare at ovulation time. Sixty ovaries collected from 30 clinically healthy mares of slaughterhouse meat production with clinically normal reproductive tracts after slaughtering were evaluated. Blood samples were taken prior to sacrifice. Follicles were classified in three categories in relation to size, as small (20-30 mm), medium (31-40 mm) and large (>41 mm), and the follicular fluid samples were extracted from each follicle. Intrafollicular and systemic serotonin (5-HT), oestradiol-17ß (E2 ) and progesterone (P4 ) were determined by means of enzyme-linked immunosorbent assay and RIA, respectively. Intrafollicular 5-HT, E2 and P4 concentrations were higher than systemic ones (p < .05). 5-HT concentrations increased in larger compared to medium follicles, without differences compared to small size follicles (p < .05). 5-HT and E2 (r = .79) and 5-HT and P4 (r = .79; p < .05) were positively correlated. 5-HT and P4 concentrations in follicular fluid increased progressively with the increase in follicular size (p < .05). Follicle diameter and E2 (r = .85) and P4 (r = .68) were correlated (p < .05). Since serotonin interacts with steroids, its role on steroidogenesis during growth of the dominant follicle may be suggested.

Effect of transportation on the sympatho-adrenal system responses in horses.

The objective of current study was to evaluate the effect of transportation stress on the circulating adrenaline (A), noradrenaline (NA) and dopamine (DA) responses of stallions, according to the different distances. Forty-two stallions were studied before and after road transportation of 100, 200 and 300km, for a period of 1-3h. An increase in plasma A after 100km (P<0.001) was observed. A similar result was seen in plasma NA after 100km (P<0.001), and 300km (P<0.001). Increases in plasma DA after 100 and 200km (P<0.0001) were observed, with a decrease after 300km (P<0.0001). Significant interactions among groups and times for A (P<0.0001), NA (P<0.0006) and DA (P<0.0001) changes were observed. These results indicate that the sympatho-adrenal system response of horses was greater after short (100km), than medium-longer period of transportation (200 and 300km). This may indicate an adaptation process during transport, considering A and NA as the primary candidate neurotransmitters for the maintenance of homeostatic process to alleviate the effects of the perceived transportation stress.

Testosterone, androstenedione and dehydroepiandrosterone concentrations in pregnant Spanish Purebred mare.

Androgens modulate maternal ovarian activity, embryo implantation and correct placental development. The objective of this study was to establish reference values for testosterone (T), androstenedione (A4) and dehydroepiandrosterone (DHEA) concentrations in pregnant mares. A total of 30 healthy Spanish Purebred mares with an age range of 9.33 ±â€¯3.31 years, were studied during the 11 months of gestation. T, A4 and DHEA concentrations were determined using EIA validated specifically for equines. T increased in the 2nd and 3rd month (P < 0.05), showing a plateau between the 4th and 6th month, decreased from the 7th to the 9th month (P < 0.05) and increased in the 10th month (P < 0.05), reaching the maximum value in the last month of pregnancy (P < 0.05). A4 increased in the 2nd month (P < 0.05), reaching the maximum value in the 3rd month (P < 0.05), decreased in the 4th month, increasing in the 5th and 6th month (P < 0.05). From the 7th month the average values decreased until reaching the minimum at the end of gestation. DHEA progressively increased from the 1st to the 5th month, at which time the maximum mean value was reached (P < 0.05), after a decrease in the 6th and 7th month occurred (P < 0.05), reaching the minimum value in the last month of gestation. T, A4 and DHEA were positive and significantly correlated (P < 0.05). The androgens analyzed in this study can be used as predictive markers of pregnancy in the mare.

The contribution of total and free iodothyronines to welfare maintenance and management stress coping in Ruminants and Equines: Physiological ranges and reference values.

In order to acquire a pattern of thyroid involvement in welfare maintenance in Ruminants and Equines, this review summarizes data concerning the reference values of total and free iodothyronines and their modifications in physiological conditions and in different management conditions (pregnancy, lactation, weaning, growth, isolation, restraint, shearing, confinement and transportation). Thyroidal and extrathyroidal tissues efficiently respond to management practices, giving a differentiated contribution to circulating iodothyronine changes. The hormonal response could be mainly attributed to the intracellular deiodination of T4 to T3. Triiodothyronine (T3) and free iodothyronines (fT3 and fT4) result more responsive to management stress, showing different pattern with species and to various conditions, as to environmental conditions in which activities are performed. Intrinsic seasonal changes of iodothyronines and a significant pregnancy effect for T3 were recorded in mares. Higher, although not significant, T3 and T4 concentrations in barren than pregnant mares were observed in donkeys. A positive significant correlation between T3 and T4 was described only in pregnant donkeys. Moreover, a significant effect of season on T3 and fT3 changes was observed both in pregnant and barren donkeys. A significant lactating effect compared with nonlactating stage for T3 and T4 was recorded in mares. In growing foals, body weight (BW) and age were positively correlated with T3 and negatively correlated with T4, fT4 and fT3. Weaning effects were shown for T3 and fT4 concentrations, indicating that weaning represents a severe stress and the presence of conspecific does not reduce psychological stress in this phase. Lambs showed significant decreased T3 and elevated T4 concentrations two weeks after weaning, with higher concentrations in both males and females compared to 24 h. Significant positive correlations were observed between BW and T4, fT3 and fT4 concentrations in lambs. A T3 decrease was detected after isolation, such as induced by confinement and weaning in lambs. Higher T3 concentration after restraint and shearing than after isolation and significant increases in T4, fT3 and fT4 values after restraint and shearing were recorded. The basal concentrations of fT3 in both the inexperienced and experienced transported horses were significantly higher than in untransported experienced horses. Moreover, increases of T3, T4 and fT4 after short road transportation, and significant correlations between T3 and rectal temperature (RT), body weight (BW) and heart rate (HR), confirmed their important role in coping strategy. Thyroid responsiveness to short transport is similar in domestic donkeys and horses, with a preferential release of T3 in horses. A greatest and constant release of T3 and T4, although differentiated, after simulated transportation and after conventional transport of horses confirmed that the degree of stress induced by confinement and additional stressful stimuli associated to road transportation could differently influence the iodothyronine release. Temperamental Limousin young beef bulls showed lower T4 and fT4 concentrations after prolonged transportation than calm subjects, and a concomitant decrease of circulating ACTH, cortisol, T3 and fT3 concentrations, probably induced by down regulation of HPA axis and cortisol negative feedback. These data reinforce the importance of taking into account the evaluation of iodothyronines, and notably of T3, as markers of welfare and stress and their role in ensuring energy homeostasis and productive and reproductive performances in Ruminants and Equines.

Total and free iodothyronine changes in response to transport of Equidae (Equus asinus and Equus caballus).

In this study the effects of short distance road transport on total and free iodothyronine changes in 12 stallions (Equus asinus and Equus caballus) were evaluated. Donkeys (n = 6) and horses (n = 6) were transported for a distance of 50 km. Blood samples were collected 1 week before transport in basal conditions, 1 week later immediately before loading, and after transport and unloading. After transport, donkeys showed significant increases in circulating T4 (P≤0.01), fT3 (P≤0.001), and fT4 (P≤0.01) levels; while horses had significant increases in circulating T3, fT3 and fT4 (P≤0.01) levels. Compared to donkeys' values, horses showed lower T4 values in basal condition, before and after transport (P≤0.001); higher fT3 values in basal condition and before (P≤0.001), and lower values (P≤0.001) after transport; higher fT4 values (P≤0.001) in basal condition. The results indicate that short road transport of donkeys and horses induces the activation of the thyroid gland, with the same release of fT3 and fT4 iodothyronines, but with different preferential release of T3 in horses and T4 in donkeys after transport.

Seasonal thyroid and lipid profiles in Thoroughbred pregnant and nonpregnant mares (Equus caballus).

The purpose of this study was to determine which physiological seasonal thyroid and lipid panel (triglyceride, [TG], total cholesterol, [tCho], high-density lipoprotein cholesterol, [HDL Cho], LDL cholesterol, phospholipids, [PDs]) changes occur in 12 Thoroughbred pregnant and six nonpregnant mares over a period of 12 months, from April to March, by into account the lactation stage. Serum total triiodothyronine (T3) and thyroxine (T4) levels were measured using immunoenzymatic assay kits; the serum lipid panel was analyzed using the enzymatic colorimetric method. One-way ANOVA reported a significant effect of time over 12 months for T3 (F = 5.46; P < 0.001) and T4 (F = 3.98; P < 0.001), in both pregnant and nonpregnant mares, and for triglyceride (F = 2.57; P < 0.01), tCho (F = 2.32; P < 0.01), HDL Cho (F = 5.60; P < 0.0001), and PDs (F = 7.01; P < 0.0001) in pregnant mares. Two-way ANOVA repoted a significant lactating effect compared with nonlactating stage for T3 (F = 8.33; P = 0.006), T4 (F = 6.43; P = 0.003), tCho (F = 10.38; P = 0.0078), HDL Cho (F = 7.64; P = 0.020), and PDs (F = 5.07; P = 0.048). Thus, it appears that under similar environmental condition, nutrition regime, and management system, lactation stage plays a significant role in the seasonal thyroid and lipid profiles in Thoroughbred pregnant mares, with higher T3, T4, and PDs values, and lower tCho and HDL Cho in lactating than nonlactating mares. The physiological values obtained in this study may be used as additional resources to evaluate thyroid and lipid profiles in Thoroughbred pregnant and nonpregnant mares, generating a wished numbers of observation, especially when the additional breed and physiological conditions are considered.

Do Handling and Transport Stress Influence Adrenocortical Response in the Tortoises (Testudo hermanni)?

The goal of this study was to analyze circulating cortisol levels from tortoises (Testudo hermanni) to establish reference intervals and to develop guidelines for the interpretation of the effect of handling and transport stress. Blood samples were obtained from the caudal venous from 23 healthy juvenile tortoises (9 males and 14 females), aged 8-20 years, in basal condition, four weeks prior to and four weeks following handling and short transportation. The study was carried out on the experimental group: 10 tortoises, 4 males and 6 females, and on a control group: 13 tortoises, 5 males and 8 females. Compared to basal values, circulating cortisol concentrations was higher after handling and transport (+286%; P < 0.001), with an increase of +246% (P < 0.001) in males, +236% (P < 0.005) in females, +370% (P < 0.005) in subjects aged 8-12 years, and +240% (P < 0.001) in subjects aged 13-20 years. These observations support the hypotheses that cortisol may act to mediate the effects of handling and transport stress in this species and that four weeks following handling and transport were insufficient to restore their homeostasis.

Hypothalamic-pituitary-adrenal axis responses of horses to therapeutic riding program: effects of different riders.

In order to determine whether therapeutic riding could result in higher levels of stress than recreational riding, hypothalamic-pituitary-adrenal (HPA) axis response was evaluated in six horses by monitoring circulating ß-endorphin, ACTH and cortisol concentrations. Horses were already accustomed to be trained both for therapy and riding school activity since 2004. Intervention consisted of 60-minute therapeutic sessions, two times per week for 6weeks with different riders: disabled and recreational riders (session A and B respectively). The therapeutic riders' group (A) consisted of six children with psychomotor disabilities; the recreational riders' group (B) consisted of six healthy children without any previous horse riding experience. Horses were asked to perform the same gaits and exercises at all sessions, both with disabled and healthy users. The statistical analysis showed that during both sessions the mean basal ß-endorphin and ACTH levels of horses did not show any significant changes, while the one way RM-ANOVA showed significant effects of sessions A on the cortisol (F=11.50; P<0.01) levels. Horses submitted to sessions A showed lower cortisol levels both at 5min (P<0.001) and at 30min (P<0.005) after therapeutic sessions than those after session B. Results suggest that in tested horses and for the variables settled, HPA axis was less responsive to disabled than healthy, recreational riders. Among the endocrine responses, cortisol was one of the indicators of HPA axis stress response.

Comparative endocrinological responses to short transportation of Equidae (Equus asinus and Equus caballus).

In order to evaluate the effects of short transportation on ß-endorphin, adrenocorticotropic hormone (ACTH) and cortisol changes, 12 healthy stallions of Equidae (Equus asinus and Equus caballus) were studied before and after transportation of 50 km. Blood samples were collected 1 week before transportation in basal conditions, immediately before loading and after transportation and unloading, on their arrival at the breeding station. Compared to basal and before values, donkeys showed an increase in circulating ACTH (P < 0.001) and cortisol (P < 0.0005) levels after transportation and higher ACTH (P < 0.01) levels than horses after transportation. A positive and significant correlation (r = 0.885; P < 0.01) between ACTH and cortisol levels after transportation was found. No significant differences were observed for ß-endorphin levels. Compared to basal and before values, horses showed higher cortisol (P < 0.005) levels after transportation and no significant differences were observed for ACTH and ß-endorphin levels in donkeys. Horses facing forward (direction of travel) showed higher (P < 0.01) ß-endorphin levels after transportation than donkeys; horses facing backward (the opposite direction of travel) showed lower (P < 0.05) ACTH levels after transportation. The results indicate that short transportation induces a preferential activation of the hypothalamus-pituitary-axis (HPA), with significant release of ACTH and cortisol in donkeys and only of cortisol in horses, suggesting that transportation for donkeys may be more stressful than horses.

Circulating beta-endorphin, adrenocorticotrophic hormone and cortisol levels of stallions before and after short road transport: stress effect of different distances.

BACKGROUND: Since transport evokes physiological adjustments that include endocrine responses, the objective of this study was to examine the responses of circulating beta-endorphin, adrenocorticotrophic hormone (ACTH) and cortisol levels to transport stress in stallions. METHODS: Forty-two healthy Thoroughbred and crossbred stallions were studied before and after road transport over distances of 100, 200 and 300 km. Blood samples were collected from the jugular vein: first in a single box immediately before loading (pre-samples), then immediately after transport and unloading on arrival at the breeding stations (post-samples). RESULTS: An increase in circulating beta-endorphin levels after transport of 100 km (P < 0.01), compared to basal values was observed. Circulating ACTH levels showed significant increases after transport of 100 km (P < 0.001) and 200 km (P < 0.001). Circulating cortisol levels showed significant increases after road transport over distances of 100, 200 and 300 km (P < 0.001). An effect of transport on beta-endorphin, ACTH and cortisol variations was therefore evident for the different distances studied. No significant differences (P > 0.05) between horses of different ages and different breeds were observed for beta-endorphin, ACTH and cortisol levels. CONCLUSION: The results obtained for short term transportation of stallions showed a very strong reaction of the adrenocortical system. The lack of response of beta-endorphin after transport of 200-300 km and of ACTH after transport of 300 km seems to suggest a soothing effect of negative feedback of ACTH and cortisol levels.