Rats were immunized against luteinizing hormone releasing hormone (LH-RH) and ovulation and follicular development were studied 12, 24 and 48 weeks later. The abolition of regular cyclic patterns of vaginal smears and the absence of luteal tissue in all but one of 32 rats showed that the immunization was effective in blocking ovulation. Follicular growth varied between rats and appeared to be dependent on whether the inhibition of LH-RH had been sufficient to affect the secretion of basal levels of gonadotrophins. Low levels of gonadotrophins were associated with poor follicular development, uterine atrophy and leucocytic vaginal smears, whereas levels of gonadotrophins similar to those in the dioestrous controls led to adequate follicular growth in the absence of ovulation, the production of cystic follicles, uterine stimulation and persistent vaginal oestrus. A group of rats was ovariectomized 12 weeks after immunization against LH-RH; animals with low antibody titres and large follicles responded with increases in the levels of LH and FSH in the blood, whereas in those with high antibody titres and little follicular development the concentrations of gonadotrophins remained low. The reproductive capacity of rats immunized against LH-RH was tested by caging them with normal male rats from 3 weeks after immunization. Although mating occurred in three rats during the first month, no offspring were produced. No matings occurred in the remaining 41 weeks.
H. M. FRASER and T. G. BAKER
H. M. Picton, C. G. Tsonis and A. S. McNeilly
The hypogonadotrophism model induced by the chronic administration of gonadotrophin-releasing hormone (GnRH) agonist was used to investigate the effects of different concentrations of FSH with or without LH pulses on the stimulation of follicular development in the ewe. Continuous administration of an agonist (buserelin) by osmotic minipump to thirty-six Welsh Mountain ewes from the early luteal phase for 5 weeks resulted in a sustained suppression of the plasma concentration of FSH and inhibited the pulsatile release of LH. The inhibition of gonadotrophin secretion was due to the desensitization and/or down-regulation of pituitary gonadotroph function, since the agonist-treated animals showed no response to a challenge of 1 μg GnRH.
During week 6 of agonist treatment, ewes were infused with either 4-hourly pulses of ovine LH (9 μg/pulse), low concentrations of ovine FSH (3 μg/h) or high concentrations of FSH (9 μg/h) alone or with 4-hourly pulses of LH. After 5 days of gonadotrophin infusion, there was no difference between the mean number of follicles per ewe from the animals treated with LH alone, low concentrations of FSH with or without LH pulses or the high concentration of FSH alone compared with the mean number of follicles from control ewes on day 8 of the luteal phase. Infusion of the high concentration of FSH alone stimulated the development of an increased number of large oestrogenic follicles (follicles > 2·5 mm in diameter and secreting > 3·7 nmol oestradiol/h in vitro) compared with control ewes. The addition of high-amplitude LH pulses to the infusion of the high concentration of FSH prevented follicles developing beyond 2·5 mm in diameter, but doubled the number of small follicles (≤2·5 mm) present in the ovaries.
These results show that normal follicular development can be induced by physiological concentrations of FSH alone in the absence of pulsatile LH release. The addition of high-amplitude LH pulses antagonized this stimulatory effect of FSH on follicle growth in the ewe.
Journal of Endocrinology (1990) 127, 273–283
K. Kato, M. R. Sairam and K. Ramasharma
The acute effects of pooled porcine follicular fluid (PFF), before and after various methods of processing to eliminate steroids, were studied on the luteinizing hormone releasing hormone (LH-RH)-induced release of FSH and LH by whole pituitary glands, from 34-day-old mice, incubated in vitro for 3–4 h. Charcoal treatment of PFF eliminated the steroids and reduced the inhibitory potency on gonadotrophin secretion. On the other hand, dialysis or ultrafiltration (mol. wt > 10 000) did not reduce the inhibitory activity on gonadotrophin secretion.
Of the three steroids tested, only oestradiol at a concentration of 10−10 mol/l inhibited FSH and LH secretion in vitro. This inhibitory effect was counteracted by the inclusion of the oestrogen antagonist tamoxifen in the incubation medium. The presence of tamoxifen did not decrease the suppression of FSH and LH induced by PFF, suggesting that the inhibition observed under the conditions of incubation was not due to oestrogen. Preincubation of mouse pituitary tissue for 1 h with PFF reduced the subsequent release of bioactive FSH and LH induced by LH-RH. The inhibitory effect of PFF was rapid and sustained. The continuous presence of PFF throughout the incubation period was not necessary for manifestations of the inhibitory effects on gonadotrophin release. The suppression of gonadotrophin secretion was related to the dose of PFF with the curve showing a biphasic pattern. The degree of FSH suppression was uniformly greater than that of LH, showing the preferential nature of the inhibitory effect of PFF. At high doses of PFF, the degree of FSH suppression was decreased significantly. This effect on LH release was less pronounced.
The inhibition caused by PFF in the in-vitro incubation procedure was not due to destruction of LH-RH or the released gonadotrophins.
G. A. Schuiling, N. Pols-Valkhof, H. Moes and T. R. Koiter
Suppression of gonadotrophin secretion during prolonged treatment with a high dose of LHRH was studied. Long-term ovariectomized rats were infused for 6 days with various doses of LHRH (25, 50, 100, 250 or 500 ng/h) with or without simultaneous treatment with oestradiol benzoate (OB; 3 μg/s.c. injection); a further group was treated with OB only. The effects of these treatments were studied on plasma concentrations of LH and FSH, the pituitary content of LH and FSH and on LH and FSH secretion in vitro (perifusion) in the unstimulated state and following maximal LHRH stimulation (1 μg LHRH/ml perifusion medium).
Administration of LHRH caused a dose-dependent reduction in plasma concentrations of LH and FSH and depleted the pituitary LH/FSH stores. Treatment with OB lowered the plasma concentration of LH to about 30% and that of FSH to about 65% of the control values. Administration of OB plus a low dose of LHRH (25 or 50 ng/h) markedly stimulated the secretion of LH but not of FSH, so that the plasma concentrations of LH were fully restored to the control value. With higher rates of LHRH infusion, OB caused no enhancement of the plasma concentrations of LH and FSH.
The experiments in vitro revealed a sensitizing, i.e. stimulatory effect, of OB on LHRH-stimulated LH and FSH secretion. However, the higher the rate of infusion of LHRH the smaller the sensitizing effect. Interpolation from dose–response curves showed that an LHRH infusion rate of about 150 ng/h (which would establish a plasma concentration of LHRH of about 68 pmol/l) would have no effect. With still higher rates of infusion of LHRH, the effect of OB was either inhibitory (LH) or absent (FSH).
The present experiments show that, depending on dose and length of treatment, both LHRH and OB can have variable effects on the secretion of LH and FSH: LHRH, while depleting and desensitizing the pituitary gland, stimulates the release of LH, whilst OB, which is known to suppress the secretion of LHRH by the hypothalamus, augments the rates of unstimulated and LHRH-stimulated LH and FSH secretion. The augmenting effect of OB on the latter component of LH and FSH secretion can be suppressed with high doses of LHRH. However, these experiments also show that even with doses as high as 500 ng LHRH/h, infused continuously for 6 days, plasma concentrations of LH and FSH are not reduced by more than 60–70%.
J. Endocr. (1987) 115, 469–475
P. G. Knight and R. J. Castillo
Intact and chronically ovariectomized ewes were treated for 4 days with charcoal-treated bovine follicular fluid (FF) or charcoal-treated bovine serum during the late-anoestrous period, and the effects on basal and gonadotrophin-releasing hormone (GnRH)-induced secretion of LH and FSH observed. Subsequently, ewes received s.c. implants containing a sustained-release formulation of a potent GnRH agonist d-Ser(But)6-Azgly10-LHRH (ICI 118630) to desensitize pituitary gonadotrophs to hypothalamic stimulation, and the effects of bovine FF and bovine serum were re-assessed 2 weeks later. Chronic exposure (for 2–3 weeks) to ICI 118630 significantly reduced basal levels of LH and FSH in both intact and ovariectomized ewes and completely abolished both spontaneous LH pulses as well as exogenous GnRH-induced acute increases in plasma LH and FSH levels. Treatment with bovine FF significantly reduced plasma FSH levels, but not LH levels, in both intact and ovariectomized ewes before and after chronic exposure to ICI 118630. In intact ewes before exposure to ICI 118630, treatment with bovine FF actually enhanced pulsatile LH secretion and raised mean plasma LH levels by 240% (P <0·05). No such stimulatory effect of bovine FF on LH secretion was observed in intact ewes exposed to ICI 118630 or in ovariectomized ewes before or after exposure to ICI 118630, suggesting that the effect probably involved an alteration in ovarian steroid feedback affecting hypothalamic GnRH output. Treatment with bovine FF did not significantly affect the magnitude of GnRH-induced surges of LH or of FSH observed in either intact or ovariectomized ewes before exposure to ICI 118630. These observations indicate that charcoal-treated bovine FF, a rich source of inhibin, can directly suppress pituitary FSH secretion in vivo, irrespective of whether a functionally intact hypothalamo-pituitary-ovarian axis is present.
J. Endocr. (1988) 117, 431–439
H. M. FRASER, A. GUNN, S. L. JEFFCOATE and DIANE T. HOLLAND
Autoimmunity to luteinizing hormone releasing hormone (LH-RH) in adult male rats, induced by immunization with LH-RH conjugated to bovine serum albumin, resulted in atrophy of the testes and secondary sex organs and aspermatogenesis. Both immunoreactive luteinizing hormone (LH) and follicle-stimulating hormone (FSH) in serum and the pituitary were reduced to low levels compared with those of control animals. It is suggested that antibodies to LH-RH can inhibit the action of endogenous hormone and that LH-RH is, in fact, the gonadotrophin-releasing hormone in the rat, required for the release of both LH and FSH.
BARBARA M. HARTLEY, D. B. CRIGHTON and G. E. LAMMING
A sheep pituitary incubation system was developed which may be used to study the release of both luteinizing hormone (LH) and follicle-stimulating hormone (FSH) as assayed biologically. Specificity was examined by adding at high concentration various materials present in the hypothalamus, a crude acid extract of sheep cerebral cortex or crude or partly purified extracts of ovine hypothalamic tissue.
Of the materials tested for LH and FSH releasing activity, oxytocin, adrenaline, synthetic lysine vasopressin and cerebral cortex extract failed to influence LH or FSH release. Natural vasopressin containing unknown proportions of the lysine and arginine forms did not affect FSH release but produced a significant increase in the LH content of the medium, the reason for which was not clear. Noradrenaline failed to influence FSH release but produced an apparent depression in the LH content of the medium due to an action on the LH released or upon the ovarian ascorbic acid depletion assay since the addition of noradrenaline to standard ovine LH reduced the potency in the assay to an almost identical degree.
Extracts of ovine hypothalamic tissue consistently increased the LH and FSH content of the medium.
The method described may be applicable to the detection and estimation of LH and FSH releasing activity in ovine hypothalamic tissue extracts and to detecting changes in the responsiveness to hypothalamic stimulation of sheep pituitary tissue.
R. J. ETCHES and K. W. CHENG
The changes in the binding of FSH during follicular maturation were examined in the hen using 125I-labelled bovine FSH (bFSH) and unlabelled bFSH. The binding of 125I-labelled bFSH was not inhibited by bovine LH or chicken LH but was inhibited by extracts of chicken pituitary glands. The ovarian stroma, which contained both interstitial tissue and small follicles, bound the greatest amount of FSH. As the follicles progressed through the yolk-filled hierarchy of maturation, they bound decreasing amounts of FSH. In the two largest follicles of the hierarchy, there was a significant increase in the binding of FSH 12–16 h before ovulation. There were two peaks in the concentrations of LH; a preovulatory peak occurred 4–6 h before ovulation and a second peak occurred 14–16 h before ovulation. Plasma concentrations of testosterone, oestradiol and progesterone began to rise 9, 8 and 6 h, respectively, before ovulation. These data are consistent with the hypothesis that changes in the gonadotrophin concentration and binding regulate the order of the follicular hierarchy and the onset of preovulatory steroidogenesis in the hen.
SUSAN M. EVANS, C. H. TYNDALE-BISCOE and R. L. SUTHERLAND
A heterologous radioimmunoassay for tammar wallaby FSH, using an ovine FSH antiserum and a human FSH tracer, is described. With this assay concentrations of FSH in plasma of intact female tammars are not detectable except rarely at the time of oestrus. However the assay has proved useful in studies of the control of gonadotrophin secretion in intact male and in ovariectomized tammars.
In the female tammar, concentrations of LH and FSH in plasma rose within a few days of bilateral ovariectomy. Ovariectomized tammars respond to a luteinizing hormone releasing hormone stimulus (10 μg, i.v.) with a prompt release of LH, peak levels of 16·9 ± 1·4 ng NIH-LH-S19/ml plasma (n = 12) being reached within 25 min of injection. Concentrations of LH and FSH in plasma were reduced to preoperative values in ovariectomized tammars when lutein tissue developed in ovarian cortex grafts autotransplanted under the pouch skin. Ovarian interstitial tissue was not necessary for this effect.
After lutectomy during quiescence, the female tammar ovulates again in about 14 days. Injections of progesterone (700 μg/kg per day, i.m.) for 7 days after the operation did not delay this response, but follicular development and ovulation appeared to be retarded in animals given oestradiol-17β (5 μg/kg per day, i.m.) with or without progesterone.
G. Bolet, M. Meunier, M.R. Blanc, L. Martinet and J.C. Poirier
Mating induces a surge of both LH and FSH in the blood of female rabbits, followed 10–12 h later by a surge of FSH only, which begins at the time of ovulation. We have studied the effect of suppression of ovulation on the post-ovulatory surge of FSH. In the first experiment, follicular fluid and oocytes were withdrawn from the largest follicles 8 h after coitus. In the second experiment, ovulation was inhibited by injecting the rabbits with 25 mg indomethacin/kg body weight 7·5 h after mating. Levels of serum FSH and LH were measured for 24-48 h after mating. Control rabbits ovulated normally in both experiments. The treatments did not significantly affect the levels of serum FSH in either experiment, although the second surge of FSH was slightly higher after fluid had been aspirated from the preovulatory follicles. These observations show that the post-ovulatory surge of serum FSH is not dependent upon the completion of ovulation and that it is programmed before 7·5–8 h post coitum.
J. Endocr. (1987) 112, 57–61