The present study examined the influence of post-cleavage time of nuclear donors on the development of reconstituted embryos in bovine nuclear transfer. Blastomeres of 16-cell stage embryos derived from in vitro-maturation, fertilization and culture were used as nuclear donor source. They were treated with 10 mu-M nocodazole for 12 hr. Blastomeres that cleaved within 3 hr after the removal of nocodazole were used-for the study. Metaphase II (M-II) oocytes were used as recipient cytoplasm. IN experiment 1, donor blastomeres at 6, 11 and 15 hr after the removal of nocodazole and donor blastomeres no treated with nocodazole were transferred into ethanol-exposed and enucleated oocytes. The reconstituted embryos produced by donor blastomeres oat 6 hr after the removal of nocodazole had a significantly higher developmental rate to the blastocyst stage than those at 15 hr and the untreated groups (P<0.01). In experiment 2, blastomeres at 6 hr after the removal of nocodazole used as nuclear donors were transferred into ethanol-exposed and enucleated M-II oocytes. The reconstituted embryos with ethanol-exposed and enucleated oocytes as recipient cytoplasm had a significantly higher rate of initial-cleavage (P<0.05) and development to the blastocyst stage (P<0.01) than non ethanol-exposed and enucleated M-II oocytes. These results demonstrate that the development of reconstituted embryos was improved when cleaved donor blastomeres after the removal of nocodazole were immediately transferred (at 3-6 hr post-cleavage) into activated enucleated oocytes by exposure to ethanol.

peer reviewed | Owing to technical and ethical limitations, a substantial part of the knowledge about the pathophysiologic mechanism of the human diaphragm has been obtained from studies in which phrenic nerve activation was usually carried out by direct surgical exposure of the nerves in the neck of deeply anesthetized, mechanically ventilated animals. Novel information has been gleaned from such studies, but the restrictive conditions under which it was collected preclude reliable extrapolation. We, therefore, addressed the question of whether accurate electrophysiologic evaluation of the phrenic nerve-diaphragm pathway can be performed in intact, nonanesthetized calves. Transjugular phrenic activation was well tolerated, safe, specific, and able to achieve constant symmetric and supramaximal phrenic stimulations during prolonged periods. Eighteen noninvasive cutaneous and esophageal reception circuits were tested for their ability to record the diaphragmatic evoked potential. In addition, they were compared for specificity and reproducibility of the recorded potentials during prolonged periods of tidal or stimulated respiration. The best diaphragmatic potential was recorded from surface electrodes attached to the skin of the ninth and tenth intercostal spaces, using a xyphoidian reference. We describe a method that allows easy, longterm, and reliable electrophysiologic evaluation of the phrenic nerve-diaphragm pathway in intact, conscious calves. It is hoped that such a model will produce relevant novel information regarding pathophysiology of the diaphragm.

A virologic survey was conducted on calves with diarrhea associated with bovine rotavirus (BRV) on a closed dairy farm. The BRV was detected from 32 of 219 (14.6%) fecal specimens repeatedly collected from 56 calves born during the years 1992-1993, regardless of whether they had diarrhea. Most of the 32 strains were isolated from fecal specimens obtained from 2-to 6-week-old calves. After electrophoresis of doublestranded viral RNA from the 32 strains, genomic RNA migration patterns were similar to those of the predominant BRV strains isolated at the same farm during the years 1990-1991. All representative strains were identified as G serotype 6 (G6) and P type 5 (P5) by results of the virus-neutralization test and polymerase chain reaction procedure. Thus, BRV had no change in genomic RNA electropherotypes and serologic antigenicities in a closed dairy herd over a period of several years.

Pharmacokinetic and pharmacodynamic variables of flunixin were studied in calves after IV administration of the drug at a dose rate of 2.2 mg/kg of body weight. The anti-inflammatory properties of flunixin were investigated, using a model of acute inflammation; this involved surgically implanting tissue cages at subcutaneous sites and stimulating the tissue cage granulation tissue by intracavitary injection of carrageenan. The actions of flunixin on exudate concentrations of several substances related to the inflammatory process, including proteases (metalloprotease [active and total] and cysteine and serine proteases), enzymes (lactate dehydrogenase, acid phosphatase, and beta-glucuronidase [beta-glu]), eicosanoid (prostaglandin E2 [PGE2], leukotriene B4, and serum thromboxane B2 [TXB2]) concentrations, and bradykinin (BK)-induced edema, were investigated. Flunixin had a long elimination half-life--6.87 +/- 0.49 hours--and volume of distribution was 2.11 +/- 0.37 L/kg, indicating extensive distribution of the drug in the body. Body clearance was 0.20 +/- 0.03 L/kg/h. Flunixin exerted inhibitory effects on serum TXB2 and exudate PGE2 concentrations, B-glu activity, and BK-induced swelling. Other enzymes and inflammatory mediators were not significantly affected. Pharmacokinetic/pharmacodynamic modeling of the data revealed similar mean concentration producing 50% of the maximal effect values for inhibition of exudate PGE2 and beta-glu and of BK-induced swelling (0.070 +/- 0.006, 0.064 +/- 0.040, and 0.061 +/- 0.030 microgram/ml), respectively). A lower concentration producing 50% of the maximal effect value was obtained for inhibition of serum TXB2 concentration (0.023 +/- 0.004 microgram/ml). Differences also were observed in equilibration half-life for these actions, suggesting the existence of 3 distribution compartments correlating with 3 sites of action--a central compartment and shallow and deep peripheral compartments. Pharmacokinetic/pharmacodynamic modeling proved to be a useful analytical method, providing a quantitative description of in vivo drug pharmacodynamics and indicating possible mechanisms of action.

The effect of xylazine, cisapride, and naloxone on myoelectric activity of the ileum, cecum, and proximal loop of the ascending colon (PLAC) was determined in 4 healthy Jersey cows implanted with 8 pairs of bipolar electrodes. A 4 X 4 Latin square design was used. The treatments included xylazine (0.04 mg/kg of body weight), cisapride (0.08 mg/kg), naloxone (0.05 mg/kg), and 0.9% sodium chloride solution (20 ml). All treatments were administered IV during early phase I of the migrating myoelectric complex in the ileum. Myoelectric activity was recorded for 4 hours after treatment, and data were analyzed for each hour separately. Xylazine significantly (P < 0.05) increased the duration of phase I of the first migrating myoelectric complex in the ileum to 220.72 +/- 26.89 minutes, compared with 30.91 +/- 10.11 minutes after administration of 0.9% sodium chloride solution. The number of cecocolic spikes per minute per electrode and the duration of cecocolic spike activity (percentage of recording time) were significantly (P < 0.05) decreased for the first 3 hours, and the number of propagated spike sequences in the cecum and PLAC was significantly (P < 0.05) decreased for the first 2 hours after administration of xylazine. Significant difference was not found between control and either,cisapride or naloxone treatment of healthy cows. However, during hour 1 after treatment with cisapride, number of spikes per minute, duration of spike activity, and number of propagated spike sequences were highest, compared with the other treatments. It was concluded that naloxone at the dosage used in this study was not suitable for medical treatment of cecal dilatation in cattle, when hypomotility of the cecum and PLAC must be reversed. Xylazine should not be used for relief of signs of pain in cases of cecal dilatation, because it significantly reduced myoelectric activity of the cecum and PLAC for at least 2 hours after treatment. Furthermore, results of this study indicated a trend (P > 0.05) toward increase of cecocolic myoelectric activity after administration of cisapride. It is the authors' opinion that the potential benefit of cisapride for medical treatment of cecal dilatation in cattle needs further evaluation.

The main objective of the study reported here was to determine whether signs typical of exocrine pancreatic insufficiency (EPI) are alleviated when affected dogs are fed a diet with low fat content, compared with feeding ordinary commercial dog food or food prepared by the owner. The most cost-effective amount of enzyme supplement also was estimated. The study consisted of 6 test periods. Duration of the first and third periods was 4 weeks, and that of the others was 2 weeks. During the first 2 periods, the dogs were fed their original diet. The amount of enzyme supplement was reduced by half between the first and the second period. During the last 4 periods, the dogs were fed only the low-fat diet, and amount of the enzyme supplement was reduced stepwise. During the entire study, owners were asked to assess daily the severity of 9 signs typical of EPI. A new index was established by adding the daily scores of each individual EPI sign. This index was designated the EPI index and was used as a measure of the general well-being of the dog. When the mean EPI indexes of the original diet periods were compared with those of the corresponding low-fat diet periods, there were no statistically significant differences by use of Turkeys test or the paired t-test. There was considerable variability between dogs, however. The fat content of the original diet did not correlate with the difference in EPI signs when the dogs were fed the low-fat diet. According to our study, feeding a low-fat diet to dogs with EPI did not significantly alleviate clinical signs of the disease. Decreasing the enzyme supplementation by 50% of the recommended of dose did not significantly increase severity of the cumulative EPI score. Decreasing the enzyme supplement by three-fourths of the recommended dose was excessive, and the severity of the clinical signs increased significantly (P < 0.05) The cost of the low-fat diet, compared with that of the original diet, was high.

Two techniques for evaluating argyrophilic nucleolar organizer regions (AgNOR) were compared on 74 canine mammary tumors to discriminate between benign and malignant lesions. For each lesion, direct counting of AgNOR on at least 100 cell nuclei was compared with area, perimeter, and integrated optical density AgNOR dot values determined by image analysis. Significant differences between benign and malignant tumors were observed with both methods; however, lesions determined as aggressive or proliferative by histologic evaluation were only singled out by image analysis measurements. Image analysis, in our hands, was a reliable, precise, and convenient technique to characterize malignancy in canine mammary tumors.

Using 7 penicillins (amoxicillin, ampicillin, methicillin, penicillin G, oxacillin, cloxacillin, and dicloxacillin), simultaneous and direct determination of residual penicillins in biological samples was carried out by use of bioassay and high-performance liquid chromatography with spectrophotometric or fluorometric detectors. By use of assay medium seeded with penicillin-sensitive Micrococcus luteus (ATCC No. 9341) as a test organism, we were able to detect penicillins even at low concentrations. All penicillins treated with 10 U of penicillinase/ml did not produce inhibition zones by disk testing, even at a concentration of 100 micrograms of penicillin/ml/assay plate. Using a mobile phase of acetonitrile:methanol:0.01M KH2PO4 (19:11:70, v/v/v; pH, 7.1), standard solutions of the penicillins were separated from each other by use of high-performance liquid chromatography analysis, producing symmetric peaks without tailing, each of which had a characteristic retention time. Simultaneous detection of residual penicillins in bovine serum, kidneys, and liver, for the 5 penicillins for which analysis was possible by use of the UV method, yielded recovery rates from 71.4 to 102.3%; for the 2 amino-penicillins, amoxicillin and ampicillin, which could only be detected by use of the fluorometric method, recovery rate ranged from 72.9 to 103%.

A monoclonal antibody (MAB) against equine tumor necrosis factor-alpha (Eq TNF) was used to investigate the role of TNF in cytokine, eicosanoid, and metabolic responses of Miniature Horses given endotoxin. Plasma concentrations of interleukin 6 (IL-6), lactate, thromboxane A2 metabolite, and prostacyclin metabolite (6-keto-PGF(1 alpha)) were measured in 10 Miniature Horses given 0.25 micrograms of lipopolysaccharide (LPS; Escherichia coli O55:B5)/kg of body weight. Five horses were given Eq TNF MAB and 5 were given isotype-matched MAB as control. All horses were given 1.86 mg of antibody/kg by IV infusion, 5 minutes before LPS was given IV. Blood samples were taken 20 minutes before and at multiple intervals for 24 hours after LPS was given. Interleukin 6 bioactivity in plasma was measured, using IL-6-dependent cell line (B9). Eicosanoid activities were assessed by enzyme immunoassay, and plasma lactate concentration was determined enzymatically. Data were analyzed by ANOVA and Tukey's honest significant difference test for significant (P < 0.05) effect of treatment. Horses given Eq TNF MAB had significantly (P < 0.050) lower peak mean +/- SEM IL-6 (59 +/- 29 U/ml), lactate (16 +/- 2.00 mg/dl), and 6-keto-PGF(1 alpha) (254 +/- 79 pg/ml) values then did horses given control MAB (880 +/- 375 U/ml for IL-6; 26 +/- 0.04 mg/dl for lactate; and 985 +/- 290 pg/ml for 6-keto-PGF(1 alpha)). There was no effect of anti-TNF treatment on LPS-induced thromboxane A2 metabolite production. Tumor necrosis factor mediated IL-6, lactate, and prostacyclin responses, without affecting thromboxane production in horses given LPS.

A matched case-control study design was used to assess the effects of long-term administration of a prolonged release formulation of bovine somatotropin (sometribove) on clinical lameness and limb lesions in dairy cows. Cows treated with sometribove for at least 2 lactations (cases) and nontreated dairy cows matched by herd, parity, age, and stage of lactation (controls) in 8 herds were evaluated for clinical lameness (as assessed by gait abnormality) and limb lesions by 2 observers, using a standardized scoring procedure at a single herd visit. Although a high proportion of the study cows were clinically lame (43%), an association was not detected between chronic administration of sometribove and prevalent lameness. Of 21 types of limb lesions identified, 2 were positively associated and 2 were negatively associated with long-term sometribove use. Superficial laceration of the tarsus (odds ratio [OR] = 2.1) and superficial swelling of the metatarsophalangeal joint (OR = 4.5) were positively associated with sometribove treatment, whereas femoral lesions (OR = 0.2) and superficial lacerations of the femur (OR = 0.14) were negatively associated with sometribove treatment.