Radioimmunoassay for PMSG and its application to in-vivo studies.

A double-antibody radioimmunoassay for PMSG, especially for meauring PMSG in cattle blood after exogenous application, has been developed. A rabbit antiserum against PMSG and pure PMSG for radioiodination were used. There was a strong cross-reaction against equine LH and FSH, but the slight cross-reaction against bovine LH and FSH could be eliminated by adding bovine LH to each tube during the assay. Unspecific, interfering influences of equine or cow serum could be eliminated by adding a constant amount of PMSG-free serum to each tube. PMSG added to 200 microliter of serum could be recovered by this method with a mean of 90 . 5 +/- 9 . 9%. Inhibition curves obtained with pregnant mare serum or cow serum after administration of PMSG were parallel to those obtained with the PMSG standard preparation. The intra-assay coefficient of variation (CV) was 6 . 9%. The inter-assay CV was 12 . 6%. Sensitivity of the assay was 1 mi.u. PMSG/tube. Values of PMSG measured in the serum of pregnant mares by this assay were comparable with those obtained by a bioassay on the same samples. PMSG was still measurable in blood serum about 10 days after injection of 1500-3000 i.u. PMSG. After infusion of 12,000 i.u. PMSG for 3 h (2 heifers), the half-life of PMSG was found to have two components, one of 51 or 40 h and a slower one of 123 or 118 h.     

Effect of diethylstilboestrol on the relationship between LH, PMSG and progesterone during pregnancy in the mare.

Two studies were conducted to determine the relationship between LH and progesterone and between PMSG and progesterone during pregnancy in mares. In the first, samples of jugular blood were collected daily from 7 mares from the first day of oestrus until Day 28 of pregnancy, and in the second, samples were collected weekly from 14 mares from Day 35 of gestation until parturition. In an attempt to prolong secretion of progesterone from accessory corpora lutea, 7 of these 14 mares were injected with increasing doses (2--10 mg) of diethylstilboestrol (DES) between Days 84 and 142 of gestation. The remaining 7 mares received injections of vehicle. Concentrations of LH, PMSG and progesterone in serum were determined by radioimmunoassay. From the onset of oestrus until Day 4 of gestation, serum concentrations of LH and progesterone were negatively correlated (r = 0.67, P less than 0.01), but from Days 5 to 28 a positive correlation (r = 0.80, P less than 0.01) was noted. Likewise, serum concentrations of PMSG and progesterone were highly correlated between Days 35 and 196 in mares injected with DES (r = 0.72, P less than 0.01) and the vehicle (r = 0.75, P less than 0.01). Injections of DES did not influence serum concentrations of LH, PMSG or progesterone, or affect the length of gestation. It was concluded that DES does not influence the maintenance of pregnancy in the mare.     

Effect of gonadotropin-releasing hormone,stage of sexual maturation and 6-methoxybenzoxazolinone on plasma gonadotropins,ovarian development and uterine weight in prepuberal gilts

Two experiments were conducted with prepuberal gilts at 60, 120 and 160 days of age to a) determine the effect of 6-methoxybenzoxazolinone 6-MBOA) on reproductive plasma hormone concentrations and organ development, and b) determine how plasma follicle-stimulating hormone (FSH) and luteinizing hormone (LH) concentrations before and after injection of gonadotropin-releasing hormone (GnRH) or 6-MBOA varied in relation to ovarian development. In Exp. 1, 12 gilts were used in a 4×4 Latin square design. Four gilts/age group were injected once with: 1) vehicle, 2.5% propylene glycol in 50% ethanol, 2) 2 μg of GnRH/kg body weight (BW), 3) 0.2 mg of 6-MBOA/kg BW, and 4) 2 mg of 6-MBOA/kg BW on four successive days in random order. Blood was collected via indwelling vena cava catheters. Injection of GnRH into gilts increased plasma FSH and LH at each age compared with vehicle (P<0.05). Hormone profiles for FSH and LH differed among age groups (P<0.01), but area under curves did not differ significantly among age groups. Injection of 6-MBOA did not significantly affect plasma FSH and LH. Plasma FSH and LH before the GnRH injection or on days when GnRH was not injected were greater at 60 than at 120 and 160 days (FSH, 128 vs 54 and 42 ng/ml; LH, 0.38 vs 0.16 and 0.13 ng/ml for 60, 120 and 160 days, respectively (P<0.05). In Exp. 2, vehicle, 0.2 or 2 mg of 6-MBOA/kg BW were injected once daily for 7 days in 19 gilts. Injections of 6-MBOA had no detectable effects on gonadotropin secretion, ovarian development or uterine weight. Between 60 and 120 days of age, vesicular follicles developed, ovarian weight increased 20-fold, and uterine weight increased 10-fold (P<0.05); basal concentrations of plasma FSH and LH decreased three- and twofold, respectively.     

Continuous administration of low-dose GnRH in mares II. Pituitary and ovarian responses to uninterrupted treatment beginning near the autumnal equinox and continuing throughout the anovulatory season

We tested the hypothesis that continuous subcutaneous treatment with low-dose GnRH, administered to mares from late September/early October through March, would prevent the development of seasonal anovulation. Quarter Horse mares (n=20) were stratified by age and body condition score and assigned randomly to either a saline control (n=9) or a GnRH (n=11) treatment group. Gonadotropin-releasing hormone was delivered continuously via osmotic minipumps, with sham pumps placed in control mares. Initial pumps were inserted on Day 3 following ovulation or during the follicular phase if the next anticipated ovulation did not occur by 9 October. Delivery rate of GnRH was 2.5 microg/h (60 microg/day) for the first 60 days, followed by 5.0 microg/h (120 microg/day) thereafter. Pumps were replaced every 30 days. Eighty and 100% of all mares had become anovulatory by 1 November and 1 December, respectively, and remained anovulatory through the end of February. Neither serum concentrations of LH throughout the study nor total releasable pools of LH in March differed between groups. Although control mares that exhibited ovulatory cycles after study onset had greater (P<0.05) mean concentrations of LH during the follicular phase and metestrus compared to GnRH-treated mares, neither size of ovulatory follicles nor interovulatory intervals differed between groups. Serum concentrations of FSH were not affected by treatment, but were lowest (P<0.05) from November through January. Continuous infusion of low-dose GnRH, beginning soon after autumnal equinox and continuing until just after vernal equinox, failed to prevent the occurrence of or to hasten transition from seasonal anovulation.     

Establishment and maintenance of pregnancy after embryo transfer in ovariectomized mares treated with progesterone

Pregnancy was established and maintained after embryo transfer in 3 ovariectomized mares treated with progesterone only. Four ovariectomized mares were used as recipients, and 7 transfers were performed. Progesterone in oil, 300 mg i.m. daily, was given starting 5 days before transfer of a 7-day embryo. If the mare was pregnant at 20 days, progesterone treatment was continued to 100 days of gestation. The 3 pregnant mares carried to term and delivered live foals with normal parturition, lactation and maternal behaviour. No differences were seen between pregnant and non-pregnant ovariectomized mares in jugular plasma concentrations of oestrogen, LH or FSH from day of transfer (Day 7) to Day 20. Pregnant ovariectomized mares showed a rise in LH, reflecting production of horse CG, starting at Day 36. Oestrogen values remained low until Day 50.     

Effect of administration of an alpha 2-adrenergic agonist, xylazine, on pulsatile gonadotrophin secretion in anoestrous horse mares

Two experiments were performed to test the hypothesis that the seasonal suppression of gonadotrophin pulse frequency in anoestrous horse mares reflects inhibitory neural mechanisms. In a preliminary experiment (Exp. 1) conducted in February, 4 anoestrous mares were sedated by repeated intravenous injections of xylazine, an alpha 2-adrenergic receptor agonist. On the day of treatment, 1-2 LH pulses were observed in xylazine-treated mares. In contrast, during a 12-h period only 1/8 untreated control mares exhibited a LH pulse. In Exp. 2, the effect of xylazine-induced sedation on pulsatile gonadotrophin release was examined in 4 anoestrous mares on two occasions before (18 November and 9 December) and after (23 December and 6 January) an abrupt, artificial increase in day length. Treatment with xylazine was associated with an overall increased FSH (P less than 0.01) and LH (P less than 0.05) pulse frequency, compared with that observed during 12-h pretreatment periods. To evaluate an effect of treatment at the various time during the experimental period, the change in FSH pulse frequency was analysed, since occasionally FSH pulses were unaccompanied by a change in serum LH values indicative of a LH pulse. On two occasions before increased daylength only 1/4 and 3/4 mares exhibited an increase in FSH pulses; in contrast, 14 days after increased daylength (23 December), 4/4 mares exhibited increased FSH pulse frequency associated with treatment. After 27 days of increased daylength (6 January), endogenous FSH pulse frequency was greater than before increased daylength and treatment with xylazine was unaccompanied by a further increase.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effectiveness of an antagonist to gonadotrophin releasing hormone on the FSH and LH response to GnRH in perifused equine pituitary cells,and in seasonally acyclic mares

We wish to use a gonadotrophin-releasing hormone (GnRH) antagonist in the mare as a tool for investigating the control of the oestrous cycle. The aim of this study was to test the effectiveness of the antagonist cetrorelix by testing both in vitro, using perifused equine anterior pituitary cells, and in vivo in seasonally acyclic mares. Pituitary cells were prepared and after 3-4 days incubation, loaded onto columns and given four pulses of GnRH (at 0, 30, 60 and 90 min; dose-response study). After the second GnRH pulse, infusion of cetrorelix began (0, 100, 1000 and 2000 pmol/l) and continued until the end of the experiment. To mimic luteal phase conditions, cells were pre-incubated and perifused with progesterone (25 nmol/l) and GnRH pulses given at 0, 90, 180 and 270 min. Cetrorelix (0 or 1000 pmol/l) began after the second GnRH pulse. Follicle stimulating hormone (FSH) and luteinizing hormone (LH) concentrations were measured in 5 min fractions. Both FSH and LH response areas (above baseline) after GnRH were inhibited by 1000 pmol/l cetrorelix (P < 0.01, P < 0.01, respectively) but not by 100 pmol/l cetrorelix. Similarly, in the presence of progesterone, cetrorelix inhibited the FSH (P < 0.001) and LH (P = 0.0002) response area. Seasonally acyclic mares, pre-treated for 3 days with progesterone (150 mg i.m. per day) were given cetrorelix as (i) a loading dose of 1 microg/kg then infusion at 2.2 ng/(kg min) for 90 min, (ii) a s.c. injection at 20 microg/kg, (iii) infusion at 2.2 ng/(kg min) for 48 h, and (iv) no cetrorelix (control mares). At 90 min, 6, 24 and 48 h after cetrorelix was first administered, mares were given a bolus injection of GnRH (22.2 ng/kg i.v.) and the FSH and LH responses measured. All doses of cetrorelix inhibited the FSH response at 90 min. The response was no longer suppressed at 6 h in the 90 min infusion group, showing a rapid recovery from inhibition. At 24 h, the FSH responses in the injected and 48 h infusion group were suppressed. The LH concentrations were low and showed no significant changes. This study has defined the time course and dose of cetrorelix with respect to its effect on FSH in the horse. It is concluded that cetrorelix could be used to elucidate the role of FSH in follicular development in cyclic mares.     

Selective release of follicle-stimulating hormone and luteinizing hormone by 5 alpha-dihydroprogesterone and 3 alpha, 5 alpha-tetrahydroprogesterone in pregnant mare's serum gonadotropin-primed immature rats exposed to constant light

The effects of 5 alpha-dihydroprogesterone (5 alpha-DHP) and 3 alpha, 5 alpha-tetrahydroprogesterone (3 alpha, 5 alpha-THP) on follicle-stimulating hormone (FSH) and luteinizing hormone (LH) release were examined in the pregnant mare's serum gonadotropin (PMSG)-primed immature female rat (8 IU PMSG at 28 days of age) maintained in constant light. Control rats kept in 14L:10D conditions exhibited proestrous-like surges of LH and FSH release with peak levels attained at 1800 h on the second day after PMSG treatment. In rats exposed to constant light, the PMSG-induced surges of LH and FSH were not only delayed until 1000 h on the third day after PMSG, resulting in a delay in ovulation, but were also significantly attenuated when compared to the gonadotropin surges that occurred on Day 2 in rats kept under normal light-dark conditions. The administration of 5 alpha-DHP significantly enhanced the release of FSH at 1000 h on Day 3 when compared to constant light-exposed controls, but had no effect on LH. Treatment with 3 alpha, 5 alpha-THP selectively potentiated the release of LH at 1000 h on Day 3 and had an attenuating effect on FSH release on Days 2 and 3. These observations confirm earlier findings in the immature ovariectomized estrogen-primed rat and suggest that 5 alpha-DHP and 3 alpha, 5 alpha-THP may have significant roles in the regulation of FSH and LH secretion.     

Effects of unilateral castration on serum luteinizing hormone, follicle stimulating hormone, and testosterone concentrations in one-, two-, and three-year-old stallions

The endocrine control of compensatory hypertrophy was investigated in 12 Morgan stallions, four each at one, two and three years of age. Half were assigned to be unilaterally castrated (UC) in January and half to remain intact (IN). Nine blood samples were taken from each stallion at half-hour intervals 30, 90, and 150 d after unilateral castration for radioimmunoassay of serum concentrations of luteinizing hormone (LH), follicle stimulating hormone (FSH), and testosterone. Mean serum LH concentration was greater (P<0.06) in UC than IN stallions; however, the difference was greatest at 30 d and least at 150 d. Serum LH was greater (P<0.01) in two- and three-year-olds than in one-year-olds. The mean log(10) for serum FSH concentration was greater (P<0.06) in UC than IN stallions. Mean serum testosterone concentrations were similar in UC and IN stallions for all sample days, suggesting that the single testes of the UC stallions produced as much testosterone as the two testes of the IN stallions. Two- and three-year-old stallions had greater (P<0.01) serum testosterone than one-year-old stallions. Unilateral castration of stallions was associated with a significant increase in serum LH and FSH concentrations and, perhaps, higher intratesticular testosterone, which may explain, in part, the compensatory hypertrophy noted in the remaining testis.     

Inhibin concentrations in mares with granulosa cell tumors

The hormone-producing equine granulosa cell tumor (GCT) may secrete high levels of inhibin. Measurement of inhibin concentrations may be useful in the diagnosis and conformation of mares with GCT. Inhibin may be measured using RIA, which recognizes dimeric alphabetaA-inhibin as well as the monomeric (free) inhibin alpha-subunit, or using a two-site immunoradiometric assay (IRMA) specific for alphabetaA-inhibin. The objective of this study was to examine concurrent relationships among alpha-inhibin (as measured using RIA), alphabetaA-inhibin (as measured using IRMA), and other hormones (testosterone, estradiol, LH, FSH) in mares with GCT. Hormone concentrations were measured in single serum or plasma samples obtained from 22 mares with GCT and from 31 normal cycling mares. One GCT mare had blood samples collected at 12-h intervals for 21 days, and at 15-min intervals for two 6-h periods during that time. Results showed that in GCT mares alpha-inhibin was increased to a greater extent, was more uniformly elevated, and had a less variable secretory pattern than did alphabetaA-inhibin. Concentrations of alpha-inhibin and tumor mass were positively correlated (P < 0.01). Concentrations of LH were higher (P < 0.02) in GCT mares than control mares and were positively associated with testosterone concentrations (P = 0.05). Concentrations of FSH tended to be lower in GCT than control mares and were inversely related with alphabetaA-inhibin in GCT mares. Testosterone and estradiol concentrations were variable. It was concluded that immunoreactive alpha-inhibin reflected detection of both alphabetaA-inhibin and free a-subunit. Free alpha-subunit was evidently secreted at a relatively steady rate dependent upon mass of the GCT, whereas secretion of alphabetaA-inhibin was more responsive to FSH regulation. Determination of alpha-inhibin using RIA appeared to be a more reliable indicator of the presence of a GCT than specific measurement of alphabetaA-inhibin using IRMA.     

The use of radioreceptor assay for the detection of pregnancy in the mare

Jugular blood samples were collected between 42-45 days from the last breeding for measurement of pregnant mare serum gonadotropin (PMSG) and progesterone in 46 pregnant mares. A radioreceptor assay (RRA) was developed to measure human chorionic gonadotropin (hCG) and subsequently modified to measure PMSG. Highly purified hCG was iodinated using a lactoperoxidase enzymatic procedure and served as the labeled antigen for both the hCG and PMSG RRA. Standard curves were generated using purified hCG or PMSG. Bovine corpora lutea served as the receptor source. The assay was conducted at 37 degrees C for one hour with a total elapsed time from preparation of the luteal cell homogenate until final results were calculated of 2.5 hours. Twelve of the 46 mares failed to maintain their pregnancy, returned to estrus and reovulated after 45 days post-breeding (non-foaling mares). Thirty-four of the 46 mares delivered foals following a gestation of normal length. Mean concentrations of PMSG in the foaling mares was higher than in non-foaling mares. A concentration of 6.9 I.U. of PMSG/ml was used as the lowest concentration necessary for the confirmation of pregnancy. Five of the mares delivering foals had low concentrations of PMSG and were called non-pregnant. Thus, the incidence of false negatives by RRA was 14.7%. All of the non-foaling mares were correctly diagnosed non-pregnant for an error rate (false positive) of 0.0. Mare Immunological Pregnancy (MIP) tests on the 12 non-foaling mares indicated three false positives - an error rate of 25%. Of the 34 foaling mares, the MIP test indicated 8 inconclusive or false negatives, an error rate of 23.5%. At day 42-45, there was no significant difference in progesterone concentrations (determined by RIA) between foaling and non-foaling mares. RRA is a quick, accurate and quantitative method for measuring PMSG in the mare and can be used to diagnose pregnancy at 42-45 days post-ovulation. Plasma progesterone concentrations at this time were not associated with subsequent pregnancy maintenance as were plasma PMSG concentrations.     

Evaluation of TSH secretion after GnRh in patients with primary hypothyroidism

6 women (mean age 38 years) with high Thyroid Stimulating Hormone (TSH) serum levels because affected from primary hypothyroidism were studied. 6 healthy women (mean age 31 years) represented the control group. All subjects underwent evaluation of serum TSH, Follicle Stimulating Hormone (FSH), Luteinizing Hormone (LH), basally and 20, 30, 60, 120 minutes after administration of Gonadotropin Releasing Hormone (GnRH: 100 meg. IV). Seric FSH and LH show a large increase 30 minutes after GnRH either in healthy or in hypothyroid subjects. TSH is unresponsive to GnRH in normal condition, while shows a clear decrease (-78%) 30 minutes after GnRH in primary hypothyroidism. Rarely the hypothalamic releasing hormones possess an inhibitory effect on anteipophyseal secretions. Previously a GnRH inhibitory effect on prolactin (PRL) release from PRL secreting tumors in rat. The GnRH inhibitory effect on TSH release in pathological conditions such as primary hypothyroidism is difficult to explain: it may be that GnRH acts on Central Nervous System or at pituitary level: in the last case it could bind sites which are not quite different in the different glycoprotein secreting cells.     

An inexpensive method for assessing pituitary response to luteinizing hormone releasing hormones: analysis of serum gonadotropins in pooled samples

Sixty-two children, aged 2-18 years, suspected of an endocrine disorder were given one of several IM injections of the LHRH and the blood was drawn prior to and at 30, 60, 90, 120, 240 minutes after the injection. An aliquot of .2 ml of serum was taken from each post-LHRH specimen and pooled. The gonadotropins were measured in all the individual and pooled samples. A high correlation (r = .974, LH; r = .981, FSH) between the peak and the pooled sample suggests that the analysis of gonadotropins in a pooled sample gives information comparable to that obtained by the gonadotropin analysis in multiple serum samples. A formula to calculate the peak LH and FSH from the pooled specimen taken from only four post-LHRH injections is: peak LH = 3.015 + 1.049 times the pooled LH; peak FSH = 3.153 + 1.072 times the pooled FSH value. The correlation coefficient between the observed and the calculated LH was .98; the same for the FSH was .987. I suggest that the pooled sample, due to it's integrated response, is a better reflection of the pituitary response to the LHRH.     

Opioidergic control of gonadotropin secretion in the female rabbit: divergent effects of morphine on secretion of follicle-stimulating hormone and luteinizing hormone

The nature of secretion of luteinizing hormone (LH) and follicle-stimulating hormone (FSH) was followed in female rabbits on a daily basis from age 36 to 60 days by sequential 5-min blood sampling over 1- to 2-h periods each day. Both LH and FSH were found to be secreted in a pulsatile manner. The mean LH pulse amplitude over the 25 days was 0.95 +/- 0.32 ng/mL and for FSH it was 10.15 +/- 1.11 ng/mL. Mean plasma LH levels were significantly increased from 1.46 +/- 0.08 ng/mL in 36 to 42-day-old rabbits to 1.89 +/- 0.12 ng/mL in 43 to 50-day-old rabbits and remained elevated from 50 to 60 days. FSH levels during the same periods also rose significantly from 14.93 +/- 0.79 to 19.57 +/- 2.05 ng/mL. To examine the influence of endogenous opioid peptides on the release of LH and FSH in 36 to 60-day-old female rabbits, morphine sulfate at 0.2, 0.5, 2.0, and 5.0 mg/kg was administered subcutaneously after 30 min baseline sampling, and blood was taken for another 60-120 min. Morphine at all doses and at all ages inhibited the amplitude and frequency of LH pulses but had no effect on FSH secretion. To determine whether the effects of morphine on LH secretion could be reversed with naloxone, females aged 82-114 days were used. Naloxone administered 1 h after morphine reversed the inhibitory effects of morphine, whereas the simultaneous administration of naloxone with morphine had variable effects but seemed to delay the LH increase.(ABSTRACT TRUNCATED AT 250 WORDS)     

Bimodal effects of luteinizing hormone and role of androgens in modifying superovulatory responses of rats to infusion with purified porcine follicle-stimulating hormone

Prepubertal (28-30 days old) female rats were infused s.c. over a 60-h period with a purified porcine pituitary follicle-stimulating hormone (FSH) preparation having FSH specific activity 8.4 times that of NIH-FSH-S1 and luteinizing hormone (LH) specific activity less than 0.005 times that of NIH-LH-S1, based on radioreceptor assays. When the FSH infusion rate of this preparation was increased over the range of 0.5-2 units/day (mg NIH-FSH-S1 equivalent), an all-or-none response was observed, with the threshold dose for superovulation being between 1 and 2 units/day. Eleven of twelve rats receiving the 2 units/day dose ovulated a mean +/- SEM of 67 +/- 8 oocytes on the morning of the third day after the beginning of FSH infusion. Addition of human chorionic gonadotrophin (hCG), as a source of LH activity, to a subthreshold (1 U/day) FSH infusion rate resulted in 20% of rats ovulating at an hCG dosage of 50 mIU/day; increasing the hCG infusion to 200 mIU/day concomitant with the subthreshold FSH infusion rate increased ovulation rate to a mean of 69 +/- 8/rat, with 100% of rats ovulating. To determine the effect of varying both FSH infusion rates and LH:FSH ratios, FSH was infused at several rates, with hCG added to give varying hCG:FSH ratios for each FSH infusion rate. Administration of hCG alone was ineffective in causing ovulation except at the highest infusion rates. Adding hCG to FSH to reach a ratio of 0.2 IU hCG/U FSH significantly increased the superovulatory response to an intermediate, 1 U/day FSH dose, but not to the low, 0.5 U/day dose.(ABSTRACT TRUNCATED AT 250 WORDS)     

Pituitary and ovarian hormone secretion and ovulation in gilts injected with gonadotropins during and after oral administration of progesterone agonist (Altrenogest)

We determined changes in plasma hormone concentrations in gilts after treatment with a progesterone agonist, Altrenogest (AT), and determined the effect of exogenous gonadotropins on ovulation and plasma hormone concentrations during AT treatment. Twenty-nine cyclic gilts were fed 20 mg of AT/(day X gilt) once daily for 15 days starting on Days 10 to 14 of their estrous cycle. The 16th day after starting AT was designated Day 1. In Experiment 1, the preovulatory luteinizing hormone (LH) surge occurred 5.6 days after cessation of AT feeding. Plasma follicle-stimulating hormone (FSH) increased simultaneously with the LH surge and then increased further to a maximum 2 to 3 days later. In Experiment 2, each of 23 gilts was assigned to one of the following treatment groups: 1) no additional AT or injections, n = 4; 2) no additional AT, 1200 IU of pregnant mare's serum gonadotropin (PMSG) on Day 1, n = 4); 3) AT continued through Day 10 and PMSG on Day 1, n = 5, 4) AT continued through Day 10, PMSG on Day 1, and 500 IU of human chorionic gonadotropin (hCG) on Day 5, n = 5; or 5) AT continued through Day 10 and no injections, n = 5. Gilts were bled once daily on Days 1-3 and 9-11, bled twice daily on Days 4-8, and killed on Day 11 to recover ovaries. Termination of AT feeding or injection of PMSG increased plasma estrogen and decreased plasma FSH between Day 1 and Day 4; plasma estrogen profiles did not differ significantly among groups after injection of PMSG (Groups 2-4). Feeding AT blocked estrus, the LH surge, and ovulation after injection of PMSG (Group 3); hCG on Day 5 following PMSG on Day 1 caused ovulation (Group 4). Although AT did not block the action of PMSG and hCG at the ovary, AT did block the mechanisms by which estrogen triggers the preovulatory LH surge and estrus.     

Gonadotropin-releasing hormone treatment induces follicular growth and ovulation in seasonally anestrous mares

A study was conducted to evaluate the effectiveness of gonadotropin-releasing hormone (GnRH) pulse infusion to stimulate follicular development and induce ovulation in seasonally anestrous standardbred mares. Seventeen mares were selected for use in this experiment, on the basis of a previous normal reproductive history, and were housed under a photoperiod of 8L:16D beginning one week prior to the start of the experiment (second week in January). Mares were infused with 20 micrograms (n = 7) or 2 micrograms (n = 6) GnRH/h, or were subjected to photoperiod treatment only (controls, n = 4). Serum concentrations of luteinizing hormone (LH), follicle-stimulating hormone (FSH), and progesterone did not vary, and neither significant follicular development nor ovulation was observed in any control mare throughout the experimental period (greater than 60 days). By contrast, both groups of GnRH-treated mares showed elevated serum concentrations of LH and FSH within one day after the start of infusion. Mares infused with 20 micrograms GnRH/h had at least one follicle greater than or equal to 25 mm in 7.4 +/- 1.3 (mean +/- SEM) days following the start of infusion, and ovulated in 12.0 +/- 0.7 days. In the 2-microgram-GnRH/h treatment group, a 25-mm follicle was detected in 5.7 +/- 0.7 days, and ovulation occurred after 10.0 +/- 0.3 days of infusion. Ovulation in every instance was followed by a functional luteal phase, as indicated by the profiles of progesterone secretion.(ABSTRACT TRUNCATED AT 250 WORDS)     

Use of deslorelin short-term implants to induce ovulation in cycling mares during three consecutive estrous cycles

Alternatives to human chorionic gonadotropin (hCG) for inducing ovulation in cycling mares over several consecutive cycles were explored. Placebo, one, three or five short-term implants each containing 2.2 mg of gonadotropin-releasing hormone (GnRH) analogue (deslorelin) were administered to cycling mares after identification of a follicle over 30 mm. Mares were treated over three consecutive cycles, and artificially inseminated during the third cycle only. Serum was assayed for concentrations of luteinizing hormone (LH), follicle-stimulating hormone (FSH), and progesterone during each cycle. All deslorelin doses decreased the interval to ovulation (4.0 days, 2.6 days, 2.4 days and 2.0 days), increased the proportion of mares ovulating within 48 h (20.0%, 83.3%, 73.3% and 85.7%), and decreased the diameter of the largest follicle at ovulation (45.8 mm, 38.0 mm, 41.0 mm and 37.2 mm) for the placebo, 2.2 mg, 6.6 mg and 11.0 mg groups, respectively (P < 0.05). The interovulatory interval was lengthened in the 11.0 mg group compared with all other groups (21.2 days, 21.8 days, 26.4 days and 32.7 days for the placebo, 2.2 mg, 6.6 mg and 11.0 mg groups, respectively, P < 0.05). No differences (P > 0.05) were detected in serum concentrations of progesterone or pregnancy rate among the groups. Cycle number had no effect (P > 0.05) on the reproductive parameters or serum concentrations of progesterone. Concentrations of LH were higher in Cycle 3 in the 6.6 mg and 11.0 mg groups, and FSH concentrations were decreased in Cycles 2 and 3 in treated mares. Deslorelin was efficacious for inducing ovulation in cycling mares with no diminished activity over three consecutive cycles. However, ovarian suppression and changes in serum concentrations of gonadotropins were noted at higher doses.     

Induction of reproductive function in anestrous mares using a dopamine antagonist

We investigated the role of dopamine in the regulation of seasonal reproductive activity in mares. Nine seasonal anestrous mares, maintained under a natural photoperiod, were treated daily with a dopamine D2 antagonist, [-]-sulpiride (200 mg/mare, im), beginning February 5 (day of year = 36) until the first ovulation of the year or for a maximum of 58. Nine untreated anestrous mares were maintained under the same conditions. The ovaries were examined by ultrasonography twice a week, and blood was collected three times a week for progesterone, LH, FSH and prolactin determinations. Mean day of first ovulation was significantly advanced for [-]-sulpiride-treated mares than control mares (mean day of year +/- SEM = 77.3 +/- 7.9 and 110.0 +/- 6.8, respectively; P < 0.01). Eight mares ovulated during [-]-sulpiride treatment while one mare failed to ovulate. Ovulation occurred 91 d after the start of treatment or on Day 127. All mares continued to have normal estrous cycles after the first ovulation. First cycle length and luteal progesterone concentrations did not differ between [-]-sulpiride-treated and control mares. Plasma prolactin concentrations were significantly increased at 2 and 9 h after [-]-sulpiride administration (P < 0.05), and had returned to basal levels by 24 h. At the time of the LH surge associated with the first ovulation, mean LH and FSH secretion was significantly higher in [-]-sulpiride-treated mares than in control mares (P < 0.05). These results suggest that dopamine plays a role in the control of reproductive seasonality in mares and exerts a tonic inhibition on reproductive activity during the anovulatory season.     

The effect of exogenous progestins on endogenous progesterone secretion in pregnant mares

Twenty-four pregnant, light horse mares were used in a study to determine if an exogenous progestin or progesterone would alter serum concentrations of progesterone. On day 40 of gestation, mares were randomly assigned to one of three administration groups: 1) 250 mg of progesterone in oil every other day, 2) 22 mg of Altrenogest (Regumate, American Hoechst, Somerville, NJ 08876) orally every day, or 3) 10 ml of neobee oil (control) orally every day. The treatment period was from day 40 to 105. Pregnancy status was monitored on days 40, 60, 80, 100 and 105 and a single jugular blood sample was obtained daily from days 40 to 46, 69 to 75 and 99 to 105. Serum concentrations of progesterone were determined by radioimmunoassay. Concentrations of progesterone were similar (P>0.05) among groups at each sampling period. Overall concentrations of progesterone increased (P<0.001) from days 40 to 46. Injection of 250 mg of progesterone in oil failed (P>0.05) to maintain concentrations of progesterone in serum above baseline for 24 hr. Thus higher doses and/or more frequent injections would be needed in order to increase concentrations of progesterone above those seen in untreated controls. In summary, Altrenogest was found to be a nonstressful, convenient method of administering progestins to pregnant mares without altering their endogenous secretion of progesterone.