Pain management with flunixin meglumine at dehorning of calves.

Dehorning (DH) of calves is a common procedure on commercial dairy farms. Pain management of calves has been investigated in several studies. It is generally accepted that the use of local anesthesia before DH is essential for pain management. Postoperative inflammatory pain should be treated by using a nonsteroidal antiinflammatory drug. The objective of this controlled, randomized, and blinded clinical trial was to determine the effects of the nonsteroidal antiinflammatory drug flunixin meglumine before DH on cortisol concentrations in sera of 5- to 9-wk old calves. Furthermore, selected behavioral characteristics and heart and respiratory rate were examined to assess pain in the hours after dehorning. A total of 80 calves were allocated to 4 groups. In each of 20 replicates, 4 calves were randomly assigned to the following groups: in 3 treatment groups, calves received a local anesthetic (10 mL of procain hydrochloride) and a first treatment (i.v.) with flunixin meglumine or a placebo 20 min before hot-iron dehorning, and a second treatment with flunixin meglumine or a placebo (0.9% saline) 3 h after DH. Calves in the control (CON) group were not dehorned and did not receive any treatment. Groups received 2.2 mg of flunixin meglumine/kg followed by a placebo (FP), 2.2 mg of flunixin meglumine/kg for both treatments (FF), or a placebo for both treatments (PP). Blood samples were collected from all calves, including CON calves, 20 min before restraint in a headlock for DH, 2 min after DH, as well as 30 min and 1, 2, 4, 6, and 8 h after DH. Samples were analyzed for concentration of cortisol by enzyme immunoassay. It was found that concentration of cortisol, calculated as area under the curve, was greater in PP compared with FF and tended to be greater compared with FP. Significant differences between PP and FF were detected at 30 min and 2 h after DH. Throughout the observation period, cortisol concentrations were in both flunixin meglumine-treated groups at a similar level as in the CON group. The heart and respiratory rates showed neither difference between the CON group and the 3 dehorned groups nor between the treatment groups.

Measurement of fecal glucocorticoid metabolites and evaluation of udder characteristics to estimate stress after sudden dry-off in dairy cows with different milk yields.

Sudden dry-off is an established management practice in the dairy industry. But milk yield has been increasing continuously during the last decades. There is no information whether the dry-off procedure, which often results in swollen and firm udders, causes stress, particularly in high-producing dairy cows. Therefore, we evaluated the effect of a sudden dry-off on extramammary udder pressure and the concentration of fecal glucocorticoid metabolites (i.e., 11,17-dioxoandrostane, 11,17-DOA) as an indirect stress parameter. Measurements were carried out within the last week before dry-off and until 9d after dry-off considering 3 groups of milk yield (i.e., low: <15 kg/d, medium: 15-20 kg/d, and high: >20 kg/d). Udder pressure increased in all yield groups after dry-off, peaked at d 2 after dry-off and decreased afterwards. Pressures were highest in high-yielding cows and lowest in low-yielding cows. But only in high-yielding cows was udder pressure after dry-off higher than before dry-off. Baseline 11,17-DOA concentrations depended on milk yield. They were highest in low-yielding (121.7 ± 33.3 ng/g) and lowest in high-yielding cows (71.1 ± 30.0 ng/g). After dry-off, 11,17-DOA increased in all yield groups and peaked at d 3. Whereas in medium- and high-yielding cows 11,17-DOA levels differed significantly from their respective baseline during the whole 9-d measuring period, low-yielding cows showed elevated 11,17-DOA levels only on d 3 after dry-off. However, especially the increase in 11,17-DOA after dry-off between the 3 yield groups was considerably different. Mean 11,17-DOA increase from baseline to d 3 was highest in high-yielding cows (129.1%) and considerably lower in low-yielding cows (40.1%). The highest fecal 11,17-DOA concentrations were measured on d 3 after dry-off, indicating that the stress was most intense on d 2, which is due to an 18-h time lag; at about the same time, udder pressure peaked. Our results showed a negligible effect of a sudden dry-off on low-yielding cows. High-yielding cows, however, faced high extramammary pressures and increased glucocorticoid production. Considering animal welfare aspects, a review of the current dry-off strategies might be warranted.

Physiological and behavioral stress parameters in calves in response to partial scrotal resection, orchidectomy, and Burdizzo castration.

Establishing artificial cryptorchids by partial scrotal resection without removing the testicles is a technique for castration of bull calves that recently has gained new interest. In contrast to orchidectomy and Burdizzo castration, the stress response of calves to shortening of the scrotum is unknown. In this study, partial scrotal resection in bull calves was compared with orchidectomy, Burdizzo castration, and controls without intervention (n=10 per group, ages 56 ± 3 d). Procedures were performed under xylazine sedation and local anesthesia. We hypothesized that partial scrotal resection is least stressful. Salivary cortisol, heart rate, heart rate variability, behavior, and locomotion were analyzed. Cortisol concentration peaked 60 min after start of the procedures. Cortisol release was at least in part xylazine induced and none of the experimental procedures released additional cortisol. Heart rate increased in calves of all groups with initial handling, but immediately after xylazine sedation decreased to 30% below initial values and was not modified by surgical procedures. The heart rate variability variables standard deviation of beat-to-beat interval and root mean square of successive beat-to-beat differences increased when calves were placed on the surgery table but effects were similar in calves submitted to surgeries and control calves. Locomotion increased, whereas lying time decreased in response to all surgeries. Locomotion increase was most pronounced after orchidectomy. Plasma fibrinogen concentrations increased after orchidectomy only. With adequate pain medication, orchidectomy, Burdizzo castration, and partial scrotal resection do not differ with regard to acute stress and, by inference, pain. Partial scrotal resection when carried out under xylazine sedation and local anesthesia thus is an acceptable castration technique in bull calves.

Cortisol release, heart rate and heart rate variability, and superficial body temperature, in horses lunged either with hyperflexion of the neck or with an extended head and neck position.

Bringing the head and neck of ridden horses into a position of hyperflexion is widely used in equestrian sports. In our study, the hypothesis was tested that hyperflexion is an acute stressor for horses. Salivary cortisol concentrations, heart rate, heart rate variability (HRV) and superficial body temperature were determined in horses (n = 16) lunged on two subsequent days. The head and neck of the horse was fixed with side reins in a position allowing forward extension on day A and fixed in hyperflexion on day B. The order of treatments alternated between horses. In response to lunging, cortisol concentration increased (day A from 0.73 ± 0.06 to 1.41 ± 0.13 ng/ml, p < 0.001; day B from 0.68 ± 0.07 to 1.38 ± 0.13 ng/ml, p < 0.001) but did not differ between days A and B. Beat-to-beat (RR) interval decreased in response to lunging on both days. HRV variables standard deviation of RR interval (SDRR) and RMSSD (root mean square of successive RR differences) decreased (p < 0.001) but did not differ between days. In the cranial region of the neck, the difference between maximum and minimum temperature was increased in hyperflexion (p < 0.01). In conclusion, physiological parameters do not indicate an acute stress response to hyperflexion of the head alone in horses lunged at moderate speed and not touched with the whip. However, if hyperflexion is combined with active intervention of a rider, a stressful experience for the horse cannot be excluded.

Environmental influences on Adelie penguin breeding schedules, endocrinology, and chick survival.

To understand how the social and physical environment influences behaviour, reproduction and survival, studies of underlying hormonal processes are crucial; in particular, interactions between stress and reproductive responses may have critical influences on breeding schedules. Several authors have examined the timing of breeding in relation to environmental stimuli, while others have independently described endocrine profiles. However, few studies have simultaneously measured endocrine profiles, breeding behaviour, and offspring survival across seasons. We measured sex and stress hormone concentrations (oestrogens, testosterone, and corticosterone), timing of breeding, and chick survival, in Adelie penguins (Pygoscelis adeliae) at two colonies in two different years. Clutch initiation at Cape Bird South (CBS; year 1, ~14,000 pairs) occurred later than at Cape Crozier East (CCE; year 2, ~ 25,000 pairs); however, breeding was more synchronous at CBS. This pattern was probably generated by the persistence of extensive sea ice at CBS (year 1). Higher corticosterone metabolite and lower sex hormone concentrations at CBS correlated with later breeding and lower chick survival compared to at CCE - again, a likely consequence of sea ice conditions. Within colonies, sub-colony size (S, 50-100; M, 200-300; L, 500-600; XL, >1000 pairs) did not influence the onset or synchrony of breeding, chick survival, or hormone concentrations. We showed that the endocrine profiles of breeding Adelie penguins can differ markedly between years and/or colonies, and that combining measures of endocrinology, behaviour, and offspring survival can reveal the mechanisms and consequences that different environmental conditions can have on breeding ecology.

Female androgen patterns and within-pair testosterone compatibility in domestic geese (Anser domesticus).

For successfully raising offspring, long-term monogamous pair partners need to be behaviorally and hormonally coordinated. In the monogamous, biparental greylag geese (Anser anser) a dyadic pairbond-specific measure, 'within-pair testosterone compatibility' (TC) indicated how closely synchronized are seasonal androgen levels, which co-varied with reproductive output. Males, in particular, were assumed to respond to their females' hormonal and fecundity phases. We now present experiments with biparental domestic geese (Anser domesticus) kept as pairs to ask whether TC occurs also in these generally polygynous animals. We further ask how different conditions of mate choice affect TC and whether established TC is maintained during a polygynous flock situation. We measured androgen metabolites (AM) non-invasively from individual droppings. In females, AM was related with gonadal activity as it increased after GnRH but not ACTH challenge. Females with preferred partners had higher maximum AM during egg laying and higher rates of initiating incubation than randomly paired females. Domestic ganders had seasonal AM patterns typical for polygynous males. Within-pair TC ranged from almost perfectly positive to non-correlated in domestic geese but mate choice did not explain TC variation. TC of previous pairs was generally reduced in the flock situation, probably confounded by factors of the social environment, i.e. mating opportunity and availability of multiple partners. On top of the underlying reproductive physiology our results suggest two episodic components of TC: a female androgen responsiveness to the preferred partner at least during egg formation, and the male's facultative potential to respond to her readiness to breed.

Gestagens and glucocorticoids in chicken eggs.

Avian eggs contain a variety of steroid hormones, which have been attributed as a tool for maternal phenotypic engineering. The majority of studies focuses on androgens, but also significant amounts of progesterone as well as other steroid hormones have been measured. The question if corticosterone is also present in eggs of chickens is currently under debate. The only analytical validation performed so far has failed to demonstrate corticosterone in the yolk of chickens, suggesting that antibodies for corticosterone measurement cross-react with other steroids present in the yolk. In order to investigate this assumption and to characterise potential cross-reacting hormones in more detail, we performed high-performance liquid chromatographic (HPLC) analyses of chicken yolk extracts and determined the concentration of immunoreactive corticosterone, progesterone and cortisol. The progesterone antibody revealed several immunoreactive substances, including progesterone, pregnenolone and two substances with lower polarity. The corticosterone enzyme immunoassay detected immunoreactive substances at exactly the same elution positions as the progesterone assay and a very small peak at the elution position of corticosterone. Immunoreactive cortisol was not found. In addition, inner and outer regions of the yolk sphere were analysed separately via HPLC. We found different concentrations of immunoreactive substances between the inner and outer yolk regions, probably reflecting the steroidogenic activity of the follicle cells during oocyte growth. We conclude that in homogenised yolk extracts without previous clean-up, the measured corticosterone concentrations may actually reflect those of progesterone and its precursors, most probably being 5 alpha- and 5 beta-pregnanes and pregnenolone.

Effect of altrenogest-treatment of mares in late gestation on adrenocortical function, blood count and plasma electrolytes in their foals.

REASONS FOR PERFORMING STUDY: Mares with compromised pregnancies are often treated with altrenogest to prevent abortion. However, there is only limited information about effects on the foal when altrenogest treatment is continued during final maturation of the fetus. OBJECTIVES: To determine effects of altrenogest treatment during late gestation in mares on maturity, haematology changes, adrenocortical function and serum electrolytes in their newborn foals. METHODS: Six mares were treated with altrenogest (0.088 mg/kg bwt) once daily from Day 280 of pregnancy until foaling and 7 mares served as controls. RESULTS: Foals born to altrenogest-treated mares had a significantly lower neutrophil/lymphocyte ratio on the first day after birth than control foals (P<0.05). Basal plasma cortisol concentrations immediately after birth were higher in foals of altrenogest-treated mares than in control foals (P<0.05). Cortisol release in response to exogenous adrenocorticotropic hormone (ACTH)--except for higher values 15 min after ACTH injection in foals of altrenogest-treated mares on Day 1--revealed no differences in adrenocortical function between the groups of foals. Plasma potassium concentration in foals from altrenogest-treated mares compared to control foals was significantly lower immediately after birth (P<0.05) and plasma ionised calcium concentration was significantly lower 3 h after birth (P = 0.01). CONCLUSIONS AND POTENTIAL RELEVANCE: Altrenogest treatment of pregnant mares prolonged labour had no major effects on adrenocortical function in foals. A reduced neutrophil/lymphocyte ratio in these foals may suggest either immunomodulatory effects of altrenogest or dysmaturity of the foals.

Social context rather than behavioral output or winning modulates post-conflict testosterone responses in Japanese quail (Coturnix japonica).

Testosterone regulates the expression of sexual and aggressive behavior in male vertebrates and treatments with testosterone may promote territorial aggression and winning in dyadic contests. Conversely, individual testosterone levels respond to sexual or aggressive interactions and the social environment. Post-conflict testosterone in winner males though appears to be more complex than simply reflecting conflict outcome. Expression and degree of post-conflict testosterone responses may adapt to additional modulators such as repeated winning experience, audience presence, opponent's fighting ability, and self-assessment. We present simulated intrusion experiments with male Japanese quail using mirror-elicited aggression and fights with real opponents ('direct challenge'). We recorded agonistic behavior and measured immunoreactive testosterone metabolites (TM) non-invasively from individual droppings. Frequencies of initiated agonistic behavior were similar whether towards the mirror or in direct challenge tests, although some of the males were behaviorally non-responsive to the mirror ('mirror submissives'). However, there was no TM response to the mirror test in both, mirror fighters and mirror submissives, thus independently of behavioral output. After direct challenges TM levels were elevated in all males (focal males winning or conflict unresolved after 30 min), hence independently of conflict outcome. Thus, in male quail a combination of physical stimuli and the individual perception of own and opponent's fighting ability explained the expression of post-conflict TM responses rather than behavioral performance, conflict outcome, or any of these factors alone. In sum, our results emphasize that the degree of androgen responsiveness to agonistic behavior is fine-tuned by components related with social context and environment.

Corticosterone in chicken eggs.

Birds are discussed as models for prenatal stress. In this study, several experiments were conducted to gain basic knowledge of if, how, and when maternal adrenocortical activity is reflected by corticosterone concentrations in the egg. Radiolabeled corticosterone was administered to 10 laying hens to investigate the uptake into as well as the distribution within the eggs. The yolk was dissected in concentric layers and analyzed. Less than 1% of the administered radioactivity entered the egg but was, however, not evenly distributed. On the day after injection, highest radioactivity (Bq/g) was detected in the albumen and the outmost layer, whereas concentration peaked 4-7 days later in the inner layers. In two other experiments, increased plasma levels of corticosterone were induced by injection of adrenocorticotropic hormone (ACTH) or feeding of corticosterone. Again, yolk disks were cut in layers and analyzed with a corticosterone enzyme immunoassay. No effect of the ACTH administration was detected, whereas feeding of corticosterone resulted in increased immunoreactive corticosterone concentrations in the yolk. Straight-phase high-performance liquid chromatographic (HPLC) separations were also performed to characterize immunoreactive steroids in the yolk. Two close-eluting peaks at the approximate elution position of corticosterone could be observed after the feeding experiment, whereas in untreated control eggs they were absent. It was concluded that transfer from plasma to egg is low for corticosterone and that further investigations concerning the transport mechanisms and the exact nature of yolk steroids are necessary.

Stress hormones in mammals and birds: comparative aspects regarding metabolism, excretion, and noninvasive measurement in fecal samples.

A multitude of endocrine mechanisms are involved in coping with challenges. Front-line hormones to overcome stressful situations are glucocorticoids (GCs) and catecholamines (CAs). These hormones are usually determined in plasma samples as parameters of adrenal activity and thus of disturbance. GCs (and CAs) are extensively metabolized and excreted afterwards. Therefore, the concentration of GCs (or their metabolites) can be measured in various body fluids or excreta. Above all, fecal samples offer the advantages of easy collection and a feedback-free sampling procedure. However, large differences exist among species regarding the route and time course of excretion, as well as the types of metabolites formed. Based on information gained from radiometabolism studies (reviewed in this paper), we recently developed and successfully validated different enzyme immunoassays that enable the noninvasive measurement of groups of cortisol or corticosterone metabolites in animal feces. The determination of these metabolites in fecal samples can be used as a powerful tool to monitor GC production in various species of domestic, wildlife, and laboratory animals.

Heart rate and salivary cortisol concentrations in foals at birth.

Heart rate (HR), HR variability (HRV) and salivary cortisol concentrations were determined in foals (n = 13) during the perinatal phase and until 5 months of age. In the fetus, HR decreased from 77 ± 3 beats/min at 120 min before birth to 60 ± 1 beats/min at 5 min before birth (P <0.01). Within 30 min of birth, HR increased to 160 ± 9 beats/min (P <0.01). Salivary cortisol concentrations immediately after birth were 11.9 ± 3.6 ng/mL and within 2 h increased to a maximum of 52.5 ± 12.3 ng/mL (P <0.01). In conclusion, increases in HR and salivary cortisol concentrations in foals are not induced during parturition, but occur immediately after birth.

Social, state-dependent and environmental modulation of faecal corticosteroid levels in free-ranging female spotted hyenas.

Little is known about to what extent the sensitivity of the hypothalamic-pituitary-adrenal (HPA) axis may be state dependent and vary in the same species between environments. Here we tested whether the faecal corticosteroid concentrations of matrilineal adult female spotted hyenas are influenced by social and reproductive status in adjacent ecosystems and whether they vary between periods with and without social stress. Females in the Serengeti National Park frequently become socially subordinate intruders in other hyena territories by undertaking long-distance foraging trips to migratory herds, whereas in the Ngorongoro Crater they usually forage inside their own small territories on resident prey. The faecal corticosteroid concentrations in Serengeti females were significantly higher than in Ngorongoro females. Energy expenditure by lactation is exceptionally high in spotted hyenas and this may be reflected in their corticosteroid levels. The faecal corticosteroid levels in both populations were higher in lactating than in non-lactating females. During periods of social stability, faecal corticosteroid concentrations increased in non-lactating females but not in lactating females as social status declined. Lactating Serengeti females had significantly higher faecal corticosteroid concentrations during periods with acute severe social stress than during periods without, indicating that the HPA axis is sensitive to social stimuli even in lactating females. So far few studies have used non-invasive monitoring methods for assessing social stress in freeranging animals. This study demonstrates for the first time, to the authors' knowledge, that corticosteroid concentrations may differ between periods with and without social stress for a free-ranging female mammal and that the modulating effect of social status may depend on reproductive status.

Stimulation of androgen and oestrogen concentrations in plasma of cows after administration of a synthetic glucocorticoid (flumethasone) at the end of gestation.

Administration of flumethasone (3.5 mg i.m.) to six cows on day 260 of pregnancy induced parturition in only one animal. In the other five cows circulating concentrations of epitestosterone and conjugated oestrogens increased to reach maximum values (epitestosterone, 13.73 +/- 2.81 nmol/l; conjugated oestrogens, 33.59 +/- 6.87 nmol/l) 2-3 days after treatment. Concentrations of these steroids were raised as long as the synthetic glucocorticoid was present in the circulation (as judged by depression of cortisol concentrations). After clearance of the drug, concentrations of these steroids declined to values present in the control group. Concentrations of unconjugated oestrogens were only slightly raised after flumethasone. In contrast to the effect observed after administration on day 260, treatment on day 270 induced parturition in four of six treated animals. In these cows, mean circulating concentrations of epitestosterone, unconjugated and conjugated oestrogens increased to 9.50 +/- 2.96, 9.62 +/- 1.48 and 36.51 +/- 4.8 nmol/l respectively to reach concentrations observed in the control group at parturition. After parturition the concentrations of epitestosterone and oestrogens declined rapidly in all groups.

Plasma levels of androstenedione, epitestosterone, testosterone and oestrogens in cows at parturition.

The concentration of total unconjugated oestrogens in the peripheral plasma of cows increased from 2.00 +/- 0.20 (S.E.M.) ng/ml 7 days before to 3.87 +/- 0.50 ng/ml 1 day before parturition and declined to values below 0.30 ng/ml on day 2 post partum. The concentrations of androstenedione, epitestosterone and testosterone remained approximately constant at 0.90 +/- 0.10, 0.92 +/- 0.08 and 0.40 +/- 0.20 ng/ml respectively during the last week of pregnancy. After parturition the concentration of the three androgens declined rapidly. Similar to data obtained in goats we did not find a marked increase in androgen levels in the peripheral blood of cows before parturition.

Immunohistochemical distribution of oestrogen and progesterone receptors and tissue concentrations of oestrogens in the cervix of non-pregnant cows.

An immunohistochemical study of the expression of oestrogen (ER) and progesterone receptors (PR) in different regions along the longitudinal and vertical axes of the cervix of non-pregnant cows was performed. Animals were separated into two groups depending on the presence or absence of a functional corpus luteum in their ovaries, as indicated by blood progesterone concentrations. The high progesterone group (HP4) had serum progesterone concentrations > 2.0 ng mL(-1) (n = 6) and the low progesterone group (LP4) had serum progesterone concentrations < or = 0.5 ng mL(-1) (n = 4). Significantly higher concentrations of oestrogen were found in the cervical tissue of animals in the LP4 group than those in the HP4 group (473 +/- 53 v.149 +/- 46 pg g(-1) wet weight; P < 0.01). Furthermore, there was a significant effect of tissue layer (epithelium to deep stroma) on the number of ER (P < 0.01) and PR (P < 0.05) immunoreactive nuclei per 1000 cells. For both ER and PR the proportion of cells expressing the receptor increased from epithelium to subepithelial stroma (P < 0.01) and from subepithelium to deep stroma (ER P < 0.05; PR P =0.061). When the number of receptor-positive cells were expressed per mm2 tissue, differences between the subepithelial stroma and the deep stroma became even more marked. In addition, the vaginal part of the cervix had significantly more (P < 0.01) ER and PR immunoreactive nuclei per 1000 cells than the uterine part, but these differences were no longer apparent when a correction was made for cell density. There was no relationship between progesterone status of the animals, nor local tissue oestrogen concentrations and ER or PR immunoreactivity in the cervix of these non-pregnant cows. Instead, a strong relationship between both longitudinal and vertical positioning of tissue in the cervix and expression of both receptor types was shown. In addition, a strong correlation between ER and PR expression in the subepithelial stroma (R = 0.85, P < 0.01) and the deep stroma (R = 0.83 P < 0.01) was evident. In conclusion, these results demonstrate that in studies of steroid hormone receptor expression in the cervix, careful description of sampling site and depth are necessary if the results are to be interpreted meaningfully.

Hormones as indicators of stress.

Animal welfare is of increasing importance and absence of chronic stress is one of its prerequisites. During stress, various endocrine responses are involved to improve the fitness of the individual. The front-line hormones to overcome stressful situations are the glucocorticoids and catecholamines. These hormones are determined as a parameter of adrenal activity and thus of disturbance. The concentration of glucocorticoids (or their metabolites) can be measured in various body fluids or excreta. Above all, fecal samples offer the advantage that they can be easily collected and this procedure is feedback free. Recently, enzyme immunoassays (EIA) have been developed and successfully tested, to enable the measurement of groups of cortisol metabolites in animal feces. The determination of these metabolites in fecal samples is a practical method to monitor glucocorticoid production.

Conversion of steroids in bovine blood in vitro.

Blood samples collected from eight Braunvieh cows between the sixth and eighth month of gestation were allowed to stand with and without anticoagulant at 20 degrees C and 0 degrees C for different time periods. In these samples the degree of in vitro conversion of gestagens, androgens and estrogens was investigated. The concentrations were measured by radioimmunoassay. After 24 h at 20 degrees C, the levels of pregnenolone, progesterone, 17alpha-hydroxyprogesterone, androstenedione, dehydroepiandrosterone and estrone decreased to 62, 29, 25, 10, 34 and 44%, respectively, of the initial value and those of 17alpha, 20beta-dihydroxyprogesterone, epitestosterone and estradiol-17alpha increased to 385, 800 and 852%, respectively. The conversion was slower in clotted blood. The concentrations of testosterone and estradiol-17beta were consistent over the 24 h period. There was no marked decrease of the steroid concentration after 24 h of incubation of whole blood at 0 degrees C and of plasma at 20 degrees C. After the addition of (3)H-steroids, conversion could be demonstrated by thin-layer chromatography and autoradiography. These results demonstrate that all investigated hormones except testosterone and estradiol-17beta were metabolized by bovine blood cells.

Coping capacity of dairy cows during the change from conventional to automatic milking.

In conventional milking systems, dairy cows are driven to the milking stall twice or thrice daily, whereas in automatic milking systems (AMS), the cows enter the milking stall voluntarily. In this study, noninvasive methods were used to analyze the physiological reaction of 17 cows toward the changeover from conventional to automatic milking. Milk yield and composition were analyzed. Heart rate was recorded continuously, and feces was sampled twice daily to determine cortisol metabolites (11, 17-dioxoandrostanes) for a period of 2 wk. During the first visit to the AMS (without milking), heart rate was elevated compared with parlor milking by 35 +/- 3 beats per minute (bpm) above basal heart rate (P < 0.05). Heart rate during the first milking in AMS (eighth visit) was already similar to the heart rate previously measured during milking in the parlor (18.1 +/- 2.2 bpm above basal level). Concentration of fecal cortisol metabolites was unchanged during the change-over compared with parlor milking. A decreased (P < 0.05) milk yield of 68 +/- 7% relative to previous parlor yield during the first AMS milking indicated a disturbance of milk ejection in most cows. Individual yields ranged from 8 to 96% of the previous parlor yield. To examine the relationship between adrenal cortex sensitivity and the coping process, an ACTH challenge experiment was performed after the changeover period. Cows that released more cortisol after ACTH injection, indicating a higher adrenal cortex sensitivity, had a less enhanced heart rate and a near normal milk ejection during the first AMS milkings (P < 0.05). In conclusion, the reactions toward the changeover to AMS milking varied widely within cows. Adaption to the AMS was easier in animals with a higher adrenal cortex sensitivity to ACTH.

Measurement of cortisol metabolites in faeces of ruminants.

Twenty-one metabolites were detected in faecal samples collected after infusion of (14C)cortisol into the jugular vein of sheep. Using high-performance liquid chromatography/radiometric analysis plus mass spectrometry. One group of metabolites had molecular weights of between 302 and 308, and another group of 350, which indicates that the substances have a C19O3 or a C21O4 structure. Therefore, an enzyme immunoassay against 5beta-androstane-3alpha-ol-11,17-dione-17-CMO:BSA was established. Faecal samples were collected from 10 cows immediately after transport and then during a course in which non-invasive diagnostic procedures were being taught (course 1). For comparison, faeces were sampled from another 5 cows that were being used for teaching invasive procedures (course 2). Six cows from a university farm served as controls. In the animals used in course 1, the highest concentrations of cortisol metabolites were measured immediately after transport to the university (median value: 2.2 micromol/kg faeces). During the first 5 days at the university, the concentrations decreased to 0.52 micromol/kg (median) and remained at this level during the rest of the course. The median concentration in the samples that were taken during coursc 2 (collected about 2 months after transport) was 0.48 micromol/kg. There was no significant difference in the excretion of cortisol metabolites between these cows and the controls. We conclude from these data that, using the enzyme immunoassay against 5beta-androstane-3alpha-ol-11,17-dione-17-CMO, we were able to detect transport/novel environment stress but not the potential disturbance that cows experience during diagnostic procedures.