Charcoal-extracted Bovine Follicular Fluid Research Articles

Effect of “inhibin” from bovine testicular and follicular fluids on attainment of puberty in male rabbits

For this study, 15 young male rabbits of same breeds, with age range of 11-12 weeks old and weighing 1.0 ±0.2kg were used. These were randomly assigned to three treatments, each consisting of five rabbits per treatment. The treated groups(Treatments B and C) and the control group (Treatment A)were injected intramuscularly with charcoal-extracted bovine follicular fluid (FF), charcoal extracted bovine testicular fluid (TF) and charcoal treated distilled water (DW) respectively, at the rate of 0.2ml per rabbit on every other day and on three different occasions. Duration of the treatment lasted six days. Data were analyzed using descriptive statistics and ANOVA. After administration of the different treatments, body weight (BW), testes weight (TW) and histometric studies (HS) were conducted. Histometric studies included: seminiferous tubule diameter (STD), cellular elements (CE), stages of the cycle of seminiferous epithelium (CSE), determination of daily sperm production (DSP) and daily sperm production per gram testis (DSP/g). Figures obtained pertaining to their weights showed that they grew significantly over their initial weights with animals treated with TF showing highly significant ( p 0.05 ) in paired TW among the three different groups. STD of control group was significantly ( p 0.05 ) differences among the three different treatments in the frequency occurrence of stages 2 and 7 of the CSE, DSP and DSP/g testes. However, the frequency of occurrence of stages 1, 4 and 8 of the CSE differ significantly ( p<0.01 ) between control group and treated groups. There was a highly significant spermatozoa decrease ( p<0.01 ) in the treated groups when compared with control group and this supports the ability of the inhibin from bovine follicular and testicular fluids to inhibit the onset of active spermatogenesis.

Effects of bovine follicular fluid and passive immunization against gonadotropin-releasing hormone (GnRH) on messenger ribonucleic acid for GnRH receptor and gonadotropin subunits in ovariectomized ewes.

In cultured ovine pituitary cells, inhibin increases concentrations of mRNA encoding GnRH receptor and numbers of GnRH receptors. The objective of this study was to test the hypothesis that inhibin increases concentrations of ovine GnRH receptor mRNA in vivo. Ovariectomized ewes were used to eliminate effects of endogenous ovarian hormones, and passive immunization against GnRH was employed to avoid possible confounding influences of GnRH on GnRH receptor gene expression. Two groups of ewes (n = 5/group) were treated with 50 ml GnRH antiserum on Days 0 and 3 of the experiment. One group of immunized ewes received 10 ml charcoal-extracted bovine follicular fluid (bFF) as a source of inhibin every 8 h for 48 h on Days 4-6 of the experiment. A third group of ewes was not passively immunized and was treated only with bFF, and control ewes received no treatments. Anterior pituitary glands were collected from all ewes on Day 6. Passive immunization against GnRH, alone or in combination with treatment with bFF, decreased mean concentrations of LH (p < 0.01) and LH pulse amplitude (p < 0.001). In ewes treated only with GnRH antiserum, number of LH pulses was also reduced (p < 0.03). Circulating concentrations of FSH tended to be lower (p = 0.06) in passively immunized ewes compared to controls. Treatment with bFF, alone or in combination with GnRH antiserum, reduced circulating concentrations of FSH (p < 0.02) and amounts of FSHbeta subunit mRNA (p < 0.001) to less than 30% and 10% of control values, respectively. Despite effects of bFF on concentrations of FSHbeta mRNA and secretion of FSH, concentrations of GnRH receptor mRNA were similar among controls, ewes treated with bFF alone, and passively immunized ewes treated with bFF. Passive immunization against GnRH did not affect concentrations of GnRH receptor mRNA but resulted in a reduction (p < 0.05) in amount of LHbeta mRNA. Treatment with bFF did not affect amounts of either alpha subunit or LHbeta subunit mRNA except when combined with treatment with antiserum, when amounts of both alpha and LHbeta subunit mRNA were reduced (p < 0.05). These results do not support the hypothesis that inhibin increases concentrations of GnRH receptor mRNA in the ewe, and they provide evidence that inhibin is not an acute regulator of ovine GnRH receptor gene expression in vivo.

Impairment in Follicle-Stimulating Hormone Secretion by Different Treatments with Follicular Fluid as a Source of Inhibin Blocks the Ability of Pulsatile Luteinizing Hormone to Induce Preovulatory Surges of Gonadotropins in Prepubertal Ewe Lambs1

This study was conducted to investigate whether FSH can affect the ability of pulsatile exogenous LH to induce a preovulatory surge of gonadotropins in prepubertal ewe lambs. March-born ewe lambs were randomly assigned in November to the following treatments: 1) control; 2) 1.5 ml charcoal-extracted bovine follicular fluid (bFF) for 240 h; 3) 1.5 ml bFF for 72 h; and 4) 3 ml bFF for 72 h. Follicular fluid was administered as a source of inhibin to reduce secretion of FSH. Starting at 0 h, all animals were given (i.v.) 10 micrograms ovine LH every h for a 48-h period. Lambs in the control group did not receive further treatment. Females in group 2 were administered (s.c.) 1.5 ml bFF every 8 h from -184 h to 56 h. In groups 3 and 4, lambs received (s.c.) 1.5 ml bFF, respectively, every 8 h from -16 h to 56 h. Preovulatory surges of LH and FSH occurred concomitantly in 4 of 6 control lambs at a mean time of 34 h (range 20-42 h). In contrast, none of the females in groups 2, 3, or 4 showed a preovulatory surge of gonadotropins in response to pulsatile LH (p < 0.05). At -16 h, FSH was reduced in the ewe lambs of group 2 (p < 0.01). Thereafter, mean concentrations and area under the curve for FSH were lower (p < 0.001), and variability in the pattern of secretion was reduced (p < 0.05), in lambs of groups 2, 3, and 4 as compared to controls. LH, apart from the preovulatory surge, was not affected. Females in groups 2, 3, and 4 secreted less estradiol than controls (p < 0.01). Mean concentrations and area under the curve for estradiol during the 20 h preceding preovulatory surges (20 h = earliest surge) in controls were greater than those during the 20 h preceding 34 h (34 h = mean time for surges) in lambs that did not show a surge (p < 0.001). The induced luteal phases were of short duration, and no animal continued to cycle. Ovarian follicular development was evident in the majority (8 of 10) of the randomly selected lambs examined by laparoscopy during the expected midluteal phase. In summary, long- and short-term administration of bFF resulted in reduced concentrations of FSH, preventing the ability of pulsatile LH to induce a preovulatory surge of gonadotropins in prepubertal ewe lambs. The decrease in estradiol associated with the bFF-induced reduction in FSH may have been the mechanism responsible for the lack of LH and FSH surges. It is proposed that unimpaired secretion of FSH is required for treatment with LH to induce preovulatory surges of gonadotropins in this model.

Effects of bovine follicular fluid and luteinizing-hormone-releasing hormone antagonist on the dominant follicle maintained by a norgestomet ear implant in heifers

The effects of BFF and LHRHa on the dominant follicle, maintained by 9-d synthetic progestin (norgestomet) ear implants inserted at proestrus, were studied in cattle. From day 4 of implant insertion, heifers (n = 18) were injected twice daily with saline (control, n = 6) or charcoal-extracted bovine follicular fluid, 10 mL i.v. for 4 d (BFF, n = 6) or LHRHa, 0.8 mg i.v. for 3 d (LHRHa, n = 6). Follicular changes were monitored daily by ultrasonography. Plasma estradiol-17β, serum LH and FSH were measured by radioimmunoassays. The dominant follicle was maintained during the treatment period, and it ovulated after implant removal in control heifers. In BFF group, the dominant follicle either ovulated after implant removal (two of six heifers) or gradually regressed until day 10 in the post-implant period (other four of six heifers). In the LHRHa group, the dominant follicle regressed during or after the implant period in 5 of 6 heifers. The dominant follicle ovulated in 1 of 6 heifers. Growth rate of the dominant follicle, until it reached its maximum diameter, was lower in BFF group than the control group (0.4 ± 0.1 vs 0.7 ± 0.1 mm d−1; P &lt; 0.01). Both BFF and LHRHa groups showed high regression rates in those follicles that did regress (2.2 ± 0.1 (n = 4) and 1.9 ± 0.8 mm d−1 (n = 5) respectively; P &lt; 0.01), compared with no regression in the control group. The BFF injections failed to suppress serum FSH levels consistently in all the heifers treated. LHRHa-treated heifers showed a significant decline (P &lt; 0.01) in the mean serum LH concentration. In conclusion, deprivation of LH causes atresia of the dominant follicle. Key words: Follicular fluid, LHRH antagonist, follicular dynamics, gonadotropins, estradiol-17β, ultrasound, regression

The efficiency of different gonadotrophin preparations on the superovulatory responses of holstein cows

This study examined the effects of altered serum FSH concentration on subsequent ovarian response to superovulation. Synchronized heifers were assigned randomly on Day 1 of the cycle (estrus = Day 0) to three pretreatment groups that consisted of 6-d of saline (7ml, s.c., b.i.d.; Group I), FSH-P (0.5 mg, i.m., b.i.d.; Group II) or charcoal-extracted bovine follicular fluid (BFF; 7 ml, s.c., b.i.d.; Group III) injections. Superovulation was initiated on Day 7 and consisted of FSH-P in decreasing dosages over 4 d (4,3,2,1 mg; i.m., b.i.d.), with cloprostenol (500 μg) on the morning of the third day. A second replicate with 14 heifers was conducted using the same protocol but twice the pretreatment dosage of FSH-P (1 mg) and BFF (14 ml). Endogenous plasma FSH decreased during BFF and FSH-P pretreatments compared to controls (P < 0.02). Endogenous FSH concentrations in both primed groups (II and III) were similar to control values (Group I) 12 h after the start of superovulation. Basal LH concentrations were not different between pretreatment groups. The interval from cloprostenol treatment to the preovulatory LH surge in Group III was 21.3 and 23.9 h longer (P < 0.0001) than it was in Groups I and II. The postovulation progesterone rise was delayed in Group III. The number of corpora lutea (CL) was lowest in the BFF-primed group (4.2 ± 0.8) compared with the FSH-primed (7.4 ± 1.3) and the control (12.0 ± 1.8; P < 0.003) groups. In the FSH-primed group (0.68 ± 0.06 cm3), CL volumes were larger than in the control group (0.45 ± 0.03 cm3), whereas in the BFF-primed group (0.27 ± 0.02 cm3) CL volumes were smaller compared with the control group (P < 0.0001). Mean FSH concentrations for 48 h preceding superovulation and the number of CL per cow were positively correlated (r = 0.55; P < 0.004; n = 26). We concluded that both FSH-P and BFF pretreatments decreased the superovulatory response of heifers to FSH-P. The mechanism for this would appear to be associated with reduced endogenous FSH prior to the start of superovulation.

Ovarian follicular development and endocrine responses in follicular-fluid-treated and hemi-ovariectomized heifers.

The effects of charcoal-extracted bovine follicular fluid (BFF) on endocrine profiles and follicular development in intact and hemiovariectomized postpubertal heifers were examined. Oestrus-synchronized heifers received Norgestomet implants on day 1 and 7 of treatment and were then injected s.c. with 11 ml saline (control) or 11 ml BFF twice a day for 12 days. The ovary bearing the largest follicle (OV1) was removed on day 7 and the remaining ovary (OV2) was collected on day 13. Follicles were observed by daily ultrasonography and were classified according to diameter (size 1: 2-3 mm; size 2: 4-6 mm; size 3: 7-10 mm; size 4: > 10 mm). After ovariectomy they were classified by diameter and histologically as normal or atretic. Intact control heifers had increased numbers of size 4 follicles on OV1 on days 6 and 7; no increase was observed in BFF-treated heifers (P < 0.03). In BFF-treated heifers, the mean basal LH concentration was higher (P < 0.05) and that of FSH was lower (P < 0.04) than in controls. FSH concentrations in BFF-treated heifers decreased from 0.60 +/- 0.08 ng ml-1 (day 1) to 0.22 +/- 0.05 ng ml-1 (day 7; P < 0.04). The concentration of oestradiol increased in control heifers, but not in BFF-treated heifers (P < 0.001). After hemicastration, OV2 underwent compensatory hypertrophy in control heifers, with an increase in the number of size 2, 3 and 4 follicles (P < 0.05), whereas BFF-treated heifers did not. Thus, total follicular volume was much lower in BFF-treated than in control heifers on day 13 (92.2 +/- 15.4 versus 1393.8 +/- 276.6 mm3; P < 0.0002). A transient increase in FSH (P < 0.006) and oestradiol (P < 0.01) concentrations occurred after hemiovariectomy in control but not in BFF-treated animals. In control heifers, an analysis of temporal relationships showed negative correlations between the volume of size 3 and size 4 follicles, and between FSH concentrations and the volume of size 3 and 4 follicles. A positive correlation was found between the mean diameter of the largest follicle and the concentration of oestradiol, whereas negative relationships were found between the concentrations of FSH and oestradiol, and between FSH and the mean diameter of the largest follicle. Analysis of the histological data showed that the number and volume of follicles > 8.57 mm was lower in the BFF-treated OV1 ovary, whereas no differences were found for follicles < or = 8.57 mm.(ABSTRACT TRUNCATED AT 250 WORDS)

Development of a persistent ovarian follicle influences response to superovulation in heifers

This study examined the effects of altered serum FSH concentration on subsequent ovarian response to superovulation. Synchronized heifers were assigned randomly on Day 1 of the cycle (estrus = Day 0) to three pretreatment groups that consisted of 6-d of saline (7ml, s.c., b.i.d.; Group I), FSH-P (0.5 mg, i.m., b.i.d.; Group II) or charcoal-extracted bovine follicular fluid (BFF; 7 ml, s.c., b.i.d.; Group III) injections. Superovulation was initiated on Day 7 and consisted of FSH-P in decreasing dosages over 4 d (4,3,2,1 mg; i.m., b.i.d.), with cloprostenol (500 μg) on the morning of the third day. A second replicate with 14 heifers was conducted using the same protocol but twice the pretreatment dosage of FSH-P (1 mg) and BFF (14 ml). Endogenous plasma FSH decreased during BFF and FSH-P pretreatments compared to controls (P < 0.02). Endogenous FSH concentrations in both primed groups (II and III) were similar to control values (Group I) 12 h after the start of superovulation. Basal LH concentrations were not different between pretreatment groups. The interval from cloprostenol treatment to the preovulatory LH surge in Group III was 21.3 and 23.9 h longer (P < 0.0001) than it was in Groups I and II. The postovulation progesterone rise was delayed in Group III. The number of corpora lutea (CL) was lowest in the BFF-primed group (4.2 ± 0.8) compared with the FSH-primed (7.4 ± 1.3) and the control (12.0 ± 1.8; P < 0.003) groups. In the FSH-primed group (0.68 ± 0.06 cm3), CL volumes were larger than in the control group (0.45 ± 0.03 cm3), whereas in the BFF-primed group (0.27 ± 0.02 cm3) CL volumes were smaller compared with the control group (P < 0.0001). Mean FSH concentrations for 48 h preceding superovulation and the number of CL per cow were positively correlated (r = 0.55; P < 0.004; n = 26). We concluded that both FSH-P and BFF pretreatments decreased the superovulatory response of heifers to FSH-P. The mechanism for this would appear to be associated with reduced endogenous FSH prior to the start of superovulation.

Effect of bovine follicular fluid on gonadotropin secretion and ovulation rate in the cycling ewe with subsequent passive immunization of prepuberal mice with bovine follicular fluid antisera

The response to administration of charcoal-extracted bovine follicular fluid on gonadotropin secretion, ovulation rate, and antibody formation in the cycling ewe was investigated. Twelve ewes were randomly assigned to three treatment groups of four ewes each, and received injections twice daily (10 ml, intradermal) from Day 1 to Day 11 (the first day of estrus, following synchronization, was designated Day 0) (with Freund's complete adjuvant on Days 3, 6 and 9). Group 1 received whole charcoal-extracted bovine follicular fluid (bFF), Group 2 received heat-inactivated charcoal-extracted bovine follicular fluid (IFF), and Group 3 received saline (SAL). Follicle stimulating hormone (FSH) and luteinizing hormone (LH) were assayed from serum on Days 1–17. Follicle stimulating hormone levels were significantly reduced from Day 2 to Day 12, as compared with controls, in both the bFF and IFF groups and, from the bFF-treated group only, a post-treatment rebound in FSH levels was observed between Days 13 and 14. While LH levels in bFF- and IFF-treated animals were significantly lower than in controls on Day 3, overall LH levels were not substantially affected by these treatments. Laparoscopies performed after Day 21 showed a trend toward increased ovulation rates for bFF-treated animals and significantly increased ovulation rates for IFF-treated animals as compared with controls. The presence of bFF-reactive antibodies was detected from both bFF- and IFF-treated animals using an immunodiffusion assay after Day 17, and again after the subsequent estrous cycle. To further investigate possible effects of endogenous antibody production on ovulation rate, immature female mice were given a single, 0.1 ml intraperitoneal injection of antiserum collected from bFF-treated ewes. This treatment resulted in significantly higher ovulation rates compared with controls given normal ovine serum. These results indicate that treatment of ewes with bFF initially suppresses FSH levels and results in a post-treatment rebound to higher than normal FSH levels. Moreover, these findings also indicate that whole bFF or bFF antisera (produced in the ewe), will increase ovulation rate. Under these conditions, the inhibin control on FSH secretion may have been relieved, thus resulting in an elevated ovulatory rate.

Effects of suppressing plasma FSH on ovarian follicular dominance in cattle.

The importance of FSH during development of the dominant follicle of the first wave of follicular growth in cattle was assessed by injecting heifers twice daily with 20 ml saline (n = 6) or 20 ml charcoal-extracted bovine follicular fluid (bFF; n = 10), containing inhibin, on days 6 and 7 of the oestrous cycle. All animals received a luteolytic dose of PGF2 alpha on day 8. The interval to oestrus after PGF2 alpha in controls was 2.6 +/- 0.2 days. Five of ten bFF-treated heifers exhibited oestrus at the same time as controls (2.7 +/- 0.1 days), whereas time to oestrus in the remaining five bFF-treated heifers was significantly longer (6.8 +/- 0.6 days; P < 0.05). Treatment with bFF suppressed plasma FSH during the 48 h injection period (P < 0.05). Concentrations of FSH were not different in bFF-treated animals that did not display delayed oestrus compared with those in animals that exhibited delayed oestrus. In controls, the dominant follicle of the first wave continued growing during treatment and ovulated after injection of PGF2 alpha. In all ten bFF-treated animals, growth of the dominant follicle of the first wave was arrested during the treatment period. In bFF-treated animals that did not exhibit delayed oestrus, the dominant follicle resumed growth after luteolysis and ovulated. In bFF-treated animals that displayed delayed oestrus, the dominant follicle regressed after luteolysis and the ovulatory follicle was selected from a newly recruited (second) wave of follicular growth.(ABSTRACT TRUNCATED AT 250 WORDS)

Effects of bovine follicular fluid and partially purified bovine inhibin on FSH and LH release by bovine pituitary cells in culture

We have established a dispersed bovine pituitary cell culture system to study the effects of charcoal-extracted bovine follicular fluid (BFF) or bovine inhibin, partially purified by immunoaffinity chromatography (IPI), on the spontaneous release of follicle-stimulating hormone (FSH) and luteinizing hormone (LH). Pituitary cells were plated at 0.25, 0.5 or 1 x 10(6) viable cells/well (c/w) and incubated for 48 h. The medium was replaced and BFF (0, 0.54, 2.7, 13.7, 68.7 or 343.5 micrograms protein) or IPI (0, 0.01, 0.06, 0.29, 1.45 or 7.25 micrograms protein) added to the cultures and the incubation was continued for 48 h. Concentrations of FSH and LH in spent medium were determined by RIA and data analyzed by ANOVA with means compared by Student-Neuman-Keuls (SNK) test. We have shown an increase in spontaneous FSH and LH release attributable to both number of bovine pituitary cells plated and to the length of incubation. The addition of BFF reduced spontaneous FSH release over 48 h incubation. The dose-dependent inhibition curves observed in culture in which different numbers of cells were plated, indicates that inhibition was greater when 1 x 10(6) c/w were plated compared to 0.25 or 0.5 x 10(6) c/w. Bovine follicular fluid at 0.45 micrograms of protein (equivalent to 0.01 microliters of BFF) incubated with 1 x 10(6) c/w, suppressed FSH release by 10.6% compare to control. Maximal suppression of 34.1% was obtained with 50 micrograms (equivalent to 1.56 microliters of BFF). Immunopurified bovine inhibin at 1.45 micrograms tended to suppress FSH release and at 7.25 micrograms significantly reduced FSH release. Neither BFF nor IPI had a measurable effect on LH release. We conclude that BFF and IPI suppress the spontaneous release of FSH from bovine pituitary cells in culture in a dose-dependent manner, without concomitant suppression of LH release.

Suppression of the secondary FSH surge with bovine follicular fluid is associated with delayed ovarian follicular development in heifers

Holstein heifers were given 5 injections (twice/day) of 10 ml charcoal-extracted bovine follicular fluid (bFF; N = 6) or 10 ml saline (N = 5) beginning 12 h after the onset of oestrus. Blood samples were collected for determination of plasma concentrations of FSH, LH, progesterone and oestradiol-17 beta. Treatment with bFF suppressed the secondary FSH surge (P less than 0.01). Cessation of bFF injections was followed by a rebound period during which FSH was elevated compared with controls (P less than 0.01). Daily ultrasonographic examinations revealed that follicular growth occurred in waves, with 4 of 5 control heifers exhibiting 3 waves and the other 2 waves. In contrast, 5 of 6 bFF-treated animals exhibited 2 waves and the other 3 waves. Appearance of follicles in the first wave was delayed in bFF-treated heifers (Day 3.3 +/- 0.3 compared with Day 1.4 +/- 0.2; P less than 0.0001) and appearance of the dominant follicle of the first wave was delayed (Day 4.5 +/- 0.3 compared with Day 1.8 +/- 0.2; P less than 0.0001). Follicles in the second wave appeared later in animals treated with bFF (Day 12.7 +/- 0.4 compared with Day 10.4 +/- 0.6; P less than 0.01), and the dominant follicle of this wave also appeared later (Day 13.0 +/- 0.5 compared with Day 10.6 +/- 0.5; P less than 0.01). Oestradiol-17 beta increased during the early luteal phase, but this increase occurred later in heifers treated with bFF (peak concentrations on Day 6.3 +/- 0.6 compared with Day 4.2 +/- 0.2; P less than 0.05). LH, progesterone and cycle length were not affected by bFF. Delayed follicular growth associated with suppression of FSH suggests that the secondary FSH surge is important in the initiation of follicular development early in the bovine oestrous cycle, and thus may play a role in the regulation of ovarian follicular dynamics.

Endocrine and superovulatory responses in heifers pretreated with FSH or bovine follicular fluid

This study examined the effects of altered serum FSH concentration on subsequent ovarian response to superovulation. Synchronized heifers were assigned randomly on Day 1 of the cycle (estrus = Day 0) to three pretreatment groups that consisted of 6-d of saline (7ml, s.c., b.i.d.; Group I), FSH-P (0.5 mg, i.m., b.i.d.; Group II) or charcoal-extracted bovine follicular fluid (BFF; 7 ml, s.c., b.i.d.; Group III) injections. Superovulation was initiated on Day 7 and consisted of FSH-P in decreasing dosages over 4 d (4,3,2,1 mg; i.m., b.i.d.), with cloprostenol (500 μg) on the morning of the third day. A second replicate with 14 heifers was conducted using the same protocol but twice the pretreatment dosage of FSH-P (1 mg) and BFF (14 ml). Endogenous plasma FSH decreased during BFF and FSH-P pretreatments compared to controls (P < 0.02). Endogenous FSH concentrations in both primed groups (II and III) were similar to control values (Group I) 12 h after the start of superovulation. Basal LH concentrations were not different between pretreatment groups. The interval from cloprostenol treatment to the preovulatory LH surge in Group III was 21.3 and 23.9 h longer (P < 0.0001) than it was in Groups I and II. The postovulation progesterone rise was delayed in Group III. The number of corpora lutea (CL) was lowest in the BFF-primed group (4.2 ± 0.8) compared with the FSH-primed (7.4 ± 1.3) and the control (12.0 ± 1.8; P < 0.003) groups. In the FSH-primed group (0.68 ± 0.06 cm 3), CL volumes were larger than in the control group (0.45 ± 0.03 cm 3), whereas in the BFF-primed group (0.27 ± 0.02 cm 3) CL volumes were smaller compared with the control group (P < 0.0001). Mean FSH concentrations for 48 h preceding superovulation and the number of CL per cow were positively correlated (r = 0.55; P < 0.004; n = 26). We concluded that both FSH-P and BFF pretreatments decreased the superovulatory response of heifers to FSH-P. The mechanism for this would appear to be associated with reduced endogenous FSH prior to the start of superovulation.

Effects of Bovine Follicular Fluid on Pulsatile Secretion of Gonadotrophin‐Releasing Hormone and Gonadotrophins in Ovariectomized Ewes

Abstract Ovariectomized ewes were injected with charcoal-extracted bovine follicular fluid (n = 5) or with hypophysectomized ewe serum (n = 5) after which hypophyseal portal blood samples were taken to monitor the release of gonadotrophin-releasing hormone (GnRH). Peripheral blood samples were also taken to monitor plasma luteinizing hormone and follicle-stimulating hormone (FSH) concentrations. Bovine follicular fluid treatment caused a 50% decrease in plasma FSH concentrations whereas hypophysectomized ewe serum did not significantly alter plasma FSH levels. The frequency and amplitude of luteinizing hormone and GnRH pulses was similar in both experimental groups. It is concluded that inhibin activity in bovine follicular fluid selectively suppresses the release of FSH by direct action on the pituitary gland, with no effect on GnRH secretion.

Depression of follicle stimulating hormone in bulls injected with follicular fluid

Eight bulls were divided into two groups and injected with either charcoal-extracted steer blood serum or charcoal-extracted bovine follicular fluid (bFF). Ten-milliliter injections were given subcutaneous every 12 h for 4 wk. Jugular blood collected before, during and after the injection period was analyzed for follicle-stimulating hormone (FSH) and luteinizing hormone (LH) by radioimmunoassay. All bulls were exposed to restrained, estrual heifers for 15 min every 2 wk for 16 wk starting 4 wk before the first injection. The number of mounts and services by each bull was recorded. Semen was collected with an artificial vagina and evaluated on alternate weeks during the same period. The concentration of FSH in serum decreased (P < 0.05) by 12 h after the first injection and remained 61% lower than that of serum-injected bulls during the injection period. The concentration of FSH increased (P < 0.05) by 3 d after the last injection. Injections of bFF did not affect the concentration of LH in serum. Bovine follicular fluid injections significantly depressed FSH; however, libido, serving capacity, and semen characteristics were unchanged.

Follicle Stimulating Hormone Pattern and Luteal Function in Ewes Receiving Bovine Follicular Fluid during Three Stages of the Estrous Cycle2

The objectives of this study were to determine 1) the ability of charcoal-extracted bovine follicular fluid (bFF) to suppress endogenous follicle stimulating hormone (FSH) at various stages of the estrous cycle and 2) the effects of suppression of FSH on luteal function and lengths of the current and subsequent estrous cycles. Twenty-six mature ewes were assigned randomly to receive 5 ml of either bFF or saline, subcutaneously, at 8-h intervals on d 1 through 5 (bFF n = 6; saline n = 3), d 6 through 10 (bFF n = 6; saline n = 3) or d 11 through 15 (bFF n = 6; saline n = 2) of the estrous cycle (d 0 = estrus). Blood was collected daily beginning at estrus and continued until the third estrus (two estrous cycles) or 40 d; more frequent samples were collected 2 h prior to initiation of treatment (0600), hourly for the first 8 h of treatment, then every 4 h until 0800 on the first day after treatment, and finally at 1600 and 2400 on that day. Plasma concentrations of FSH were lower (P less than .001) in bFF-treated than in saline-treated ewes. Treatment with bFF reduced (P less than .05) plasma concentrations of progesterone during the current but not during the subsequent estrous cycle. Treatment with bFF did not affect plasma concentrations of estradiol-17 beta. Administration of bFF on d 11 through 15 of the estrous cycle lengthened the interval from the decline in progesterone to estrus and the inter-estrous interval by approximately 3 and 4 d, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)

The roles of inhibin and gonadotrophin-releasing hormone in the control of gonadotrophin secretion in the ewe.

The respective roles and relative importance of ovarian inhibition and hypothalamic stimulation in the differential control of the secretion of FSH and LH were studied in the ewe. In the first experiment two groups of ten intact ewes were injected i.v. twice daily with 9 ml charcoal-extracted bovine follicular fluid (bFF), a preparation rich in inhibin (3.65 ku./ml), throughout the luteal phase of the oestrous cycle. Compared with the control ewes, this treatment significantly reduced pituitary and plasma FSH concentrations and increased the frequency and amplitude of the LH pulses, but did not affect pituitary LH concentrations. In a second experiment, five control and five bFF-treated ewes from experiment 1 were ovariectomized and the injection regime was altered to 2.5 ml s.c. every 8 h. This treatment was maintained for 21 days. In control ewes, plasma FSH concentrations rose significantly within 12 h and continued to rise for 3-4 days. Treatment with bFF abolished this increase and maintained plasma FSH concentrations below those observed in intact ewes. The rise in mean plasma LH concentrations evoked by ovariectomy was also partially inhibited in the bFF-treated ewes. The response to the gonadotrophin-releasing hormone (GnRH) agonist buserelin (5 micrograms i.v.) was measured 6, 12 and 18 days after ovariectomy. In control ewes the agonist consistently evoked large surges of both hormones but in bFF-treated ewes the FSH response was completely blocked and the initial phase of the LH response (the first 'pool') was greatly reduced. In experiment 3, six ewes were ovariectomized and passively immunized against GnRH 3 days after oestrus. The increase in plasma LH which normally follows ovariectomy was completely abolished and mean concentrations remained very low and did not change over the following 14 days. In contrast, mean FSH concentrations rose significantly within 12 h of ovariectomy and continued to rise until the third day, after which they fell gradually. Treating three of the ewes with bFF (2.5 ml s.c. every 8 h) 8 days after ovariectomy and immunization further reduced the FSH concentrations. When the ewes were injected repeatedly (200 ng i.v., hourly for 5 h) with [D-penicillamine-(But)6]-GnRH(1-9)nonapeptide-ethylamide, a synthetic GnRH analogue which does not bind to the antiserum, there was a rapid rise in the secretion of LH in both control and bFF-treated animals but, as with the responses to buserelin, the initial response was significantly lower in bFF-treated than in control ewes.(ABSTRACT TRUNCATED AT 400 WORDS)

Treatment of ovariectomized ewes with bovine follicular fluid decreases secretion of FSH without changing secretion of LH

Ovariectomized ewes were injected with 0, 0.25, 0.5, 1.0 or 2.0 ml of charcoal-extracted bovine follicular fluid. Treating ewes with 2 ml of follicular fluid resulted in a decrease in circulating concentrations of FSH to 72.8% of the pretreatment value. With smaller doses of follicular fluid, the magnitude of the decrease was less. Concentrations of LH did not change significantly. Pretreatment of ovariectomized ewes with estradiol and/or progestogen did not alter the magnitude of the FSH decrease. This action of follicular fluid extract fits the effect of the non-steroidal substance known as inhibin or folliculostatin.

Effects of charcoal-extracted, bovine follicular fluid on gonadotropin concentrations, the onset of estrus and luteal function in heifers.

A study was conducted to determine the effect of charcoal-extracted, bovine follicular fluid (CFF) on plasma follicle stimulating hormone (FSH) and luteinizing hormone (LH) concentrations, the interval from luteolysis to estrus, and subsequent luteal function in heifers. Fifteen Angus, Simmental and Hereford heifers were allotted by age, weight and breed to a control (C, n = 8) or a CFF (n = 7) group. Heifers received injections of saline or CFF (iv, 8 ml/injection) every 12 h from d 1 (d 0 estrus) through d 5 of the estrous cycle. On d 6, each heifer was injected (im) with 25 mg of prostaglandin F2 alpha (PGF2 alpha). Blood samples were collected every 12 h by venipuncture starting just before the first saline or CFF injection and continuing until estrus. Thereafter, blood samples were collected every other day during the subsequent estrous cycle and assayed for FSH, LH, estradiol-17 beta and progesterone by radioimmunoassay. Injections of CFF had no effect (P greater than .05) on circulating FSH or LH concentrations from d 1 to 5 relative to the C group; however, there was a transient rise (P less than .05) in FSH concentrations 24 h following cessation of CFF injections. This transient rise in FSH was not immediately followed by an increase in plasma estradiol-17 beta concentrations. Although CFF injections did not interfere with PGF2 alpha-induced luteolysis, the interval from PGF2 alpha injection to estrus was delayed (P less than .05) by 5 d in the CFF group compared with the C group.(ABSTRACT TRUNCATED AT 250 WORDS)

Effect of unilateral ovariectomy and injection of bovine follicular fluid on gonadotropin secretion and compensatory ovarian hypertrophy in prepuberal heifers.

A study was conducted to determine whether compensatory ovarian hypertrophy occurred in prepuberal heifers, and if so, could compensatory ovarian hypertrophy be prevented by injecting charcoal-extracted bovine follicular fluid (CFF). An additional objective was to determine the effect of CFF injections on follicle stimulating hormone (FSH) and luteinizing hormone (LH) concentrations following unilateral ovariectomy (ULO). Thirty-one prepuberal Angus, Hereford, and Angus X Hereford heifers were assigned by weight and breed to the following groups: Control-saline injections (C), unilateral ovariectomy-saline injections (ULO) and unilateral ovariectomy-CFF injections (ULO-CFF). In group C, a sham operation was performed on d 0 and both ovaries were removed on d 7. In groups ULO and ULO-CFF, either the right or left ovary was removed on d 0 and the remaining ovary was removed on d 7. From d 0 to 7, each heifer received an injection (iv, 7 ml/injection) of saline or CFF every 12 h. Jugular blood samples were collected immediately before each saline injection from d 0 to 7 and analyzed for FSH and LH. Unilateral ovariectomy increased ovarian (P less than .08) and follicular fluid (P less than .05) weights compared with groups C and ULO-CFF; however, lyophilized ovarian weight was similar (P greater than .05) for all groups. Injections of CFF blocked the increase in ovarian and follicular fluid weight following ULO. The total number of visible follicles per ovary was similar for all groups; however in the ULO group there was an increase (P less than .01) in those 5 to 6 mm and greater than or equal to 9 mm.(ABSTRACT TRUNCATED AT 250 WORDS)