Effect of the peripubertal pattern of LH and FSH secretion on in-vitro oestradiol-17 beta secretion and follicular growth within juvenile rat ovaries

Juvenile rat ovaries were placed in perfusion culture and exposed to (1) no gonadotrophin, (2) tonic NIH-FSH (200 ng RP-1 equiv./ml) or (3) NIH-FSH + NIH-LH pulses (2/h, amplitude = 80 ng RP-1 equiv./ml). After 3 h of perifusion, the ovaries were prepared for histological analysis and the perifusate assayed for oestradiol-17 beta. Since the NIH-FSH preparation is contaminated with LH, a second experiment was conducted using recombinant bovine LH and FSH. Ovaries were perifused for 3 h with (1) no hormones, (2) recombinant FSH (200 ng/ml) or (3) recombinant FSH plus 25 ng recombinant LH/ml. NIH-FSH alone increased the number of mid-size antral follicles (P less than 0.05) and decreased the number of small antral follicles (P less than 0.05). Pulsatile LH in the presence of FSH increased the number of mid-size antral follicles without reducing the number of small antral follicles. Studies with recombinant FSH and LH demonstrated that both FSH and LH are necessary to stimulate follicles to grow, indicating that the growth-promoting property of the NIH-FSH is due to the presence of both FSH and LH. Regardless of whether NIH or recombinant gonadotrophins were used, follicular growth was induced without increasing oestradiol secretion. These results demonstrate that enhanced oestradiol secretion is not essential for the induction of follicular growth, while both LH and FSH are necessary to stimulate small antral follicles to grow into mid-size antral follicles.(ABSTRACT TRUNCATED AT 250 WORDS)     

Changes in rat ovaries of specific binding for LH, FSH and prolactin during the oestrous cycle and pregnancy.

In cyclic rats, the highest ovarian specific binding for LH was 6-0+/- 2-2% inpro-oestrus. During pregnancy, the specific binding of 125I-labelled bovine LH by rat ovaries increased gradually and reached a maximum of 24-1+/-4-9% between Days 14 and 18 of gestation; a slight decrease in binding was observed at Day 20 of pregnancy. Ovarian specific binding for FSH was also highest in pro-oestrus (8-9+/-2-1%), decreasing to about 50% in oestrus and metoestrus, but staying relatively constant during pregnancy. For prolactin, the specific binding in rat ovaries was highest (7-1+/-1-6%) in pro-oestrus, quite high in metoestrus and dioestrus and low in oestrus. Specific binding increased gradually only after Day 14 of pregnancy. Serum concentrations of rat LH, FSH and prolactin at different stages of the oestrous cycle and during pregnancy were determined by radioimmunoassays, and no obvious correlation was observed between levels of circulating hormones and the specific binding of these hormones in ovarian tissues. Affinity constants (Ka) for the hormones were very similar between ovaries from pro-oestrous rats and late-pregnant rats, being 0-31 X 10(9) M-1 for LH, 0-65 X 10(10)M-1 for FSH, and 1-14 X 10(10)M-1 for prolactin. Increases in specific binding for different hormones were due to increases of total binding sites in the ovary under different physiological states.     

Effect of p-chlorophenylalanine on LH, FSH, prolactin and ovulation in the rat

The effect of p-chlorophenylalanine (PCPA: 300 mg/kg) on the rate of ovulation and plasma LH, FSH and prolactin secretion has been studied in rats at preovulatory periods (18th hour of diestrus) and post-ovulatory periods (9th hour of metaestrus). In both experimental groups, results showed that administration of PCPA caused an increase in both prolactin concentration and number of mature ovarian follicles (p less than 0.001). No changes were observed in FSH levels. LH concentration, however, decreased (p less than 0.001) and ovulation became totally inhibited. Rats treated at the 9th hour of metaestrus exhibited a marked luteinization as well as an increased number of corpus luteum in the ovaric tissue (p less than 0.001), whereas those treated at the 18th hour of diestrus underwent no luteinization and merely showed a greater number of mature ovarian follicles (p less than 0.001). PCPA, therefore, seems not to have a double effect on ovulation, LH, FSH, and prolactin secretion regardless of the pre or post-ovulatory periods. Changes observed in the ovaric tissue might be due to an increase in plasma prolactin concentration which appears earlier in the preovulatory than in the post-ovulatory treated animals. This difference may explain the double effect that has been attributed to the ovaric cycle and reproductive behavior.     

Relationships between LH, FSH and prolactin secretion and reproductive activity in the weaned sow

Blood samples were collected from primiparous sows via indwelling jugular cannulae at 15-min intervals for 12 h before and for 24 h (2 sows) or 48 h (10 sows) after weaning and then every 4 h until behavioural oestrus. Weaning to oestrus intervals ranged from 3 to 10 days and 2 sows showed no signs of oestrus and had not ovulated by Days 11 and 16 after weaning. Prolactin concentrations in plasma decreased significantly (P less than 0.001) and reached basal levels 1-2 h after weaning in all sows whilst plasma progesterone concentrations remained basal until approximately 30 h after the preovulatory LH surge in sows that ovulated. Elevated concentrations of prolactin or progesterone during the post-weaning period were, therefore, not responsible for delayed restoration of cyclicity. Overall, mean LH concentrations rose significantly (P less than 0.001) from 0.22 +/- 0.02 during the 12-h period before weaning to 0.38 +/- 0.03 ng/ml during the 12-h post-weaning period. After weaning, pulsatile and basal LH secretions were markedly increased for sows that showed an early return to oestrus (less than or equal to 4 days) compared with sows showing a longer weaning to oestrus interval but a correlation did not exist between either of these LH characteristics and the time taken to resume cyclicity. Mean LH concentrations before weaning were, however, inversely related (r = -0.649; P less than 0.05) to the weaning to oestrus interval. Overall, mean FSH concentrations rose significantly (P less than 0.001) from 151.1 +/- 6.2 (s.e.m.) ng/ml in the 12-h period immediately before weaning to 187.7 +/- 9.7 ng/ml in the subsequent 12-h period but there was no correlation between FSH concentrations, before or after weaning, and the interval from weaning to oestrus. However, a significant correlation was apparent between ovulation rate and peak concentrations of the rise in FSH after weaning (r = 0.746; P less than 0.05) and overall mean FSH values (r = 0.645; P less than 0.05). It is concluded that both LH and FSH concentrations in peripheral blood rose in response to removal of the suckling stimulus at weanling. The increase in LH pulse frequency associated with weaning was not directly related to the weaning to oestrus interval although a specific pattern of LH secretion was observed in sows showing an early return to oestrus (less than or equal to 4 days).(ABSTRACT TRUNCATED AT 400 WORDS)     

Comparison of the effects of two GnRH antagonists on LH and FSH secretion,follicular growth and ovulation in the mare

The effects of two GnRH antagonists were tested in order to delay and/or synchronise ovulation in mares. Five mares received Antarelix (0.01 mg.kg(-1)), 5 mares received Cetrorelix (the same dose), 5 mares (control mares) received the vehicle intravenously, twice daily, for 8 days from the day the largest follicle reached 22 mm following prostaglandin administration. Ovulation was postponed in all mares injected with Antarelix (19.4 +/- 1.2 days after the beginning of the treatment) and in 2/5 mares injected with Cetrorelix (20 +/- 1 days) vs. 6.2 +/- 0.4 days in control mares. During the treatment, LH concentrations were strongly depressed in Antarelix and in Cetrorelix mares (1.6 +/- 0.1 and 3.8 +/- 0.5 ng.mL(-1) respectively vs. 21 +/- 2.5 ng.mL(-1) in control mares). In the 3 Cetrorelix mares which ovulated during the treatment. 2 initiated their LH surge at this moment. FSH concentrations were not affected in Antarelix or in Cetrorelix mares during the treatment (11.4 +/- 1.3 and 7.9 +/- 0.8 ng.mL(-1) respectively vs. 10.5 +/- 0.8 ng.mL(-1) in control mares). In conclusion, Antarelix seems more efficient than Cetrorelix for postponing ovulation in mares. The role of LH in antral follicular development before the preovulatory stage is confirmed.     

Modulation of testicular receptors for LH, FSH and prolactin by the administration of ovine prolactin in mature rat

To elucidate the role of prolactin on testicular function, we treated mature rats with ovine prolactin (oPRL) and investigated the dose and time-dependent changes in testicular LH, FSH and prolactin receptors as well as in serum gonadotropin and steroid levels. Twelve week-old rats were injected sc with a single dose of various amounts of oPRL (0.2, 1 and 5 IU) and killed on the first, second and third days after the treatment. Testicular LH receptor decreased to 59% of the control level as a function of time while prolactin receptor increased to 244% maximally of the control level on the second day. In contrast, FSH receptor changed in a different fashion. Smaller amounts of oPRL (0.2 and 1 IU) raised the receptor level to 193% of the control level on the first day whereas a larger amount (5 IU) did not change the receptor, which tended to remain in a low level throughout the experimental period. The serum FSH level significantly increased in every group on the second day, then returned to the control range by the third day. On the other hand, the serum testosterone level changed in a characteristic manner, decreased significantly in every group on the first day though not in a dose-dependent fashion, returned to normal on the second day and significantly increased in the 0.2 IU group on the third day (p less than 0.01). Similarly, the serum estradiol level decreased in the oPRL-treated groups on the first day and was restored on the second day.(ABSTRACT TRUNCATED AT 250 WORDS)     

Endothelin-3 inhibits prolactin and stimulates LH, FSH and TSH secretion from pituitary cell culture

The influence of endothelin-3 (ET-3) on anterior pituitary hormone secretion was investigated over a wide range of concentrations (from 10(-14) to 10(-6) M) and incubation times (from 4 to 48 hours). ET-3 elicited a concentration-dependent inhibition of prolactin (PRL) secretion and stimulated the release of luteinizing hormone (LH), follicle stimulating hormone (FSH) and thyroid stimulating hormone (TSH) from primary monolayer cultures of anterior pituitary cells derived from female rats. The responsiveness of different pituitary cells to ET-3 differs markedly in terms of onset and duration: the maximal inhibition of PRL secretion occurred after 12 hours and the stimulation of LH, FSH and TSH reached the maximum after 4, 48 and 48 hours of incubation, respectively. These data corroborate the concept that ET-3 has an important role as a neuroendocrine modulator. Moreover, the data presented suggest different intracellular mechanisms underlying ET-3 actions.     

Role of LH pulse amplitude and the rate of LH increase on oestradiol-17 beta secretion by immature rat ovaries in perfusion culture

Immature rat ovaries were placed in perifusion culture and exposed to LH in a variety of patterns, while tonic levels of FSH (200 ng/ml) were maintained. Perifusate was collected every 5 min and assayed for oestradiol-17 beta. In Exp. I, LH was delivered at 50 ng/ml for 30 min then reduced to 0 ng/ml for an additional 30 min (50 ng/ml LH step) or as linear increasing concentrations reaching 50 ng/ml by 30 min of culture then linear decreasing concentrations reaching 0 ng/ml by 30 min of culture (50 ng/ml LH ramp). Although both delivery systems exposed the tissue to 250 ng LH over the 1-h culture period, only the 50 ng/ml step significantly increased oestradiol-17 beta secretion rates (P less than 0.05). In Exp. II, ovaries were not exposed to LH for the first 30 min of culture but then LH was administered at a concentration of 25 or 50 ng/ml for 30 min. The LH concentration was reduced to 0 ng/ml for an additional 30 min of culture. Oestradiol-17 beta secretion rates were increased 50% by both treatments. These results suggest that the mechanism by which an LH pulse stimulates the immature ovary to secrete oestradiol-17 beta is, in part (1) 'threshold'-dependent and not solely dependent on the amount of LH administered, and (2) dependent on the time required to achieve a 'threshold' level.     

Effects of the intraventricular administration of 5,7-dihydroxytryptamine on FSH, LH and prolactin secretion in neonatally estrogenized male rats

The role of the serotoninergic system in the control of LH, FSH and prolactin secretion was analyzed in control and neonatally estrogenized male rats. Animals injected s.c. with 500 micrograms of estradiol benzoate (EB) on day 1 of life, or their corresponding sham-treated controls, were divided on day 75 into the following groups: (1) orchidectomized; (2) injected intraventricularly with 5,7-dihydroxytryptamine (5,7-DHT); (3) orchidectomized and treated with 5,7-DHT, and (4) sham operated. 15 days later, the animals were decapitated and their FHS, LH and prolactin plasma values measured by specific RIA systems. After the treatment with 5,7-DHT, control animals showed a decline in basal prolactin levels but no modification in basal LH and FSH values. After castration, 5,7-DHT-treated animals showed a reduced LH increase and a more marked prolactin decrease. In neonatal estrogen-treated animals, the 5,7-DHT injection did not change FSH, LH or prolactin levels but did partially or completely abolish the post-castration rise in FSH and LH levels, respectively. These data seem to indicate that neonatal estrogenization induced a modification of the serotoninergic role in the control of LH, FSH and prolactin.     

Effects of oestradiol on LH, FSH and prolactin in ovariectomized ewes

This study was conducted to test the hypothesis that the rate (dose/time) at which oestradiol-17 beta (oestradiol) is presented to the hypothalamo-pituitary axis influences secretion of LH, FSH and prolactin. A computer-controlled infusion system was used to produce linearly increasing serum concentrations of oestradiol in ovariectomized ewes over a period of 60 h. Serum samples were collected from ewes every 2 h from 8 h before to 92 h after start of infusion, and assayed for oestradiol, LH, FSH and prolactin. Rates of oestradiol increase were categorized into high (0.61-1.78 pg/h), medium (0.13-0.60 pg/h) and low (0.01-0.12 pg/h). Ewes receiving high rates of oestradiol (N = 11) responded with a surge of LH 12.7 +/- 2.0 h after oestradiol began to increase, whereas ewes receiving medium (N = 15) and low (N = 11) rates of oestradiol responded with a surge of LH at 19.4 +/- 1.7 and 30.9 +/- 2.0 h, respectively. None of the surges of LH was accompanied by a surge of FSH. Serum concentrations of FSH decreased and prolactin increased in ewes receiving high and medium rates of oestradiol, when compared to saline-infused ewes (N = 8; P less than 0.05). We conclude that rate of increase in serum concentrations of oestradiol controls the time of the surge of LH and secretion of prolactin and FSH in ovariectomized ewes. We also suggest that the mechanism by which oestradiol induces a surge of LH may be different from the mechanism by which oestradiol induces a surge of FSH.     

On the correlation between FSH, LH and prolactin serum levels.

In 188 males FSH, LH, and prolactin serum levels determined from a single blood sample were found to be closely correlated. No correlation appeared to testosterone levels. The same correlation is observed, if serum levels of FSH, LH, and prolactin are measured after stimulation with LH-RH and TRH. In order to explain the close correlation, in five young men hormone levels were measured at 2-min-intervals over a period of 2 hours. Peaks of prolactin often correspond to those of FSH and LH, and a statistical correlation was found in two cases between FSH and prolactin. Results suggest a common releasing mechanism, which is superposed to the main mediating mechanism.     

Reconstituted basement membrane influences prolactin, LH, and FSH secretion from adult and fetal adenohypophyseal cells in vitro.

These investigations tested the hypothesis that secretion of prolactin (PRL), LH, and FSH in vitro is influenced by the substratum on which adult or fetal adenohypophyseal cells are cultured. Adenohypophyses were removed from adult male Golden Syrian hamsters and from fetal hamsters on day 16 of gestation. The glands were dissociated and cultured in a 1:1 mixture of Dulbecco's Modified Eagle's Medium (DMEM) and Ham's F-12 medium containing 10% fetal bovine serum (FBS), 25 mM Hepes, and antibiotics. The cells were cultured on three substrata: glass, laminin, and the reconstituted basement membrane of the Engelbreth-Holm-Swarm (EHS) tumor (Matrigel). Medium was collected and replaced every 48 h for 14-22 days. Concentrations of PRL, LH, and FSH in medium were measured by RIA. The substratum influenced hormone secretion. PRL concentrations were elevated in cultures of adult cells on Matrigel in each of four experiments. Adenohypophyseal cells on Matrigel maintained a rounded shape longer than cells on glass or laminin. In studies using fetal adenohypophyseal cells, PRL concentrations were elevated significantly in medium from cultures on Matrigel at and after 2 days as were concentrations of LH and FSH after 6 days. Additional experiments showed that the higher PRL concentrations in medium surrounding adult cells plated on Matrigel were not due to the release of soluble factors from Matrigel, differential cell attachment on Matrigel, the differential presence of adenohypophyseal fibroblasts, nor differential rates of cell proliferation. The results show that Matrigel maintains the secretion of PRL from adult adenohypophyseal cells in vitro more effectively than glass or laminin substrata and support the hypothesis that cell-matrix interactions mediate the observed differences. The results also show that in long-term cultures (14-22 days), fetal adenohypophyseal cells secrete significantly more PRL, LH, and FSH on Matrigel than they secrete when cultured on glass or laminin. Thus, Matrigel influences the function and possibly the maturation of adenohypophyseal cells in vitro. Furthermore, although laminin is the most abundant component in Matrigel, the effects of Matrigel on lactotrophs and gonadotrophs in vitro are probably not attributable solely to its laminin content.     

Effects of FSH and LH on ovarian and follicular blood flow, follicular growth and oocyte developmental competence in young and old mares

Objectives of the experiment were to determine the effects of mare age and gonadotropin treatments on dominant follicle vascularity, ovarian blood flow and dominant follicle growth and to associate follicular vascularity with oocyte developmental capacity. Growing follicles >30mm from young (4-9 years) and old (>20 years) mares were assessed for blood flow using color Doppler ultrasonography before maturation induction with recombinant equine LH (eLH) and immediately prior to oocyte collection at 20-24h after eLH. Pulsed Doppler was used to obtain resistance indices of ovarian arteries ipsilateral to preovulatory follicles. For eFSH-treated estrous cycles, eFSH administration was started after detection of a cohort of follicles ≥20 to <25mm and continued until a follicle >30mm. Oocytes were harvested using transvaginal, ultrasonic-guided aspirations and cultured and injected with sperm at 40±1h after eLH. Presumptive zygotes were incubated, and rates of cleavage (≥2 cells) and blastocyst formation were obtained. Embryos were transferred nonsurgically into recipients' uteri, and pregnancy rates were assessed. Vascularity (number of color pixels per total pixels) was higher (P=0.003) in the follicles of old compared to young mares, with no significant interaction of eFSH or eLH. Effects of eFSH and time from eLH on follicle vascularity were not significant. The vascularity of follicles associated with oocytes that did compared to those that did not form blastocysts was greater (P=0.048), although follicular vascularity was less (P=0.02) for follicles associated with oocytes that did compared to those that did not develop into pregnancies. Resistance indices were not different for age, eFSH treatment, time after eLH administration and oocyte developmental potential. Growth of the dominant follicle was not associated with vascularity, although advanced age tended (P=0.09) to have a negative effect on follicle growth.     

Low serum levels of FSH, LH and prolactin in luteal phase inadequacy

In order to elucidate the relationship between prolactin (PRL) levels and corpus luteum function in humans, assessment of temporal relationship between levels of PRL, LH, FSH, estradiol and progesterone was made in eleven normal cycling women and six short luteal women. All hormones were determined by specific radioimmunoassay. The mean PRL level in the luteal phase was higher than that in the follicular phase in normal women. On the other hand, no difference mean was seen between the PRL levels of follicular and luteal phases in short luteal women. In addition, follicular and luteal phase secretion of PRL in the short luteal phase (SLP) was lower than that in the normal control. LH and FSH in the follicular and luteal phases, estradiol secretion in the late follicular and early to mid-luteal phases in SLP were also lower than those in the control. These observations were consistent with the hypothesis that SLP is a sequel to aberrant folliculogenesis. In addition, it is inferred that low PRL levels in the SLP might be due to inadequate augmentation by estrogen, rather than giving PRL any positive controlling role in the maintenance of corpus luteum function.