In vivo Ultrasonography and Ovariectomy for Evaluating Individual Follicles Superovulated on Different Stage of Estrous Cycles in Japanese Black Cows

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In order to clarify the ovarian response to the superovulation treatment carried out with FSH in different estrous cycles, a consecutive ultrasonography for tracing the individual follicular development and an ovariectomy for follicular morphology were performed.The superovulation treatment accompanied with multiple intramuscular injections of p-FSH (28 AU) executed in decreasing doses at 12 hr intervals for 4 days, was initiated on Day 7 (3 cows) and on Day 10 (4 cows). Ovaries were monitored with ultrasonography at 12 hr intervals after an estrus previously synchronized estrus by PGF_<2a> injection until the end of superovulation treatment. All the follicles more than 4mm in diameter were monitored in freezing ultrasonography images with built-in calipers as previously described. And then, the ovariectomy was carried out at 1OO hr after the 1^<st> FSH injection.In both groups, the diameter of dominant follicle (DF) became significantly larger than the largest subordinate follicle (LS … More F) since Day 4. In Day 7 group, DF reached the maximum diameter (7.4 mm) on Day 7, while in Day 10 group it reached that (12.7 mm) on Day 8, ceasing to grow on Day 10 (12.4 mm). In Day 7 group, the number of S-size follicles decreased from 30.7 at the first FSH to 17.0 at the last FSH, while L-sized ones increased from 1.0 to 15.7 mm. In Day 10 group, S-sized ones decreased from 32.8 mm at first FSH to 17.5 mm at last FSH, and L-sized ones increased 1.3 mm to 31.5 mm. In Day 7 group, the number of follicles (>4mm in diameter) recruited in response to FSH injections showed maximum at 48 hr (5.0 follicles) after first FSH injection, while it showed that earlier at 24 hr (8.0 follicles) in Day 10 group. The plasma E_2 profile showed significantly higher levels at and after 36 hr, during superovulation treatment in Day 10 group, compared those in Day 7 group.Out of 223 follicles (>4 mm in diameter) observed with ultrasonography during the superovulation treatment, 170 follicles were individual dissected (73% recovery), 123 oocytes were collected under stereomicroscope (72%o recovery). Between the Day 7 and Day 10 groups, there was no statistic difference in the follicular fluid steroid concentration, however the follicles that emerged later tended to increase the E/P ratio of follicular fluid steroids. A total of 92 oocytes subjected for fluorochroming showed a positive viable color against Calcein-AM.There was no significant difference in the follicular morphological characteristics depending on the treatment.These results indicate that the ovarian responses were apt to be made more increased by the superovulation treatment initiated at Day 10 after DF loss its dominance effect than by the treatment executed at Day 7 when DF was still in dominant phase. Less