Effect of rapamycin treatment during post‐activation and/or in vitro culture on embryonic development after parthenogenesis and in vitro fertilization in pigs

This study investigated the effects of early induction of autophagy on embryonic development in pigs. For this, oocytes or embryos were treated with an autophagy inducer, rapamycin (RP), during post‐activation (Pa), in vitro fertilization (IVF) and/or in vitro culture (IVC). When parthenogenesis (PA) embryos were untreated (control) or treated with various concentrations of RP for 4 hr during Pa, 100 nm RP showed a higher blastocyst formation (48.8 ± 2.7%) than the control (34.6 ± 3.0%). When PA embryos were treated during the first 24 hr of IVC, blastocyst formation was increased (p < .05) by 1 and 10 nm RP (61.9 ± 3.0 and 59.6 ± 3.0%, respectively) compared to the control (43.2 ± 1.8%) and 100 nm RP (47.8 ± 3.2%), with a higher embryo cleavage in response to 10 nm RP (87.3 ± 2.4%) than the control (74.1 ± 3.2%). RP treatment during IVC and Pa + IVC showed increased blastocyst formation (44.7 ± 2.5 and 44.1 ± 2.0%, respectively) compared to the control (33.2 ± 2.0%). In addition, RP treatment during Pa and/or IVC increased glutathione content and inversely reduced reactive oxygen species. In IVF, RP treatment for 6 hr during IVF significantly increased embryonic development (34.0 ± 2.6%) compared to the control (24.8 ± 1.6%), but treatment during IVC for 24 hr with RP did not (23.0 ± 3.8%). Autophagy was significantly increased in PA oocytes by the RP treatment during Pa but not altered by the treatment during the first 24 hr of IVC. Overall, RP treatment positively regulated the pre‐implantation development of pig embryos, probably by regulating cellular redox state and stimulating autophagy.     

Simulated small intestinal volvulus in the anesthetized horse

Experimental closed loop small intestinal volvulus was studied in the anesthetized horse. Volvulus was simulated by ligation of the mesenterial veins to a segment of small intestine. Physical signs and hemodynamic, hematologic, clinical chemical, bacteriologic and peritoneal fluid values were examined. Compared to conscious horses anesthesia highly delayed and modified the clinical signs of shock (changes in mucosal colour, dehydration, decreased skin temperature, elevated pulse rate, low blood pressures) and of small intestinal volvulus (altered peristalsis, gastric dilation). Plasma glucose response to shock was also modified by unconsciousness. However, a dose response relationship was indicated between the extent of small intestinal damage and clinical symptoms. The same was applicable to changes in blood pressures, blood acid-base balance, lactate, potassium, chloride, glucose, inorganic phosphorus, creatinine, creatine kinase, red blood cell and total white blood cell counts and serum total protein. The relationship was also indicated in the following peritoneal fluid values: volume, lactate, pH, total white cell counts, alkaline phosphatase and bacteriology. Changes related to shock (insufficient tissue perfusion) were low blood pressures and metabolic acidosis due to anaerobic glycolysis with accumulation of lactic acid. Also low plasma glucose and elevated plasma potassium, creatinine, inorganic phosphorus and creatine kinase were regarded as consequences of shock.     

Pharmacokinetics of amoxicillin after oral administration in recently weaned piglets with experimentally induced Escherichia coli subtype O149:F4 diarrhea

OBJECTIVE: To measure the effect of Escherichia coli subtype O149:F4-induced diarrhea on the pharmacokinetics of orally administered amoxicillin in affected piglets relative to that of uninfected piglets. ANIMALS: 22 healthy 4-week-old recently weaned Danish crossbred piglets. PROCEDURE: 12 piglets were orally inoculated through gastric intubation with 10(9) CFUs of an E. coli O149:F4 strain and responded by developing diarrhea 12 to 16 hours later. Piglets were dosed with amoxicillin trihydrate solution (20 mg/kg) by gastric intubation. A control group of 10 age-matched piglets without signs of diarrhea was dosed similarly. Blood samples were obtained before amoxicillin administration and at 0.5, 1, 1.5, 2, 3, 6, 12, and 24 hours after amoxicillin administration. The plasma concentration of amoxicillin was analyzed by high-performance liquid chromatography. RESULTS: A significant 39% decrease in the area under the plasma concentration versus time curve of amoxicillin was observed in piglets with diarrhea relative to that of control piglets. The maximum plasma concentration (Cmax) was significantly (52%) lower in piglets with diarrhea, compared with control piglets, while the elimination rate constant, time to reach Cmax, and elimination half-life were unchanged. CONCLUSIONS AND CLINICAL RELEVANCE: Escherichia coli-induced diarrhea may decrease systemic bioavailability of amoxicillin. Escherichia coli bacteria attach to the intestinal epithelial cells. Because it is assumed that the concentration of the antimicrobial at the site of infection reflects the systemic concentration, higher doses of amoxicillin in the treatment of piglets with E. coli O149:F4-induced diarrhea may be appropriate.     

The effects of oral and intramuscular administration and dose escalation of enrofloxacin on the selection of quinolone resistance among Salmonella and coliforms in pigs

The effect of route of administration and dose of enrofloxacin (Baytril) on the development of fluoroquinolone resistance in Salmonella and Escherichia coli in the intestinal tract of pigs was investigated. Healthy pigs at the age of 8-10 weeks were infected with a mixture of susceptible wild-type (MICciprofloxacin = 0.03 microg/ml) and a mutant Salmonella typhimurium with reduced susceptibility to fluoroquinolones (MICciprofloxacin = 0.5 microg/ml) (in the ratio 99:1) and treated with 2.5 mg/kg bwt enrofloxacin by either intramuscular (i.m.) or oral (p.o.) administration at time points either 4 or 24 h after the infection. The treatment via the intramuscular route of administration (24 h after the infection) was carried out with elevated doses of 7.5 and 15 mg/kg bwt as well. Emergence of resistance during a 3-day treatment period and persistence up to 13 days after treatment, was monitored by counting the resistant and total number of coliforms and Salmonella in faeces of the pigs. High frequencies of fluoroquinolone resistance developed rapidly among the coliform flora independent of route of administration, dose or time of initiation of the treatment. Selection for resistance among the artificially introduced Salmonella was reduced by using the intramuscular route and by escalating the dose 3 or 6 times the recommended dose of 2.5 mg/kg bwt, which also resulted in shortening of the period, in which the pigs were shedding Salmonella. The resistance among the coliform flora persisted for at least 2 weeks. The Salmonella infection was cleared in all cases during the 2 weeks independent of frequency of resistance. The study showed that resistance is very easily selected by treatment with enrofloxacin at the recommended dose 2.5 mg/kg bwt, but also that the intensity of selection can be reduced by using intramuscular dosing (instead of oral dosing) and by escalating that i.m. dose. The results obtained with Salmonella also showed that even very small changes in the active drug concentrations might completely change the intensity of selection.     

Local anaesthetics attenuates spinal nociception and HPA-axis activation during experimental laparotomy in pigs

The effect of local anaesthetics on spinal nociception and activation of the hypothalamic-pituitary-adrenal axis (HPA-axis) was examined in a porcine model of abdominal surgery. A standardised laparotomy without visceral involvement was performed on 24 pigs. One group received a unilateral infiltration of mixed lidocaine and bupivacaine in skin, muscle and peritoneum of the surgical area prior to surgery (n=12), while local anaesthetics were replaced by isotonic saline in a second group (n=12). A sham group was subjected to anaesthesia (n=8), but did not undergo surgery. Two hours after surgery, half of the pigs from each group were perfused with formalin and the spinal cord was taken out for stereological quantification of the total number of Fos-like-immunoreactive (Fos-LI) neurones in the dorsal horn. Surgery with saline gave rise to a significant increase in the number of Fos-LI neurones ipsilaterally (107,001+/-16,548; p<0.001) as well as contralaterally (12,766+/-3,842; p<0.01) compared to the sham group. In animals undergoing surgery with LA, the number of Fos-LI neurones ipsilaterally was not significantly different from the sham group (p=0.78), and was reduced significantly both ipsilaterally (6960+/-1662; p<0.001) and contralaterally (3974+/-1131; p<0.05) compared to the saline group. In the other half of each group, blood samples, for determination of ACTH, cortisol, C-reactive protein and interleukin-6 concentrations, were drawn prior to and at predetermined time-points during and after surgery. Surgery with saline gave rise to dramatic increases in plasma ACTH and cortisol (p<0.01 and p<0.001, respectively) within 15 min of incision. In contrast, no changes from the initial concentrations of ACTH and cortisol were observed in pigs receiving local anaesthetics. No changes in plasma concentrations of C-reactive protein or interleukin-6 were observed in either of the groups. These results indicate that spinal nociception and HPA-axis activation caused by laparotomy in pigs can be attenuated by use of infiltration and incisional local anaesthetics prior to surgery. The present model provides a valuable tool in the evaluation of analgesic treatment during surgery, offering objective measures of both nociception and stress.     

Immunity to Escherichia coli in pigs: Serum Gamma Globulin Levels, Indirect Hemagglutinating Antibody Titres and Bactericidal Activity Against E. coli in pigs up to five Weeks of Age

Serum gamma globulin levels, indirect hemagglutinating antibody titres and bactericidal activity against the 0149:K91;K88ac:H10 Serotype of Escherichia coli were determined in pigs up to five weeks of age from vaccinated and non-vaccinated sows.

Gamma globulin levels at two days of age were approximately twice adult levels, by three weeks of age they were one quarter of adult levels and remained so until five weeks of age. Indirect hemagglutinating antibody activity was highest at two days of age, fell until three weeks of age and then rose. Little or no indirect hemagglutinating antibody activity was detected in sera taken at two days of age from pigs from non-vaccinated sows. Only three of 26 two day old pigs had demonstrable bactericidal activity; by three weeks of age 16 of 26 had bactericidal activity. Serum from piglets of vaccinated sows had no more bactericidal activity than did sera from non-vaccinated sows.

     

Immunity to Escherichia coli in Pigs: Anti-enterotoxins in Colostrum and Serum from Vaccinated and Non-vaccinated Sows

Colostrum from non-vaccinated sows did not contain naturally occurring antibodies to heat labile Escherichia coli enterotoxin. Vaccination of sows by either the intramuscular or intramammary routes with a live formalinized E. coli vaccine resulted in the production of colostrum capable of neutralizing the heat labile toxin. Intramammary vaccination resulted in the production of colostrum which significantly reduced the enterotoxigenic effects of the vaccine strain of E. coli organisms but not that of a heterologous strain.

Vaccination of the sows resulted in the production of serum antibodies to heat labile enterotoxin. Antibodies to heat stable enterotoxin were not demonstrable in the colostrum of either non-vaccinated or vaccinated sows.

     

Pharmacokinetics of hexobarbital, sulphadimidine and chloramphencoliin neonatal and young pigs.

Half-life and apparent specific volume of distribution of hexobarbital, sulphadimidine and chloramphenicol were investigated in newborn, 1, 3, 5 and 8 weeks old pigs. Hexobarbital sleeping time and plasma concentration of hexobarbital at recovery were measured in the same age groups. The half-life of hexobarbital and chloramphenicol was long in newborn pigs but decreased fast during the first week after birth. From 1 to 8 weeks after birth the decrease was less pronounced. The half-life of sulphadimidine increased during the first 3 weeks of life, but in 1 and 3 weeks old pigs the amount of N⁴-acetylated sulphadimidine in plasma at 200 min. after the injection was higher than in the newborn pigs.The apparent specific volume of distribution of hexobarbital, sulphadimidine and chloramphenicol was changed in different ways from birth to 8 weeks of age.The hexobarbital sleeping time was very long in the newborn pigs and decreased until 3 weeks of age. The concentration of hexobarbital in plasma at recovery was unchanged from birth to 8 weeks of age.The concentration of chloramphenicol metabolites in plasma 100 min. after the injection increased very fast during the 8 weeks of observation. The concentration of N⁴-acetylated sulphadimidine in plasma at 200 min. after the injection increased from birth to 1 week of age, then it decreased.The data are stressing that the neonatal pig is a convenient model for pharmacokinetic testing of drugs used as pharmacotherapeutics in neonatal life.