The effects of food restriction on the perception of hunger, metabolic state and stress responsiveness in sheep

Sheep in temperate regions may be exposed to periods of limited food availability and a range of environmental conditions. Insufficient dietary intake may change live weight (LW) and body condition score (BCS) resulting in an increase in hunger and alterations in metabolic state, which may influence the welfare of sheep. In Chapter 2, a behavioural demand methodology to measure feeding motivation was refined and improved; five different walking distances (costs) were used and the number of rewards consumed, the total distance walked, Pmax (maximum price) and Omax (maximum expenditure) were used as measures of motivation. Sixteen non-pregnant ewes were subjected to two feeding levels (24 h fasted and ad libitum fed control); fasted ewes consumed more rewards and walked a greater total distance compared to control ewes. The methodology provided good measures of feeding motivation and was a potential indicator of hunger. However, the range of costs used was too narrow for accurate assessment of Pmax and Omax. In Chapter 3, the effects of body condition score (BCS) on feeding motivation and endocrine and metabolic responses in twin-bearing ewes were investigated to identify and quantify indicators of hunger and to assess the metabolic state of twin-bearing ewes with different BCS. Twenty-two ewes were divided into low BCS (LBC), medium BCS (MBC) and high BCS (HBC) treatments. Feeding motivation was assessed between day 91 and 105 of pregnancy and blood samples were collected once every 2 weeks between day 35 and 133 of pregnancy. The number of rewards consumed and Omax were significantly lower in the HBC ewes compared to LBC and MBC ewes, while the MBC ewes tended to show a lower motivation compared to the LBC ewes. LBC ewes were in a metabolically catabolic state and had low plasma concentrations of leptin, insulin and insulin-like growth factor-1. In Chapter 4, it was investigated whether a fast loss in BCS would increase hunger and alter the metabolic state more compared to a slow loss in BCS. Twenty-six twin-bearing ewes were assigned to Slow Loss (1 BCS loss in 13 weeks, SL), Fast loss (1 BCS loss in 10 weeks, FL) or Control (maintain BCS) treatments. Feeding motivation was assessed between day 42 and 67 of pregnancy and blood samples were collected once every 2 weeks between day 35 and 140 of pregnancy. SL and FL ewes had not lost much BCS at the start of the motivation period and no effects on feeding motivation were found. SL ewes were in a similar metabolic state as Control ewes. However, FL ewes were in a catabolic state in mid-pregnancy. In Chapter 5, it was investigated whether BCS affects the ability to maintain energy homeostasis and induce adaptive metabolic and stress responses to an environmental challenge. Eighteen shorn single-bearing ewes were divided into low, medium and high BCS treatments. Ewes were exposed to a 6 h acute cold challenge between days 85-87 of pregnancy. LBC ewes were able to maintain body temperature, but were slower to mobilise energy substrates and had a delayed and reduced cortisol response compared to MBC and HBC ewes. In conclusion, feeding motivation appeared to be an indicator of hunger in ewes; fasting and a loss in BCS markedly increased hunger. A loss in BCS induced a metabolically catabolic state. However, ewes were metabolically better able to adapt to a slow than a fast loss in BCS. Finally, low BCS ewes may have a reduced ability to respond to acute stressors

Supplementation of Lactobacillus early in life alters attention bias to threat in piglets

Gut microbes play an important role in regulating brain processes and influence behaviour, cognition and emotional states in humans and rodents. Nevertheless, it is not known how ingestion of beneficial microbes modulates emotional states in piglets and whether it can improve welfare. Here we use an attention bias task to assess the effects of Lactobacillus reuteri ATCC-PTA-6475 and Lactobacillus plantarum L1-6 supplementation early in life on emotional states in 33 piglets compared to 31 placebo supplemented piglets. We hypothesized that Lactobacillus supplementation would reduce vigilance behaviour (head at shoulder height or higher) and attention (head oriented towards the threat) in response to an auditory threat. The results showed that the control group increased vigilance behaviour in response to the threat, but there was no increase in the probiotics group. Despite the increased vigilance, the control group paid less attention to the threat. One explanation may be that control piglets avoided looking in the direction of the threat just because they perceived it as more threatening, but further research is necessary to confirm this. In conclusion, Lactobacillus supplementation may be a suitable tool to reduce anxiety, promote a more appropriate response to a challenge and so improve welfare

Can’t see the forest for the trees? Investigating attentional scope as a novel indicator of emotional state in animals

In humans, contrasting emotional states can lead to a broadening or narrowing of attentional scope. Whether this is also the case in animals has yet to be investigated. If confirmed, measurement of attentional scope has potential as a novel cognitive method of welfare assessment. In this study, we therefore aimed to investigate a test of attentional scope as a measure of emotional state in animals. We did this by inducing four putatively different emotional states in dogs (N = 10), varying in valence (positive, negative) and arousal (high, low), in two different reward contexts (food rewards in Experiment 1, social rewards in Experiment 2) and then assessing dogs' behavioural responses in a test of attentional scope. We also recorded heart rate variability (HRV) parameters as additional confirmatory affective indicators. In Experiment 1, the dogs showed a narrowing of attentional scope after the induction of both positively valenced emotional states. That dogs were in a positive state was supported by the reduced Standard Deviation of normal-to-normal R-R intervals (SDNN) and the reduced Low Frequency (LF) and Very Low Frequency (VLF) HRV. In Experiment 2, when responses to social rewards were examined, we did not detect any statistically significant differences in attentional scope between the emotional states, but dogs had a slightly narrow attentional scope in the negatively valenced emotional states. The LF tended to be reduced in the high arousal positive treatment. In conclusion, our study provides the first indication that emotional states can also alter attentional scope in animals. The results justify further investigation of this approach for use in animal welfare assessment, although additional studies are needed to refine predictions

Investigating attentional scope as a novel indicator of emotional state in animals

In humans, contrasting emotional states can lead to a broadening or narrowing of attentional scope. Whether this is also the case in animals has yet to be investigated. If confirmed, measurement of attentional scope has potential as a novel cognitive method of welfare assessment. In this study, we therefore aimed to investigate a test of attentional scope as a measure of emotional state in animals. We did this by inducing four putatively different emotional states in dogs (N = 10), varying in valence (positive, negative) and arousal (high, low), in two different reward contexts (food rewards in Experiment 1, social rewards in Experiment 2) and then assessing dogs’ behavioural responses in a test of attentional scope. We also recorded heart rate variability (HRV) parameters as additional confirmatory affective indicators. In Experiment 1, the dogs showed a narrowing of attentional scope after the induction of both positively valenced emotional states. That dogs were in a positive state was supported by the reduced Standard Deviation of normal-to-normal R-R intervals (SDNN) and the reduced Low Frequency (LF) and Very Low Frequency (VLF) HRV. In Experiment 2, when responses to social rewards were examined, we did not detect any statistically significant differences in attentional scope between the emotional states, but dogs had a slightly narrow attentional scope in the negatively valenced emotional states. The LF tended to be reduced in the high arousal positive treatment. In conclusion, our study provides the first indication that emotional states can also alter attentional scope in animals. The results justify further investigation of this approach for use in animal welfare assessment, although additional studies are needed to refine predictions

Validation of the LUMIPULSE automated immunoassay for the measurement of core AD biomarkers in cerebrospinal fluid

Altres ajuts: Swedish Research Council (#2017-00915); Alzheimer Drug Discovery Foundation (ADDF), USA (#RDAPB-201809-2016615); Swedish Alzheimer Foundation (#AF-742881); Hjärnfonden, Sweden (#FO2017-0243); Swedish State Under the Agreement Between the Swedish Government and the County Councils, the ALF-Agreement (#ALFGBG-715986); National Program of Sustainability II (MEYS CR); Ministry of Health of the Czech Republic (grant no. 19-04-00560); ZonMW (part of the Dutch national 'Deltaplan for Dementia'; Selfridges Group Foundation; National Institutes of Health, USA (grant number 5R01NS104147-02); Alzheimerfonden (AF-930934); Åhléns-stiftelsen; Stiftelsen för Gamla tjänarinnor; Canadian Institutes of Health Research (CIHR) (MOP-11-51-31; RFN 152985, 159815, 162303); Canadian Consortium of Neurodegeneration and Aging (CCNA; MOP-11-51-31 -team 1); Weston Brain Institute, the Alzheimer's Association (NIRG-12-92090, NIRP-12-259245); Brain Canada Foundation (CFI Project 34874; 33397); Fonds de Recherche du Québec - Santé (FRQS; Chercheur Boursier, 2020-VICO-279314); Wallenberg Scholar supported by grants, Swedish Research Council (#2018-02532); Swedish State Support for Clinical Research (#ALFGBG-720931); Alzheimer Drug Discovery Foundation (ADDF), USA (#201809-2016862); Dementia Research Institute at UCL.Objectives: The core cerebrospinal fluid (CSF) biomarkers; total tau (tTau), phospho-tau (pTau), amyloid β 1-42 (Aβ 1-42), and the Aβ 1-42/Aβ 1-40 ratio have transformed Alzheimer's disease (AD) research and are today increasingly used in clinical routine laboratories as diagnostic tools. Fully automated immunoassay instruments with ready-to-use assay kits and calibrators has simplified their analysis and improved reproducibility of measurements. We evaluated the analytical performance of the fully automated immunoassay instrument LUMIPULSE G (Fujirebio) for measurement of the four core AD CSF biomarkers and determined cutpoints for AD diagnosis. Methods: Comparison of the LUMIPULSE G assays was performed with the established INNOTEST ELISAs (Fujirebio) for hTau Ag, pTau 181, β-amyloid 1-42, and with V-PLEX Plus Aβ Peptide Panel 1 (6E10) (Meso Scale Discovery) for Aβ 1-42/Aβ 1-40, as well as with a LC-MS reference method for Aβ 1-42. Intra- and inter-laboratory reproducibility was evaluated for all assays. Clinical cutpoints for Aβ 1-42, tTau, and pTau was determined by analysis of three cohorts of clinically diagnosed patients, comprising 651 CSF samples. For the Aβ 1-42/Aβ 1-40 ratio, the cutpoint was determined by mixture model analysis of 2,782 CSF samples. Results: The LUMIPULSE G assays showed strong correlation to all other immunoassays (r>0.93 for all assays). The repeatability (intra-laboratory) CVs ranged between 2.0 and 5.6%, with the highest variation observed for β-amyloid 1-40. The reproducibility (inter-laboratory) CVs ranged between 2.1 and 6.5%, with the highest variation observed for β-amyloid 1-42. The clinical cutpoints for AD were determined to be 409 ng/L for total tau, 50.2 ng/L for pTau 181, 526 ng/L for β-amyloid 1-42, and 0.072 for the Aβ 1-42/Aβ 1-40 ratio. Conclusions: Our results suggest that the LUMIPULSE G assays for the CSF AD biomarkers are fit for purpose in clinical laboratory practice. Further, they corroborate earlier presented reference limits for the biomarkers

Reduced Cortisol and Metabolic Responses of Thin Ewes to an Acute Cold Challenge in Mid-Pregnancy: Implications for Animal Physiology and Welfare

Background: Low food availability leading to reductions in Body Condition Score (BCS; 0 indicates emaciation and 5 obesity) in sheep often coincides with low temperatures associated with the onset of winter in New Zealand. The ability to adapt to reductions in environmental temperature may be impaired in animals with low BCS, in particular during pregnancy when metabolic demand is higher. Here we assess whether BCS affects a pregnant animal’s ability to cope with cold challenges. Methods: Eighteen pregnant ewes with a BCS of 2.760.1 were fed to attain low (LBC: BCS2.360.1), medium (MBC: BCS3.260.2) or high BCS (HBC: BCS3.660.2). Shorn ewes were exposed to a 6-h acute cold challenge in a climate-controlled room (wet and windy conditions, 4.460.1uC) in mid-pregnancy. Blood samples were collected during the BCS change phase, acute cold challenge and recovery phase. Results: During the BCS change phase, plasma glucose and leptin concentrations declined while free fatty acids (FFA) increased in LBC compared to MBC (P,0.01, P,0.01 and P,0.05, respectively) and HBC ewes (P,0.05, P,0.01 and P,0.01, respectively). During the cold challenge, plasma cortisol concentrations were lower in LBC than MBC (P,0.05) and HBC ewes (P,0.05), and FFA and insulin concentrations were lower in LBC than HBC ewes (P,0.05 and P,0.001, respectively). Leptin concentrations declined in MBC and HBC ewes while remaining unchanged in LBC ewes (P,0.01). Glucose concentrations and internal body temperature (Tcore) increased in all treatments, although peak Tcore tended to be higher in HBC ewes (P,0.1). During the recovery phase, T4 concentrations were lower in LBC ewes (P,0.05). Conclusion: Even though all ewes were able to increase Tcore and mobilize glucose, low BCS animals had considerably reduced cortisol and metabolic responses to a cold challenge in mid-pregnancy, suggesting that their ability to adapt to cold challenges through some of the expected pathways was reduced

The gut microbiota and microbial metabolites are associated with tail biting in pigs

Tail biting is an abnormal behaviour that causes stress, injury and pain. Given the critical role of the gut-microbiota in the development of behavioural problems in humans and animals, the aim of this study was to determine whether pigs that are biters, victims of tail biting or controls (nine matched sets of pigs) have a different microbiota composition, diversity and microbial metabolite profile. We collected faecal and blood samples from each individual for analysis. The gut microbiota composition was most different between the biter and the control pigs, with a higher relative abundance of Firmicutes in tail biter pigs than the controls. Furthermore, we detected differences in faecal and plasma short chain fatty acids (SCFA) profiles between the biter and victim pigs, suggesting physiological differences even though they are kept in the same pen. Thus, in addition to supporting an association between the gut microbiota and tail biting in pigs, this study also provides the first evidence of an association between tail biting and SCFA. Therefore, further research is needed to confirm these associations, to determine causality and to study how the SCFA profiles of an individual play a role in the development of tail biting behaviour

Effect of Morphine Administration on Social and Non-Social Play Behaviour in Calves

The objective of this study was to evaluate the effect of morphine on social and non-social play behaviour in calves. Twelve calves experienced four treatments in a cross over 2 × 2 factorial design: Calves received an intravenous injection of morphine or saline 10 min prior to being tested individually or in pairs in an arena for 20 min. Play behaviour was continuously recorded in the arena test. Lying times were recorded in the home pen. Cortisol concentrations were measured before and after testing. In the arena test, calves given morphine tended to perform more social play events than calves given saline, however, morphine administration had no effect on locomotor play. Calves given morphine spent less time lying than calves given saline during the first 4 h after returning to the home pen. Cortisol concentrations were suppressed in calves given morphine. Administration of morphine appeared to increase social play but had no effect on locomotor play in calves. This study highlights the importance of investigating different aspects of play behaviour in animals as some may be more indicative of a positive affective state than others. More studies investigating the effects of morphine on play are needed to confirm the results found in this study

Chronic stress influences attentional and judgement bias and the activity of the HPA axis in sheep.

INTRODUCTION:Environmental challenges are part of everyday life for most domestic animals. However, very little is known about how animals cope emotionally and physiologically with cumulative challenges. This experiment aimed to determine the impact of long-term exposure to environmental challenges on the affective state and hypothalamic-pituitary-adrenal (HPA) axis responses to a subsequent additional acute shearing challenge. METHODS:Sheep were exposed to either a long-term environmental challenge (rest disruption and individual housing) in order to induce chronic stress (chronic stress group) or control conditions (group housing in a field with low stress handling and daily feed rewards, control group). Judgement and attention bias were assessed as measures of the emotional state following several days of the challenge or control treatment (pre-shearing tests). In addition, the responsiveness of the HPA-axis was evaluated using a combined Corticotropin Releasing Hormone and Arginine Vasopressin (CRH/AVP) challenge. Finally, all animals were exposed to an acute shearing challenge, then judgement bias (post-shearing test), HPA-axis and internal body temperature responses were determined. RESULTS:In the pre-shearing judgement bias test, the chronic stress group slightly increased optimism compared to the control treatment. In the attention bias test, the chronic stress group showed reduced vigilance behaviour towards a predator threat and a quicker approach to the food compared to the control treatment. The chronic stress group also had lower plasma ACTH concentrations in response to the CRH/AVP challenge compared to the control group, no differences in cortisol concentrations were found. In the post-shearing judgement bias test, differences in optimism were no longer evident between the chronic stress and control groups. Plasma ACTH concentrations and body temperatures showed a greater increase in response to shearing in the chronic stress group compared to the control group. CONCLUSION:Our results suggest that long-term exposure to challenges biased cognitive measures of the affective state towards an increased expectation of a reward and reduced attention towards a threat. The exaggerated ACTH responses in the chronic stress group may be indicative of HPA-axis dysregulation. Despite a period of challenge exposure in the chronic stress group, judgement bias responses to the shearing challenge were similar in the chronic stress and control groups; the reasons for this need further investigation. The altered affective state together with signs of HPA-axis dysregulation may indicate an increased risk of compromised welfare in animals exposed to long-term environmental challenges

Opioid control of behaviour in sheep: Effects of morphine and naloxone on food intake, activity and the affective state

The affective states of animals are important determinants of welfare, yet they are poorly understood. Here we investigate opioid involvement in the regulation of behaviours that may be indicative of the arousal and valence components of affective states in sheep. Ewes treated with sterile water (C), a low or high dose of the opioid agonist morphine (M1, M2) or opioid antagonist naloxone (N1, N2; 'n' = 8 per treatment) were exposed to a range of different situations, including availability of concentrate feed, grazing in a paddock, novel object test and isolation box test; behavioural indicators, activity and ear postures were assessed. Morphine treated ewes crossed more zones (78.1 ± 5.4 for M1, 'P' = 0.025 and 99.3 ± 5.4 for M2, 'P' = 0.01) compared to C ewes (37.3 ± 5.4) and vocalized more (49.9 ± 6.5 for M1, 'P