Population estimate of the preantral follicles and frequency of multioocyte follicles in prepubertal and adult bitches.

Oocytes from preantral follicles could be an alternative for in vitro maturation because most follicles are at the preantral stage. There are few studies that have sought to estimate the number of preantral follicles in bitches. Therefore, the aims of this study were to estimate the population of preantral follicles in the ovaries of small- and medium-sized prepubertal and adult bitches and compare the population of preantral follicles between the right and left ovaries and evaluate the frequency of multioocyte follicles (MOF). Eighty ovaries were collected by elective ovariohysterectomy from 40 healthy bitches. The bitches were divided into four groups: small-size prepubertal bitches (<10 kg, n = 20), medium-size prepubertal bitches (10-20 kg, n = 20), small-size adult bitches (<10 kg, n = 20), and medium-size adult bitches (10-20 kg, n = 20). Immediately after surgery, the ovaries were fixed in Bouin's solution and processed for histology. For each specimen, 70 histologic sections were cut and mounted on slides; then, the number of preantral follicles was estimated using a correction factor. The preantral follicles were classified according to the developmental stage. The data were analyzed using the Kruskal-Wallis test followed by Dunn's test for comparison between groups, and Fisher's exact test was used to evaluate the frequency of MOF (P ≤ 0.05). Considering the population of preantral follicles from the pair of ovaries, medium-size prepubertal bitches had the highest (P < 0.05) population of preantral follicles compared with the small and medium-size adult groups. There was a large variation in the numbers of preantral follicles among individuals of the same weight and within each group. There were differences between medium-size prepubertal and adult bitches regarding the population of preantral follicles in the right ovaries (145,482 ± 110,712 vs. 49,500 ± 44,821; P = 0.02); however, no differences were observed between the groups on the basis of comparisons of the number of preantral follicles in the left ovaries (P > 0.05). The prevalence of primordial MOF was higher in prepubertal bitches (47% vs. 28%), whereas adult bitches had a higher frequency of secondary MOF (49% vs. 25%; P < 0.05). We conclude that medium-size prepubertal bitches had the highest population of preantral follicles compared with small and medium-size adult bitches, and the use of only one ovary per bitch implied contrasting result. The presence of primordial MOF was higher in prepubertal bitches and at the secondary stage in adult bitches.

Comparison of two different extenders for cryopreservation of epididymal dog sperm.

The collection of epididymal sperm is an option for preservation of germplasm of genetically superior animals that need to be orchiectomized or have died. The extender type used to freeze sperm is important to avoid spermatozoal membrane damage and to preserve semen quality after cryopreservation. The objective of this study was to verify the effects of a commercial bovine extender (Bovimix(®); Nutricell, Campinas) and a traditional TRIS-citric acid-glucose-egg yolk-7% glycerol extender on cryopreservation of canine epididymal sperm. The testes of 13 adult dogs were kept at 5 °C for 24 h in saline solution, and epididymal sperm was recovered in Ringers solution without lactate and were evaluated for motility. Samples with ≥ 80% motility were pooled and then divided before dilution and packaging in 0.5 ml plastic straws, equilibration at 4 °C for 1 h, freezing in nitrogen vapour for 20 min and storing at -196 °C. The straws were thawed at 56 °C for 10 s and were evaluated for motility by computer assisted analysis (CASA). The semen parameters, sperm movement index, linearity, total motility and rapid progressive motility were statistically higher in Bovimix(®) than TRIS. In contrast, amplitude of lateral head displacement, slow sperm and static sperm were lower in Bovimix(®). Despite the high percentage of sperm defects in epididymal cells, regardless of the extender, we concluded that Bovimix(®) is a viable alternative for the freezing of canine epididymal sperm.