Temporal interrelationships among luteolysis, FSH and LH concentrations and follicle deviation in mares

The effect of altered LH concentrations on the deviation in growth rates between the 2 largest follicles was studied in pony mares. The progestational phase was shortened by administration of PGF2alpha on Day 10 (Day 0=ovulation; n=9) or lengthened by daily administration of 100 mg of progesterone on Days 10 to 30 (n=11; controls, n=10). All follicles > or = 5 mm were ablated on Day 10 in all groups to initiate a new follicular wave. The interovulatory interval was not altered by the PGF2alpha treatment despite a 4-day earlier decrease in progesterone concentrations. Time required for growth of the follicles of the new wave apparently delayed the interval to ovulation after luteolysis. The FSH concentrations of the first post-ablation FSH surge were not different among groups. A second FSH surge with an associated follicular wave began by Day 22 in 7 of 11 mares in the progesterone group and in 0 of 19 mares in the other groups, indicating reduced functional competence of the largest follicle. A prolonged elevation in LH concentrations began on the mean day of wave emergence (Day 11) in the prostaglandin group (19.2 +/- 2.2 vs 9.0 +/- 0.7 ng/mL in controls; P<0.05), an average of 4 d before an increase in the controls. Concentrations of LH in the progesterone group initially increased until Day 14 and then decreased so that by Day 18 the concentrations were lower (P<0.05) than in the control group (12.9 +/- 1.6 vs 20.2 +/- 2.6 ng/mL). Neither the early and prolonged increase nor the early decrease in LH concentrations altered the growth profile of the second-largest follicle, suggesting that LH was not involved in the initiation of deviation. However, the early decrease in LH concentrations in the progesterone group was followed by a smaller (P<0.05) diameter of the largest follicle by Day 20 (26.9 +/- 1.7 mm) than the controls (30.3 +/- 1.7 mm), suggesting that LH was necessary for continued growth of the largest follicle after deviation.     

Selection of the dominant follicle in cattle: establishment of follicle deviation in less than 8 hours through depression of FSH concentrations

Deviation in follicle diameter in cattle is characterized by continued growth of the largest follicle of a follicular wave and a reduction or cessation of growth of the smaller follicles. Deviation begins when the largest follicle reaches about 8.5 mm. Two experiments were done to test the hypothesis that the deviation mechanism is established in < 8 h, as indicated by the temporal relationships between follicle removal and an increase in FSH concentrations (Experiment 1) and between a decrease in FSH concentrations and follicle inhibition (Experiment 2). In Experiment 1, the role of the first follicle to reach 8.5 mm was studied by follicle ablation (Hour 0). The combined mean FSH concentrations for the control group (n = 8) and ablation group before ablation (n = 7) progressively decreased (P < 0.02) over two 8-h intervals before the largest follicle reached > or = 8.5 mm (Hour-16, 1.77 +/- 0.11 ng/mL; Hour 0, 1.49 +/- 0.08 ng/mL). In controls, the concentrations continued to decrease (P < 0.02) until Hour 10 (1.21 +/- 0.09 ng/mL). Ablation of the largest follicle at > or = 8.5 mm resulted in increased (P < 0.02) circulating FSH concentrations between Hours 5 (1.34 +/- 0.04 ng/mL) and 8 (1.61 +/- 0.09 ng/mL). Growth rate of the second-largest follicle between Hours 0 and 8 was greater (P < 0.05) in the ablation group than in the controls, and the second largest follicle became dominant in 7 of 7 heifers following ablation of the largest follicle. In Experiment 2, a minimal single injection of a depressant of FSH concentrations (4.4 mL of steroid-reduced follicular fluid) was given when the largest follicle was a mean of 8.4 mm (Hour 0; controls, n = 4; treated, n = 4). An interaction of group and hour (P < 0.005) for FSH concentrations was attributable to an FSH decrease (P < 0.002) by Hour 6 and an increase (P < 0.002) between Hours 9 and 12 in the treated group. The growth rate of the largest follicle between Hours 0 and 12 was less (P < 0.05) in the treated group (0.2 +/- 0.2 mm/12 h) than in the control group (1.2 +/- 0.4 mm/12 h). The reduced diameter was recorded within 6 h after suppression of FSH concentrations, supporting the hypothesis. Our preferred interpretation is that when the largest follicle reaches a critical diameter of about > or = 8.5 mm, FSH concentrations continue to decrease and become lower than required by the smaller follicles but not the largest follicle. The results further indicate that a close temporal coupling between a change in FSH concentrations and the follicular response could establish the deviation mechanism in < 8 h or before the second largest follicle reaches a similar critical diameter.     

Intrafollicular effect of IGF1 on development of follicle dominance in mares

The effect of an injection of a supraphysiologic dose of rhIGF1 into the second-largest ovarian follicle (F2) at the expected beginning of deviation (F1, > or =20 mm; Day 0) on development of dominance by F2 was studied in mares (n=16; controls, n=8). F1 became dominant (> or =28 mm) in 8 of 8 and 15 of 16 follicles in the controls and treated groups, respectively. The incidence of dominance (P<0.001) and ovulation (P<0.02) for F2 was greater for the IGF1 group (13 of 16 and 10 of 16) than for the controls (1 of 8 and 1 of 8). There were day effects but no group effects or group-by-day interactions for systemic FSH, LH, estradiol, or ir-inhibin during the 4 days after treatment. In another experiment, treatment of every follicle, excluding F1, when it reached > or =20mm after the expected beginning of deviation resulted in dominance by 8 of 12 follicles treated with rhIGF1 on Days 1-3 (n=8 mares). Results demonstrated that the IGF1 system plays a pivotal intrafollicular role in the deviation mechanism without altering systemic concentrations of the gonadotropins and ovarian follicular hormones.     

Response of estradiol and inhibin to experimentally reduced luteinizing hormone during follicle deviation in mares

The increase in LH concentrations at the time of the decrease in FSH concentrations during follicle deviation in mares was studied to determine the role of LH in the production of estradiol and immunoreactive inhibin (ir-inhibin). Ten days after ovulation, all follicles > or =6 mm were ablated, prostaglandin F(2 alpha) was given, and either 0 mg (control group, n = 15) or 100 mg of progesterone in safflower oil (treated group, n = 16) was given daily for 14 days, encompassing the day of diameter deviation. The follicular and hormonal data were normalized to the expected day of the beginning of diameter deviation when the largest follicle first reached > or =20 mm (Day 0). The experimentally induced decrease in LH concentrations during follicle deviation beginning on Day -4 delayed and stunted the increase in circulating concentrations of ir-inhibin and estradiol beginning on Days -3 and -1, respectively, but did not alter the predeviation FSH surge and the initiation of diameter deviation between the two largest follicles. Combined for both groups, the interval to the expected day of deviation was 16.6 days after ovulation when the largest follicle was a mean of 21.6 mm. After deviation, the largest follicle started to regress in the treated group beginning on Day 1 and was associated with decreased concentrations of ir-inhibin and estradiol, and increased concentrations of FSH. The negative influence of the dominant follicle on the postdeviation decrease in FSH observed in the control group was alleviated and concentrations resurged in the treated group. Apparently this is the first in vivo evidence that the increase in LH that precedes follicle deviation has a positive effect in supporting the production of inhibin during diameter deviation. It was concluded that the increase in LH concentrations before diameter deviation played a role in the production of estradiol and inhibin by the largest follicle during deviation.     

Mechanism of follicle deviation in monovular farm species

Diameter deviation is a distinctive change in growth rates among the follicles of a wave, heralding the formation of a dominant follicle and subordinate follicles. When the follicles are about 5mm in cattle and 13 mm in horses, the wave-stimulating FSH surge reaches peak concentrations. Follicle and FSH manipulation studies in both species have shown that the declining portion of the surge before the beginning of deviation is a function of multiple growing follicles that require the decreasing FSH. During this time, all follicles of the wave have the potential for future dominance. Deviation begins when the two largest follicles on average are 8.5 and 7.7 mm in cattle and 22.5 and 19.0 mm in horses or about 3 days after the FSH peak in both species. The FSH/follicle relationship is close so that a change in one event soon causes a detectable change in the other. Thus, the difference in diameter between the two largest follicles at the beginning of deviation is compatible with rapid establishment of the destiny of the two follicles before the second-largest follicle can also show dominance. The deviation mechanism is initiated when FSH concentrations are low and the most advanced follicle reaches a specific developmental stage. In cattle, the future dominant follicle develops greater LH-receptor expression than the other follicles about 8 h before the beginning of diameter deviation. Estradiol and free IGF-1 begin to establish higher concentrations in the future dominant follicle than in other follicles and activin-A is transiently elevated in both follicles a few hours before the beginning of diameter deviation. In horses, estradiol, free IGF-1, activin-A, and inhibin-A begin to increase differentially in the future dominant follicle about 1 day before deviation. These changes underlie a greater responsiveness to LH and FSH by the developing dominant follicle than for other follicles, thereby accounting for deviation. Results of in vitro studies, although frequently done in other species, support this conclusion.     

Effect of presence of a dominant follicle on the superovulatory response in buffalo (Bubalus bubalis)

Ten buffalo were superovulated by administration of 8 doses of FSH in a descending schedule spread over 4 d (5.5/5.5, 4.5/4.5, 3.5/3.5 and 2.5/2.5 mL, i.m.; total dose of 64 AU in 32 mL) beginning on Day 10 of an unstimulated estrous cycle, and 30 and 20 mg Lutalyse was given alongwith the 5th and 6th injections of FSH, respectively, to induce luteolysis. The number of corpora lutea (CL) was determined on 6 d post estrus. The ovaries were examined daily by ultrasonography from Day -5 to Day 5 (Day 0 = day of start of superovulation). The animals were retrospectively classified into 2 groups depending upon the presence (n = 4) or absence of a dominant follicle (n = 6). The mean diameter of the largest follicle (F1) increased from 8.25 +/- 0.48 mm on Day -5 to 10.75 +/- 0.25 mm on Day 0 in the dominant group, whereas in the nondominant group the F1 follicle exhibited a progressive decrease from 9.00 +/- 0.45 mm to 7.00 +/- 0.65 mm during the same period, the difference in profiles between the 2 groups was significant (P = 0.042). The profile of the diameter of the second largest follicle (F2) and the difference in diameters between largest and second largest follicles (F1-F2) were not significantly different between the 2 groups. The profile of mean number of large (> or = 10 mm diameter), but not small (2 to 5 mm diameter) or medium (6 to 9 mm diameter) follicles differed significantly (P = 0.001) between the 2 groups from Day -5 to Day 5 (P = 0.030). The number of CL was not significantly different between nondominant (4.00 +/- 0.97) and dominant groups (3.25 +/- 1.31). The number of CL was positively correlated (P < 0.01) with the number of medium follicles and the total number of follicles on the day of initiation of superovulation, but not with follicles of any size category or total number of follicles on any previous day. The results of this study indicate that following the use of morphological criteria based on the size of the largest follicle alone, the superovulation response is not affected by the presence of a dominant follicle at the initiation of superovulation in buffalo.     

Follicle selection in monovular species

Follicle deviation is proposed to be the eminent event in follicle selection in monovular species. At deviation, the largest follicle establishes dominance apparently before the second-largest follicle can reach a similar diameter. In cattle, based on diameters of the two follicles at the beginning of deviation, the mechanism becomes established in <8 h. An FSH:follicle-coupling hypothesis has been supported as the essence of follicle selection. According to the hypothesis, the growing follicles cause the FSH decline from the peak of the wave-stimulating FSH surge until deviation, even though the follicles continue to require FSH (two-way functional coupling involving multiple follicles). During multiple-follicle coupling, inhibin is the primary FSH suppressant. Near the beginning of deviation, the largest follicle secretes increased estradiol, and apparently both estradiol and inhibin contribute to the continuing FSH decline; only the more-developed largest follicle is able to utilize the low FSH concentrations (single-follicle coupling). Deviation is encompassed by a transient elevation in LH in heifers and by a component, often distinct, of the long ovulatory LH surge in mares. In heifers, receptors for LH appear in the granulosa cells of the future dominant follicle about 8 h before the beginning of deviation. The LH stimulates the production of estradiol and insulin-like growth factor-1. These intrafollicular factors and perhaps others account for the responsiveness of the largest follicle to the low concentrations of FSH. The smaller follicles have not reached a similar developmental stage and because of their continued and close dependency on FSH become susceptible to the low concentrations. Thereby, follicle selection is established.     

Emergence and deviation of follicles during the development of follicular waves in cattle

The nature of emergence and deviation of follicles during follicular waves in cattle was studied in 3 experiments by re-examining data from previous projects. Wave emergence was defined as the day or examination (when more than 1 examination per day) the future dominant follicle was 4 mm (Day 0 or Examination 0). Deviation was defined as the beginning of the greatest difference in growth rates between the 2 largest follicles and between 2 consecutive examinations. The search for deviation in an individual wave was done retrospectively from the examination with the maximum diameter of the second largest follicle. In Experiment 1, follicles were assessed ultrasonically for 28 waves every 8 h. The number of examinations that encompassed the emergence of all growing 3-mm follicles was 10.0 +/-0.5 (mean +/-SEM; equivalent to 3.3 d) and extended from mean Examination -3.1 +/-0.3 to mean Examination 6.0 +/-0.6. A mean of 24 growing 3-mm follicles was found, and the maximal attained diameters were 4 mm (46%), 5 mm (25%), and >/=6 mm (29%). More (P<0.05) 3-mm follicles at Examinations -2 and -1 grew to >/=6 mm than to 4 or 5 mm, whereas more 3-mm follicles at Examinations 2 to 6 grew to only 4 mm. On average, the future dominant follicle appeared as a 3-mm follicle (Examination -2.1 +/-0.2) 6 and 10 h earlier (P<0.03) than for the largest (Examination -1.4 +/-0.3) and second-largest (Examination -0.8 +/-0.4) future subordinates, respectively. This result supported the hypothesis that the future dominant follicle has, on the average, an early developmental advantage. In Experiment 2 (n=33 waves), data were normalized to the day at the beginning of deviation (Day 2.8 +/-0.2) when the mean diameters of the dominant and largest subordinate follicle were 8.5 +/-0.2 mm and 7.2 +/-0.2 mm, respectively. This result suggests that the follicle selected to become dominant, as manifested by deviation, is the first follicle to develop to a decisive stage. In Experiment 3 (n=19 waves), FSH concentrations were lower (P<0.05) on the day at the beginning of deviation (8.5 +/-0.5 ng/ml) than on the day before (10.1 +/-0.8 ng/ml), with no continuing decrease after deviation. This temporal result suggests that the attainment of approximate basal levels of FSH is a component of the deviation mechanism.     

Follicle selection in cattle: role of luteinizing hormone

The circulating concentrations of LH were reduced by administration of 50 mg of progesterone every 8 h for 72 h, beginning when the largest follicle was 6.0 mm (experiment 1; n = 10). Progesterone treatment prevented the transient increase in LH that accompanied deviation (partitioning into dominant and subordinate categories) in control heifers (n = 10). The reduced LH concentrations were associated with reduced growth of the largest follicle, beginning a mean of 31 h after deviation, but did not alter the time of deviation or the growth and regression of the second-largest follicle. In experiment 2, 0 mg (controls) or 50 mg of progesterone was given every 8 h for three injections, beginning when the largest follicle was 7.0 mm (predeviation group) or 9.0 mm (postdeviation group; n = 8 for each of the four groups). Blood samples from the jugular vein and follicular-fluid samples from the two largest follicles were taken 8 h after the last treatment when the largest follicle was a mean of 8.7 mm in the predeviation group and 10.8 mm in the postdeviation group. In the controls, follicular-fluid concentrations of estradiol and free insulin-like growth factor (IGF)-1 in the largest follicle and IGF binding protein (IGFBP)-2 in the second-largest follicle were higher (P: < 0.05) in the postdeviation group than in the predeviation group. Progesterone treatment lowered (P: < 0.006) the circulating LH concentrations to a similar extent in both groups. In the predeviation group, progesterone treatment did not have a significant effect on any of the characteristics of the largest follicle. In the postdeviation group, the largest follicle of the progesterone-treated heifers had significant reductions in diameter and in follicular-fluid concentrations of estradiol and free IGF-1. Follicular-fluid concentrations of immunoreactive inhibin were not different for any of the comparisons. The results supported the hypothesis that LH has a positive effect on diameter of the largest follicle but not until after the beginning of diameter deviation. In addition, the results indicated that LH is involved in the production of estradiol by the largest follicle and that free IGF-1 concentrations increase in the largest follicle during deviation.     

Experimental assumption of dominance by a smaller follicle and associated hormonal changes in mares.

A two-follicle model was used to study the nature of selection of the dominant follicle in mares by ablating neither or one of the two follicles on the day the larger follicle reached >/= 20 mm (Day 0). The larger follicle became the dominant follicle in all mares in which both follicles (n = 8) or only the larger follicle (n = 10) was retained. When only the smaller follicle (n = 9) was retained, it became dominant and ovulated in six mares and became atretic in three mares; the difference in diameter between the two follicles on Day 0 was less (p < 0.01) in mares in which the retained smaller follicle grew and ovulated (2.2 +/- 0.6 mm) than in the mares in which the follicle became atretic (5.9 +/- 1.2 mm). A decline (p < 0. 0001) in FSH concentrations occurred over Days -4 (8.4 +/- 0.7 ng/ml) to 0 (5.9 +/- 0.3 ng/ml), averaged over all groups, and the decline continued for several more days in the groups with both follicles or with only the larger follicle retained. In the group with only the smaller follicle retained, compared to the group with both follicles retained, FSH concentrations and diameter of the smaller follicle increased between Days 0 and 1 (significant interaction for each end point). After Day 1, FSH concentrations continued to increase when the smaller retained follicle became atretic; concentrations decreased when the smaller retained follicle became dominant. An increase (p < 0.0001) in LH concentrations occurred over Days -4 (12.2 +/- 1.1 pg/ml) to 0 (21.1 +/- 2.0 pg/ml), averaged over the three groups. In 23 of 27 mares, a transient peak in LH concentrations occurred within 2 days of Day 0. In the groups with both follicles or with only the larger follicle retained, an increase (p < 0.0001) in systemic estradiol concentrations occurred between Day 0 (5.3 +/- 0.6 pg/ml) and Day 2 (7.5 +/- 0.4 pg/ml). When only the smaller follicle was retained, estradiol did not begin to increase until Day 2, and it increased only when the retained follicle grew and became dominant. The beginning of an increase in estradiol and continued decrease in FSH at the expected beginning of deviation were attributable to the future dominant follicle; there was no indication that the smaller follicle was involved.     

Systemic and intraovarian effects of dominant follicles on ovine follicular growth

The objective was to study the endocrine activity in sheep with large ovarian follicles and the effects of dominant follicles on other follicles, looking for possible intraovarian differences. Induction of dominant follicles was achieved using controlled exogenous LH pulses every 90 min over 14 days in eight Scottish Blackface ewes. During this period, follicular development was assessed by daily transrectal ultrasonography and jugular venous blood samples were collected every 12 h for FSH, LH inhibin and oestradiol assay. The exogenous LH pulses caused the appearance of large follicles in all the ewes, which reached a maximum mean diameter of 7.2 +/- 0.5 mm on Day 5.5 +/- 2.6 after first detection. In the presence of a dominant follicle, no other follicle grew to a diameter larger than 4 mm and there was a decrease in the number of new growing follicles (P < 0.05) and in the number of smaller follicles (P < 0.01). This effect of dominance was mediated by changes in FSH concentration, since FSH level decreased (P < 0.05) as dominant follicles grew and the decrease in FSH levels was related to a decline in the number of remaining follicles (P < 0.05). However, the greatest decrease in the number of small follicles growing to larger sizes was observed in the ovary ipsilateral to the dominant follicle (P < 0.05). These data confirm that the presence of a large follicle depresses the recruitment and growth of other follicles by systemic factors and provide some evidence of local inhibitors blocking the final development of other putative large follicles.     

Follicular and hormonal dynamics during the first follicular wave in heifers

A few days after the first follicular wave emerges as 4-mm follicles, follicular deviation occurs wherein 1 follicle of the wave continues to grow (dominant follicle) while the others regress. The objectives of this study were to characterize follicle growth and associated changes in systemic concentrations of gonadotropins and estradiol at 8-h intervals encompassing the time of follicle deviation. Blood samples from heifers (n = 11) were collected and the ovaries scanned by ultrasound every 8 h from 48 h before to 112 h after the maximal value for the preovulatory LH surge. The follicular wave emerged at 5.8 +/- 5.5 h (mean +/- SEM) after the LH surge, and at this time the future dominant follicle (4.2 +/- 0.8 mm) was larger (P < 0.001) than the future largest subordinate follicle (3.6 +/- 0.1 mm). There was no difference in growth rates between the 2 follicles from emergence to the beginning of the deviation (0.5 mm/8 h for each follicle), indicating that, on average, the future dominant follicle maintained a size advantage over the future subordinate follicle. Deviation occurred when the 2 largest follicles were 8.3 +/- 0.2 and 7.8 +/- 0.2 mm in diameter, at 61.0 +/- 3.7 h after wave emergence. Diameter deviation was manifested between 2 adjacent examinations at 8-h intervals. Mean concentrations of FSH decreased, while mean concentrations of LH increased 24 and 32 h before deviation, respectively, and remained constant (no significant differences) for several 8-h intervals encompassing deviation. In addition to the increase and decrease in circulating estradiol concentrations associated with the preovulatory LH surge, an increase (P < 0.05) occurred between the beginning of deviation and 32 h after deviation. The results supported the hypotheses that deviation occurs rapidly (within 8 h), that elevated systemic LH concentrations are present during deviation, and that deviation is not preceded by an increase in systemic estradiol.     

Follicular and hormonal response to experimental suppression of FSH during follicle deviation in cattle

A near steroid-free fraction of bovine follicular fluid was used to suppress FSH concentrations at the expected time of follicle deviation or when the largest follicle of Wave 1 reached > or = 8.0 mm (actual mean diameter, 8.4 mm; Hour 0). It was hypothesized that the low concentrations of FSH associated with deviation are inadequate for the smaller follicles but are needed for continued growth of the largest follicle. Control heifers (n=8) received 10 mL of saline, and treated heifers (n=16) received either 8.8 mL or 13.3 mL of the follicular-fluid fraction at Hours 0, 12, and 24. Between Hours -48 and 0, FSH concentrations decreased (P<0.05) and diameters of the 4 largest follicles increased (Hour effect, P<0.0001) similarly between groups. Concentrations of LH in the controls increased (P<0.05) between Hours -24 and -12 and decreased (P<0.05) between Hours 8 and 36, demonstrating a transient LH surge encompassing the expected beginning of deviation. In the treated group, a comparable increase in LH occurred before deviation but a decrease did not occur until after Hour 48. By Hour 4.5, the FSH concentrations in the treated group decreased (P<0.05) to below the concentrations in the controls. Suppressed diameter (P<0.001) of the largest follicle was detected at the first post-treatment examination (Hour 12; 7.5 h after FSH suppression) and was accompanied by reduced (P<0.04) systemic estradiol concentrations. The mean growth rates of the 3 smaller follicles in both the treated and control groups began to decrease at Hours -12 to 24 and were not different between groups during Hours 0 to 36. Concentrations of FSH in the treated group returned to control concentrations by Hour 24 (hour of last treatment). A rebound (P<0.05) in concentrations of FSH to >100% above control concentrations occurred by Hour 48 and was accompanied by resumed growth of the largest follicle in 75% of the heifers between Hours 48 and 72. The results demonstrated that the low concentrations of FSH associated with deviation can be further reduced by treatment with a nonsteroidal factor of follicular origin. Transient reduction of FSH concentrations to below the already low control concentrations inhibited the largest follicle but did not further inhibit the smaller follicles. These results support the hypothesis that the low FSH concentrations associated with follicle deviation are below the minimal requirements of the smaller or subordinate follicles but are needed for continued growth of the largest or dominant follicle in cattle.     

Selection of the dominant follicle in cattle: role of two-way functional coupling between follicle-stimulating hormone and the follicles

The functional coupling between the declining portion of the FSH surge and the growing follicles of a wave was studied by treating heifers with a minimal dose of estradiol to decrease FSH concentrations without an associated change in LH concentrations. Estradiol treatment when the largest follicle reached >/= 6.0 mm (Hour 0) resulted in depression of both FSH concentrations and diameter of the largest follicle by Hour 8. The smaller follicles were also inhibited. These results supported the hypothesis that FSH continues to be needed by the growing follicles even when the FSH concentrations are decreasing during the declining portion of the FSH surge. Estradiol treatment when the largest follicle was >/= 8.5 mm (expected time of follicular deviation) also resulted in a transient decrease in both FSH concentrations and diameter of the largest follicle, but the diameters of the smaller follicles were not affected. These results supported the hypothesis that the low concentrations of FSH at the expected time of deviation, although inadequate for the smaller follicles, were required for continued growth of the largest follicle. In another study, ablation (Hour 0) of the largest follicle was done at >/= 7.5 mm vs. >/= 8.5 mm. The mean FSH concentrations for the 8.5-mm groups were greater for the ablation group than for the control group at Hours 8 and 12, but there was no difference between the 7.5-mm groups at any hour. These results supported the hypothesis that by the time the largest follicle reaches the expected beginning of deviation it has developed a greater capacity for suppressing FSH. It is postulated that the essence of the selection of a dominant follicle is a close two-way functional coupling between changing FSH concentrations and follicular growth.     

The effect of recombinant bovine somatotropin on ovarian function in heifers: follicular populations and peripheral hormones.

The objective of this study was to investigate the possible effect of recombinant bovine somatotropin (BST) on ovarian folliculogenesis and ovulation rate. Twelve Hereford x Friesian heifers received daily injections of either 25 mg BST (6 heifers) or vehicle (6 heifers) for a period of two estrous cycles until slaughter. Blood samples were collected three times a week for measurements of peripheral growth hormone (GH), insulin-like growth factor I (IGF-I), FSH, LH, estradiol, and progesterone. Serial blood samples were also taken every 10 min for 8 h on Days 12 and 19 of the second estrous cycle to monitor GH, IGF-I, FSH, and LH profiles. At the end of treatment (Day 7 of the third estrous cycle), the heifers were killed and their ovaries were collected. Ovulation rate was determined by counting the number of fresh corpora lutea (CL). All antral follicles greater than or equal to 2 mm in diameter were dissected to assess antral follicle populations. Granulosa and thecal cells from the three largest follicles and CL from each heifer were collected for FSH and LH binding measurements. All heifers had a single ovulation. The treated heifers had significantly more antral follicles (60.2 +/- 6.7) than did the animals in the control group (33.2 +/- 3.2) (p less than 0.001). When follicles were grouped according to diameter, the mean numbers of follicles greater than 10 mm, 5-10 mm, and 2-5 mm in diameter were 0.8 +/- 0.2, 6.8 +/- 1.4, and 52.5 +/- 6.5 for the treated group, and 0.8 +/- 0.2, 6.5 +/- 1.0, and 25.8 +/- 2.7 for controls.(ABSTRACT TRUNCATED AT 250 WORDS)     

Supplementation of equine early spring transitional follicles with luteinizing hormone stimulates follicle growth but does not restore steroidogenic activity

This study was conducted to test the hypothesis that supplementation of growing follicles with LH during the early spring transitional period would promote the development of steroidogenically active, dominant follicles with the ability to respond to an ovulatory dose of hCG. Mares during early transition were randomly assigned to receive a subovulatory dose of equine LH (in the form of a purified equine pituitary fraction) or saline (transitional control; n = 7 mares per group) following ablation of all follicles >15 mm. Treatments were administered intravenously every 12 h from the day the largest follicle of the post-ablation wave reached 20 mm until a follicle reached >32 mm, when an ovulatory dose of hCG (3000 IU) was given. Saline-treated mares during June and July were used as ovulatory controls. In a preliminary study, injection of this pituitary fraction (eLH) to anestrus mares was followed by an increase in circulating levels of LH (P < 0.01) but not FSH (P > 0.6). Administration of eLH during early transition stimulated the growth of the dominant follicle (Group x Day, P < 0.00001), which attained diameters similar to the dominant follicle in ovulatory controls (P > 0.1). In contrast, eLH had no effect on the diameter of the largest subordinate follicle or the number of follicles >10 mm during treatment (P > 0.3). The numbers of mares that ovulated in response to hCG in transitional control, transitional eLH and ovulatory control groups (2 of 2, 3 of 5 and 7 of 7, respectively) were not significantly different (P > 0.1). However, after hCG-induced ovulation, all transitional mares returned to an anovulatory state. Circulating estradiol levels increased during the experimental period in ovulatory controls but not in transitional eLH or transitional control groups (Group x Day, P = 0.013). In addition, although progesterone levels increased after ovulation in transitional control and transitional eLH groups, levels in these two groups were lower than in the ovulatory control group after ovulation (Group, P = 0.045). In conclusion, although LH supplementation of early transitional waves beginning after the largest follicle reached 20 mm promoted growth of ovulatory-size follicles, these follicles were developmentally deficient as indicated by their reduced steroidogenic activity.     

Regulation of circulating gonadotropins by the negative effects of ovarian hormones in mares

The functional and temporal relationships between circulating gonadotropins and ovarian hormones in mares during Days 7-27 (ovulation = Day 0) was studied using control, follicle ablation, and ovariectomy groups (n = 6 mares/group). In the follicle-ablation group, all follicles > or = 6 mm were ablated on Day 7, and every 2 days thereafter, newly emerging follicles were also ablated. Estradiol concentrations decreased (P < 0.01) similarly in the controls and the follicle-ablation group between Days 7 and 11 and by Day 15 began to increase in the controls and continued to decrease in the follicle-ablation group. Concentrations of progesterone were not affected by follicle ablation, but diameter of the corpus luteum was greater (P < 0.05) by Day 21 in the follicle-ablation group; these results indicated that the follicles were involved in morphologic luteolysis, but not in functional luteolysis. Concentrations of LH were higher (P < 0.05) on Days 15 and 16 in the follicle-ablation group than in the controls, indicating an initial negative effect of follicles on LH. Immunoreactive inhibin and estradiol decreased (P < 0.0001) and FSH and LH increased (P < 0.05) within 1 or 2 days after ovariectomy; these changes occurred more slowly in the follicle-ablation group. The maximum value for an FSH surge in each control mare was below the lower 95% confidence limit in the ovariectomy group. Maximum concentration for the periovulatory LH surge in the controls was not different from the mean maximum LH concentrations in the ovariectomy group. Our interpretation is that the gonadotropin surges resulted from changes in the magnitude of the negative effects of ovarian hormones on the positive effects of extraovarian control. There was no indication of a positive ovarian effect on either FSH or LH.     

Suppression of circulating concentrations of FSH and LH by inhibin and estradiol during the initiation of follicle deviation in mares

The role of estradiol and inhibin in suppression of FSH and LH during the initiation of follicle deviation was examined in mares. In Experiment 1, the two largest follicles (F1, F2) were retained during a wave and the rest were ablated as they reached > or =10 mm. The largest follicle was left intact (control, n=12) or was ablated when it reached > or =20.0 mm (Day 0; expected beginning of deviation). The second largest follicle continued growing (n=9) or regressed (n=4) after F1 ablation. Circulating estradiol and total inhibin decreased after Day 0 in the F2-regressing group, whereas estradiol increased after Day 0.5 and inhibin was unaltered in the control and F2-growing groups. Circulating FSH decreased in the control group and increased in the F2-regressing group after Day 0. In the F2-growing group, FSH increased between Days 0 and 0.5 and then decreased. Circulating LH increased between Days 0 and 2 in the F2-regressing group and between Days 0 and 0.5 in the F2-growing group. In Experiment 2, 0 or 1 follicle was retained in a wave followed by administration of 0 or 1 mg of estradiol at the expected beginning of deviation (Hour 0; 2 x 2 factorial design, n=4-6/group). Circulating total inhibin was higher and FSH was lower at Hour 0 in the 1-follicle than in the 0-follicle groups. Follicle-stimulating hormone decreased after Hour 0 in the 1-mg but not in the 0-mg groups, and the decrease in the 0-follicle/1-mg group was not to the level of that in the 1-follicle/1-mg group. Circulating LH was not affected by follicle number but was reduced by estradiol. Results supported the hypotheses that F1 near the beginning of deviation produces inhibin and estradiol and that the increase in circulating estradiol at the beginning of deviation induces FSH suppression in combination with other follicle substances (presumably inhibin). Results also indicated that the increase in estradiol induces suppression of LH.     

Follicular deviation and acquisition of ovulatory capacity in bovine follicles.

Selection of dominant follicles in cattle is associated with a deviation in growth rate between the dominant and largest subordinate follicle of a wave (diameter deviation). To determine whether acquisition of ovulatory capacity is temporally associated with diameter deviation, cows were challenged with purified LH at known times after a GnRH-induced LH surge (experiment 1) or at known follicular diameters (experiments 2 and 3). A 4-mg dose of LH induced ovulation in all cows when the largest follicle was > or =12 mm (16 of 16), in 17% (1 of 6) when it was 11 mm, and no ovulation when it was < or =10 mm (0 of 19). To determine the effect of LH dose on ovulatory capacity, follicular dynamics were monitored every 12 h, and cows received either 4 or 24 mg of LH when the largest follicle first achieved 10 mm in diameter (experiment 2). The proportion of cows ovulating was greater (P < 0.05) for the 24-mg (9 of 13; 69.2%) compared with the 4-mg (1 of 13; 7.7%) LH dose. To determine the effect of a higher LH dose on follicles near diameter deviation, follicular dynamics were monitored every 8 h, and cows received 40 mg of LH when the largest follicle first achieved 7.0, 8.5, or 10.0 mm (experiment 3). No cows with a follicle of 7 mm (0 of 9) or 8.5 mm (0 of 9) ovulated, compared with 80% (8 of 10) of cows with 10-mm follicles. Thus, follicles acquired ovulatory capacity at about 10 mm, corresponding to about 1 day after the start of follicular deviation, but they required a greater LH dose to induce ovulation compared with larger follicles. We speculate that acquisition of ovulatory capacity may involve an increased expression of LH receptors on granulosa cells of the dominant follicle and that this change may also be important for further growth of the dominant follicle.     

Follicle diameters and hormone concentrations in the development of single versus double ovulations in mares

Relationships between double ovulations and plasma hormone concentrations were compared between 18 single ovulating and 6 double ovulating mares. The study began when the first follicle reached >or=30 mm, and ultrasound scanning and blood sampling were done every 12h to Day 3 (ovulation=Day 0). Data were analyzed for 2.5 d after the largest follicle was >or=30 mm and after Day -2.5 to encompass the mean 5-d interval between a >or=30 mm follicle and Day 0. During the 2.5 d after >or=30 mm, the increasing diameter of the largest follicle was less pronounced and plasma FSH concentrations were lower (approached significance) in the double ovulators than in the single ovulators. By Day -2.5, the largest follicle was smaller (P<0.01) and plasma FSH was lower (P<0.04) in the double ovulators. Plasma estradiol concentrations were higher (P<0.001) during the 2.5 d after >or=30 mm in the double ovulators and the correlation between estradiol and FSH was negative (r=-0.39, P<0.0001). In double ovulators, compared to single ovulators, the largest follicle was smaller, FSH was lower and estradiol was higher on most occasions between Days -2.5 and -0.5 (P<0.05), but plasma concentrations of LH and ir-inhibin were not significantly different. In conclusion, smaller preovulatory follicles in double ovulators were a response to lower FSH concentrations, due to higher estradiol concentrations from two preovulatory follicles; preovulatory differences in hormone concentrations between single and double ovulators were an effect rather than a cause of the double ovulations.