Spermatic and oxidative profile of domestic cat (Felis catus) epididymal sperm subjected to different cooling times (24, 48 and 72 hours)

Cooling stored epididymal samples for several days allows facilities to transport and process genetic material post‐mortem. Improvements to this practice allow the preservation of sperm from domestic cats, which are the ideal study model for wild felids. However, the modifications in spermatic features and the oxidative profile are not fully understood in cats. This information is necessary for the development of biotechniques, such as new extenders for cryopreservation. Therefore, the purpose of this study was to evaluate the spermatic and oxidative profile in samples from the epididymal cauda of domestic cats cooled at 5°C for 24, 48 and 72 hr. Spermatozoa were collected from the epididymis cauda. Evaluations consisted of computer‐assisted sperm analysis (CASA), plasma membrane integrity (eosin/nigrosin), acrosome integrity (fast green/rose bengal), sperm morphology, sperm DNA integrity (toluidine blue), mitochondrial activity (3′3 diaminobenzidine), activity of the antioxidant enzymes glutathione peroxidase (GPx) and superoxide dismutase (SOD), measurement of lipid peroxidation (TBARS) and protein oxidation. A decrease in sperm motility parameters was observed after 72 hr of cooling (i.e. total and progressive) with a higher percentage of minor (37.7 ± 6.3%) and total defects (53.4 ± 6.3%). Additionally, a decrease in high mitochondrial activity (Class I: 16.6 ± 2.2%) occurred after 72 hr. The decrease in motility rates after a long cooling time probably was caused by the increase in sperm abnormalities. A long cooling time causes cold shock and mitochondrial exhaustion, but there was no observed change with the oxidative stress condition. Therefore, cat epididymal sperm stored at 5°C appear to maintain a high quality for up to 48 hr of cooling time.     

A fast,low‐cost and efficient method for the diagnosis of sperm DNA fragmentation in several species

Sperm DNA fragmentation is a condition that interferes directly in the reproductive efficiency. Currently, there are several methods for assessing the sperm DNA integrity, such as Alkaline Comet, TUNEL and Sperm Chromatin Structure Assay. However, many of these techniques are laborious and require high‐precision equipment. Thus, the development of new techniques can optimize the evaluation of sperm DNA damage. Therefore, the aim of this study was to standardize the toluidine blue (TB) stain technique for the analysis of DNA fragmentation of dog, cat, bull, stallion and ram spermatozoa. For this purpose, we used six animals of each specie (n = 30), in reproductive age. Sperm was collected by different methods according to the particularities of each species, and such samples were divided into two aliquots: a sperm sample was kept at 5°C (considered as intact sperm DNA), and the remaining samples were submitted to the induction of DNA fragmentation by exposure to ultraviolet light for 4 hr. Samples were then mixed with the intact sample to obtain known and progressive proportions of sperm with fragmented DNA (0%, 25%, 50%, 75% and 100%). Semen smears were performed and subjected to staining with TB. Blue‐stained spermatozoa were considered to have DNA fragmentation. We observed high linear regression coefficients between the expected proportion of damaged DNA and the results of TB for dog, cat, ram, bull and stallion samples. In conclusion, TB stain was considered a fast and effective technique for the study of spermatozoa DNA in several species.     

Kisspeptin,c‐Fos and CRFR Type 2 Co‐expression in the Hypothalamus after Insulin‐Induced Hypoglycaemia

Normal reproductive function is dependent upon availability of glucose and insulin‐induced hypoglycaemia is a metabolic stressor known to disrupt the ovine oestrous cycle. We have recently shown that IIH has the ability to delay the LH surge of intact ewes. In the present study, we examined brain tissue to determine: (i) which hypothalamic regions are activated with respect to IIH and (ii) the effect of IIH on kisspeptin cell activation and CRFR type 2 immunoreactivity, all of which may be involved in disruptive mechanisms. Follicular phases were synchronized with progesterone vaginal pessaries and at 28 h after progesterone withdrawal (PW), animals received saline (n = 6) or insulin (4 IU/kg; n = 5) and were subsequently killed at 31 h after PW (i.e., 3 h after insulin administration). Peripheral hormone concentrations were evaluated, and hypothalamic sections were immunostained for either kisspeptin and c‐Fos (a marker of neuronal activation) or CRFR type 2. Within 3 h of treatment, cortisol concentrations had increased whereas plasma oestradiol concentrations decreased in peripheral plasma (p < 0.05 for both). In the arcuate nucleus (ARC), insulin‐treated ewes had an increased expression of c‐Fos. Furthermore, the percentage of kisspeptin cells co‐expressing c‐Fos increased in the ARC (from 11 to 51%; p < 0.05), but there was no change in the medial pre‐optic area (mPOA; 14 vs 19%). CRFR type 2 expression in the lower part of the ARC and the median eminence was not altered by insulin treatment. Thus, disruption of the LH surge after IIH in the follicular phase is not associated with decreased kisspeptin cell activation or an increase in CRFR type 2 in the ARC but may involve other cell types located in the ARC nucleus which are activated in response to IIH.     

Genetic differentiation and gene flow in Atlantic salmon,Salmo salar L.: a case study of the River Shannon system in Ireland

An analysis of the effect of an extensive restocking programme on genetic differentiation and gene flow of the Atlantic salmon, Salmo solar L., in the River Shannon system in Ireland is presented, using allozyme electrophoresis to estimate population differentiation. The observed differentiation in the River Shannon is attributed to random genetic drift, caused by small effective population sizes in some of the tributaries, coupled with the confounding effect of extensive stocking of indigenous fish which were largely derived from a single tributary. The methods of analysis are assessed by applying numerical resampling techniques to determine confidence levels for each of the estimates.     

Vascular Response of the Equine Radius to Cerclage Devices

The arterial blood supply to the diaphysis of the radius, and the vascular reaction when three cerclage devices (cerclage wire, Parham-Martin band, and a partially contacting band) were applied 2 cm apart, were studied in six adult ponies. The opposite radial diaphysis served as the control. Ponies were euthanatized at weeks 4 and 8. A centrifugal blood flow pattern was demonstrated by microangiography. Larger numbers of medullary arteries exist in equine than in canine and human long bones. Changes in centrifugal blood supply were not identified beneath any of the cerclage devices.     

'No-mucus skin disease' of rainbow trout, Oncorhynchus mykiss (Walbaum): a case report

Abstract A skin disease of intensively reared salmonids in Ontario hatcheries, known to the farmers as'no-mucus skin disease', is reported for the first time. It was characterized by erosive and ulcerative lesions found mainly on the flanks of fingerlings, which resulted in exposure of the tips of the scales. Associated with these lesions were colonies of bacteria seen in the SEM to be clustered round the mucous cell pores and under-running the margins of epithelial cells. The cause of this condition is unknown, although the response of fish to formalin treatment, and the presence of bacilli seen in skin scrapings and in the SEM, suggest that bacteria are responsible.     

Radiographic and Histologic Changes After Thoracolumbar Disc Curettage

Forty-eight thoracolumbar disc spaces were approached by a dorsolateral muscle-separating technique in six mature dogs. Twenty-four of the discs were fenestrated and curetted, and the changes were followed radiographically and histologically for 16 weeks. Disc space collapse was visible radiographically immediately after the surgical procedure, and those disc spaces remained narrow throughout the observation period. Histologically, there was evidence of chondrocyte activation within 2 weeks in the excavated disc spaces. By 16 weeks, fibrocartilage had filled the void in the curetted disc spaces.     

The pathology of chronic ammonia toxicity in rainbow trout, Salmo gairdneri Richardson

Abstract. Two groups of rainbow trout fingerlings were exposed for 90 days to 0.2 and 0.4 mg/1 of un-ionized ammonia, respectively. Several fish exposed to the higher concentration soon developed clinical signs suggestive of a neurological dysfunction but subsequently recovered. No lesion attributable to ammonia was seen in the gills of any of the fish. This raises questions about the precise role of ammonia in the production of gill diseases in intensively cultured trout.