Effects of dihydrotestosterone administration with and without estradiol pretreatment on gonadotropin secretion in ovariectomized pony mares

Twenty ovariectomized pony mares were used to determine if dihydrotestosterone propionate (DHTP) administration, with or without estradiol benzoate (EB) pretreatment, would have the same effects on follicle stimulating hormone (FSH) and luteinizing hormone (LH) secretion as testosterone propionate (TP) administration. All mares were given an initial injection of gonadotropin releasing hormone (GnRH) to characterize their LH and FSH response, and then two groups of mares (n = 4/group) were administered EB (22 micrograms/kg of body weight), two groups were administered vehicle (safflower oil) and a fifth group was administered TP (175 micrograms/kg of body weight) daily for 10 days. Following a second injection of GnRH, one group of EB-treated mares and one group of oil-treated mares were administered DHTP (175 micrograms/kg of body weight) daily for 10 days; the other EB- and oil-treated mares were administered oil and the TP-treated mares were continued on the same dose of TP for 10 days. A final injection of GnRH was then given. Treatment with EB increased (P less than .01) concentrations of LH in daily blood samples and increased (P less than .05) the LH response to exogenous GnRH. Administration of TP or DHTP reduced (P less than .05) both daily LH concentrations and the LH response to exogenous GnRH. Concentrations of FSH in daily blood samples were reduced (P less than .05) and the FSH response to exogenous GnRH was increased (P less than .05) by administration of EB alone, DHTP alone or TP.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effects of dihydrotestosterone benzoate administration on gonadotropin secretion in ovariectomized pony mares

Eight long-term ovariectomized pony mares were treated with either dihydrotestosterone (DHT) benzoate (400 micrograms/kg body weight) in safflower oil or an equivalent amount of oil every other day for 21 d to determine the effects of DHT on follicle stimulating hormone (FSH) and luteinizing hormone (LH) concentrations in blood samples drawn once daily and after administration of three successive injections of gonadotropin releasing hormone (GnRH). The GnRH injections were given at 4-h intervals on the day following the last DHT or oil injection. Treatment with DHT benzoate did not alter (P greater than .10) concentrations of FSH or LH in daily blood samples relative to controls. The FSH and LH response, assessed by areas under the GnRH curves, decreased (P less than .05) from the first to third injection of GnRH when averaged over both groups of mares. There was no effect of DHT treatment on FSH response to GnRH. There was an interaction (P less than .05) between treatment and GnRH injection for LH areas; areas decreased (P less than .05) for DHT-treated mares from the first to third GnRH injection but were unchanged for control mares. It seems that DHT alone cannot mimic the stimulatory effects of testosterone on FSH production and secretion as observed in previous experiments with ovariectomized and intact mares. Moreover, because intact mares have been shown previously to respond to DHT treatment with an increase in GnRH-induced FSH secretion, it appears that some mechanism is lost in long-term ovariectomized mares, making them unresponsive to DHT treatment.     

Gonadotropin secretion in ovariectomized pony mares treated with dexamethasone or progesterone and subsequently with dihydrotestosterone

Twelve long-term ovariectomized (OVX) pony mares were used to determine the effects of dexamethasone (DEX) or progesterone (PR) on concentrations of follicle stimulating hormone (FSH) and luteinizing hormone (LH) in daily blood samples and after administration of gonadotropin releasing hormone (GnRH). All mares were subsequently administered dihydrotestosterone (DHT) to determine if DEX or PR treatment altered the FSH or LH response to this androgen. Daily blood sampling was started on day 1. After a pretreatment injection of GnRH on day 5, four mares were administered DEX at 125 micrograms/kg of body weight (BW), four mares were administered PR at 500 micrograms/kg of BW and four mares were administered vehicle. Injections were given subcutaneously in vegetable shortening daily through day 14. After a second injection of GnRH on day 15, all mares were administered DHT in shortening at 150 micrograms/kg of BW. Injections of DHT were given daily through day 24. A final injection of GnRH was given on day 25. Treatment of mares with DEX 1) reduced (P less than .01) daily LH secretion and briefly increased (P less than .05) daily FSH secretion and 2) increased (P less than .01) the FSH response to exogenous GnRH. Treatment of mares with PR had no effect on daily LH secretion but increased (P less than .05) daily FSH secretion and increased (P less than .01) the FSH response to exogenous GnRH.(ABSTRACT TRUNCATED AT 250 WORDS)     

Relationships among LH, FSH and prolactin secretion, storage and response to secretagogue and hypothalamic GnRH content in ovariectomized pony mares administered testosterone, dihydrotestosterone, estradiol, progesterone, dexamethasone or follicular fluid

Thirty-five ovariectomized pony mares were used to study the relationships among luteinizing hormone (LH), follicle stimulating hormone (FSH) and prolactin (PRL) concentrations in blood (secretion), in pituitary (storage) and in blood after secretagogue administration, as well as the content of gonadotropin releasing hormone (GnRH) in hypothalamic areas, under various conditions of steroidal and nonsteroidal treatment. Five mares each were treated daily for 21 d with vegetable shortening (controls), testosterone (T; 150 micrograms/kg of body weight, BW), dihydrotestosterone (DHT; 150 micrograms/kg BW), estradiol (E2; 35 micrograms/kg BW), progesterone (P4; 500 micrograms/kg BW), dexamethasone (DEX; 125 micrograms/kg BW) or charcoal-stripped equine follicular fluid (FF; 10 ml). Secretagogue injections (GnRH and thyrotropin releasing hormone, TRH, at 1 and 4 micrograms/kg of BW, respectively) were given one d prior to treatment and again after 15 d of treatment. Relative to controls, treatment with T, DHT and DEX reduced (P less than .05) LH secretion, storage and response to exogenous GnRH, whereas treatment with E2 increased (P less than .05) these same characteristics. Treatment with P4 reduced (P less than .05) only LH secretion. Treatment with T, DHT, E2 and DEX reduced (P less than .05) FSH secretion, whereas treatment with P4 increased (P less than .05) it and FF had no effect (P greater than .1). All treatments increased (P less than .05) FSH storage, whereas only treatment with T and DHT increased (P less than .05) the FSH response to exogenous GnRH. Other than a brief increase (P less than .05) in PRL secretion in mares treated with E2, secretion of PRL did not differ (P greater than .1) among groups. Only treatment with E2 increased (P less than .01) PRL storage, yet treatment with T or DHT (but not E2) increased (P less than .05) the PRL response to exogenous TRH. Content of GnRH in the body and pre-optic area of the hypothalamus was not affected (P greater than .1) by treatment, whereas treatment with T, E2 and DEX increased (P less than .1) GnRH content in the median eminence. For LH, secretion, storage and response to exogenous GnRH were all highly correlated (r greater than or equal to .77; P less than .01). For FSH, only storage and response to exogenous GnRH were related (r = .62; P less than .01). PRL characteristics were not significantly related to one another. Moreover, the amount of GnRH in the median eminence was not related (P greater than .1) to any LH or FSH characteristic.     

Testosterone effects on gonadotropin response to GNRH: cows and pony mares

Effects of testosterone propionate (TP) treatment on plasma concentrations of luteinizing hormone (LH) and follicle-stimulating hormone (FSH) before and after an injection of gonadotropin releasing hormone (GnRH) were studied using ovariectomized cows and pony mares. An initial injection of GnRH (1 microgram/kg of body weight) was followed by either TP treatment or control injections for 10 (cows) or 11 (ponies) d. A second GnRH injection was administered 1 d after the last TP or oil injection. Concentrations of LH and FSH were determined in samples of plasma taken before and after each GnRH injection. Control injections did not alter the response to GnRH (area under curve) nor the pre-GnRH concentrations of LH and FSH in ovariectomized cows or ponies. Testosterone treatment increased (P less than .01) the FSH release in response to GnRH in ovariectomized mares by 4.9-fold; there was no effect in cows, even though average daily testosterone concentrations were 59% higher than in pony mares. Testosterone treatment reduced the LH release in response to GnRH by 26% in ovariectomized mares (P less than .05) and by 17% in ovariectomized cows (P approximately equal to .051). These results are consistent with a model that involves ovarian androgens in the regulation of FSH secretion in the estrous cycle of the mare, but do not support such a model in the cow.     

Effects of active immunization against gonadotropin releasing hormone on gonadotropin secretion after ovariectomy and testosterone propionate administration to mares

Five lighthorse mares were actively immunized against gonadotropin releasing hormone (GnRH) conjugated to bovine serum albumin (BSA) to study the involvement of GnRH in luteinizing hormone (LH) and follicle stimulating hormone (FSH) secretion following ovariectomy (OVX) and after administration of testosterone propionate (TP). Five mares immunized against BSA served as controls. Immunizations were started on November 1, and OVX was performed in June (d 1). All mares were treated with TP from d 50 to 59 after OVX. On the day of OVX, concentrations of LH were lower (P less than .05) in GnRH-immunized mares than in BSA-immunized mares and were generally nondetectable; FSH concentrations were reduced (P less than .05) by 50% in GnRH-immunized mares relative to BSA-immunized mares. In contrast to BSA-immunized mares, plasma concentrations of LH or FSH did not increase after OVX in GnRH-immunized mares. The LH response to GnRH analog (less than .1% cross-reactive with GnRH antibodies) on d 50 was reduced (P less than .05) by 97% in GnRH-immunized mares relative to BSA-immunized mares, whereas the FSH response was similar for both groups. Treatment with TP for 10 d reduced (P less than .01) the LH response and increased (P less than .01) the FSH response to GnRH analog in BSA-immunized mares, but it had no effect (P greater than .1) on the response of either gonadotropin in GnRH-immunized mares.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effect of active immunization against estrogen on gonadotropin response to testosterone propionate treatment in ovariectomized pony mares

An experiment was conducted to determine whether partial neutralization of estrogens via active immunization alters testosterone propionate (TP)-induced increases in FSH secretion after GnRH administration in ovariectomized pony mares. Twenty mares were used in a 2 X 2 factorial arrangement of treatments (n = 5/group). Factor 1 was long-term active immunization against either bovine serum albumin (BSA) or estrone-17-oxime-BSA. Factor 2 was 11-d administration of either vehicle (vegetable oil) or TP (175 micrograms/kg BW). Plasma concentrations of FSH were not affected (P greater than .1) by either factor. As expected, the FSH response to exogenous GnRH was threefold greater (P less than .05) in BSA-immunized mares treated with TP than in BSA-immunized mares receiving oil. However, immunization against estrogens reduced (P less than .05) this TP-induced increase in FSH response by 52%. Plasma concentrations of LH were decreased (P less than .08) by TP; this effect was not altered (P greater than .1) by immunization against estrogen. The LH response to exogenous GnRH was not affected (P greater than .1) by either factor. We conclude that aromatization of testosterone to estrogen is partially responsible for the increased FSH response to exogenous GnRH in TP-treated mares. In contrast, suppression of LH concentrations by TP appears to involve only the androgenic effect of TP.     

Plasma concentrations of cortisol, prolactin, luteinizing hormone, and follicle-stimulating hormone in stallions after physical exercise and injection of secretagogue before and after sulpiride treatment in winter

Ten lighthorse stallions were used to determine 1) whether prolactin (PRL) and cortisol responses previously observed after acute exercise in summer would occur in winter when PRL secretion is normally low, 2) whether subsequent treatment with a dopamine receptor antagonist, sulpiride, for 14 d would increase PRL secretion and response to thyrotropin-releasing hormone (TRH) and exercise, and 3) whether secretion of LH, FSH, and cortisol would be affected by sulpiride treatment. On January 11, blood samples were drawn from all stallions before and after a 5-min period of strenuous running. On January 12, blood samples were drawn before and after an i.v. injection of GnRH plus TRH. From January 13 through 26, five stallions were injected s.c. daily with 500 mg of sulpiride; the remaining five stallions received vehicle. The exercise and secretagogue regimens were repeated on January 27 and 28, respectively. Before sulpiride injection, concentrations of both cortisol and PRL increased (P less than .05) 40 to 80% in response to exercise; concentrations of LH and FSH also increased (P less than .05) approximately 5 to 10%. Sulpiride treatment resulted in (P less than .05) a six- to eightfold increase in daily PRL secretion. The PRL response to TRH increased (P less than .05) fourfold in stallions treated with sulpiride but was unchanged in control stallions. Sulpiride treatment did not affect (P greater than .05) the LH or FSH response to exogenous GnRH.(ABSTRACT TRUNCATED AT 250 WORDS)     

Testosterone propionate treatment of stallions: Effects on secretion of luteinizing hormone and follicle stimulating hormone in daily samples and after administration of gonadotropin releasing hormone

Ten stallions were used to determine if the stallion responds to administration of testosterone propionate (TP) with an increase in follicle stimulating hormone (FSH) secretion after administration of gonadotropin releasing hormone (GnRH) as has been previously observed for geldings and intact and ovariectomized mares. Five stallions were treated with TP (350 μg/kg of body weight) in safflower oil every other day for 11 days; control stallions received injections of safflower oil. The response to GnRH (1.0 μg/kg of body weight) was determined for all stallions before the onset of treatment (GnRH I) and at the end of treatment (GnRH II). Blood samples were also withdrawn daily from 3 days prior to treatment through GnRH II. Treatment with TP decreased (P<.10) concentrations of FSH in daily blood samples. However, treatment with TP did not affect (P>.10) the GnRH-induced secretion of FSH. Concentrations of luteinizing hormone (LH) decreased (P<.05) in daily blood samples averaged over both groups of stallions and were lower (P<.10) in TP-treated stallions than in controls during the latter days of treatment. We conclude that TP administration to stallions does not alter the FSH response to GnRH as has been observed for geldings and for mares of several reproductive states.     

Prolactin and Gonadotropin Responses in Geldings to Injections of Estradiol Benzoate in Oil, Estradiol Benzoate in Biodegradable Microspheres, and Estradiol Cypionate

We previously reported success in inducing early ovulation in seasonally anovulatory mares with a combination of estradiol pretreatment followed by daily administration of a dopamine antagonist (sulpiride). Although every-other-day injections of estradiol benzoate (EB) were effective in that experiment, practical application of this technology would require simplification of the treatment regimen. The current experiment was designed to compare, in a gelding model, the biologic responses of two alternative, one-injection regimens for estradiol delivery to the established EB treatment used previously. Fifteen long-term geldings were sampled via jugular venipuncture from November 5 to 7, 2006, and were then administered intramuscular injections of vegetable oil (n = 4); EB, 11 mg in oil (n = 4; controls); EB in biodegradable microspheres (300 mg; n = 3); or estradiol cypionate, 100 mg in oil (n = 4). Injections of EB in oil were repeated every other day for a total of 10 injections, as was done in our previous experiment. Jugular blood samples were drawn from all geldings at 3, 6, 12, 24, 36, and 48 hours relative to injections, and then on the mornings of days 3, 4, 6, 8, 10 to 18, 22, 26, and 30. On days 10 through 13, all geldings received subcutaneous injections of 125 mg sulpiride, a dopamine receptor antagonist, to stimulate prolactin secretion. On day 12, each gelding received an intravenous injection of 30 μg gonadotropin-releasing hormone (GnRH) analog and 3 mg thyrotropin-releasing hormone (TRH); frequent blood samples were drawn to characterize the luteinizing hormone (LH), follicle-stimulating hormone (FSH), and prolactin responses. Relative to geldings receiving oil, all geldings receiving estradiol injections had a rise (P < .05) in estradiol concentrations lasting at least 12 days. Daily LH concentrations increased (P < 0.01) in all treated groups, but the response was delayed approximately 14 days in the geldings receiving EB in microspheres. Daily FSH concentrations decreased (P < .01) in all treated groups, with the greatest response in the geldings receiving EB in microspheres. Prolactin in daily samples increased (P < .01) similarly in all estradiol-treated groups after injection of sulpiride. The LH response to GnRH analog was greatest (P < .05) in geldings receiving EB in oil and estradiol cypionate; the FSH response was not altered by treatment. The prolactin response to TRH was greater (P < .01) in estradiol-treated geldings relative to controls, but did not differ among groups. Compared with the responses to every-other-day EB injections in oil, as we used previously, a single injection of 100 mg estradiol cypionate gave the most similar and consistent responses. Because of these similar responses in this gelding model, it is likely that a single injection of 100 mg estradiol cypionate can be used in lieu of every-other-day injections of EB in oil in the treatment regimen we reported previously for stimulating ovarian activity in seasonally anovulatory mares.     

Effects of gonadotropin-releasing hormone infused in a pulsatile or continuous fashion on serum gonadotropin concentrations and ovulation in the mare.

Studies were conducted to compare continuous vs pulsatile i.v. infusion of GnRH on serum gonadotropin concentrations and ovulation in seasonally anestrous mares and in cycling mares. Anestrous mares (Exp. 1) received no treatment (control; n = 3), 2, or 20 micrograms of GnRH/h continuous infusion (CI) (n = 4 and n = 6, respectively), or 20 micrograms of GnRH/h pulsatile infusion (PI) (n = 5). After initiation of GnRH infusion, serum LH levels increased earlier, and to a greater extent, in the PI group than in other groups (P less than .05). In contrast, serum FSH concentrations did not differ among groups. The number of days to development of the first 35-mm follicle was not different among GnRH treatment groups; however, mares receiving PI ovulated on d 9.4 of treatment, 2.8 d earlier than those receiving 20 micrograms of GnRH/h CI (P less than .05). Mares given 2 micrograms of GnRH/h CI failed to ovulate spontaneously after 16 d of treatment, but each one ovulated within 2 to 4 d after injection of 2,000 IU of hCG on d 16. Control mares did not ovulate or show any significant follicular development throughout the experiment. Cycling mares (Exp. 2) received no treatment (control; n = 6), 20 micrograms of GnRH/h CI, or 20 micrograms of GnRH/h PI (n = 4) beginning on d 16 of an estrous cycle (d 0 = day of ovulation). Serum LH concentrations in all groups increased after initiation of treatment; however, on the day of ovulation LH concentrations were lower in the CI group than in the PI or control groups (P less than .05).(ABSTRACT TRUNCATED AT 250 WORDS)     

Pituitary responsiveness of mares challenged with GnRH at various stages of the transition into the breeding season

Four groups of mares, representing anestrus (AN; n = 8), early transition (ET; n = 7), late transition (LT; n = 8) and estrus (EST; n = 12) were used to examine release of luteinizing hormone (LH) and follicle stimulating hormone (FSH) after a bolus injection of gonadotropin releasing hormone (GnRH) during the transition from anestrus into the breeding season. Estrous mares received GnRH on d 2 or 3 of estrus in the cycle immediately preceding slaughter. Anestrous, ET and LT mares received GnRH exactly 1 wk prior to slaughter. A single injection of GnRH (Sigma LHRH, L-0507, 2.0 micrograms/kg body weight in .9% saline, iv) was given to each mare. Blood samples were collected at -2, h, -1 h, directly prior to GnRH, then 15, 30, 45, 60, 90, 120, 150, 180, 210, 240, 300, 360, 420 and 480 min post-injection. Maximum release of LH and FSH was observed within 30 min after injection of GnRH. Except for the LH response in EST mares, concentrations of both hormones had returned to pre-injection baseline levels within 8 h. Group means for area under the curve (AUC) of concentrations of LH in serum, and the maximum amount (MAX) of LH quantified in serum, post-GnRH, increased (P less than .05) progressively from AN to the breeding season. The AUC and MAX responses for FSH showed a reverse pattern, decreasing (P less than .05) from AN to the breeding season.(ABSTRACT TRUNCATED AT 250 WORDS)     

The effect of pulsatile gonadotropin-releasing hormone and estradiol administration on luteinizing hormone and follicle-stimulating hormone concentrations in pituitary stalk-sectioned ovariectomized pony mares

Hourly pulses of gonadotropin-releasing hormone (GnRH) or bi-daily injections of estradiol (E2) can increase luteinizing hormone (LH) secretion in ovariectomized, anestrous pony mares. However, the site (pituitary versus hypothalamus) of positive feedback of estradiol on gonadotropin secretion has not been described in mares. Thus, one of our objectives involved investigating the feedback of estradiol on the pituitary. The second objective consisted of determining if hourly pulses of GnRH could re-establish physiological LH and FSH concentrations after pituitary stalk-section (PSS), and the third objective was to describe the declining time trends of LH and FSH secretion after PSS. During summer months, ovariectomized pony mares were divided into three groups: Group 1 (control, n = 2), Group 2 (pulsatile GnRH (25 μg/hr), n = 3), and Group 3 (estradiol (5 mg/12 hr), n = 3). All mares were stalk-sectioned and treatment begun immediately after stalk-section. Blood samples were collected every 30 min for 8 h on the day before surgery (DO) and 5 d post surgery (D5) to facilitate the comparison of gonadotropin levels before and after pituitary stalk-section. Additionally, jugular blood samples were collected every 12 hr beginning the evening of surgery, allowing for evaluation of the gonadotropin secretory time trends over the 10 d of treatment. On Day 10, animals were euthanized to confirm pituitary stalk-section and to submit tissue for messenger RNA analysis (parallel study). Plasma samples were assayed for LH and FSH by RIA. Mean LH secretion decreased from Day 0 to Day 5 in Groups 1 and 3, whereas LH secretion tended (P < 0.08) to decrease in Group 2 mares. On Day 5, LH was higher (P < 0.01) in Group 2 (17.26 ± 3.68 ng/ml; LSMEANS ± SEM), than either Group 1 (2.65 ± 4.64 ng/ml) or group 3 (4.28 ± 3.68 ng/ml). Group 1 did not differ from Group 3 on Day 5 (P < 0.40). Similarly, mean FSH levels decreased in all groups after surgery, yet Group 2 mares had significantly (P < 0.001) higher FSH concentrations (17.66 ± 1.53 ng/ml) than Group 1 or Group 3 (8.34 ± 1.84 and 7.69 ± 1. 63 ng/ml, respectively). Regression analysis of bi-daily LH and FSH levels indicated that the time trends were not parallel. These findings indicate: 1) Pituitary stalk-section lowered LH and FSH to undetectable levels within 5 d after surgery, 2) pulsatile administration of GnRH (25 μg/hr) maintained LH and FSH secretion, although concentrations tended to be lower than on Day 0, and 3) E2 did not stimulate LH or FSH secretion.     

Secretion of luteinizing hormone and follicle stimulating hormone in intact and ovariectomized mares in summer and winter

Sequential samples of blood were drawn via jugular catheters every 15 min for 24 h from four mares in each of five reproductive states: intact anestrous mares in winter, intact diestrous mares in summer, intact estrous mares in summer, ovariectomized mares in winter and ovariectomized mares in summer. Estrous mares were sampled on d 4 or 5 of estrus and diestrous mares on d 10 or 11 of diestrus. Each sample of plasma was assessed for concentrations of luteinizing hormone (LH) and follicle stimulating hormone (FSH) in two independent radioimmunoassays. A computer program was developed that determined peak hormone concentrations based on assay sensitivity, assay variability and repeatability of peaks in both independent assays. Peaks in LH and FSH were observed for mares in all five reproductive states, except for FSH concentrations in estrous mares. High frequency peaks of short duration were observed only in ovariectomized mares. Low frequency peaks of relatively long duration were observed in both intact and ovariectomized mares in both seasons. With the exception of estrous mares, there was variation among mares in the patterns of LH and(or) FSH within any one group; all estrous mares exhibited high, variable LH concentrations and low, constant FSH concentrations. In general, peaks in both gonadotropins occurred simultaneously. Ovariectomized mares exhibited more (P less than .05) peaks/24 h than intact mares for both gonadotropins. Ovariectomized mares also exhibited more (P less than .05) FSH peaks/24 h in summer than in winter.     

Testosterone administration to mares during estrus: duration of estrus and diestrus and concentrations of LH and FSH in plasma

To study the possible role of ovarian androgens in regulation of follicle stimulating hormone (FSH) secretion in the cycling mare, five mature, intact mares were treated with testosterone (20 micrograms/kg of body weight) daily during estrus; five control mares received safflower oil on the same schedule. Mares were teased for estrus and samples of jugular blood were drawn daily through one full estrous cycle. Concentrations of FSH in plasma were measured by a newly developed radioimmunoassay based on anti-ovine FSH serum and radioiodinated equine FSH. Testosterone treatment during estrus had no effect on duration of estrus, diestrus or the total cycle. Concentrations of FSH in plasma during estrus were unaffected by testosterone treatment. However, FSH concentrations in testosterone-treated mares were elevated (P less than .05) compared with controls during mid-diestrus (d 6 through 11). The magnitude and timing of the LH peaks were unaffected by treatment, as was the day on which the first elevated progesterone concentration occurred. These data are consistent with a model of FSH secretion in which ovarian androgens cause an accumulation of FSH in the pituitary during estrus in preparation for the surges that occur in FSH secretion during diestrus.     

Responses of seasonally anovulatory mares to daily administration of thyrotropin-releasing hormone and(or) gonadotropin-releasing hormone analog

Seventeen seasonally anovulatory light horse mares were treated daily, starting January 5 (d 1), for 28 d with GnRH analog (GnRH-A; 50 ng/kg BW) and(or) thyrotropin-releasing hormone (TRH; 5 microg/kg BW) in a 2 x 2 factorial arrangement of treatments to test the hypothesis that combined treatment may stimulate follicular growth and development. Ovaries were examined via ultrasonography and jugular blood samples were collected every 3 d. Frequent blood samples were collected after treatment injections on d 1, 2, 4, 7, 11, 16, and 22; on d 29, all mares received an i.v. mixture of GnRH, TRH, sulpiride, and EP51389 (a growth hormone secretagogue) to assess pituitary responsiveness. No consistent effects (P > 0.1) of treatment were observed for plasma LH, FSH, prolactin, or thyroxine concentrations in samples collected every 3 d. The only effect on ovarian follicle numbers was a reduction in number of follicles 11 to 19 mm in diameter due to TRH treatment (P = 0.029). No mare ovulated during treatment. On the days of frequent sampling, mean LH (P = 0.0001) and FSH (P = 0.001) concentrations were higher in mares receiving GnRH-A and tended to increase from d 1 through 7. In contrast, mean prolactin (P = 0.001) and thyroid-stimulating hormone (P = 0.0001) concentrations were high in mares receiving TRH on d 1 but rapidly decreased thereafter. When mares were administered the secretagogue mixture on d 29, the LH response was greater (P = 0.0002) in mares that had previously received GnRH-A but the FSH response was not affected (P > 0.1); the prolactin response was greater (P = 0.014) and the TSH response was smaller (P = 0.0005) in mares that had previously received TRH. Surprisingly, an immediate growth hormone response to EP51389 was absent in all mares. In conclusion, daily GnRH-A treatment stimulated plasma LH and FSH concentrations immediately after injection; although no long-term elevation in preinjection concentrations was achieved, the responses gradually increased over time, indicating a stimulation of gonadotropin production and storage. Daily treatment with TRH stimulated plasma TSH and prolactin concentrations, but the response diminished rapidly and was minimal within a few days, indicating a depletion of pituitary stores and little or no stimulation of production. There was no beneficial effect of adding TRH treatment to the daily GnRH-A regimen.     

Effects of pituitary stalk-transection and type of barrier on pituitary and luteal function during the estrous cycle of the ewe

Effects of pituitary stalk-transection on plasma concentrations of luteinizing hormone (LH), follicle stimulating hormone (FSH) prolactin (PRL) and progesterone were investigated during the estrous cycle of ewes. Pituitary stalk (SS) or sham (SH) transection was performed on day 1 (estrus = day 0) of the estrous cycle. A Teflon or Silastic barrier was placed between the cut ends of the stalk to prevent reorganization of the portal vasculature. Immediately following surgery, pulsatile administration of gonadotropin releasing hormone (GnRH, 200 ng/hr) or .9% NaCl was initiated and continued for the duration of the experiment. Estradiol benzoate (EB, 50 μg im) was administered to all ewes on day 3. Mean concentrations of LH were greater in SS ewes than in SH ewes (P<.05). There was a trend (P=.06) for the concentration of LH to be higher in ewes with Teflon compared with Silastic barriers between the cut ends of the stalk. Infusion of GnRH elevated concentrations of LH in both SS and SH ewes (P<.05). Concentrations of progesterone were reduced (P<.01) in saline-infused SS ewes while infusion of GnRH in SS ewes maintained concentrations of progesterone similar to saline-infused SH ewes. The concentrations of FSH or PRL were unaffected by SS, type of barrier or treatment with GnRH. Administration of EB failed to induce a surge of LH except in a SH ewe infused with GnRH. Ewes were more responsive to infusion of GnRH following SS than after SH as reflected by increased plasma concentrations of LH and progesterone.     

Growth hormone in mares and stallions: pulsatile secretion, response to growth hormone-releasing hormone, and effects of exercise, sexual stimulation, and pharmacological agents.

Short-term patterns of growth hormone (GH) secretion and factors affecting it were studied in mares and stallions. In Exp. 1, hourly blood samples were collected from three mares and three stallions in summer and winter. Although GH concentrations varied in a pulsatile manner in all horses, there was no effect of sex or season (P greater than .1) on plasma GH concentrations and no indication of a diurnal pattern of GH secretion. In Exp. 2, 10-min blood samples were drawn for 8 h from 12 mares; after 6 h, porcine GH-releasing hormone (GHRH) was administered i.v. at 0, 45, 90, or 180 micrograms/mare (three mares per dose). Pulsatile secretion of GH occurred in all mares and averaged 2.4 +/- .3 peaks/6 h; amplitudes were variable and ranged from 2.6 to 74.4 ng/mL. Eight of nine mares responded within 20 min to GHRH injection, but there was no difference (P greater than .1) among the three doses tested. In Exp. 3, plasma GH concentrations in stallions increased (P less than .05) 8- to 10-fold after 5 min of acute physical exercise or exposure to an estrual mare. Restraint via a twitch (5 min) and epinephrine administration (3 mg i.v.) also increased (P less than .05) plasma GH concentrations by approximately fourfold. In Exp. 4 and 5, administration of either .4, 2, or 10 mg of thyrotropin-releasing hormone (TRH) or 100 or 500 mg of sulpiride (a dopamine receptor antagonist) increased (P less than .01) plasma prolactin concentrations but had no effect (P greater than .1) on GH concentrations during the same period of time.(ABSTRACT TRUNCATED AT 250 WORDS)     

Pituitary responsiveness to GnRH in mares following deslorelin acetate implantation to hasten ovulation

The present experiment characterized the pituitary responsiveness to exogenous GnRH in the first 10 d after ovulation following commercially available deslorelin acetate implantation at the normal dosage for hastening ovulation in mares. Twelve mature, cyclic mares were assessed daily for estrus and three times weekly for ovarian activity starting May 1. Mares achieving a follicle at least 25 mm in diameter or showing signs of estrus were checked daily thereafter for ovarian characteristics. When a follicle >30 mm was detected, mares were administered either a single deslorelin acetate implant or a sham injection and then assessed daily for ovulation. On d 1, 4, 7, and 10 following ovulation, each mare was challenged i.v. with 50 microg GnRH, and blood samples were collected to characterize the LH and FSH responses. The size of the largest follicle on the day of treatment did not differ (P = 0.89) between groups. The number of days from treatment to ovulation was shorter (P < 0.001) by 2.0 d for the treated mares indicating a hastening of ovulation. The size of the largest follicle present on the days of GnRH challenge was larger in the treated mares on d 1 (P = 0.007) but smaller on d 10 (P = 0.02). In addition, the interovulatory interval was longer (P = 0.036) in the treated mares relative to controls by 4.4 d. Concentrations of FSH in plasma of the treated mares were lower (P < 0.05) than control concentrations from d 3 to 12; LH concentrations in the treated mares were lower (P < 0.05) relative to controls on d 0 to 5, d 7, and again on d 20 to 23. Progesterone values were the same (P = 0.99) for both groups from 2 d before ovulation though d 23. There was an interaction of treatment, day, and time of sampling (P < 0.001) for LH and FSH concentrations after injection of GnRH. Both the LH and FSH responses were suppressed (P < 0.009) in the treated mares relative to controls on d 1, 4, and 7; by d 10, the responses of the two groups were equivalent. In conclusion, deslorelin administration in this manner increased the interovulatory interval, consistently suppressed plasma LH and FSH concentrations, and resulted in a complete lack of responsiveness of LH and FSH to GnRH stimulation at the dose used during the first 7 d after the induced ovulation. Together, these results are consistent with a temporary down-regulation of the pituitary gland in response to deslorelin administered in this manner.     

Effects of deslorelin acetate implants in horses: single implants in stallions and steroid-treated geldings and multiple implants in mares

Three experiments were performed to test the following hypotheses: 1) stallions and/or progesterone-estradiol-treated geldings could serve as models for the effects of a single implant of the GnRH analog, deslorelin acetate, on LH and FSH secretion by mares; and 2) multiple implants of deslorelin acetate could be used as a means of inducing ovarian atrophy in mares for future study of the mechanisms involved in the atrophy observed in some mares after a single implant. In Exp. 1, nine light horse stallions received either a single deslorelin implant (n = 5) or a sham injection (n = 4) on d 0. In Exp. 2, 12 geldings received daily injections of progesterone on d -20 through -4, followed by twice-daily injections of estradiol on d -2 to 0. On the morning of d 0, geldings received either a single deslorelin implant (n = 6) or a sham injection (n = 6). Daily injections of progesterone were resumed on d 2 through 15. In Exp. 1, plasma LH and FSH were elevated (P < 0.05) in the treatment group relative to controls at 4, 8, and 12 h after implant insertion. In the treated stallions, FSH was decreased (P < 0.05) on d 3 to 13, and LH was decreased on d 6 to 13. In Exp. 2, plasma LH and FSH were elevated (P < 0.05) at 4,8, and 12 h after deslorelin implant insertion. Plasma LH was suppressed (P < 0.05) below controls on d 2 to 7, 9, and 11 to 15; plasma FSH was suppressed (P < 0.05) on d 4 to 15. In Exp. 3, 21 mares were used to determine whether multiple doses of deslorelin would cause ovarian atrophy. Mares received one of three treatments: 1) sham injections; 2) three implants on the first day; or 3) one implant per day for 3 d (n = 7 per group). Treatment with multiple implants increased (P < 0.05) the interovulatory interval by 14.8 d and suppressed (P < 0.01) LH and FSH concentrations for approximately 25 d; no mare exhibited ovarian atrophy. In conclusion, after an initial short-term increase in LH and FSH secretion, deslorelin implants caused long-term suppression of both gonadotropins in stallions as well as in geldings treated with progesterone and estradiol to mimic the estrous cycle. It is likely that either of these models may be useful for further study of this suppression in horses. Although multiple implants in mares suppressed gonadotropin secretion longer than a single implant, the lack of ovarian atrophy indicates that the atrophy observed after a single implant in previous experiments was likely due to the susceptibility of individual mares.