Bernd Stabenow

Consequences of repeated early isolation in domestic piglets (Sus scrofa) on their behavioural, neuroendocrine, and immunological responses.

Stress in the form of intermittent maternal deprivation and social isolation during early postnatal life in rats and monkeys produces persistent changes in physiology and behaviour. In farm animals physiological consequences of disrupting mother-infant interactions with respect to health and animal welfare are relatively unknown. Therefore, the aim of the present study was to investigate the behavioural, neuroendocrine and immunological consequences of a 2 h daily social isolation from day 3 to day 11 of age in domestic piglets as well as potential long-term effects on the brain-endocrine-immune regulation. Repeated social isolation resulted in significantly decreased open-field activity (locomotion, vocalization) during the isolation period, increased basal cortisol concentrations and decreased lymphocyte proliferation in response to concanavalin A and pokeweed mitogen one day after the isolation. There was also a significant increase of interleukin-1beta (IL-1beta) concentration in hippocampus in isolated piglets compared to controls at this time. Six weeks after isolation significant enhanced basal ACTH concentrations as well as higher IL-1beta content and glucocorticoid receptor (GR) binding in hippocampus were found. These endocrine and immune responses were associated with decreased CRH levels in the hypothalamus and increased CRH content in the amygdala. The present data indicate that early social isolation in pigs may cause changes in behavioural, neuroendocrine, and immune regulation and produce long-term effects not only on the activity of the hypothalamic-pituitary-adrenal (HPA) system, but also on the immune-brain circuitry with possible negative consequences in health and welfare of commercial pigs. Using the pig as a suitable animal model, the finding of this study may also have some implications for the etiology of anxiety and depression in humans.

Effects of postnatal social isolation on hormonal and immune responses of pigs to an acute endotoxin challenge

Social stress during early postnatal life often results in long-term effects on neuroendocrine and immune adaptation mechanisms. Therefore, the aim of the present study was to determine the influence of a 2-h daily social isolation from Day 3 to Day 11 on the acute and long-term proinflammatory and neuroendocrine responses of piglets challenged with the bacterial endotoxin lipopolysaccharide (LPS; 100 microg/kg body weight). Peripheral LPS administration significantly increased plasma concentrations of tumor necrosis factor-alpha (TNF-alpha), ACTH and cortisol in isolated and control pigs. However, the activity of the hypothalamic-pituitary-adrenal (HPA) axis after LPS stimulation was not significantly affected by isolation treatment, whereas the prior social isolation diminished the plasma TNF-alpha response to LPS 1 day as well as 45 days after the isolation period. The hippocampal TNF-alpha concentration in response to LPS was also reduced in priorly isolated pigs compared to control animals. Furthermore, the significant increase of TNF-alpha in the spleen caused by LPS was associated with a dramatic decrease in glucocorticoid receptor (GR) binding. The GR binding in hippocampus was increased in isolated pigs and was significantly decreased after LPS injection. In addition, the repeated isolation stressor was shown to increase hippocampal levels of interleukin-1beta (IL-1beta). The present results indicate that repeated social isolation of neonatal pigs may cause long-term effects on proinflammatory regulation at the periphery and in the brain following immune challenge with particular importance of TNF-alpha in mediating these interactions.

Limited and excess dietary protein during gestation affects growth and compositional traits in gilts and impairs offspring fetal growth

The aim of this study was to investigate whether dietary protein intake during gestation less than or greater than recommendations affects gilts growth and body composition, gestation outcome, and colostrum composition. German Landrace gilts were fed gestation diets (13.7 MJ of ME/kg) containing a low (n = 18; LP, 6.5% CP), an adequate (n = 20; AP, 12.1%), or a high (n = 16; HP, 30%) protein content corresponding to a protein:carbohydrate ratio of 1:10.4, 1:5, and 1:1.3, respectively, from mating until farrowing. Gilts were inseminated by semen of pure German Landrace boars and induced to farrow at 114 d postcoitum (dpc; Exp. 1). Energy and protein intake during gestation were 33.3, 34.4, and 35.8 MJ of ME/d (P < 0.001) and 160, 328, and 768 g/d, respectively, in LP, AP, and HP gilts (P < 0.001). From insemination to 109 dpc, BW gain was least in LP (42.1 kg), intermediate in HP (63.1 kg), and greatest in AP gilts (68.3 kg), whereas increase of backfat thickness was least in gilts fed the HP diet compared with LP and AP diets (3.8, 5.1, 5.0 mm; P = 0.01). Litter size, % stillborn piglets, and mummies were unaffected (P > 0.28) by the gestation diet. Total litter weight tended to be less in the offspring of LP and HP gilts (14.67, 13.77 vs. 15.96 kg; P = 0.07), and the percentage of male piglets was greater in litters of HP gilts (59.4%; P < 0.01). In piglets originating from LP and HP gilts, individual birth weight was less (1.20, 1.21 vs. 1.40 kg; P = 0.001) and birth weight/crown-rump length ratio was reduced (45.3, 46.4 vs. 50.7 g/cm; P = 0.003). Colostrum fat (7.8, 7.4 vs. 8.1%) and lactose concentrations (2.2, 2.1 vs. 2.6%) tended to be reduced in LP and HP gilts (P = 0.10). In Exp. 2, 28 gilts (LP, 10; AP, 9; HP, 9) were treated as in Exp. 1 but slaughtered at 64 dpc. At 64 dpc, LP gilts were 7% lighter than AP gilts (P = 0.03), whereas HP gilts were similar to AP gilts. Body composition was markedly altered in response to LP and HP feeding with less lean (P < 0.01) and greater fat content (P = 0.02 to 0.04) in LP and less fat content (P = 0.02 to 0.04) in HP gilts. Fetal litter weight and number, and embryonic survival at 64 dpc were not affected by the diets. These results indicated that gestation diets containing protein at 50 and 250% of recommendations and differing in protein:carbohydrate ratio led to marked changes in protein and fat metabolism in gilts resulting in fetal growth retardation of 15%, which mainly occurred during the second half of gestation.

Neuroendocrine and Immune Responses to Acute Endotoxemia in Suckling and Weaned Piglets

The objective of this study was to characterize effects of weaning stress on behavioural, endocrine and immune responses to acute peripheral lipopolysaccharide (LPS) challenge in neonatal pigs. Weaning in 28-day-old piglets was accompanied by a significant increase in ACTH concentrations (p = 0.0378) and an increase in basal cortisol level (p = 0.0135). There was also a significant suppressive effect on lymphocyte proliferation in response to concanavalin A (p = 0.0048) in newly weaned piglets. Peripheral administration of LPS induced vomiting, diarrhoea and somnolence in both suckling and weaned piglets. The frequency of these signs of sickness was significantly higher in weaned piglets compared with suckling piglets (p = 0.0049). Additionally, LPS significantly increased plasma concentrations of TNF-α, cortisol and ACTH. While weaned piglets reacted to LPS with a higher release of ACTH (p = 0.0239) and cortisol (p = 0.0015) than suckling piglets there was no significant effect of weaning on the magnitude of TNF-α. The present data indicate that weaning suppresses the lymphocyte function, causes changes in endocrine regulation and has a substantial effect on the behavioural and endocrine response to an acute peripheral LPS challenge; consequently it could increase disease susceptibility.

Effects of limited and excess protein intakes of pregnant gilts on carcass quality and cellular properties of skeletal muscle and subcutaneous adipose tissue in fattening pigs

The aim of this study was to investigate whether dietary protein intake of gilts during gestation below (50%) or above (250%) recommendations affects body composition, carcass and meat quality, and properties of skeletal muscle and subcutaneous adipose tissue (SCAT) in offspring at d 83 and 188 of age. German Landrace gilts were fed isoenergetic gestation diets (~13.7 MJ of ME/kg) containing a low (LP, 6.5%; n = 18), an adequate (AP, 12.1%; n = 20), or a high (HP, 30%; n = 16) protein content from mating until farrowing. Within 48 h of birth, offspring were cross-fostered to sows fed a standard diet. On d 83 of age, no effects of the LP diet on BW and body composition were detected, whereas HP pigs showed a slight growth delay (P = 0.06) associated with increased relative weights of small intestine (P < 0.01) and brain (P = 0.08), and reduced relative thymus weight (P < 0.01). On d 188 of age, BW was not different among the dietary groups. However, the carcass of LP pigs contained less (P = 0.01) lean and more (P = 0.07) fat compared with AP and HP pigs, which was only pronounced in pigs originating from large litters (P < 0.05). Like skeletal muscles (P = 0.06), the heart muscle weighed less (P = 0.02) in LP than AP pigs. Compared with AP pigs, LP pigs exhibited a fewer (P = 0.09) total number of myofibers in semitendinosus muscle plus LM both at d 83 and 188 of age, whereas total muscular DNA was less (P = 0.02) at d 188 only. The mRNA abundance of IGF2 measured on d 188 was reduced in SCAT (P = 0.03) and LM (P = 0.07) of LP compared with AP pigs. No changes in muscular fiber type frequency, capillary density, or creatine kinase activity, as well as SCAT adipocyte size and number, were observed at either stages of age. Meat quality characteristics remained unchanged at d 83, whereas Warner-Bratzler shear force value in LM was decreased (P = 0.03) in LP compared with AP pigs on d 188 of age. The results suggest that the maternal LP diet impairs prenatal myofiber formation, reduces the potential of postnatal lean growth related to reduced IGF2 mRNA expression and myonuclear accumulation, and consequently changes carcass quality toward reduced lean proportion and improved tenderness at market weight. In contrast, except for a slight transient growth delay, excess dietary protein during gestation seems to have little effect on the fetal programming of postnatal muscle and adipose tissue phenotype of the progeny.

Physiological and behavioral effects of different success during social confrontation in pigs with prior dominance experience

The impact of a 10-h social confrontation on behavior, plasma adrenaline, noradrenaline and cortisol was studied in 14 domestic pigs (eight castrated males and six females) with prior dominance experience. Prior to the experiments, animal groups, each consisting of nine growing pigs (12 weeks old) from different litters, were composed randomly. After 5 days, the pig with the highest rank from each group was removed, provided with a jugular vein catheter and kept in single housing for 2-3 weeks. After this period, each test animal was returned into its familiar group for a 10-h social confrontation. The reintroduction of the test animals into the familiar groups caused frequent agonistic interactions during the first 30 min. Seven animals succeeded to win most of their encounters during the confrontation test and were classified as High Success (HS) animals, whereas seven other animals lost most of their encounters and were classified as Low Success (LS) animals. The reintroduction of the test animals into the groups provoked also marked changes in behavioral and physiological measures. The frequent fighting behavior during the first 30 min was accompanied by a rapid increase of plasma catecholamines and a delayed increase of cortisol. Immediately after introduction, LS pigs tended to show higher plasma adrenaline and noradrenaline concentrations than HS pigs. There was also a tendency for a sustained increase of noradrenaline/adrenaline ratios in HS pigs, whereas the ratios of LS pigs remained nearly unchanged. No significant differences between both groups were found in cortisol levels and in the frequency of agonistic interactions. However, LS animals showed less locomotion, more lying and spent less time exploring the pen or other animals. These results show that during a social confrontation the experience of defeats for formerly high-ranking pigs is accompanied by increased submissive or passive behavior and a higher sympathoneural and adrenomedullary reaction, which may indicate more emotional distress and fear compared to successful animals.

Temporal relationship of hippocampal liquor- and plasma-cortisol-concentrations in the domestic pig challenged with synthetical adreno-corticotrophic hormone

The concentration of plasma and intracerebral cortisol in pigs challenged with 0.5 IU adrenocorticotrophic hormone/kg body weight was simultaneously measured at a 30 min rate. The pigs (n=5) were provided with indwelling jugular vein catheters and guide canullas for insertion of a hippocampal microdialysis probe. Every five blood samples taken at 6 min intervals were pooled to simulate the 30 min collection time of the microdialysis. Stimulation resulted in an increment of cortisol of roughly 200% in plasma and 190% in the dialysate compared to basal value. The average time lag of hippocampal to plasma peak was 48.75 min. Considering the temporal resolution of the protocol it is inferred that delays of at least 30 min occur between variations of peripheral and central nervous cortisol.

Interference of stress with the somatotropic axis in pigs – lights on new biomarkers

The acceptance of animal products is increasingly associated with standardized animal welfare, which relates to appropriate animal husbandry from birth to slaughter. In particular, shipment to the slaughterhouse is considered as a critical process exposing the animals to a number of, in part severe, stressors. New biomarkers may be useful for the assessment of animal welfare. The IGF-system has been assessed in a commercial pig transport in conjunction with established markers of stress response. Furthermore, the effect of repeated restraint as an experimental model for repeated acute stress was investigated. During shipment from farm to slaughterhouse, plasma concentrations of IGFBP-3 and IGFBP-2 were significantly reduced (p < 0.01). After shipment, the plasma concentrations of IGFBP-5, glucocorticoids and IL-2 increased but decreased after lairage (p < 0.05) whereas IGF-1 decreased after shipment (p < 0.01). Repeated acute stress increased concentrations of IGFBP-3 and IGF-1 in exsanguination blood (p < 0.05). Differential IGF- signatures can indicate altered endocrine or metabolic control and thus contain complex animal-related information. The somatotropic axis may be of particular interest when established biomarkers such as cortisol, glucose, or lactate cannot be used for the assessment of animal stress or welfare.