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

为了明确在不同的发情周期中对FSH进行超排处理的卵巢反应，进行了连续的超声检查以追踪个体卵泡发育和卵巢切除术以观察卵泡形态，超排处理伴随多次肌内注射p-FSH（28 Au），以递减剂量，间隔12小时，持续4天，在第7天（3头奶牛）和第10天（4头奶牛）开始。用超数排卵处理结束前，在注射前列腺素F（PGF_2）同步发情后每隔12小时用超声检查卵巢<2a>。如前所述，使用内置卡尺在冷冻超声图像中监测所有直径大于4 mm的卵泡。注射1 μ FSH后100小时行卵巢切除<st>术，两组中优势卵泡直径（DF）均明显大于最大次级卵泡直径（LS ...更多信息 F）自第4天起。在第7天组中，DF在第7天达到最大直径（7.4 mm），而在第10天组中，DF在第8天达到最大直径（12.7 mm），在第10天停止生长（12.4 mm）。在第7天组中，S型卵泡从第一次FSH时的30.7个减少到最后一次FSH时的17.0个，而L型卵泡从1.0增加到15.7 mm。在第10天组中，S型卵泡从第一次FSH时的32.8 mm减少到最后一次FSH时的17.5 mm，而L型卵泡从1.3 mm增加到31.5 mm。第1次注射FSH后48小时，卵泡数（直径&gt; 4 mm）最多（5.0个），而第10天组在24小时（8.0个）出现最早。超排处理第10天组的血浆E_2水平在36小时后显著高于第7天组，超排处理过程中超声观察到223个直径&gt;4 mm的卵泡，其中170个卵泡被单独解剖（回收率为73%），体视显微镜下采集到123个卵母细胞（回收率为72%）。在第7天和第10天组之间，卵泡液类固醇浓度没有统计学差异，但是出现较晚的卵泡倾向于增加卵泡液类固醇的E/P比值。共92个卵母细胞进行荧光染色，显示出对钙黄绿素的阳性活性颜色。这些结果表明，在DF失去优势效应后第10天开始的超排处理比在DF仍处于优势效应的第7天开始的超排处理更容易提高卵巢的反应性。主导阶段少

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