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Ewes are more attentive to their offspring experiencing pain but not stress
Applied Animal Behaviour Science 132 (2011) 114–120
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Applied Animal Behaviour Science journal homepage: www.elsevier.com/locate/applanim
Ewes are more attentive to their offspring experiencing pain but not stress Sophie Hild a,∗ , Corinna C.A. Clark b , Catherine M. Dwyer c , Joanna C. Murrell b , Mike Mendl b , Adroaldo J. Zanella a,d a b c d
Department of Production Animal Clinical Sciences, Norwegian School of Veterinary Science, P.O. Box 8146 Dep., 0033 Oslo, Norway University of Bristol, Department of Clinical Veterinary Science, Langford House, Langford BS40 5DU, United Kingdom Sustainable Livestock Systems Group, Scottish Agricultural College, West Mains Road, Edinburgh EH9 3JG, Scotland, United Kingdom Department of Animal and Aquacultural Sciences, Agricultural University of Norway, P.O. Box 5003, 1432 Ås, Norway
a r t i c l e
i n f o
Article history: Accepted 3 April 2011 Available online 2 May 2011 Keywords: Pain Stress Maternal behaviour LPS Lambs Welfare
a b s t r a c t The goal of this experiment was to detect if maternal care by ewes could be effective in mitigating psychological or physiological stress or pain in their offspring. We hypothesised that ewes are able to recognise when their offspring undergo an adverse experience and will adapt their maternal behaviour to buffer the effect of such events. Thirty-one pairs of 2-day-old lambs were subjected to a baseline clinical examination that consisted of measuring body temperature, bodyweight and mechanical pressure pain threshold. On day 3, physiological or psychological stress was applied to one sibling, induced by intravenous injection of LPS or social isolation for 10 min after a further baseline clinical examination, while the other twin experienced only the baseline clinical examination. On day 4, one sibling experienced pain induced by tail docking (females) or tail docking and castration (males) after the baseline clinical examination, while the other twin experienced only the clinical examination. Maternal sniffing, licking, glancing and nursing were recorded for 20 min post-treatment on each day. On day 4, pain behaviours of the treated lamb were also recorded, as well as mother–young inter-individual distances. Measures obtained on day 3 indicated no difference in maternal behaviour directed to the offspring that experienced the stress treatments compared to their twin lamb or to the same lamb on day 2. On day 4, ewes showed more attention to the lamb that had experienced the pain stimulus compared to the other twin lamb and to the same lamb on day 2. The amount of maternal behaviour observed on day 4 was positively correlated with pain behaviours expressed by the treated lamb, suggesting that the ewe was responding to the behaviour of the lamb. We cannot confirm whether the change in maternal behaviour may have buffered the effects of the painful stimuli but the possibility that such a phenomenon occurs later cannot be ruled out. © 2011 Published by Elsevier B.V.
1. Introduction Under production conditions, lambs are likely to encounter adverse events. Sheep are a highly gregarious
∗ Corresponding author. Tel.: +47 22 59 74 82; fax: +47 22 59 70 83. E-mail addresses: [email protected], [email protected] (S. Hild). 0168-1591/$ – see front matter © 2011 Published by Elsevier B.V. doi:10.1016/j.applanim.2011.04.003
species and being isolated is one of the most stressinducing events (Boissy et al., 2005). Psychological distress is provoked by separation from the dam, in particular in the very beginning of life (Dwyer, 2008). Diseases can also provoke a systemic reaction that activates the stress axis (Elenkov et al., 2005) and disrupts the display of normal behaviours. Watery mouth disease, infectious polyarthritis and omphalophlebitis, for example, are common condi-
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tions that impair the wellbeing of the newborn lamb and may threaten survival if not treated in time (Roger, 2008). Experimental injection of LPS (lipopolysaccharide) mimics infection by Escherichia coli but without causing the actual infection. It induces immune stress and symptoms of sickness through a cascade of cytokine synthesis and release (Elmquist et al., 1997). LPS induces fever and reduces food intake and activity, for instance in rats (Harden et al., 2006). In pigs, LPS injection was shown to activate the HPA axis and increase cortisol release (Wright et al., 2000). In several countries, tail docking is commonly carried out in young lambs to prevent fleece soiling or fly strike infections (French et al., 1994). Castration is also a common husbandry practice performed on male lambs in several parts of the world to avoid unwanted mating, to modify the males’ temperament (i.e. decreased aggressiveness; Jewell, 1997) and to change carcass characteristics (Field, 1971; Seideman et al., 1982). Both tail-docking and castration are procedures that cause severe somatic and visceral pain in lambs (Molony et al., 2002). It is not known in sheep if the dam is able to recognise and differentiate indicators of pain and psychological or physiological stress from the lamb and what type of behavioural response these will trigger in the mother. Stress and pain recognition in the offspring could be important from an adaptive perspective, as it could allow the mother to reduce discomfort in the offspring and therefore enhance the chances of the young to avoid predation or resume normal activities, e.g. feeding. It is thus expected that ewes would recognize lambs experiencing difficulties. In our previous study, lambs that were less sensitive to thermal pain stayed closer to their dams (Hild et al., 2010) and a potential analgesic effect linked to proximity with the mother was hypothesised. Mothers from other mammalian species have been shown to modulate pain responses in their young. In the human newborn, skin to skin contact was reported to offer some analgesic effects (Johnston et al., 2003; Gray et al., 2000), while in rats, more licking and grooming of the pups increased their tolerance to a painful stimulus (Blass et al., 1995; Walker et al., 2003, 2008). Landa (2003) observed that suckling, which has been found to be analgesic in other species (humans: Blass and Hoffmeyer, 1991; rats: Ren et al., 1997), was not related to a reduction in pain behaviours after tail docking in lambs, though glucose intake was found to act as an analgesic in their study. It was argued that pain was so intense that any analgesic effect from suckling would be too low to be detected. Nevertheless, it cannot be excluded that another feature of the mother–young relationship may be linked to a lower pain experience in some individuals. The overall hypothesis of this study was that ewes are able to recognise signs of pain and psychological or physiological stress in their offspring and that they respond by showing more maternal care. The first goal was to assess maternal behaviour with the assumption that ewes would show more attention towards lambs displaying signs of stress and pain compared to a situation where pain and stress are not present or compared to a control twin. In a second step, we observed the amount of pain behaviours expressed by the lambs with the prediction that if the ewes expressed more care towards the lamb in pain, it would
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result in less pain-related behaviours from the lambs, i.e. maternal behaviours will be negatively correlated with lamb pain behaviours. 2. Materials and methods The data presented here are the result of an auxiliary study to a larger project of which the aims and experimental design can be found in Clark et al. (2011). 2.1. Animals, housing and feeding The experiment took place at the Department of Clinical Veterinary Science, University of Bristol, Langford, UK. Thirty-one multiparous ewes and their twin lambs were studied between birth and four days of life (Table 1). The ewes were North Country Mules, which is a crossbreed between a Swaledale ewe and a Blueface Leicester ram. All lambs were sired by the same Suffolk ram. Six weeks prior to parturition, the ewes were divided into groups of up to six ewes and housed in 15 m2 pens with straw bedding in a shed open to the exterior along one side. Ewes were fed concentrate and hay twice a day throughout the whole experiment to meet maintenance dietary requirements. Ewes lambed in the group pen. After parturition, the ewe and her offspring were isolated from the other ewes in the group using portable metal fences. Within 24 h, the ewe and her lambs were moved together to individual pens (1.8 m2 ). 2.2. Experimental protocol On the first day post partum, no human intervention occurred. On day 2, a baseline clinical examination was carried out and all lambs were weighed. Lambs were between 22 h and 46 h old. The “baseline clinical examination” (BCE) was carried out similarly on days 3 and 4. It consisted of measurement of body temperature using a rectal thermometer and mechanical pressure pain sensitivity testing in the lambs. Procedures and results from the BCE are detailed in Clark et al. (2011). 2.2.1. Treatments Lambs were part-randomly allocated (balanced for weight) to each of the different treatments (Table 1). Stress treatments were applied on day 3, when lambs were between 46 h and 70 h old, and pain treatments were applied on day 4, when lambs were between 70 h and 94 h old. Some lambs received the stressor alone (day 3), the pain stimulus alone (day 4), both stimuli (days 3 and 4), or neither (Table 1). Pairs of siblings were included in the study where, on particular treatment days, one sibling received a treatment and the other did not. This facilitated comparison of the ewe’s behaviour, as one lamb in each pair acted as the control lamb (Table 1). 2.2.2. Stress treatments, day 3 Baseline clinical examination was carried out before the treatments. The treatments consisted of either intravenous injection of LPS (lipopolysaccharide) or social isolation. These treatments reflect some of the challenges likely
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Table 1 Distribution of the 31 pairs of siblings used in total according to their treatment. Grey boxes indicate where no comparison in maternal behaviours was possible as twins both received the same treatment or no treatment. Sibling pairs
Day 3
Day 4
Number of pairs
1
Controla Stressb
No stimulus Stress
No stimulus No stimulus
3
2
Control Painc
No stimulus No stimulus
No stimulus Pain
6
3
Control Stress and pain
No stimulus Stress
No stimulus Pain
8
4
Stress Pain
Stress No stimulus
No stimulus Pain
6
5
Pain Stress and pain
No stimulus Stress
Pain Pain
7
6
Stress Stress and pain
Stress Stress
No stimulus Pain
1
a
Lambs were used as controls where no stimulus was applied after the baseline clinical examination. The stress treatments (n = 24 pairs of lambs used) consisted of either lipopolysaccharide injection intravenously (n = 17: nine females, eight males) or social isolation (n = seven males). c The pain treatment (n = 21 pairs of lambs used) consisted in either tail docking (n = five females) or tail docking and castration (n = 16 males). b
encountered by lambs (see Section 2.5). Lambs in the LPS treatment group were injected with 0.2 g/kg of LPS into their jugular vein (E. coli serotype 0127:B8, Sigma–Aldrich in a solution concentration of 1 g LPS/ml saline). The effects of LPS were generally lying or reduced activity and were most noticeable 30–60 min after injection. Social isolation was only applied to male lambs. The isolated lambs were separated from the dam and sibling, put in a trolley and moved to a sheltered place 20 m from the experimental shed. The lamb was returned to the home pen after 10 min. Vocal signals were not prevented.
3. The ewe partially followed her offspring and separated from the other ewes but did not leave the pen. 4. The ewe followed the lambs out of the pen for 5 m with more than 30 cm distance from the lambs. 5. The ewe followed the lamb out of the pen for 5 m in contact or within 30 cm distance from the lambs.
2.2.3. Pain treatments, day 4 Baseline clinical examination was carried out before the pain treatments. Lambs in the pain treatment group were either tail-docked, in case of females, or tail-docked and castrated in case of male lambs. The procedures were carried out without the use of anaesthetics which is in line with common husbandry practice. A latex rubber ring was applied using an elastrator to the neck of the scrotum and at the end of the hairless skin area under the tail (Molony et al., 2002).
2.3.3. Treatments days (days 3 and 4) On the stress day, behaviours (Table 2) were recorded continuously for 20 min, starting 10 min after injection of LPS in the treated twin and immediately after return of the isolated twin to the home pen. On the pain day, behaviours listed in Table 2 were recorded continuously for 20 min, starting 10 min after the pain treatment. In addition to these behaviours, scan sampling of the distances between the lamb in pain and the ewe were estimated by eye and recorded as follows every minute (21 scans in total): Contact: contact to 10 cm; Close: 10–50 cm; Apart: >50 cm.
2.3.2. Baseline day (day 2) After the baseline clinical examination and 10 min after the experimenters left the pen, behaviours were recorded for 20 min. The recorded behaviours are listed in Table 2.
2.3. Behaviours recorded 2.4. Statistics 2.3.1. Follow score A follow score was given to the ewe in a test carried out within the first 24 h post partum when the ewe was moved with her offspring from the group pen to the individual pen. The protocol was modified from Dwyer (2008) to test the motivation of the ewe to follow her offspring being carried away by a human experimenter. A human carried the lambs at knee height and slowly backed out of the group pen for a distance of approximately 5 m. A score from 1 to 5 was then given to the ewe according to her behaviour: 1. The ewe did not follow or orient towards her offspring. 2. The ewe did not follow her offspring but oriented towards the lambs and may bleat.
All statistical tests were performed using JMP version 8 (SAS institute). The level of significance was set at P < 0.05 and a trend at P < 0.1. On the treatment days, only the twins where one lamb was treated and the sibling was a control were used for analysis. Wilcoxon matched-pairs signed ranks tests were performed to detect differences within the same lamb between test points (baseline vs. stress days and baseline vs. pain days) and two-sample Wilcoxon rank sum tests (equivalent to the Mann–Whitney U for independent samples) for differences between siblings (control vs. stress lambs on stress day or control vs. pain lambs on pain day). Spearman rank correlation test was used to test for correlations between the ewe sniffing or glancing, the
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Table 2 Behaviours continuously recorded for 20 min on days 1–3. Indicators of mother–young relationship were exploring, glancing and teat seeking. Indicators of pain behaviours (recorded only on day 4) were restlessness, active pain avoidance and pain-related postures. These behaviours were only recorded with the pain experience, as they rarely occurred on the stress day. Behaviour
Description
Ewe
Explores (sniffs/licks) Glances
Number of times the head of the ewe is directed to the lamb, the nose within 5 cm of the lamb showing a brief body immobility, usually accompanied by a stretch of the neck Number of times the ewe shows a brief movement of the head towards a lamb and paused for less than a second
Lamb
Teat seeking
Time spent (s) with the head in a correct sucking-position towards the udder, with or without actual teat seeking and sucking. Reflects “nursing” behaviour by the ewe Number of changes between lying down and standing up Sum of occurrences for foot stamping, kicking, easing quarters by pushing or stretching the limbs while standing or lying, jumping, rolling from one side to another, head turning to look at or touch the source of pain and tail wagging (rapid side-to-side tail movement (tail wags while teat seeking was not recorded) Total duration of time (s) spent in abnormal standing/walking (tucked abdomen, hind limbs slightly apart and further back than normal, walking unsteadily, backwards or on knees, falling) and abnormal lying position (abnormal ventral or lateral recumbency with the hind legs partially or fully extended and “dog sitting” to avoid contact between the scrotal region and the ground)
Restlessness Active pain avoidance Pain-related postures
lambs’ teat-seeking on stress or pain days, the behavioural responses of the lambs to the painful procedures (Table 2: sum of all pain related postures, active pain avoidance, and restlessness), and ewe-lamb distances (Contact, Close and Apart). Distances as well as pain-related behaviours and postures are given as medians with 95% confidence interval. Due to very little variation in the follow scores (80% of the ewes obtained 5 or 4), they were not included in the analyses. Gender of the offspring did not affect the ewe behaviour on baseline day. Similarly, maternal behaviour on day 4 (pain day) was not affected by prior experience of the stressors in some of the pain lambs (from comparison of lambs that experienced stress vs. those that were controls on day 3). These factors were thus not included in our analysis. Both stress treatments on day 3 (LPS and isolation) and both pain treatments on day 4 (tail-docking alone or tail-docking plus castration) were pooled together in the analysis to observe a wider range of stress and pain responses in the lambs and to increase the probability of detecting important associations. 2.5. Ethical considerations All procedures were completed under UK Home Office License number 30/2420. The procedures were carried out to develop and improve knowledge about whether ewes can diminish stress and pain experience in their offspring. We deliberately chose stimuli which simulate the type of events that young lambs might encounter under commercial husbandry conditions. Intravenous injection of LPS is relevant to mimic an E. coli infection, which is responsible for e.g. watery mouth disease in lambs under normal husbandry conditions. The dose used was low and the effects were short-lived. Tail docking and castration without use of anaesthetics are normal husbandry practice in the UK and several other countries despite numerous studies providing evidence that these procedures cause severe acute pain in the lambs (Mellor and Murray, 1989; Molony and Kent, 1997; Molony et al., 2002).
3. Results 3.1. Comparisons of behaviour between and within days 3.1.1. Comparison between baseline day and treatment days Stress lambs sought the teat significantly more on stress day than on baseline day (one-sample Wilcoxon signedrank test: P = 0.016), although this tended to be true for control lambs as well (P = 0.073; Fig. 1). No significant differences were detected between baseline and stress day for ewe exploring or ewe glancing (Fig. 1). On the pain day, the ewe explored significantly more often (P = 0.003) and glanced more often at pain lambs (P < 0.001; Fig. 2) than on baseline day. Control lambs showed teat seeking behaviour for a longer duration on pain day than on baseline day (P = 0.029; Fig. 2) but this was not observed in the pain lambs. 3.1.2. Comparison between treated lambs and control lambs on the same day No significant difference was detected between control lambs and stress lambs for any of the observed behaviours on the stress day (Fig. 1). On the pain day, ewes explored (2-sample Wilcoxon test: P = 0.025), and glanced at pain lambs (P < 0.001; Fig. 2) significantly more often than control lambs. No significant difference was detected between pain and control lambs for teat seeking behaviour (Fig. 2). 3.2. Correlations between maternal behaviours, distances and pain responses Lambs showed active pain avoidance behaviours 82 [68.9–132.6] times and were observed in a pain-related posture for 789 [620.2–952.0] s during the 20 min of observation. Measures of restlessness were obtained by counting the 16 [12.5–20.0] times that the lamb stood up and lay down. Active pain avoidance behaviours in the lambs were significantly positively correlated with the duration of time that the ewe spent exploring the lamb and the number
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Fig. 1. Maternal behaviours towards control lambs (C) and stress lambs (S) on the baseline day and on the stress day. Box plots express median, 25–75% quartiles, minimum, maximum, and outliers, respectively. Probabilities: comparison between days using a one-way Wilcoxon test and comparison within days using a two-sample Wilcoxon test: t: P < 0.1; *: P < 0.05.
of glances directed at the lamb (Table 3). The glancing behaviour also tended to be positively correlated with the duration of time spent in a pain-related posture by the lamb and its restlessness (number of lying down and standing up, Table 3). No other measures, including distances, were found to be significantly correlated with the pain behaviours of the lamb. Overall, the ewe and her lamb in pain were observed in Contact: 14.3 [4.8–28.6]% of total observations, Close: 38.1 [26.0–50.2]% of total observations and Apart: 38.1 [25.9–59.7]% of total observations.
4. Discussion In this study, we observed the ewe maternal behaviour towards her twin lambs when one of them underwent an immune or psychological stress and/or a painful experience. Ewes expressed similar attention towards their lamb experiencing immune or psychological stress as towards the control lambs but they showed more attention to the lambs undergoing the pain treatment than to the control sibling. Ewes nursed both lambs in a similar manner, as
Fig. 2. Maternal behaviours towards control lambs (C) and pain lambs (P) on baseline day and on pain day. Box plots express median, 25–75% quartiles, minimum, maximum, and outliers, respectively. Probabilities: comparison between days using a one-way Wilcoxon test and comparison within days using a two-sample Wilcoxon test: *: P < 0.05. **: P < 0.01. ***: P < 0.001.
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Table 3 Spearman correlations between maternal behaviours and pain related behaviours of lambs experiencing tail docking or tail docking and castration on day 4 (r: Spearman rho, P: probability level).
Restlessness Active pain avoidance Pain related postures
Exploring the lamb in pain
Glancing at the lamb in pain
Teat seeking by lamb in pain
r
P
r
P
r
P
0.0 0.6 0.3
ns 0.008* ns
0.4 0.5 0.4
0.083 0.018* 0.066
0.1 −0.1 −0.3
ns ns ns
ns: P > 0.1. * P < 0.05.
measured by teat seeking by the lambs, on both treatment days and not differently than on baseline day, which is likely to be related to the rare occurrence of the teat seeking behaviour. Interestingly, the amount of attention given by the ewe to a lamb was positively correlated to the amount of pain-related behaviour expressed by the tail-docked and castrated lambs. These results validated the first hypothesis: based on their attention behaviour, the ewes were able to discriminate between a lamb experiencing pain and its control twin, or the same lamb on the baseline day. The positive correlation between maternal behaviours and pain-related behaviours in the lamb suggests that more attention from the ewe was drawn by more pain-related behaviours. Our hypothesis that more care from the dam would result in less expression of pain in the young during the observed period is thus not supported. The proximate mechanism, or immediate factor responsible for the observed results, could be curiosity, drawing the ewe’s attention to signals of severe pain in the offspring. Sheep possess acute eyesight which enables efficient vigilance behaviour among other abilities (Hutson, 2000) and likely predisposes them to detecting subtle behavioural changes in conspecifics. Due to technical difficulty to differentiate vocalisations expressed by the studied individuals from those produced in the rest of the flock, vocalisations were not included here. To understand better which features of the young may have raised the dam’s attention, it would be useful to record the vocal activity of both lambs and ewes. Indeed, it was shown in ewes that stress can be conveyed through variations in the bleating frequency (Sèbe et al., 2009). In addition, chemical signals may increase arousal in individuals, e.g. in rats (Inagaki et al., 2008) and should also be investigated. Processes such as emotional contagion, which designates the emotional convergence of an individual towards the emotional state of another individual, may have been involved (Preston and de Waal, 2002). Such processes have been demonstrated in nonhuman primates (de Waal, 2008) and mice (Langford et al., 2006) when the individuals were familiar to each other. It may have clear adaptive value in this context (cf. de Waal, 2008). The stressed lambs did not elicit a detectable response in the ewe. This may be due to a more discrete expression of stress behaviours by the lambs that could have failed to attract their mother’s attention. We cannot ignore the possibility that the sensitivity of the behavioural measuring systems may not have been high enough and that ewes may have expressed more subtle behaviours, such as differences in ear postures, which could indicate an increased negative emotional valence (Reefmann
et al., 2009) and which were not recorded in the current study. Behaviours likely to relieve pain in the young included nursing and licking, as observed in mice where dams showing more licking and grooming of their pups decreased their pups’ sensitivity to thermal pain (Walker et al., 2008), or physical proximity, as we reported in a previous study (Hild et al., 2010) that lambs that were less sensitive to thermal pain were observed to be closer to their dam. These behaviours were not expressed differently according to the treatment of the offspring in the current study. In our protocol, behaviours were only observed up to 30 min after application of stress or pain stimuli. It should be tested whether, when given enough time, the correlation between maternal behaviours and the lamb pain related behaviours would be reversed. As the frequency of active pain behaviours begins to decrease 30–60 min after castration and tail-docking (Graham et al., 1997), the collection of physiological data such as cortisol (Graham et al., 1997), heart rate variability (Stubsjøen et al., 2009) or an index of fatigue (e.g. plasma glucose or temperature; Molony and Kent, 1997) could be implemented. Alternatively, it may not make sense for the ewe to show these potentially pain mitigating behaviours in the artificial testing conditions in which we conducted our observations. It is possible that in a more “ecological” setting, e.g. allowing the ewe to move further away from the lamb or including predatory threats, we may have observed a different range of maternal responses to an offspring in pain. In conclusion, this study represents a preliminary investigation of the impact of adverse events experienced by lambs on the dam’s response. Ewes showed more attention to their offspring experiencing pain but not stress. Whether the ewes could recognise that their lamb was in pain is not resolved and emotional contagion should be investigated in the future. No alleviation of pain corresponding to increased maternal attention and care was detected within the observation period. To complete testing this hypothesis, more sensitive methods of assessment and longer observation periods should be used. Acknowledgements The authors wish to thank Jennifer Jamieson and Ashley Arnone for experimental help, Stuart Pope for technical support, the veterinarians from Langford clinic, the project funder BBSRC, the Research Council of Norway (Forskningsrådet – project 185163), the Norwegian Committee for Research and Ethics (Utvalg for Forskning og Etikk) for
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financial support to Sophie Hild and Dr Inger Lise Andersen for commenting on the manuscript. References Blass, E.M., Hoffmeyer, L.B., 1991. Sucrose as an analgesic for newborninfants. Pediatrics 87, 215–218. Blass, E.M., Shide, D.J., Zawmon, C., Sorrentino, J., 1995. Mother as shield – differential-effects of contact and nursing on pain responsivity in infant rats – evidence for nonopioid mediation. Behav. Neurosci. 109, 342–353. Boissy, A., Fisher, A.D., Bouix, J., Hinch, G.N., Le Neindre, P., 2005. Genetics of fear in ruminant livestock. Livest. Prod. Sci. 93, 23–32. Clark, C., Mendl, M., Jamieson, J., Arnone, A., Waterman-Pearson, A., Murrell, J., 2011. Do psychological and physiological stressors alter the acute pain response to castration and tail docking in lambs? Vet. Anaesth. Analg. 38, 134–145, doi:10.1111/j.1467-2995.2011.00594.x. de Waal, F.B.M., 2008. Putting the altruism back into altruism: the evolution of empathy. Annu. Rev. Psychol. 59, 279–300. Dwyer, C.M., 2008. Genetic and physiological determinants of maternal behavior and lamb survival: implications for low-input sheep management. J. Anim. Sci. 86, 246–258. Elenkov, I.J., Iezzoni, D.G., Daly, A., Harris, A.G., Chrousos, G.P., 2005. Cytokine dysregulation, inflammation and well-being. Neuroimmunomodulation 12, 255–269. Elmquist, J.K., Scammell, T.E., Saper, C.B., 1997. Mechanisms of CNS response to systemic immune challenge: the febrile response. Trends Neurosci. 20, 565–570. Field, R.A., 1971. Effect of castration on meat quality and quantity. J. Anim. Sci. 32, 849–858. French, N.P., Wall, R., Morgan, K.L., 1994. Tail docking of lambs in the control of flystrike. Vet. Rec. 135, 47. Graham, M.J., Kent, J.E., Molony, V., 1997. Effects of four analgesic treatments on the behavioural and cortisol responses of 3-week-old lambs to tail docking. Vet. J. 153, 87–97. Gray, L., Watt, L., Blass, E.M., 2000. Skin-to-skin contact is analgesic in healthy newborns. Pediatrics, 105. Harden, L.M., du Plessis, I., Poole, S., Laburn, H.P., 2006. Interleukin-6 and leptin mediate lipopolysaccharide-induced fever and sickness behavior. Physiol. Behav. 89, 146–155. Hild, S., Andersen, I.L., Zanella, A.J., 2010. The relationship between thermal nociceptive threshold in lambs and ewe-lamb interactions. Small Ruminant Res. 90, 142–145. Hutson, G.D., 2000. Behavioural principles of sheep handling. In: Grandin, T. (Ed.), Livestock Handling and Transport. CAB International North America, p. 175. Inagaki, H., Kiyokawa, Y., Kikusui, T., Takeuchi, Y., Mori, Y., 2008. Enhancement of the acoustic startle reflex by an alarm pheromone in male rats. Physiol. Behav. 93, 606–611. Jewell, P.A., 1997. Survival and behaviour of castrated Soay sheep (Ovis aries) in a feral island population on Hirta, St. Kilda, Scotland. J. Zool. 243, 623–636.
Johnston, C.C., Stevens, B., Pinelli, J., Gibbins, S., Filion, F., Jack, A., Steele, S., Boyer, K., Veilleux, A., 2003. Kangaroo care is effective in diminishing pain response in preterm neonates. Arch. Pediatr. Adolesc. Med. 157, 1084–1088. Landa, L., 2003. The effect of milk suckling from the dam or glucose administration on the behavioural responses to tail docking in lambs. Acta Vet. Brno 72, 175–182. Langford, D.J., Crager, S.E., Shehzad, Z., Smith, S.B., Sotocinal, S.G., Levenstadt, J.S., Chanda, M.L., Levitin, D.J., Mogil, J.S., 2006. Social modulation of pain as evidence for empathy in mice. Science 312, 1967–1970. Mellor, D., Murray, L., 1989. Effects of tail docking and castration on behaviour and plasma cortisol concentration in young lambs. Res. Vet. Sci. 46, 387–391. Molony, V., Kent, J., 1997. Assessment of acute pain in farm animals using behavioral and physiological measurements. J. Anim. Sci. 75, 266–272. Molony, V., Kent, J.E., McKendrick, I.J., 2002. Validation of a method for assessment of an acute pain in lambs. Appl. Anim. Behav. Sci. 76, 215–238. Preston, S.D., de Waal, F.B.M., 2002. Empathy: its ultimate and proximate bases. Behav. Brain Sci. 25, 1–20. Reefmann, N., Kaszas, F.B., Wechsler, B., Gygax, L., 2009. Ear and tail postures as indicators of emotional valence in sheep. Appl. Anim. Behav. Sci. 118, 199–207. Ren, K., Blass, E.M., Zhou, Q.Q., Dubner, R., 1997. Suckling and sucrose ingestion suppress persistent hyperalgesia and spinal Fos expression after forepaw inflammation in infant rats. Proc. Natl. Acad. Sci. U.S.A. 94, 1471–1475. Roger, P.A., 2008. The impact of disease and disease prevention on sheep welfare. Small Ruminant Res. 76, 104–111. Sèbe, F., Poindron, P., Reby, D., Nowak, R., Blache, D., 2009. Encoding and genetic selection of psycho-acoustic markers of emotions in sheep. In: Proceedings of the 43rd Congress of the International Society for Applied Ethology , p. 133. Seideman, S.C., Cross, H.R., Oltjen, R.R., Schanbacher, B.D., 1982. Utilization of the intact male for red meat production—a review. J. Anim. Sci. 55, 826–840. Stubsjøen, S.M., Flø, A.S., Moe, R.O., Janczak, A.M., Skjerve, E., Valle, P.S., Zanella, A.J., 2009. Exploring non-invasive methods to assess pain in sheep. Physiol. Behav. 98, 640–648. Walker, C.D., Kudreikis, K., Sherrard, A., Johnston, C.C., 2003. Repeated neonatal pain influences maternal behavior, but not stress responsiveness in rat offspring. Dev. Brain Res. 140, 253–261. Walker, C.D., Xu, Z.F., Rochford, J., Johnston, C.C., 2008. Naturally occuring variations in maternal care modulate the effects of repeated neonatal pain on behavioral sensitivity to thermal pain in the adult offspring. Pain 140, 167–176. Wright, K.J., Balaji, R., Hill, C.M., Dritz, S.S., Knoppel, E.L., Minton, J.E., 2000. Integrated adrenal, somatotropic, and immune responses of growing pigs to treatment with lipopolysaccharide. J. Anim. Sci. 78, 1892–1899.
Contents lists available at ScienceDirect
Applied Animal Behaviour Science journal homepage: www.elsevier.com/locate/applanim
Ewes are more attentive to their offspring experiencing pain but not stress Sophie Hild a,∗ , Corinna C.A. Clark b , Catherine M. Dwyer c , Joanna C. Murrell b , Mike Mendl b , Adroaldo J. Zanella a,d a b c d
Department of Production Animal Clinical Sciences, Norwegian School of Veterinary Science, P.O. Box 8146 Dep., 0033 Oslo, Norway University of Bristol, Department of Clinical Veterinary Science, Langford House, Langford BS40 5DU, United Kingdom Sustainable Livestock Systems Group, Scottish Agricultural College, West Mains Road, Edinburgh EH9 3JG, Scotland, United Kingdom Department of Animal and Aquacultural Sciences, Agricultural University of Norway, P.O. Box 5003, 1432 Ås, Norway
a r t i c l e
i n f o
Article history: Accepted 3 April 2011 Available online 2 May 2011 Keywords: Pain Stress Maternal behaviour LPS Lambs Welfare
a b s t r a c t The goal of this experiment was to detect if maternal care by ewes could be effective in mitigating psychological or physiological stress or pain in their offspring. We hypothesised that ewes are able to recognise when their offspring undergo an adverse experience and will adapt their maternal behaviour to buffer the effect of such events. Thirty-one pairs of 2-day-old lambs were subjected to a baseline clinical examination that consisted of measuring body temperature, bodyweight and mechanical pressure pain threshold. On day 3, physiological or psychological stress was applied to one sibling, induced by intravenous injection of LPS or social isolation for 10 min after a further baseline clinical examination, while the other twin experienced only the baseline clinical examination. On day 4, one sibling experienced pain induced by tail docking (females) or tail docking and castration (males) after the baseline clinical examination, while the other twin experienced only the clinical examination. Maternal sniffing, licking, glancing and nursing were recorded for 20 min post-treatment on each day. On day 4, pain behaviours of the treated lamb were also recorded, as well as mother–young inter-individual distances. Measures obtained on day 3 indicated no difference in maternal behaviour directed to the offspring that experienced the stress treatments compared to their twin lamb or to the same lamb on day 2. On day 4, ewes showed more attention to the lamb that had experienced the pain stimulus compared to the other twin lamb and to the same lamb on day 2. The amount of maternal behaviour observed on day 4 was positively correlated with pain behaviours expressed by the treated lamb, suggesting that the ewe was responding to the behaviour of the lamb. We cannot confirm whether the change in maternal behaviour may have buffered the effects of the painful stimuli but the possibility that such a phenomenon occurs later cannot be ruled out. © 2011 Published by Elsevier B.V.
1. Introduction Under production conditions, lambs are likely to encounter adverse events. Sheep are a highly gregarious
∗ Corresponding author. Tel.: +47 22 59 74 82; fax: +47 22 59 70 83. E-mail addresses: [email protected], [email protected] (S. Hild). 0168-1591/$ – see front matter © 2011 Published by Elsevier B.V. doi:10.1016/j.applanim.2011.04.003
species and being isolated is one of the most stressinducing events (Boissy et al., 2005). Psychological distress is provoked by separation from the dam, in particular in the very beginning of life (Dwyer, 2008). Diseases can also provoke a systemic reaction that activates the stress axis (Elenkov et al., 2005) and disrupts the display of normal behaviours. Watery mouth disease, infectious polyarthritis and omphalophlebitis, for example, are common condi-
S. Hild et al. / Applied Animal Behaviour Science 132 (2011) 114–120
tions that impair the wellbeing of the newborn lamb and may threaten survival if not treated in time (Roger, 2008). Experimental injection of LPS (lipopolysaccharide) mimics infection by Escherichia coli but without causing the actual infection. It induces immune stress and symptoms of sickness through a cascade of cytokine synthesis and release (Elmquist et al., 1997). LPS induces fever and reduces food intake and activity, for instance in rats (Harden et al., 2006). In pigs, LPS injection was shown to activate the HPA axis and increase cortisol release (Wright et al., 2000). In several countries, tail docking is commonly carried out in young lambs to prevent fleece soiling or fly strike infections (French et al., 1994). Castration is also a common husbandry practice performed on male lambs in several parts of the world to avoid unwanted mating, to modify the males’ temperament (i.e. decreased aggressiveness; Jewell, 1997) and to change carcass characteristics (Field, 1971; Seideman et al., 1982). Both tail-docking and castration are procedures that cause severe somatic and visceral pain in lambs (Molony et al., 2002). It is not known in sheep if the dam is able to recognise and differentiate indicators of pain and psychological or physiological stress from the lamb and what type of behavioural response these will trigger in the mother. Stress and pain recognition in the offspring could be important from an adaptive perspective, as it could allow the mother to reduce discomfort in the offspring and therefore enhance the chances of the young to avoid predation or resume normal activities, e.g. feeding. It is thus expected that ewes would recognize lambs experiencing difficulties. In our previous study, lambs that were less sensitive to thermal pain stayed closer to their dams (Hild et al., 2010) and a potential analgesic effect linked to proximity with the mother was hypothesised. Mothers from other mammalian species have been shown to modulate pain responses in their young. In the human newborn, skin to skin contact was reported to offer some analgesic effects (Johnston et al., 2003; Gray et al., 2000), while in rats, more licking and grooming of the pups increased their tolerance to a painful stimulus (Blass et al., 1995; Walker et al., 2003, 2008). Landa (2003) observed that suckling, which has been found to be analgesic in other species (humans: Blass and Hoffmeyer, 1991; rats: Ren et al., 1997), was not related to a reduction in pain behaviours after tail docking in lambs, though glucose intake was found to act as an analgesic in their study. It was argued that pain was so intense that any analgesic effect from suckling would be too low to be detected. Nevertheless, it cannot be excluded that another feature of the mother–young relationship may be linked to a lower pain experience in some individuals. The overall hypothesis of this study was that ewes are able to recognise signs of pain and psychological or physiological stress in their offspring and that they respond by showing more maternal care. The first goal was to assess maternal behaviour with the assumption that ewes would show more attention towards lambs displaying signs of stress and pain compared to a situation where pain and stress are not present or compared to a control twin. In a second step, we observed the amount of pain behaviours expressed by the lambs with the prediction that if the ewes expressed more care towards the lamb in pain, it would
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result in less pain-related behaviours from the lambs, i.e. maternal behaviours will be negatively correlated with lamb pain behaviours. 2. Materials and methods The data presented here are the result of an auxiliary study to a larger project of which the aims and experimental design can be found in Clark et al. (2011). 2.1. Animals, housing and feeding The experiment took place at the Department of Clinical Veterinary Science, University of Bristol, Langford, UK. Thirty-one multiparous ewes and their twin lambs were studied between birth and four days of life (Table 1). The ewes were North Country Mules, which is a crossbreed between a Swaledale ewe and a Blueface Leicester ram. All lambs were sired by the same Suffolk ram. Six weeks prior to parturition, the ewes were divided into groups of up to six ewes and housed in 15 m2 pens with straw bedding in a shed open to the exterior along one side. Ewes were fed concentrate and hay twice a day throughout the whole experiment to meet maintenance dietary requirements. Ewes lambed in the group pen. After parturition, the ewe and her offspring were isolated from the other ewes in the group using portable metal fences. Within 24 h, the ewe and her lambs were moved together to individual pens (1.8 m2 ). 2.2. Experimental protocol On the first day post partum, no human intervention occurred. On day 2, a baseline clinical examination was carried out and all lambs were weighed. Lambs were between 22 h and 46 h old. The “baseline clinical examination” (BCE) was carried out similarly on days 3 and 4. It consisted of measurement of body temperature using a rectal thermometer and mechanical pressure pain sensitivity testing in the lambs. Procedures and results from the BCE are detailed in Clark et al. (2011). 2.2.1. Treatments Lambs were part-randomly allocated (balanced for weight) to each of the different treatments (Table 1). Stress treatments were applied on day 3, when lambs were between 46 h and 70 h old, and pain treatments were applied on day 4, when lambs were between 70 h and 94 h old. Some lambs received the stressor alone (day 3), the pain stimulus alone (day 4), both stimuli (days 3 and 4), or neither (Table 1). Pairs of siblings were included in the study where, on particular treatment days, one sibling received a treatment and the other did not. This facilitated comparison of the ewe’s behaviour, as one lamb in each pair acted as the control lamb (Table 1). 2.2.2. Stress treatments, day 3 Baseline clinical examination was carried out before the treatments. The treatments consisted of either intravenous injection of LPS (lipopolysaccharide) or social isolation. These treatments reflect some of the challenges likely
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Table 1 Distribution of the 31 pairs of siblings used in total according to their treatment. Grey boxes indicate where no comparison in maternal behaviours was possible as twins both received the same treatment or no treatment. Sibling pairs
Day 3
Day 4
Number of pairs
1
Controla Stressb
No stimulus Stress
No stimulus No stimulus
3
2
Control Painc
No stimulus No stimulus
No stimulus Pain
6
3
Control Stress and pain
No stimulus Stress
No stimulus Pain
8
4
Stress Pain
Stress No stimulus
No stimulus Pain
6
5
Pain Stress and pain
No stimulus Stress
Pain Pain
7
6
Stress Stress and pain
Stress Stress
No stimulus Pain
1
a
Lambs were used as controls where no stimulus was applied after the baseline clinical examination. The stress treatments (n = 24 pairs of lambs used) consisted of either lipopolysaccharide injection intravenously (n = 17: nine females, eight males) or social isolation (n = seven males). c The pain treatment (n = 21 pairs of lambs used) consisted in either tail docking (n = five females) or tail docking and castration (n = 16 males). b
encountered by lambs (see Section 2.5). Lambs in the LPS treatment group were injected with 0.2 g/kg of LPS into their jugular vein (E. coli serotype 0127:B8, Sigma–Aldrich in a solution concentration of 1 g LPS/ml saline). The effects of LPS were generally lying or reduced activity and were most noticeable 30–60 min after injection. Social isolation was only applied to male lambs. The isolated lambs were separated from the dam and sibling, put in a trolley and moved to a sheltered place 20 m from the experimental shed. The lamb was returned to the home pen after 10 min. Vocal signals were not prevented.
3. The ewe partially followed her offspring and separated from the other ewes but did not leave the pen. 4. The ewe followed the lambs out of the pen for 5 m with more than 30 cm distance from the lambs. 5. The ewe followed the lamb out of the pen for 5 m in contact or within 30 cm distance from the lambs.
2.2.3. Pain treatments, day 4 Baseline clinical examination was carried out before the pain treatments. Lambs in the pain treatment group were either tail-docked, in case of females, or tail-docked and castrated in case of male lambs. The procedures were carried out without the use of anaesthetics which is in line with common husbandry practice. A latex rubber ring was applied using an elastrator to the neck of the scrotum and at the end of the hairless skin area under the tail (Molony et al., 2002).
2.3.3. Treatments days (days 3 and 4) On the stress day, behaviours (Table 2) were recorded continuously for 20 min, starting 10 min after injection of LPS in the treated twin and immediately after return of the isolated twin to the home pen. On the pain day, behaviours listed in Table 2 were recorded continuously for 20 min, starting 10 min after the pain treatment. In addition to these behaviours, scan sampling of the distances between the lamb in pain and the ewe were estimated by eye and recorded as follows every minute (21 scans in total): Contact: contact to 10 cm; Close: 10–50 cm; Apart: >50 cm.
2.3.2. Baseline day (day 2) After the baseline clinical examination and 10 min after the experimenters left the pen, behaviours were recorded for 20 min. The recorded behaviours are listed in Table 2.
2.3. Behaviours recorded 2.4. Statistics 2.3.1. Follow score A follow score was given to the ewe in a test carried out within the first 24 h post partum when the ewe was moved with her offspring from the group pen to the individual pen. The protocol was modified from Dwyer (2008) to test the motivation of the ewe to follow her offspring being carried away by a human experimenter. A human carried the lambs at knee height and slowly backed out of the group pen for a distance of approximately 5 m. A score from 1 to 5 was then given to the ewe according to her behaviour: 1. The ewe did not follow or orient towards her offspring. 2. The ewe did not follow her offspring but oriented towards the lambs and may bleat.
All statistical tests were performed using JMP version 8 (SAS institute). The level of significance was set at P < 0.05 and a trend at P < 0.1. On the treatment days, only the twins where one lamb was treated and the sibling was a control were used for analysis. Wilcoxon matched-pairs signed ranks tests were performed to detect differences within the same lamb between test points (baseline vs. stress days and baseline vs. pain days) and two-sample Wilcoxon rank sum tests (equivalent to the Mann–Whitney U for independent samples) for differences between siblings (control vs. stress lambs on stress day or control vs. pain lambs on pain day). Spearman rank correlation test was used to test for correlations between the ewe sniffing or glancing, the
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Table 2 Behaviours continuously recorded for 20 min on days 1–3. Indicators of mother–young relationship were exploring, glancing and teat seeking. Indicators of pain behaviours (recorded only on day 4) were restlessness, active pain avoidance and pain-related postures. These behaviours were only recorded with the pain experience, as they rarely occurred on the stress day. Behaviour
Description
Ewe
Explores (sniffs/licks) Glances
Number of times the head of the ewe is directed to the lamb, the nose within 5 cm of the lamb showing a brief body immobility, usually accompanied by a stretch of the neck Number of times the ewe shows a brief movement of the head towards a lamb and paused for less than a second
Lamb
Teat seeking
Time spent (s) with the head in a correct sucking-position towards the udder, with or without actual teat seeking and sucking. Reflects “nursing” behaviour by the ewe Number of changes between lying down and standing up Sum of occurrences for foot stamping, kicking, easing quarters by pushing or stretching the limbs while standing or lying, jumping, rolling from one side to another, head turning to look at or touch the source of pain and tail wagging (rapid side-to-side tail movement (tail wags while teat seeking was not recorded) Total duration of time (s) spent in abnormal standing/walking (tucked abdomen, hind limbs slightly apart and further back than normal, walking unsteadily, backwards or on knees, falling) and abnormal lying position (abnormal ventral or lateral recumbency with the hind legs partially or fully extended and “dog sitting” to avoid contact between the scrotal region and the ground)
Restlessness Active pain avoidance Pain-related postures
lambs’ teat-seeking on stress or pain days, the behavioural responses of the lambs to the painful procedures (Table 2: sum of all pain related postures, active pain avoidance, and restlessness), and ewe-lamb distances (Contact, Close and Apart). Distances as well as pain-related behaviours and postures are given as medians with 95% confidence interval. Due to very little variation in the follow scores (80% of the ewes obtained 5 or 4), they were not included in the analyses. Gender of the offspring did not affect the ewe behaviour on baseline day. Similarly, maternal behaviour on day 4 (pain day) was not affected by prior experience of the stressors in some of the pain lambs (from comparison of lambs that experienced stress vs. those that were controls on day 3). These factors were thus not included in our analysis. Both stress treatments on day 3 (LPS and isolation) and both pain treatments on day 4 (tail-docking alone or tail-docking plus castration) were pooled together in the analysis to observe a wider range of stress and pain responses in the lambs and to increase the probability of detecting important associations. 2.5. Ethical considerations All procedures were completed under UK Home Office License number 30/2420. The procedures were carried out to develop and improve knowledge about whether ewes can diminish stress and pain experience in their offspring. We deliberately chose stimuli which simulate the type of events that young lambs might encounter under commercial husbandry conditions. Intravenous injection of LPS is relevant to mimic an E. coli infection, which is responsible for e.g. watery mouth disease in lambs under normal husbandry conditions. The dose used was low and the effects were short-lived. Tail docking and castration without use of anaesthetics are normal husbandry practice in the UK and several other countries despite numerous studies providing evidence that these procedures cause severe acute pain in the lambs (Mellor and Murray, 1989; Molony and Kent, 1997; Molony et al., 2002).
3. Results 3.1. Comparisons of behaviour between and within days 3.1.1. Comparison between baseline day and treatment days Stress lambs sought the teat significantly more on stress day than on baseline day (one-sample Wilcoxon signedrank test: P = 0.016), although this tended to be true for control lambs as well (P = 0.073; Fig. 1). No significant differences were detected between baseline and stress day for ewe exploring or ewe glancing (Fig. 1). On the pain day, the ewe explored significantly more often (P = 0.003) and glanced more often at pain lambs (P < 0.001; Fig. 2) than on baseline day. Control lambs showed teat seeking behaviour for a longer duration on pain day than on baseline day (P = 0.029; Fig. 2) but this was not observed in the pain lambs. 3.1.2. Comparison between treated lambs and control lambs on the same day No significant difference was detected between control lambs and stress lambs for any of the observed behaviours on the stress day (Fig. 1). On the pain day, ewes explored (2-sample Wilcoxon test: P = 0.025), and glanced at pain lambs (P < 0.001; Fig. 2) significantly more often than control lambs. No significant difference was detected between pain and control lambs for teat seeking behaviour (Fig. 2). 3.2. Correlations between maternal behaviours, distances and pain responses Lambs showed active pain avoidance behaviours 82 [68.9–132.6] times and were observed in a pain-related posture for 789 [620.2–952.0] s during the 20 min of observation. Measures of restlessness were obtained by counting the 16 [12.5–20.0] times that the lamb stood up and lay down. Active pain avoidance behaviours in the lambs were significantly positively correlated with the duration of time that the ewe spent exploring the lamb and the number
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Fig. 1. Maternal behaviours towards control lambs (C) and stress lambs (S) on the baseline day and on the stress day. Box plots express median, 25–75% quartiles, minimum, maximum, and outliers, respectively. Probabilities: comparison between days using a one-way Wilcoxon test and comparison within days using a two-sample Wilcoxon test: t: P < 0.1; *: P < 0.05.
of glances directed at the lamb (Table 3). The glancing behaviour also tended to be positively correlated with the duration of time spent in a pain-related posture by the lamb and its restlessness (number of lying down and standing up, Table 3). No other measures, including distances, were found to be significantly correlated with the pain behaviours of the lamb. Overall, the ewe and her lamb in pain were observed in Contact: 14.3 [4.8–28.6]% of total observations, Close: 38.1 [26.0–50.2]% of total observations and Apart: 38.1 [25.9–59.7]% of total observations.
4. Discussion In this study, we observed the ewe maternal behaviour towards her twin lambs when one of them underwent an immune or psychological stress and/or a painful experience. Ewes expressed similar attention towards their lamb experiencing immune or psychological stress as towards the control lambs but they showed more attention to the lambs undergoing the pain treatment than to the control sibling. Ewes nursed both lambs in a similar manner, as
Fig. 2. Maternal behaviours towards control lambs (C) and pain lambs (P) on baseline day and on pain day. Box plots express median, 25–75% quartiles, minimum, maximum, and outliers, respectively. Probabilities: comparison between days using a one-way Wilcoxon test and comparison within days using a two-sample Wilcoxon test: *: P < 0.05. **: P < 0.01. ***: P < 0.001.
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Table 3 Spearman correlations between maternal behaviours and pain related behaviours of lambs experiencing tail docking or tail docking and castration on day 4 (r: Spearman rho, P: probability level).
Restlessness Active pain avoidance Pain related postures
Exploring the lamb in pain
Glancing at the lamb in pain
Teat seeking by lamb in pain
r
P
r
P
r
P
0.0 0.6 0.3
ns 0.008* ns
0.4 0.5 0.4
0.083 0.018* 0.066
0.1 −0.1 −0.3
ns ns ns
ns: P > 0.1. * P < 0.05.
measured by teat seeking by the lambs, on both treatment days and not differently than on baseline day, which is likely to be related to the rare occurrence of the teat seeking behaviour. Interestingly, the amount of attention given by the ewe to a lamb was positively correlated to the amount of pain-related behaviour expressed by the tail-docked and castrated lambs. These results validated the first hypothesis: based on their attention behaviour, the ewes were able to discriminate between a lamb experiencing pain and its control twin, or the same lamb on the baseline day. The positive correlation between maternal behaviours and pain-related behaviours in the lamb suggests that more attention from the ewe was drawn by more pain-related behaviours. Our hypothesis that more care from the dam would result in less expression of pain in the young during the observed period is thus not supported. The proximate mechanism, or immediate factor responsible for the observed results, could be curiosity, drawing the ewe’s attention to signals of severe pain in the offspring. Sheep possess acute eyesight which enables efficient vigilance behaviour among other abilities (Hutson, 2000) and likely predisposes them to detecting subtle behavioural changes in conspecifics. Due to technical difficulty to differentiate vocalisations expressed by the studied individuals from those produced in the rest of the flock, vocalisations were not included here. To understand better which features of the young may have raised the dam’s attention, it would be useful to record the vocal activity of both lambs and ewes. Indeed, it was shown in ewes that stress can be conveyed through variations in the bleating frequency (Sèbe et al., 2009). In addition, chemical signals may increase arousal in individuals, e.g. in rats (Inagaki et al., 2008) and should also be investigated. Processes such as emotional contagion, which designates the emotional convergence of an individual towards the emotional state of another individual, may have been involved (Preston and de Waal, 2002). Such processes have been demonstrated in nonhuman primates (de Waal, 2008) and mice (Langford et al., 2006) when the individuals were familiar to each other. It may have clear adaptive value in this context (cf. de Waal, 2008). The stressed lambs did not elicit a detectable response in the ewe. This may be due to a more discrete expression of stress behaviours by the lambs that could have failed to attract their mother’s attention. We cannot ignore the possibility that the sensitivity of the behavioural measuring systems may not have been high enough and that ewes may have expressed more subtle behaviours, such as differences in ear postures, which could indicate an increased negative emotional valence (Reefmann
et al., 2009) and which were not recorded in the current study. Behaviours likely to relieve pain in the young included nursing and licking, as observed in mice where dams showing more licking and grooming of their pups decreased their pups’ sensitivity to thermal pain (Walker et al., 2008), or physical proximity, as we reported in a previous study (Hild et al., 2010) that lambs that were less sensitive to thermal pain were observed to be closer to their dam. These behaviours were not expressed differently according to the treatment of the offspring in the current study. In our protocol, behaviours were only observed up to 30 min after application of stress or pain stimuli. It should be tested whether, when given enough time, the correlation between maternal behaviours and the lamb pain related behaviours would be reversed. As the frequency of active pain behaviours begins to decrease 30–60 min after castration and tail-docking (Graham et al., 1997), the collection of physiological data such as cortisol (Graham et al., 1997), heart rate variability (Stubsjøen et al., 2009) or an index of fatigue (e.g. plasma glucose or temperature; Molony and Kent, 1997) could be implemented. Alternatively, it may not make sense for the ewe to show these potentially pain mitigating behaviours in the artificial testing conditions in which we conducted our observations. It is possible that in a more “ecological” setting, e.g. allowing the ewe to move further away from the lamb or including predatory threats, we may have observed a different range of maternal responses to an offspring in pain. In conclusion, this study represents a preliminary investigation of the impact of adverse events experienced by lambs on the dam’s response. Ewes showed more attention to their offspring experiencing pain but not stress. Whether the ewes could recognise that their lamb was in pain is not resolved and emotional contagion should be investigated in the future. No alleviation of pain corresponding to increased maternal attention and care was detected within the observation period. To complete testing this hypothesis, more sensitive methods of assessment and longer observation periods should be used. Acknowledgements The authors wish to thank Jennifer Jamieson and Ashley Arnone for experimental help, Stuart Pope for technical support, the veterinarians from Langford clinic, the project funder BBSRC, the Research Council of Norway (Forskningsrådet – project 185163), the Norwegian Committee for Research and Ethics (Utvalg for Forskning og Etikk) for
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financial support to Sophie Hild and Dr Inger Lise Andersen for commenting on the manuscript. References Blass, E.M., Hoffmeyer, L.B., 1991. Sucrose as an analgesic for newborninfants. Pediatrics 87, 215–218. Blass, E.M., Shide, D.J., Zawmon, C., Sorrentino, J., 1995. Mother as shield – differential-effects of contact and nursing on pain responsivity in infant rats – evidence for nonopioid mediation. Behav. Neurosci. 109, 342–353. Boissy, A., Fisher, A.D., Bouix, J., Hinch, G.N., Le Neindre, P., 2005. Genetics of fear in ruminant livestock. Livest. Prod. Sci. 93, 23–32. Clark, C., Mendl, M., Jamieson, J., Arnone, A., Waterman-Pearson, A., Murrell, J., 2011. Do psychological and physiological stressors alter the acute pain response to castration and tail docking in lambs? Vet. Anaesth. Analg. 38, 134–145, doi:10.1111/j.1467-2995.2011.00594.x. de Waal, F.B.M., 2008. Putting the altruism back into altruism: the evolution of empathy. Annu. Rev. Psychol. 59, 279–300. Dwyer, C.M., 2008. Genetic and physiological determinants of maternal behavior and lamb survival: implications for low-input sheep management. J. Anim. Sci. 86, 246–258. Elenkov, I.J., Iezzoni, D.G., Daly, A., Harris, A.G., Chrousos, G.P., 2005. Cytokine dysregulation, inflammation and well-being. Neuroimmunomodulation 12, 255–269. Elmquist, J.K., Scammell, T.E., Saper, C.B., 1997. Mechanisms of CNS response to systemic immune challenge: the febrile response. Trends Neurosci. 20, 565–570. Field, R.A., 1971. Effect of castration on meat quality and quantity. J. Anim. Sci. 32, 849–858. French, N.P., Wall, R., Morgan, K.L., 1994. Tail docking of lambs in the control of flystrike. Vet. Rec. 135, 47. Graham, M.J., Kent, J.E., Molony, V., 1997. Effects of four analgesic treatments on the behavioural and cortisol responses of 3-week-old lambs to tail docking. Vet. J. 153, 87–97. Gray, L., Watt, L., Blass, E.M., 2000. Skin-to-skin contact is analgesic in healthy newborns. Pediatrics, 105. Harden, L.M., du Plessis, I., Poole, S., Laburn, H.P., 2006. Interleukin-6 and leptin mediate lipopolysaccharide-induced fever and sickness behavior. Physiol. Behav. 89, 146–155. Hild, S., Andersen, I.L., Zanella, A.J., 2010. The relationship between thermal nociceptive threshold in lambs and ewe-lamb interactions. Small Ruminant Res. 90, 142–145. Hutson, G.D., 2000. Behavioural principles of sheep handling. In: Grandin, T. (Ed.), Livestock Handling and Transport. CAB International North America, p. 175. Inagaki, H., Kiyokawa, Y., Kikusui, T., Takeuchi, Y., Mori, Y., 2008. Enhancement of the acoustic startle reflex by an alarm pheromone in male rats. Physiol. Behav. 93, 606–611. Jewell, P.A., 1997. Survival and behaviour of castrated Soay sheep (Ovis aries) in a feral island population on Hirta, St. Kilda, Scotland. J. Zool. 243, 623–636.
Johnston, C.C., Stevens, B., Pinelli, J., Gibbins, S., Filion, F., Jack, A., Steele, S., Boyer, K., Veilleux, A., 2003. Kangaroo care is effective in diminishing pain response in preterm neonates. Arch. Pediatr. Adolesc. Med. 157, 1084–1088. Landa, L., 2003. The effect of milk suckling from the dam or glucose administration on the behavioural responses to tail docking in lambs. Acta Vet. Brno 72, 175–182. Langford, D.J., Crager, S.E., Shehzad, Z., Smith, S.B., Sotocinal, S.G., Levenstadt, J.S., Chanda, M.L., Levitin, D.J., Mogil, J.S., 2006. Social modulation of pain as evidence for empathy in mice. Science 312, 1967–1970. Mellor, D., Murray, L., 1989. Effects of tail docking and castration on behaviour and plasma cortisol concentration in young lambs. Res. Vet. Sci. 46, 387–391. Molony, V., Kent, J., 1997. Assessment of acute pain in farm animals using behavioral and physiological measurements. J. Anim. Sci. 75, 266–272. Molony, V., Kent, J.E., McKendrick, I.J., 2002. Validation of a method for assessment of an acute pain in lambs. Appl. Anim. Behav. Sci. 76, 215–238. Preston, S.D., de Waal, F.B.M., 2002. Empathy: its ultimate and proximate bases. Behav. Brain Sci. 25, 1–20. Reefmann, N., Kaszas, F.B., Wechsler, B., Gygax, L., 2009. Ear and tail postures as indicators of emotional valence in sheep. Appl. Anim. Behav. Sci. 118, 199–207. Ren, K., Blass, E.M., Zhou, Q.Q., Dubner, R., 1997. Suckling and sucrose ingestion suppress persistent hyperalgesia and spinal Fos expression after forepaw inflammation in infant rats. Proc. Natl. Acad. Sci. U.S.A. 94, 1471–1475. Roger, P.A., 2008. The impact of disease and disease prevention on sheep welfare. Small Ruminant Res. 76, 104–111. Sèbe, F., Poindron, P., Reby, D., Nowak, R., Blache, D., 2009. Encoding and genetic selection of psycho-acoustic markers of emotions in sheep. In: Proceedings of the 43rd Congress of the International Society for Applied Ethology , p. 133. Seideman, S.C., Cross, H.R., Oltjen, R.R., Schanbacher, B.D., 1982. Utilization of the intact male for red meat production—a review. J. Anim. Sci. 55, 826–840. Stubsjøen, S.M., Flø, A.S., Moe, R.O., Janczak, A.M., Skjerve, E., Valle, P.S., Zanella, A.J., 2009. Exploring non-invasive methods to assess pain in sheep. Physiol. Behav. 98, 640–648. Walker, C.D., Kudreikis, K., Sherrard, A., Johnston, C.C., 2003. Repeated neonatal pain influences maternal behavior, but not stress responsiveness in rat offspring. Dev. Brain Res. 140, 253–261. Walker, C.D., Xu, Z.F., Rochford, J., Johnston, C.C., 2008. Naturally occuring variations in maternal care modulate the effects of repeated neonatal pain on behavioral sensitivity to thermal pain in the adult offspring. Pain 140, 167–176. Wright, K.J., Balaji, R., Hill, C.M., Dritz, S.S., Knoppel, E.L., Minton, J.E., 2000. Integrated adrenal, somatotropic, and immune responses of growing pigs to treatment with lipopolysaccharide. J. Anim. Sci. 78, 1892–1899.