Gas alternatives to carbon dioxide for euthanasia: a piglet perspective.

The search for alternative methods to euthanize piglets is critical to address public concern that current methods are not optimal. Scientific evidence supports that blunt force trauma is humane when performed correctly, but most people find it visually difficult to accept. The use of CO2 is often recommended; at the same time, it is criticized as being aversive to pigs. This research sought to 1) identify a method of scientifically determining if piglets find a gas aversive, using an approach-avoidance test, which relies on the perspective of the piglet, and 2) test different gas mixtures to determine if they are effective and humane for neonatal piglet euthanasia. Pigs were allowed to walk freely between 1 chamber filled with air and another chamber either gradually filled with gas mixtures (Exp. 1) or prefilled with gas mixtures (Exp. 2). Experiment 1 tested CO2 (90%) and air (10%), N2O (60%) and CO2 (30%), Ar (60%) and CO2 (30%), and N2 (60%) and CO2 (30%). Because piglets had to be removed when they started to flail, the test was shortest (P < 0.01) for the pigs in the CO2 treatment compared with pigs in the N2O/CO2, Ar/CO2, and N2/CO2 treatments, 3.1 ± 0.2, 8.5 ± 0.6, 9.6 ± 0.4, and 9.9 ± 0.1 min, respectively. Nonetheless, all gas mixtures adversely affected the pigs, causing the pigs to leave the test chamber. In Exp. 2, piglets were allowed to enter a chamber prefilled with N2/CO2 or N2O/CO2 (both 60/30%). Pigs exposed to the prefill chambers started to flail in fewer than 20 s, much faster in comparison with the gradual fill method, which supports that this method was more aversive. In Exp. 3, piglets were euthanized using a 2-step procedure. Pigs were first placed in a gradual fill chamber with 1 of 4 gas mixtures: 90% CO2, N2/CO2, N2O/CO2, or N2O/O2 (the last 3 mixtures at 60/30%) followed by placement into a 90% CO2 prefill chamber when the pigs started to flail or were anesthetized. All 3 gas treatments that contained CO2 killed pigs more quickly than N2O/O2 (P < 0.05). However, N2O/O2 was the only treatment that anesthetized the pigs instead of causing squeals or flailing although requiring about 12 min longer. Although longer, a 2-step procedure in which pigs are anesthetized with a mixture of N2O and O2 before being euthanized by immersion in CO2 may prove to be more humane than CO2 alone.

Effect of prenatal stress on subsequent response to mixing stress and a lipopolysaccharide challenge in pigs.

Sows subjected to prenatal stress have been found to produce offspring that have altered responses to stress. Our objective was to determine if exposing a sow to stress would alter the response of the offspring to lipopolysaccharide (LPS) at 2 mo of age or their response to mixing stress at 4 mo of age. Sow treatments consisted of intravenous injections of ACTH (1 IU/kg of BW), exposure to rough handling for a 10-min duration (rough), or no treatment (control) once per week from d 42 to 77 of gestation. At 2 mo of age, pigs from each treatment, 1 per litter (n = 21, 17, and 15 for the ACTH, rough, and control treatments, respectively), were challenged with 2 µg of LPS/kg of BW or saline, or served as a noninjected control. Their behavioral response to a human approach test and salivary cortisol were measured. At 4 mo of age, 1 pig from each treatment (n = 14, 14, and 15 for the ACTH, rough, and control treatments, respectively) was taken from its home pen and placed in a pen of unfamiliar pigs. At this time, a punch biopsy wound (6 × 6 mm) was created to measure the ability of the pig to heal the wound. At this same time, each pig received a 1-mL intramuscular injection of 20% ovine red blood cells (oRBC), and then a second injection of oRBC at 21 d postmixing. Blood samples were collected 3 times per week for 2 wk and then once a week for 4 more weeks. Blood samples were analyzed for cortisol, porcine corticosteroid-binding globulin, antibody response to oRBC, and nitric oxide production by macrophages. Behavior was recorded during the first 5 d after mixing. All pigs in the LPS challenge responded with characteristic sickness behavior; however, pigs in the rough treatment showed less sickness behavior than those in the other 2 treatments (P < 0.05). Maternal stress treatment did not affect (P < 0.43) salivary cortisol. Pigs from all treatments responded similarly to mixing stress with regard to cortisol, porcine corticosteroid-binding globulin, antibody titers, nitric oxide production, and hematology measures, and all pigs experienced the same amount of aggression in response to mixing. Without altering peripheral measures of stress responsivity, prenatal stress enhanced the ability of pigs to cope with a simulated immune challenge, which could prove to be an adaptation to challenging environments.

Toll-like receptors 2 and 4, and acute phase cytokine gene expression in dexamethasone and growth hormone treated dairy calves.

Cattle are exposed to growth hormone stimulants and to stressors that cause cortisol release. Both of these hormones affect immune responses which may reduce disease resistance. Toll-like receptors are the pattern recognition molecules of pathogens that are on immune cells. They then orchestrate the induction of the appropriate acute phase cytokines of the early innate response. The objective of this study was to determine changes in toll-like receptors and acute phase cytokines following treatment with a synthetic glucocorticoid (dexamethasone) and growth hormone (GH). Twenty-eight calves were given the control (Cnt), dexamethasone (DEX), GH, or dexamethasone and GH (Both) treatments from 3 until 56 days of age. Blood was collected by jugular venipuncture on days 14, 28, 42, and 56. On day 56, a lung lavage was performed and spleen and thymus tissues collected. Total RNA was extracted from blood leukocytes, lung lavage cells, spleen and thymus cells. Real-time reverse transcriptase-polymerase chain reaction (RT-PCR) was used to quantify interleukin-1 (IL-1), IL-1 receptor antagonist (IL-1Ra), tumor necrosis factor (TNF)-alpha, toll-like receptor 2 (TLR2), and toll-like receptor 4 (TLR4). Blood leukocytes had a time effect for IL-1Ra (P < 0.01), with a trend for a treatment effect (P = 0.07) and had a treatment by time interaction (P < 0.05). IL-1, TNF, and TLR2 and TLR4 were greatest (P < 0.05) for Cnt only at day 14. IL-1 expression of lung lavage cells was greatest (P < 0.05) for calves on the Both treatment compared to the other three treatments. However, IL-1Ra was not different among the treatments. Toll-like receptor 2 expression was enhanced with Both compared to either DEX (P < 0.05) or GH (P < 0.05) and tended to be greater than Cnt expression (P = 0.07). Expression of TLR4 tended to be reduced by Both compared to Cnt (P = 0.06). Tumor necrosis factor-alpha was greatly enhanced by Both compared to the other three treatments (P < 0.05). Spleen cell tended to have different IL-1 expression between GH and Both (P < 0.10). Interleukin-1 receptor antagonist and TLR2 and TLR4 were not different among treatments. However, TNF-alpha expression was enhanced by the DEX treatment alone compared to the GH treatment (P < 0.05), and tended (P < 0.10) to be greater than Cnt expression. None of the gene expressions were different among treatments for thymus cells. Lung lavage cell expression appears to be most susceptible to these hormones while blood leukocyte expression was only slightly affected, and thymus cells were not affected at all. These data demonstrate that TLR2 and TLR4 and acute phase cytokine expression can be altered by stress and growth hormones, which may decrease resistance of those animals to disease.