Ultrastructural Characteristics of Non-matured and In Vitro Matured Oocytes Collected from Pre-pubertal and Adult Domestic Cat Ovaries

The present study describes the ultrastructural characteristics of cat oocytes before maturation and after 12- and 24-h in vitro maturation (IVM). Oocytes were recovered from pre-pubertal and adult queen ovaries after ovariohysterectomy and a proportion were stored in glutaraldehyde at 4°C until examination by transmission electronic microscopy (TEM). Those selected for maturation were cultured before TEM in DMEM for 12 and 24 h at 38°C in a humidified environment of 5% O₂, 5% CO₂ and 90% N₂. Specimens were divided into six groups: non-matured oocytes from pre-pubertal queens (PP0), non-matured oocytes from adult queens (A0), 12-h in vitro matured oocytes from pre-pubertal queens (PP12), 12-h in vitro matured oocytes from adult queens (A12), 24-h in vitro matured oocytes from pre-pubertal queens (PP24) and 24-h in vitro matured oocytes from adult queens (A24). Across the treatment groups, it was possible to observe differences in the thickness of the perivitelline space, the penetration of cumulus cell projections forming a junctional complex, distribution and density of small vesicles, lipid droplets, microvilli, mitochondria and cortical granules and variable degrees of development of Golgi complexes. These findings demonstrated that ultrastructural analysis of oocytes matured in vitro is a valuable tool to evaluate oocyte cytoplasmic maturation and that this IVM protocol was efficient in inducing gradual morphological changes necessary for cytoplasmic maturation of pre-pubertal and adult cat oocytes.     

Evaluation of an in-house enzyme-linked immunosorbent assay for quantitative measurement of serum total thyroxine concentration in dogs and cats

OBJECTIVE: To compare serum total thyroxine (T4) concentrations obtained with an in-house ELISA and a validated radioimmunoassay (RIA). DESIGN: Laboratory trial. SAMPLE POPULATION: 50 canine and 50 feline serum samples submitted for measurement of total T4 concentration with the RIA; samples were selected to represent a wide range of concentrations (< 6 to 167 nmol/L). PROCEDURE: Results of the ELISA and RIA were compared by calculating correlation coefficients, examining linearity, determining bias and precision, and evaluating clinical interpretations. RESULTS: Correlation coefficients for results of the 2 methods were 0.84 for the canine samples and 0.59 for the feline samples. Examination of bias plots revealed large variations in ELISA results, compared with RIA results. For the feline samples, the ELISA consistently overestimated total T4 concentration obtained with the RIA. When results of the 2 methods were categorized (low, borderline low, normal, borderline high, or high), results were discordant for 24 (48%) and 29 (58%) of the canine samples and for 18 (36%) and 28 (56%) of the feline samples (depending on whether borderline high ELISA results were considered normal or high). Reliance on results of the ELISA would have led to inappropriate clinical decisions for 31 (62%) canine samples and 25 (50%) feline samples. The ELISA coefficients of variation for the pooled canine and feline samples were 18 and 28%, respectively. CONCLUSIONS AND CLINICAL RELEVANCE: Substantial discrepancies between ELISA and RIA results for T4 concentrations were detected. Thus, we concluded that the in-house ELISA kit was not accurate for determining serum total T4 concentrations in dogs and cats.     

von Willebrand's disease in Dobermann dogs in Australia

SUMMARY Over a 5-year period (1988–92), von Willebrand factor antigen (vWf:Ag) concentrations were determined on plasma samples from 614 Dobermanns. The vWf:Ag concentration was < 50 canine units (CU)/dL in 373 dogs (61%); these dogs were classified as carriers of the von Willebrand's disease (vWD) gene. In order to identify which dogs were at risk of haemorrhage due to vWD, we determined a cut-off vWf:Ag concentration below which dogs could be considered at risk. This cut-off was chosen in order to minimise the number of dogs genuinely at risk of haemorrhage, being wrongly classified as not at risk. This was done without sacrificing the specificity of the cut-off to any great extent. A vWf:Ag concentration of < 36 CU/dL was empirically chosen as the optimum cut-off concentration. In 282 dogs (76% of the carriers), the vWf:Ag concentration was below this cut-off and these dogs were, thus, classified as being at risk of haemorrhage due to vWD. Haemorrhage attributable to vWD was seen in 107 dogs (29% of the carriers, or 17% of all the dogs). Haemorrhage mostly followed trauma or surgery, but spontaneous genitourinary and gastrointestinal haemorrhages were also frequent. Of these dogs, 92 were of known age, with a median of 3 years, and 102 were of known sex, with 61% being female. In 89 dogs in which the severity of haemorrhage was subjectively assessed, mild and moderate bleeding occurred with similar frequency (48% and 43%, respectively). There were 8 cases of severe haemorrhage, with two deaths. The likelihood of haemorrhage was related to the vWf:Ag concentration: only 8% of 91 dogs with concentrations between 36 and 49 CU/dL had haemorrhage attributable to vWD, but 36% with concentrations < 36 CU/dL did so. Furthermore, dogs with haemorrhage attributable to vWD had significantly (P < 0.001) lower vWf:Ag concentrations (median 12 CU/dL, n = 107) than dogs with no such history (median 30 CU/dL, n = 132). The data indicate that vWD is a significant problem in the Dobermann breed in Australia and we accordingly recommend that steps be taken to reduce its prevalence, such as the establishment of a national testing scheme to determine the vWD status of all dogs used for breeding.     

Effects of glucocorticoids on endocrine function in the dog

There are a multitude of possible side effects when using high levels of or chronic administration of glucocorticoid treatment. Several of the studies referred to in this discussion used large amounts of glucocorticoids for rather lengthy periods. The endocrine, as well as nonendocrine, effects of glucocorticoids are minimized when the lowest effective doses are used, when treatment is terminated as soon as reasonably possible, and when an alternate-day therapy schedule is followed. However, an occasional individual may appear with a particular susceptibility to one or more of the side effects of glucocorticoid treatment even when these measures are followed.     

Adrenocortical suppression in the dog after a single dose of methylprednisolone acetate

Adrenal function was assessed in dogs after intramuscular administration of a single dose of methylprednisolone acetate (MPA). Twelve dogs were test challenged with adrenocorticotropic hormone (ACTH) and then assigned randomly to 1 of 3 groups and given MPA. Individual groups were test challenged with ACTH 2, 3, or 4 weeks later. All dogs were rechallenged 5 weeks after MPA administration. Plasma cortisol concentration was determined by radioimmunoassay. Basal plasma cortisol (time 0) was depressed on weeks 2 and 3, but not on weeks 4 and 5. Adrenal response to ACTH (increment of cortisol change) was suppressed on weeks 2, 4, and 5, but not on week 3. It was concluded that a single dose of MPA is capable of altering adrenal cortical function in dogs for at least 5 weeks.     

Effect of alternate-day prednisolone administration on hypophyseal-adrenocortical activity in dogs.

OBJECTIVE: To evaluate effect of alternate-day oral administration of prednisolone on endogenous plasma ACTH concentration and adrenocortical response to exogenous ACTH in dogs. ANIMALS: 12 Beagles. PROCEDURE: Dogs were allotted to 2 groups (group 1, 8 dogs treated with 1 mg of prednisolone/kg of body weight; group 2, 4 dogs given excipient only). During a 30-day period, blood samples were collected for determination of plasma ACTH and cortisol concentrations before, during, and after treatment with prednisolone. From day 7 to 23, prednisolone or excipient was given on alternate days. Sample collection (48-hour period with 6-hour intervals) was performed on days 1, 7, 15, 21, and 28; on other days, sample collection was performed at 24-hour intervals. Pre- and post-ACTH plasma cortisol concentrations were determined on days 3, 9, 17, 23, and 30. RESULTS: A significant difference was detected between treatment and time for group 1. Plasma ACTH concentrations significantly decreased for 18 to 24 hours after prednisolone treatment in group-1 dogs. At 24 to 48 hours, ACTH concentrations were numerically higher but not significantly different in group-1 dogs. Post-ACTH plasma cortisol concentration significantly decreased after 1 dose of prednisolone and became more profound during the treatment period. However, post-ACTH cortisol concentration returned to the reference range 1 week after prednisolone administration was discontinued. CONCLUSIONS AND CLINICAL RELEVANCE: Single oral administration of 1 mg of prednisolone/kg significantly suppressed plasma ACTH concentration in dogs for 18 to 24 hours after treatment. Alternate-day treatment did not prevent suppression, as documented by the response to ACTH.     

Evaluation of a low-dose synthetic adrenocorticotropic hormone stimulation test in clinically normal dogs and dogs with naturally developing hyperadrenocorticism.

OBJECTIVE: To determine whether low doses of synthetic ACTH could induce a maximal cortisol response in clinically normal dogs and to compare a low-dose ACTH stimulation protocol to a standard high-dose ACTH stimulation protocol in dogs with hyperadrenocorticism. DESIGN: Cohort study. ANIMALS: 6 clinically normal dogs and 7 dogs with hyperadrenocorticism. PROCEDURE: Each clinically normal dog was given 1 of 3 doses of cosyntropin (1, 5, or 10 micrograms/kg [0.45, 2.3, or 4.5 micrograms/lb] of body weight, i.v.) in random order at 2-week intervals. Samples for determination of plasma cortisol and ACTH concentrations were obtained before and 30, 60, 90, and 120 minutes after ACTH administration. Each dog with hyperadrenocorticism was given 2 doses of cosyntropin (5 micrograms/kg or 250 micrograms/dog) in random order at 2-week intervals. In these dogs, samples for determination of plasma cortisol concentrations were obtained before and 60 minutes after ACTH administration. RESULTS: In the clinically normal dogs, peak cortisol concentration and area under the plasma cortisol response curve did not differ significantly among the 3 doses. However, mean plasma cortisol concentration in dogs given 1 microgram/kg peaked at 60 minutes, whereas dogs given doses of 5 or 10 micrograms/kg had peak cortisol values at 90 minutes. In dogs with hyperadrenocorticism, significant differences were not detected between cortisol concentrations after administration of the low or high dose of cosyntropin. CLINICAL IMPLICATIONS: Administration of cosyntropin at a rate of 5 micrograms/kg resulted in maximal stimulation of the adrenal cortex in clinically normal dogs and dogs with hyperadrenocorticism.     

Serum thyroxine and triiodothyronine concentrations in canine pyoderma

In an effort to evaluate the effect of pyoderma on circulating iodothyronines, plasma triiodothyronine (T3) and thyroxine (T4) values were determined before and after thyroid stimulating hormone administration in 25 dogs with pyoderma and in 15 controls. Basal T4 values were increased in dogs with pyoderma, but neither stimulated T4 nor T3 values were altered by this condition. On the basis of low values for circulating iodothyronines, hypothyroidism was suspected in 3 dogs in the pyoderma group. The dog with the most involved lesions had extremely low T3 and T4 values as well as an autoimmune disease. It was concluded that most dogs with pyoderma do not have thyroid dysfunction.