Optimal sampling interval for characterisation of the circadian rhythm of body temperature in homeothermic animals using periodogram and cosinor analysis.

Core body temperature (T c) is a critical aspect of homeostasis in birds and mammals and is increasingly used as a biomarker of the fitness of an animal to its environment. Periodogram and cosinor analysis can be used to estimate the characteristics of the circadian rhythm of T c from data obtained on loggers that have limited memory capacity and battery life. The sampling interval can be manipulated to maximise the recording period, but the impact of sampling interval on the output of periodogram or cosinor analysis is unknown. Some basic guidelines are available from signal analysis theory, but those guidelines have never been tested on T c data. We obtained data at 1-, 5- or 10-min intervals from nine avian or mammalian species, and re-sampled those data to simulate logging at up to 240-min intervals. The period of the rhythm was first analysed using the Lomb-Scargle periodogram, and the mesor, amplitude, acrophase and adjusted coefficient of determination (R 2) from the original and the re-sampled data were obtained using cosinor analysis. Sampling intervals longer than 60 min did not affect the average mesor, amplitude, acrophase or adjusted R 2, but did impact the estimation of the period of the rhythm. In most species, the period was not detectable when intervals longer than 120 min were used. In all individual profiles, a 30-min sampling interval modified the values of the mesor and amplitude by less than 0.1°C, and the adjusted R 2 by less than 0.1. At a 30-min interval, the acrophase was accurate to within 15 min for all species except mice. The adjusted R 2 increased as sampling frequency decreased. In most cases, a 30-min sampling interval provides a reliable estimate of the circadian T c rhythm using periodogram and cosinor analysis. Our findings will help biologists to select sampling intervals to fit their research goals.

Nurse bees regulate the larval nutrition of developing workers (Apis mellifera) when feeding on various pollen types.

Nutrition has been identified as a key driver of colony health and productivity. Yet, in honey bees, relatively little is known about how the vast variety of natural pollen sources impact larval development. The impact of the nutritional quality of 4 naturally occurring pollen sources, of importance to the Western Australian beekeeping industry, was tested on honey bee (Apis mellifera L.) development. Bee packages consisting of 800 g of bees and a mated sister queen were assigned to 40 nucleus hives and randomly allocated to one of the 4 feed treatments (10 colonies each) of marri (Corymbia calophylla Lindl.), jarrah (Eucalyptus marginata Sm.), clover (Trifolium repens L.), and canola (Brassica napus L.) pollen. Emerging bees were collected once the first bees started hatching on the assigned feed sources. Newly emerging bees were weighed individually, and body composition was measured in batches according to the feed treatment groups. Food consumption was recorded for the duration of the experiment. Nurse bees successfully raised young adult workers from the larval stage until emergence when fed with one of 4 pollen patties with different nutritional qualities. There was no difference in the body composition or weight of emerging bees fed on the different pollen types. However, the body weight of bees increased over time, most likely related to colony size and structure. With the type of pollen patties having little impact on larval development, the availability of pollen may be more important than its composition, providing bees have access to all essential nutrients.

Behavioural and physiological responses to stressors in sheep with temperament classified by genotype or phenotype.

The single nucleotide polymorphism (SNP) rs107856856, located in the tryptophan hydroxylase-2 gene, is associated with the behavioural phenotype for sheep temperament measured at weaning. Here, we tested the association between that SNP and physiological and behavioural responses to stressors in adult sheep. Two groups of adult sheep, one with genotype A/A (calm genotype) and the other with G/G (nervous genotype) in rs107856856, were selected from 160 sheep and were exposed, twice, to an open-field arena and an isolation box test (IBT). During each repeat, the behaviour and physiological responses (cortisol, prolactin, dehydroepiandrosterone [DHEA], brain derived neurotrophic factor [BDNF], characteristics of the response of body temperature, and oxidative stress) were measured. The behavioural and physiological responses of the sheep were compared between genotypes and also between groups classified on their phenotype as assessed by their initial isolation box score ("low responders" and "high responders"). The SNP rs107856856 had some effects on the behavioural phenotype (IBT score) but no effects on the physiological response to stress (cortisol, prolactin, DHEA, BDNF, oxidative stress or changes in body temperature) in the adult sheep, probably because the sheep were exposed, and therefore had adapted, to human contact during their life.

Finding biomarkers of experience in animals.

At a time when there is a growing public interest in animal welfare, it is critical to have objective means to assess the way that an animal experiences a situation. Objectivity is critical to ensure appropriate animal welfare outcomes. Existing behavioural, physiological, and neurobiological indicators that are used to assess animal welfare can verify the absence of extremely negative outcomes. But welfare is more than an absence of negative outcomes and an appropriate indicator should reflect the full spectrum of experience of an animal, from negative to positive. In this review, we draw from the knowledge of human biomedical science to propose a list of candidate biological markers (biomarkers) that should reflect the experiential state of non-human animals. The proposed biomarkers can be classified on their main function as endocrine, oxidative stress, non-coding molecular, and thermobiological markers. We also discuss practical challenges that must be addressed before any of these biomarkers can become useful to assess the experience of an animal in real-life.

The 13C-bicarbonate technique as a tool for measurement of energy expenditure in overweight dogs undergoing body weight reduction and the effect of different dietary composition.

Changes in body size and composition, i.e., body weight (BW) gain or loss, affect the daily energy expenditure (EE). To ensure an appropriate BW reduction and to find an efficient strategy to reduce and maintain a target BW, regular evaluations and adjustments of energy allowance are important. This study aimed to provide a detailed knowledge about the possible changes in resting EE using the oral 13C-bicarbonate technique (o13CBT) as a research tool in 16 overweight pet dogs undergoing BW reduction. Dietary composition (i.e., in % of dry matter [DM] being a high protein [33.3], low fat [9.6], and high crude fiber [18.0] diet [LFHFibre], and a high protein [37.9], high fat [52.0], carbohydrate-free diet [HFat]) during 16 wk of energy restriction were evaluated regarding effects on resting EE, rate of BW reduction, body composition, and plasma concentrations of metabolic hormones involved in energy metabolism and appetite regulation. The mean BW loss was higher (P < 0.05) for the dogs fed the LFHFibre diet (1.1%/wk) than that for dogs fed the HFat diet (0.8%/wk), but the total BW reduction of 14.6% and 12.0% of initial BW did not differ significantly (P > 0.05). Resting EE was lower (P < 0.02) after the BW reduction; 414 kJ (99 kcal)/kg BW0.75/d at the start (week 0) and 326 kJ (78 kcal)/kg BW0.75/d at the end (week 16) of the study. The BW reduction in both groups (P > 0.05) consisted of both fat mass (FM) and fat-free mass (FFM). Energy expenditure, calculated in relation to amount of FFM, was not significantly (P > 0.05) affected by BW reduction. Dietary composition did not significantly affect (P > 0.05) plasma concentrations of insulin, leptin, and ghrelin, and no effect (P > 0.05) of BW reduction was observed on hormone concentrations. In conclusion, the o13CBT proved to be a useful research method for studying short-term EE in overweight dogs. Even though all dogs lost BW, most dogs were still overweight at the end of the study. Due to a high individual variation among dogs, a longer experimental period with a larger sample size would be desirable.

The most common nutritional disorder in dogs is overweight, and knowledge about dogs' energy requirement is therefore important to adjust daily feed allowance. Changes in body weight may affect energy expenditure (EE) and, thereby, energy requirement. This study aimed to measure such potential changes under resting conditions in overweight dogs. It was found that the minimally invasive 13C-bicarbonate technique was a useful research method for studies regarding EE during weight loss (WL) in dogs. EE decreased when the dogs lost weight, and energy allowance needed to be reduced to maintain WL. The second objective of this study was to evaluate the effects of feeding diets with different macronutrient compositions on EE, rate of WL, body composition, and plasma concentrations of hormones involved in energy metabolism and appetite regulation. The mean WL rate was slightly higher for dogs fed a diet with high protein, low fat, and high crude fiber contents than those fed a carbohydrate-free diet with a high protein and fat contents. However, diet did not affect the resting EE, measured plasma hormone concentrations, or the total WL at the end of the study.

Low-moderate dietary phytoestrogens transiently disrupt spermatogenesis and the seminal plasma proteome in the ram.

In brief: Dietary phytoestrogens disrupt a specific stage of ram spermatogenesis, causing subtle decreases in sperm quality by affecting the expression of pathways involved in the structural integrity of the spermatozoa. This paper demonstrates for the first time that ram reproduction is compromised by oestrogenic pasture, whilst also providing a longitudinal model for the impact of phytoestrogens on male fertility. Abstract: Compounds with oestrogen-like actions are now common in both the Western diet. The long-term impacts and underlying mechanisms by which oestrogenic compounds alter male reproduction, however, are unclear. To investigate this, we used a longitudinal sheep model examining the impact of oestrogenic pasture consumption on semen quality and production, testicular size, sexual behaviour and the seminal plasma proteome of Merino rams (n = 20), over a full spermatogenic cycle and in the subsequent breeding season. Throughout the study period, sexual behaviour, sperm production and motility were similar between the exposed and non-exposed rams (P > 0.05). However, between 5 and 8 weeks of exposure to dietary phytoestrogens, rams produced a higher percentage of spermatozoa with a specific malformation of the sperm midpiece and reduced DNA integrity, compared to non-exposed rams (P < 0.001). Investigation into the seminal plasma proteome revealed 93 differentially expressed proteins between phytoestrogen-exposed and control rams (P < 0.05). Exposure to phytoestrogens increased the expression of proteins involved in cellular structure development, actin cytoskeleton reorganisation, regulation of cell function and decreased expression in those related to catabolic processes. The greatest fold changes were in proteins involved in the assembly of the sperm flagella, removal of cytoplasm, spermatid development and maintenance of DNA integrity. After returning to non-oestrogenic pasture, no differences in any measure were observed between treatment groups during the subsequent breeding season. We conclude that dietary phytoestrogens can transiently disrupt specific stages of ram spermatogenesis, causing subtle decreases in sperm quality by affecting the expression of pathways involved in the structural integrity of the spermatozoa.

Concurrent measurement of serum and radiomic biomarkers in the clinical investigation of equine musculoskeletal injuries: A prospective pilot study.

The prevention of musculoskeletal injuries and their related welfare and economic impacts represent an immediate priority for the horse racing industry. This prospective pilot study aimed to evaluate a method to quantitatively analyze scintigraphic features of specific anatomical regions of the horse's appendicular skeleton in combination with secondary measures of musculoskeletal metabolism in blood. Twelve horses referred for scintigraphic assessment of lameness were enrolled. Blood samples were collected immediately prior to the administration of radiotracer. Serum concentrations associated with bone turnover were determined for the following biomarkers: C-terminal telopeptides of type I collagen, proteoglycans and sulfated glycosaminoglycans, collagen type II, osteocalcin, and procollagen II C-terminal propeptide. Scintigraphic images underwent radiomic analysis of discrete regions of the distal limbs and these data were correlated to bone turnover markers. Three lame horses demonstrated asymmetrical radiomic abnormalities. The concentration of osteocalcin in the lame horses was significantly higher when compared to the control group, while no significant changes were observed for the other screened serum biomarkers. Findings from the current study provided evidence that radiomic analysis of equine scintigraphy is feasible. This method has the potential to interrogate which serum markers are associated with musculoskeletal injuries.

A repeated measures cognitive affective bias test in rats: comparison with forced swim test.

RATIONALE: There is an urgent need to identify behaviours in animals that can provide insight into the aetiology and potential treatment of depression in humans. OBJECTIVES: This study aimed to validate a repeated measures cognitive affective bias (CAB) test in a rat model of chronic stress and compare CAB with forced swim test (FST) measures. METHODS: Male and female Sprague Dawley rats were trained to associate large and small rewards with scent, spatial, and tactile cues, and their response to an ambiguous tactile stimulus tested. Rats underwent weekly CAB testing for 4 weeks with no intervention, or for 2 weeks of chronic restraint stress (CRS), followed by 2 weeks of fluoxetine, vehicle, or no treatment. CRS rats also underwent the FST at selected timepoints. RESULTS: In control rats, CAB was positive and remained stable over the 4-week period. In CRS-fluoxetine and CRS-vehicle groups, CAB was initially positive, became negative during chronic restraint stress, and returned to positive by 2 weeks after treatment. However, in the CRS-no treatment group, CAB was variable at the outset and unstable over time. Behaviour in the FST was not affected by treatment, and there was no correlation between CAB and FST outcomes. CONCLUSIONS: Instability in the CRS-no treatment group precluded interpretation of the impact of fluoxetine on CAB post-CRS. Our results suggest that behaviour in the FST does not reflect or alter affective state and support the use of CAB tests as part of the behavioural testing repertoire for preclinical animal models of affective disorders.

Pekin ducks are motivated to lay in their preferred nest substrate.

Nest design is one factor contributing to floor-laying in farmed poultry. We investigated: (i) if ducks (Anas platyrhynchos) prefer a particular nest substrate; and (ii) how important that preference is to them, indicated by stress-induced hyperthermia, egg albumen corticosterone, and behaviour. Twelve female ducks that were trained in a push-door task had temperature data loggers implanted. Preference testing identified the most and least preferred nest substrates between sawdust, astroturf, and hemp fibres. A behavioural demand test then required the ducks to use push-doors to access nests containing either the most or least preferred substrate. The preferred substrate door was loaded with increasing weight (0-120% of bodyweight, four nights per workload) and eventually blocked to prevent nest access. The least preferred substrate door remained unweighted. The overall rank order of substrate preferences was sawdust > hemp > astroturf. Six of the 12 birds pushed all workloads and attempted to push the blocked door. The area under the curve (AUC) of hyperthermia was larger when the preferred substrate door was blocked compared with 0%. The AUC did not differ between nights 2-4 of the blocked door compared with night 1. Egg albumen corticosterone was unaffected. We conclude that laying Pekin ducks prefer manipulatable nest substrates and accessing one is important enough to pay a cost. The results indicate that a manipulatable substrate should be provided to commercially farmed nesting ducks.

Estrogenic Pastures: A Source of Endocrine Disruption in Sheep Reproduction.

Phytoestrogens can impact on reproductive health due to their structural similarity to estradiol. Initially identified in sheep consuming estrogenic pasture, phytoestrogens are known to influence reproductive capacity in numerous species. Estrogenic pastures continue to persist in sheep production systems, yet there has been little headway in our understanding of the underlying mechanisms that link phytoestrogens with compromised reproduction in sheep. Here we review the known and postulated actions of phytoestrogens on reproduction, with particular focus on competitive binding with nuclear and non-nuclear estrogen receptors, modifications to the epigenome, and the downstream impacts on normal physiological function. The review examines the evidence that phytoestrogens cause reproductive dysfunction in both the sexes, and that outcomes depend on the developmental period when an individual is exposed to phytoestrogen.

Metabolic profile and productivity of dairy Holstein cows milked by a pasture-based automatic milking system during early lactation: Effects of cow temperament and parity.

This study aimed to evaluate the influence of temperament and parity on milk production and metabolic responses of Holstein cows in a pasture-based automatic milking system. Thirty-five primiparous and 58 multiparous cows were classified as 'calm', 'intermediate' or 'reactive' in each of the temperament tests conducted: milking reactivity (MR), race time (RT), flight speed, and flight distance. The milk yield and milk composition, as well as the blood biochemistry, were measured during the first 10 weeks of lactation. Primiparous cows produced less milk than multiparous cows (24.0 ± 0.8 vs. 34.2 ± 0.6 L/d, respectively, P Ë‚ 0.0001), while the serum beta-hydroxybutyrate concentration differences according to parity were not detected. Multiparous cows that were classified as 'reactive' on MR and RT tests produced 3 kg/d more milk and had greater concentrations of milk components than 'calm' cows (P ≤ 0.05). There was no such relation in primiparous cows. Also, cows scored 'reactive' on RT showed greater milking frequency (2.50 ± 0.05 vs. 2.35 ± 0.04 milking/d; P = 0.01) and blood beta-hydroxybutyrate concentrations (1.28 ± 0.09 vs. 0.92 ± 0.10 mmol/L; P = 0.04) than RT 'calm' cows. In conclusion, the cows' temperament was associated with milk production and metabolism, which depended on parity and the temperament tests used. Although productive performance was different between parities, lipomobilization was not.

Pancreas deficiency modifies bone development in the ovine fetus near term.

Hormones have an important role in the regulation of fetal growth and development, especially in response to nutrient availability in utero. Using micro-CT and an electromagnetic three-point bend test, this study examined the effect of pancreas removal at 0.8 fraction of gestation on the developing bone structure and mechanical strength in fetal sheep. When fetuses were studied at 10 and 25 days after surgery, pancreatectomy caused hypoinsulinaemia, hyperglycaemia and growth retardation which was associated with low plasma concentrations of leptin and a marker of osteoclast activity and collagen degradation. In pancreatectomized fetuses compared to control fetuses, limb lengths were shorter, and trabecular (Tb) bone in the metatarsi showed greater bone volume fraction, Tb thickness, degree of anisotropy and porosity, and lower fractional bone surface area and Tb spacing. Mechanical strength testing showed that pancreas deficiency was associated with increased stiffness and a greater maximal weight load at fracture in a subset of fetuses studied near term. Overall, pancreas deficiency in utero slowed the growth of the fetal skeleton and adapted the developing bone to generate a more compact and connected structure. Maintenance of bone strength in growth-retarded limbs is especially important in a precocial species in preparation for skeletal loading and locomotion at birth.

Diet-altered body temperature rhythms are associated with altered rhythms of clock gene expression in peripheral tissues in vivo.

Temperature rhythms can act as potent signals for the modulation of the amplitude and phase of clock gene expression in peripheral organs in vitro, but the relevance of the circadian rhythm of core body temperature (Tc) as a modulating signal in vivo has not yet been investigated. Using calorie restriction and cafeteria feeding, we induced a larger and a dampened Tc amplitude, respectively, in male Wistar rats, and investigated the circadian expression profile of the core clock genes Bmal1, Per2, Cry1, and Rev-erbα, the heat-responsive genes heat shock protein 90 (Hsp90) and cold-inducible RNA binding protein (Cirbp), and Pgc1α, Pparα/γ/δ, Glut1/4, and Chop10 in the liver, skeletal muscle, white adipose tissue (WAT), and adrenal glands. Diet-altered Tc rhythms differentially affected the profiles of clock genes, Hsp90, and Cirbp expression in peripheral tissues. Greater Tc amplitudes elicited by calorie restriction were associated with large amplitudes of Hsp90 and Cirbp expression in the liver and WAT, in which larger amplitudes of clock gene expression were also observed. The amplitudes of metabolic gene expression were greater in the WAT, but not in the liver, in calorie-restricted rats. Conversely, diet-altered Tc rhythms were not translated to distinct changes in the amplitude of Hsp90, Cirbp, or clock or metabolic genes in the skeletal muscle or adrenal glands. While it was not possible to disentangle the effects of diet and temperature in this model, taken together with previous in vitro studies, our study presents novel data consistent with the notion that the circadian Tc rhythm can modulate the amplitude of circadian gene expression in vivo. The different responses of Hsp90 and Cirbp in peripheral tissues may be linked to the tissue-specific responses of peripheral clocks to diet and/or body temperature rhythms, but the association with the amplitude of metabolic gene expression is limited to the WAT.

Oestrogenic metabolite equol negatively impacts the functionality of ram spermatozoa in vitro.

Oestrogenic pastures are known to cause infertility in the ewe, due primarily to the oestrogen-like actions of the metabolite equol. Despite strong evidence that phytoestrogens and their metabolites compromise male reproductive function in many other species, there is little information concerning the effect of oestrogenic pastures on ram sperm quality and function. To investigate this, ram spermatozoa were exposed in vitro to physiologically relevant concentrations of either 0, 0.001, 0.01, 0.1 and 1 µM equol and incubated over 6 h. Sperm motility, viability, DNA integrity, membrane lipid disorder, mitochondrial superoxide production, lipid peroxidation and intracellular reactive oxygen species were assessed via computer assisted sperm analysis and flow cytometry at 0.5, 3 and 6 h post-equol exposure. Whilst sperm viability was decreased only at 1 µM equol at 0.5 h post-exposure, exposure to equol at concentrations of 0.1 and 1 µM reduced sperm total and progressive motility (P < 0.001), increased sperm membrane fluidity (P < 0.001), increased mitochondrial superoxide production (P < 0.001) and promoted lipid peroxidation (P < 0.001) across all timepoints. At 6 h post-exposure to 0.1 and 1 µM equol, DNA fragmentation was greater compared that of non-exposed spermatozoa (P = 0.045). Intracellular reactive oxygen species did not change between treatment groups throughout the study (P > 0.05). It is concluded that even low concentrations of equol negatively impact the functionality of ram spermatozoa, these effects likely driven through increased mitochondrial superoxide production. This work indicates that equol may exert oestrogen-like actions upon ram spermatozoa, bringing into question as to whether oestrogenic pastures could influence ram fertility.

Daily temperature cycles prolong lifespan and have sex-specific effects on peripheral clock gene expression in Drosophila melanogaster.

Circadian rhythms optimize health by coordinating the timing of physiological processes to match predictable daily environmental challenges. The circadian rhythm of body temperature is thought to be an important modulator of molecular clocks in peripheral tissues, but how daily temperature cycles affect physiological function is unclear. Here, we examined the effect of constant temperature (Tcon, 25°C) and cycling temperature (Tcyc, 28°C:22°C during light:dark) paradigms on lifespan of Drosophila melanogaster, and the expression of clock genes, heat shock protein 83 (Hsp83), Frost (Fst) and senescence marker protein-30 (smp-30). Male and female D. melanogaster housed at Tcyc had longer median lifespans than those housed at Tcon. Tcyc induced robust Hsp83 rhythms and rescued the age-related decrease in smp-30 expression that was observed in flies at Tcon, potentially indicating an increased capacity to cope with age-related cellular stress. Ageing under Tcon led to a decrease in the amplitude of expression of all clock genes in the bodies of male flies, except for cyc, which was non-rhythmic, and for per and cry in female flies. Strikingly, housing under Tcyc conditions rescued the age-related decrease in amplitude of all clock genes, and generated rhythmicity in cyc expression, in the male flies, but not the female flies. The results suggest that ambient temperature rhythms modulate D. melanogaster lifespan, and that the amplitude of clock gene expression in peripheral body clocks may be a potential link between temperature rhythms and longevity in male D. melanogaster. Longevity due to Tcyc appeared predominantly independent of clock gene amplitude in female D. melanogaster.

Calm Hu ram lambs assigned by temperament classification are healthier and have better meat quality than nervous Hu ram lambs.

The objective of this study was to evaluate the effects of temperament classification (assessed using an arena test) on health and productivity of Hu ram lambs. In experiment one, eight ram lambs classified as calm and eight classified as nervous (selected from 100 ram lambs) were fed individually for 60-days to compare food intake, food digestibility, weight gain, and biochemical indices of health. In experiment two, nine ram lambs classified as calm and nine classified as nervous (selected from 150 ram lambs) were fed in a group and slaughter traits, meat quality, and muscle histology were compared. Calm lambs had higher dry matter digestibility, lower serum TNF-α, higher total antioxidant capacity, higher total superoxide dismutase activity, higher dressing percentage, higher cross-sectional area of loin, higher myofibre density, lower ultimate pH of the meat, and higher meat redness, than nervous lambs. Selection for calm temperament could be beneficial to health, slaughter, and carcass traits in Hu ram lambs.

Pekin ducks are motivated to access their nest site and exhibit a stress-induced hyperthermia when unable to do so.

The origins of floor-laying in ducks could be low motivation for a nest, or stress related to difficulties with accessing a nest (e.g. competition). Using a behavioural demand test, we investigated if increasing the work required to access their nest impacted ducks' behaviour and two indicators of stress: egg corticosterone concentration and elevation of core body temperature (stress-induced hyperthermia (SIH)). Twelve laying Pekin ducks previously trained in an operant push-door task were required to use a push-door to access their nest. The door was loaded with increasing weight (0-160% of individual BW, four nights per workload) and eventually blocked to prevent nest access. Before testing, temperature data loggers were implanted in the abdomen. Eggs were collected daily to measure corticosterone concentrations. Behaviour towards the push-door was quantified. Three birds were excluded from the experiment at an early stage. Five of the nine remaining birds pushed all workloads up to 160% BW and attempted to pass the blocked door, with another two birds pushing up to 80 and 140% BW. For those that pushed at all workloads (n = 5) the area under the curve (AUC) of hyperthermia was larger at workloads of 80% (P < 0.001), 120% (P < 0.01), 140% (P < 0.001), 160% (P < 0.001), and when the door was blocked (P < 0.001), compared with 0%. On the first night when the door was blocked, all five birds pushed more at the door, but no attempts were made to push on the following 3 nights, yet the AUC of hyperthermia did not differ between nights 2-4 of the blocked door, compared with the first night that the door was blocked. Increasing workload and inability to access the nest had no effect on corticosterone in egg albumen. It was concluded that laying Pekin ducks were motivated to access a nest. Although it was not possible to differentiate metabolic from psychogenic stress on the first night that nest access was denied, we suggest that the occurrence of hyperthermia on the subsequent nights was due to SIH resulting from frustration at their inability to use their preferred nest. Floor-laying therefore is unlikely due solely to low nest-seeking motivation. Egg corticosterone was not a relevant indicator of acute stress. Strategies to improve nest availability (e.g. decreasing competition) could improve the welfare of commercial ducks.

Association between temperament related traits and single nucleotide polymorphisms in the serotonin and oxytocin systems in Merino sheep.

Animal temperament is defined as the consistent behavioral and physiological differences that are seen between individuals in response to the same stressor. Neurotransmitter systems, like serotonin and oxytocin in the central nervous system, underlie variation in behavioral traits in humans and other animals. Variations like single nucleotide polymorphisms (SNPs) in the genes for tryptophan 5-hydroxylase (TPH2), the serotonin transporter (SLC6A4), the serotonin receptor (HTR2A), and the oxytocin receptor (OXTR) are associated with behavioral phenotype in humans. Thus, the objective of this study was to identify SNPs in those genes and to test if those variations are associated with the temperament in Merino sheep. Using ewes from the University of Western Australia temperament flock, which has been selected on emotional reactivity for more than 20 generations, eight SNPs (rs107856757, rs107856818, rs107856856 and rs107857156 in TPH2, rs20917091 in SLC6A4, rs17196799 and rs17193181 in HTR2A, and rs17664565 in OXTR) were found to be distributed differently between calm and nervous sheep. These eight SNPs were then genotyped in 260 sheep from a flock that has never been selected on emotional reactivity, followed by the estimation of the behavioral traits of those 260 sheep using an arena test and an isolation box test. We found that several SNPs in TPH2 (rs107856757, rs107856818, rs107856856 and rs107857156) were in strong linkage disequilibrium, and all were associated with behavioral phenotype in the nonselected sheep. Similarly, rs17196799 in HTR2A was also associated with the behavioral phenotype.

Thyroid Hormone Deficiency Suppresses Fetal Pituitary-Adrenal Function Near Term: Implications for the Control of Fetal Maturation and Parturition.

Background: The fetal hypothalamic-pituitary-adrenal (HPA) axis plays a key role in the control of parturition and maturation of organ systems in preparation for birth. In hypothyroid fetuses, gestational length may be prolonged and maturational processes delayed. The extent to which the effects of thyroid hormone deficiency in utero on the timing of fetal maturation and parturition are mediated by changes to the structure and function of the fetal HPA axis is unknown. Methods: In twin sheep pregnancies where one fetus was thyroidectomized and the other sham-operated, this study investigated the effect of hypothyroidism on circulating concentrations of adrenocorticotrophic hormone (ACTH) and cortisol, and the structure and secretory capacity of the anterior pituitary and adrenal glands. The relative population of pituitary corticotrophs and the masses of the adrenal zones were assessed by immunohistochemical and stereological techniques. Adrenal mRNA abundances of key steroidogenic enzymes and growth factors were examined by quantitative polymerase chain reaction. Results: Hypothyroidism in utero reduced plasma concentrations of ACTH and cortisol. In thyroid-deficient fetuses, the mass of corticotrophs in the anterior pituitary gland was unexpectedly increased, while the mass of the zona fasciculata and its proportion of the adrenal gland were decreased. These structural changes were associated with lower adrenocortical mRNA abundances of insulin-like growth factor (IGF)-I and its receptor, and key steroidogenic enzymes responsible for glucocorticoid synthesis. The relative mass of the adrenal medulla and its proportion of the adrenal gland were increased by thyroid hormone deficiency in utero, without any change in expression of phenylethanolamine N-methyltransferase or the IGF system. Conclusions: Thyroid hormones are important regulators of the structure and secretory capacity of the pituitary-adrenal axis before birth. In hypothyroid fetuses, low plasma cortisol may be due to impaired adrenocortical growth and steroidogenic enzyme expression, secondary to low circulating ACTH concentration. Greater corticotroph population in the anterior pituitary gland of the hypothyroid fetus indicates compensatory cell proliferation and that there may be abnormal corticotroph capacity for ACTH synthesis and/or impaired hypothalamic input. Suppression of the development of the fetal HPA axis by thyroid hormone deficiency may contribute to the delay in fetal maturation and delivery observed in hypothyroid offspring.

Precision Betacarotene Supplementation Enhanced Ovarian Function and the LH Release Pattern in Yearling Crossbred Anestrous Goats.

The possible out-of-season effect of beta-carotene supplementation on ovulation rate (OR), antral follicles (AFN), and total ovarian activity (TOA = OR + AFN) as related to the LH release pattern in yearling anestrous goats was evaluated. In late April, Alpine-Saanen-Nubian x Criollo goats (n = 22, 26 N) were randomly allotted to: (1) Beta-carotene (BETA; n = 10, orally supplemented with 50 mg/goat/d; 36.4 ± 1.07 kg live weight (LW), 3.5 ± 0.20 units, body condition score (BCS) or (2) Non-supplemented (CONT; n = 12, 35.2 ± 1.07 kg LW, 3.4 ± 0.2 units BCS). Upon estrus synchronization, an intensive blood sampling (6 h × 15 min) was accomplished in May for LH quantifications; response variables included (pulsatility-PULSE, time to first pulse-TTFP, amplitude-AMPL, nadir-NAD and area under the curve-AUC). Thereafter, an ultrasonography scanning was completed to assess OR and AFN. The Munro algorithm was used to quantify LH pulsatility; if significant effects of time, treatment or interaction were identified, data were compared across time. Neither LW nor BCS (p > 0.05) or even the LH (p > 0.05); PULSE (4.1 ± 0.9 pulses/6 h), NAD (0.47 ± 0.13 ng) and AUC (51.7 ± 18.6 units) differed between treatments. Nonetheless, OR (1.57 vs. 0.87 ± 0.18 units) and TOA (3.44 vs. 1.87 ± 0.45 units) escorted by a reduced TTFP (33 vs. 126 ± 31.9 min) and an increased AMPL (0.55 vs. 0.24 ± 0.9 ng), favored to the BETA supplemented group (p < 0.05), possibly through a GnRH-LH enhanced pathway and(or) a direct effect at ovarian level. Results are relevant to speed-up the out-of-season reproductive outcomes in goats while may embrace translational applications.