Confirmation of epidural needle placement using nerve stimulation in dogs

Caudal epidural anesthesia is useful when anesthesia of the lumbar and sacral dermatomes is needed. Its success relies on the proper placement of the needle in the epidural space. However, accurate positioning of the needle can be difficult in certain patients (i.e.obesity). The purpose of this preliminary study was to document the use of nerve stimulation as a means of confirming accurate needle positioning in the epidural space prior to drug administration. Twenty large breed dogs undergoing hindlimb or perineal surgery were enrolled. Following induction of general anesthesia, patients were prepared for routine epidural drug administration. A 17 ga, 3.5” shielded Tuohy needle was used and was connected to a peripheral nerve stimulator set to deliver a current at 1 Hz, with a pulse width of 0.2 m sec. Initial current was set at 1.2 mA as the needle was advanced into position. Confirmation of epidural needle placement was confirmed when twitches were observed in the hindlimbs and/or tail. Current setting was then decreased incrementally by 0.2 mA until no further twitches were observed. Success of epidural drug placement was confirmed subjectively by motor blockade to the blocked dermatomes and clinical signs of balanced anesthesia (lack of sympathetic response to surgical stimulation while maintained at light plane of anesthesia). Lowest mean (range) current to elicit hindlimb twitches was 0.72 mA (0.4–1.0 mA). Lowest mean (range) current to elicit tail twitches was 0.58 mA (0.4–1.0 mA). Tail twitches were reliably lost at mean current of 0.37 mA (0.2–0.8). Epidural anesthesia was considered to be successful in 19/20 dogs. In only 9/20 dogs, needle placement would have been correct based on using ‘classic’ indicators alone (‘pop’ as enter epidural space, loss of resistance to injection). The results of this study suggest that nerve stimulation may be useful in confirming correct epidural needle placement prior to drug administration.     

A study of the microbial flora of the anterior vagina of normal sows during different stages of the reproductive cycle

SUMMARY Sterile guarded swabs were used to sample the anterior vaginal and cervical area of 23 normal healthy sows during various stages of the reproductive cycle. The samples were collected one week before farrowing, within 24 hours of farrowing, weekly up to weaning, at mating and at 2 and 3 weeks after mating, and then plated and incubated aerobically and anaerobically. At least one positive sample was obtained from each sow and at each stage of the reproductive cycle. Most positive samples (78.3%) were obtained on the day of farrowing and the least 3 weeks after mating (19.0%). The second highest number of positive samples (45.5%) was found immediately after mating. Although there was no significant difference among sows of different parities, there was a trend for older sows to have more positive samples after farrowing (84.6%). There was a greater decrease in positive samples after farrowing and after mating among younger sows compared with older sows. A wide range of bacteria including aerobic and anaerobic species, were recovered from 142 Isolates. The more representative bacteria were Streptococcus spp (23.2%); Escherichia coll (22.5%); Staphylococcus spp (19.0%) and Corynebacterium spp (13.4%). Of the cultures, 54.7% were pure and 45.3% were mixed. Both the percentage of bacterial isolates as well as the type of culture (pure or mixed) were similar to those frequently reported in clinical cases of vulval discharge syndrome. The results indicate that sows usually develop infections of the reproductive tract at farrowing and mating but these infections do not normally persist.     

Storage of Bovine Reproductive Tissues and RNA Extracts on Ice for 24 h or Repeated Freeze–Thaw Cycles do not Affect RNA Integrity

The aims of this study were to test (i) the effect of time of tissue and RNA extracts storage on ice and (ii) the effect of repeated freeze–thaw cycles on RNA integrity and gene expression of bovine reproductive tissues. Fragments of endometrium (ENDO), corpus luteum (CL) and ampulla (AMP) were subdivided and incubated for 0, 1, 3, 6, 12 or 24 h on ice. RNA extracts were incubated on ice for 0, 3, 12 or 24 h, or exposed to 1, 2, 4 or 6 freeze–thaw cycles. RNA integrity number (RIN) was estimated. Expression of progesterone receptor (PGR) and cyclophilin genes from RNA extracts stored on ice for 0 or 24 h, and 1 or 6 freeze–thaw cycles was measured by qPCR. Tissue and RNA extract incubation on ice, and repeated freeze–thaw cycles did not affect RIN values of RNA from ENDO, CL or AMP. Storage on ice or exposure to freeze–thaw cycles did not affect Cq values for PGR or cyclophilin genes . In conclusion, neither generalized RNA degradation nor specific RNA degradation was affected by storage of tissue or RNA extracts on ice for up to 24 h, or by up to 6 freeze–thaw cycles of RNA extracts obtained from bovine ENDO, CL and AMP.     

Late Pleistocene Desiccation of Lake Victoria and Rapid Evolution of Cichlid Fishes

Lake Victoria is the largest lake in Africa and harbors more than 300 endemic species of haplochromine cichlid fish. Seismic reflection profiles and piston cores show that the lake not only was at a low stand but dried up completely during the Late Pleistocene, before 12,400 carbon-14 years before the present. These results imply that the rate of speciation of cichlid fish in this tropical lake has been extremely rapid.     

Assessment of Extent of Apoptosis and DNA Defragmentation in Chilled Semen of Stallions During the Breeding Season

The objective of the study was to assess apoptosis and DNA defragmentation in equine semen diluted and chilled to +4°C. Semen was collected from nine fertile stallions, including four Arabian thoroughbreds and five coldbloods. Examinations were carried out immediately after semen collection (0) and at five storage times (24, 48, 72, 96 and 120 h). The basic semen evaluation was performed in terms of volume, sperm concentration, viable sperm percentage, progressive motility and morphology. Using flow cytometry, DNA defragmentation and cell membrane integrity of spermatozoa were determined. The results of basic tests did not demonstrate significant differences amongst stallions, except for progressive sperm motility, which was significantly higher (p < 0.05) in the semen of Arabian stallions. In the semen of the same stallions, a significant decrease in the percentage of alive spermatozoa was observed at 72, 96 and 120 h of storage, whereas a significant increase in the number of spermatozoa with DNA defragmentation was found after 24 h storage. In the semen of coldblood stallions, significantly reduced live spermatozoa percentage was observed at 96 and 120 h, while increased DNA defragmentation was observed at 48 h. These findings demonstrated that the semen of Arabian stallions chilled to +4°C retained original characteristics until 24 h of storage, whereas in coldbloods, these were preserved up to 48 h of storage.     

Comparison of the TBARS Assay and BODIPY C11 Probes for Assessing Lipid Peroxidation in Red Deer Spermatozoa

Several methods are used to measure lipid peroxidation (LPO) in spermatozoa. The objective of this study was comparing the thiobarbituric acid reactive species (TBARS) method and the BODIPY 581/591 C₁₁ (B581) and BODIPY 665/676 C₁₁ (B665) fluorescent probes to measure induced peroxidative damage in thawed epididymal spermatozoa from Iberian red deer. Samples from three males were thawed, pooled, diluted in PBS, incubated at room temperature and assessed at 0, 3, 6 and 24 h under different experimental conditions: Control, hydrogen peroxide (H₂O₂) 0.1 mm or 1 mm , or tert‐butyl hydroperoxide (TBH) 0.1 mm or 1 mm . LPO was assessed by the TBARS assay [malondialdehyde (MDA) detection] and by the fluorescence probes B581 and B665 (microplate fluorimeter and flow cytometry). Increasing MDA levels were only detectable at 1 mm of TBH or H₂O₂. Both fluorescence probes, measured with fluorometer, detected significant increases of LPO with time in all treatments, except Control. Flow cytometry allowed for higher sensitivity, with both probes showing a significant linear relationship of increasing LPO with time for all oxidizing treatments (p < 0.001). All methods showed a good agreement, except TBARS, and flow cytometry showed the highest repeatability. Our results show that both B581 and B665 might be used for LPO analysis in Iberian red deer epididymal spermatozoa, together with fluorometry or flow cytometry. Yet, the TBARS method offered comparatively limited sensitivity, and further research must determine the source of that limitation.