Suppression of spermatogenesis in man induced by Nal-Glu gonadotropin releasing hormone antagonist and testosterone enanthate (TE) is maintained by TE alone

GnRH antagonists plus testosterone (T) suppress LH and FSH levels and inhibit spermatogenesis to azoospermia or severe oligozoospermia. High-dose T treatment alone has been shown to be an effective male contraceptive (contraceptive efficacy rate of 1.4 per 100 person yr). Combined GnRH antagonist and T induces azoospermia more rapidly and at a higher incidence than T alone; this combination has therefore been proposed as a prototype male contraceptive. However, because GnRH antagonists are expensive to synthesize and difficult to deliver, it would be desirable to rapidly suppress sperm counts to low levels with GnRH antagonist plus T and maintain azoospermia or severe oligozoospermia with T alone. In this study, 15 healthy men (age 21-41 yr) with normal semen analyses were treated with T enanthate (TE) 100 mg im/week plus 10 mg Nal-Glu GnRH antagonist sc daily for 12 weeks to induce azoospermia or severe oligozoospermia. At 12-16 weeks, 10 of 15 subjects had zero sperm counts, and 14 of 15 had sperm counts less than 3 x 10(6)/mL. The 14 who were suppressed on combined treatment were maintained on TE alone (100 mg/week im) for an additional 20 weeks. Thirteen of 14 subjects in the TE alone phase had sperm counts maintained at less than 3 x 10(6)/mL for 20 weeks. Ten remained persistently azoospermic or had sperm concentration of 0.1 x 10(6)/mL once during maintenance. Mean LH and FSH levels in the subjects were suppressed to 0.4+/-0.2 IU/L and 0.5+/-0.2 IU/L in the induction phase, which was maintained in the maintenance phase. The 1 subject who failed to suppress sperm counts during induction had serum LH and FSH reduced to 0.3 and 0.5 IU/L, respectively. The subject who failed to maintenance had LH and FSH suppressed to 1.0 and 0.2 IU/L, respectively, during the induction phase but these rose to 1.6 and 2.1 IU/L, respectively, during maintenance. Failure to suppress or maintain low sperm counts may be related to incomplete suppression of serum LH and FSH levels. We conclude that sperm counts suppressed with GnRH antagonist plus T can be maintained with relatively low dose TE treatment alone. This concept should be explored further in the development of effective, safe, and affordable hormonal male contraceptives.     

Evidence suggesting an additional control mechanism regulating episodic secretion of luteinizing hormone and follicle stimulating hormone in pre-pubertal children and post-menopausal women

The possible differential regulation of pulsatile follicle stimulating hormone (FSH) and luteinizing hormone (LH) secretion in pre-pubertal children and in post-menopausal women was investigated. Children were studied for 4 h and post-menopausal women for 6 h; blood samples were taken every 10 min. Post-menopausal women were studied before and 21 days after administration of a single i.m. dose of gonadotrophin-releasing hormone (GnRH) analogue. Eight post-menopausal women and 18 children (nine boys and nine girls) were enrolled. The children were divided into two groups: A, at Tanner stages 0-1 (four boys and three girls); B, at Tanner stage 2-3 (five boys and six girls). Plasma LH and FSH concentrations were determined using an immunofluorimetric assay. Time series were analysed and the specific concordance (SC) index was computed to determine the degree of concordance between episodes of LH and FSH secretion. While children of group A had LH concentrations below the minimal detectable dose of 0.1 IU/l, group B showed measurable LH plasma concentrations (1.4 +/- 0.3 IU/l, mean +/- SEM). Plasma FSH concentrations were detectable in both groups. Group A showed FSH plasma concentrations significantly lower than those of group B (0.75 +/- 0.2 and 1.95 +/- 0.4 IU/l respectively; P < 0.05), but FSH pulse frequency was higher in group A (P < 0.05). Children of group B showed significant concomitance of LH and FSH secretory events at time 0 (P < 0.05).(ABSTRACT TRUNCATED AT 250 WORDS)     

Acute effects of estradiol infusion and naloxone on luteinizing hormone secretion in pubertal boys

We have shown previously in pubertal boys that testosterone (T) suppresses the nocturnal augmentation of luteinizing hormone (LH) secretion principally by decreasing LH pulse frequency. As T can be aromatised to estradiol (E2), and E2 effects on LH secretory dynamics may be separate from those of T, we examined the effects of acute E2 infusion on LH secretion in pubertal boys. Opioid receptor blockade has been reported to increase LH secretion after estradiol suppression in adult men, so we also examined whether naloxone might augment LH secretion during E2 treatment in pubertal boys. Starting at 1000 h, eight pubertal boys were given a 33 h saline infusion, followed 1 week later by an E2 infusion at 4.6 nmol/m2/h. During both infusions, four iv boluses of saline were given hourly beginning at 1200 h on the first day, and four naloxone iv boluses, 0.1 mg/kg each, were given hourly beginning at 1200 h on the second day. Blood was obtained every 15 min for LH, and every 60 min for T and E2, from 1200 h until the end of the infusion. Pituitary responsiveness to gonadotropin-releasing hormone (GnRH) was assessed after both infusions by iv administration of 250 ng/kg synthetic GnRH. Estradiol infusion increased the mean plasma E2 concentration from 23 +/- 4 to 46 +/- 6 pmol/L (P < 0.01) and suppressed mean plasma T from 4.9 +/- 1.4 to 3.0 +/- 3.5 nmol/L (saline vs. E2 infusion, P < 0.05). The overall mean LH was suppressed by E2 infusion from 3.7 +/- 0.5 to 2.2 +/- 0.4 IU/L (saline vs. E2 infusion, P < 0.01). LH pulse frequency was suppressed by 50%, whereas mean LH pulse amplitude was not different between saline and E2 infusions. Administration of naloxone did not alter the mean LH, LH pulse frequency, or amplitude during either saline or E2 infusions. Pituitary responsiveness to exogenous GnRH was similar during both infusions. These studies indicate that E2 produces its negative feedback in pubertal boys principally by suppression of LH pulse frequency, and naloxone does not reverse these suppressive effects. Thus E2 suppression of LH secretion is mediated by a decrease of hypothalamic GnRH secretion that is independent of endogenous opioid pathways.     

Erythropoietin stimulates testosterone production in man

Human recombinant erythropoietin (rHuEPO) treatment improves sexual function in end-stage renal failure patients with a still-debated mechanism. Experimental data suggested that rHuEPO was able to stimulate rat Leydig steroidogenesis; therefore, it has been suggested that rHuEPO may induce its effects in humans by acting on gonadal steroid production. Thirteen young adult males (age range, 16-28 yr) catheterized at peripheral and left internal spermatic venous levels during a contrast study for varicocele, were studied. In five subjects, rHuEPO (60 IU/kg, up to a maximum of 4000 IU total) was injected over 1 min in the cubital vein. Similarly, in other five patients, 50 micrograms GnRH were infused. In three subjects, 2 mL saline were injected, as controls. Plasma LH, FSH, and testosterone (T) levels were then determined at -15, 0, 15, 30, 45, 60, 90, and 120 min simultaneously in peripheral and spermatic venous blood. rHuEPO infusion did not have any effect on plasma LH and FSH levels in peripheral or spermatic veins. Similarly, rHuEPO infusion did not affect peripheral T concentration, but increased (approximately 400% vs. controls; P < 0.05) spermatic T levels. GnRH infusion induced an increase in plasma LH and FSH levels in both peripheral and spermatic veins. After GnRH infusion, an increase of approximately 12-fold (P = 0.05-0.001) in T was observed only at the spermatic venous level, without any peripheral T variation. These findings show that rHuEPO was able to influence testicular steroidogenesis by stimulating T production in man, whereas the absence of any effect on gonadotropin secretion suggests that rHuEPO might act directly on human Leydig cell function.     

Systematic relationships among pocket gopher chewing lice (Phthiraptera: Trichodectidae) inferred from electrophoretic data

This study was designed to explore the efficacy of gonadotrophin-releasing hormone (GnRH) to antagonize the effect of gonadotrophin surge-inhibiting factor (GnSIF) on the timing of the induction by GnRH of the maximal self-priming effect on pituitary LH responsiveness. The GnSIF levels were increased by FSH treatment and reduced after gonadectomy. Female rats were injected s.c. with 10 IU FSH or saline (control) on three occasions during the 4-day cycle. Serial i.v. injections of GnRH (500 pmol/kg body weight) were administered to intact rats on the afternoon of pro-oestrus or 15-30 min after ovariectomy. Intact male rats were given 10 IU FSH and 500 or 2000 pmol GnRH/kg body weight on an equivalent time-schedule. Endogenous GnRH release was suppressed with phenobarbital. In intact female control rats, the timing of the maximally primed LH response was delayed as the GnRH pulse-interval increased. FSH treatment of female rats induced a suppression of the initial unprimed LH response and delayed the maximally primed LH response, which showed further delay as the GnRH pulse-interval was increased. When the pulsatile administration of GnRH was started 15-30 min after ovariectomy, the priming effect of GnRH did not change as the GnRH pulse-interval was increased in the saline-treated rats. However, FSH treatment caused a suppression of the unprimed LH response, a delay in the primed LH response and decreased the delay of the maximally primed LH response to GnRH when the GnRH pulse-interval was decreased. Increasing the interval between ovariectomy and the first GnRH pulse to 4 h diminished the efficacy of the FSH treatment: GnRH-induced priming was delayed by only one pulse instead of the two pulses in control rats. In intact males but not in orchidectomized rats, a self-priming effect was demonstrated during GnRH pulses which were 1 h apart. The effect of 2 nmol GnRH/kg body weight was the most pronounced. Compared with intact female rats, the timing of the maximally primed LH response was delayed by 1 h. FSH treatment did not affect the pituitary LH response to both dose levels of GnRH. It is concluded that FSH treatment increased the release of GnSIF by the ovary, then induced a state of low responsiveness of the pituitary gland to GnRH and subsequently delayed GnRH-induced maximal self-priming. The efficacy of GnRH to prime the pituitary gland was higher when GnSIF levels were decreasing after removal of the ovaries.(ABSTRACT TRUNCATED AT 400 WORDS)     

Changes in gonadotrophin response to gonadotrophin releasing hormone in normal women following bilateral ovariectomy

OBJECTIVE: Pituitary responsiveness to GnRH varies throughout the normal menstrual cycle. We have investigated whether there are differences in the ovarian mechanisms which regulate gonadotrophin secretion between the follicular and the luteal phase of the cycle. DESIGN: Normally ovulating women were studied during the first week following hysterectomy plus bilateral ovariectomy performed either in the mid- to late follicular phase (follicle size 16 mm) or in the early to midluteal phase (5 days post LH peak). The response of LH to a single dose of 10 micrograms GnRH was investigated 2 hours before the operation and every 12 hours after the operation until postoperative day 4 and every 24 hours until day 8. PATIENTS: Fourteen normally cycling premenopausal women with normal FSH (< 10 IU/l). Seven women were ovariectomized in the follicular and 7 in the luteal phase. MEASUREMENTS: Pituitary response to GnRH was calculated as the net increase in FSH (delta FSH) and LH (delta LH) at 30 minutes above the basal value. RESULTS: Basal levels of FSH and LH before the operation were significantly lower in the luteal than the follicular phase (P < 0.05), while those of oestradiol (E2) were similar. Also, similar were delta LH and delta FSH values. Serum progesterone and immunoreactive inhibin (Ir-inhibin) concentrations before the operation were higher in the luteal than the follicular phase (P < 0.05). Following the operation, serum E2, progesterone and Ir-inhibin values declined dramatically, while basal FSH and LH as well as delta FSH values showed a gradual and significant increase. The percentage increase in FSH and LH values (mean +/- SEM) on day 8 after the operation was similar in the follicular (453 +/- 99% and 118 +/- 35% respectively) and the luteal phase (480 +/- 71% and 192 +/- 45% respectively). In contrast to delta FSH, delta LH values after a temporal increase 12 hours from the operation, remained stable in the follicular phase and declined significantly in the luteal phase up to day 4. CONCLUSIONS: Basal gonadotrophin secretion during the normal menstrual cycle is predominantly under a negative ovarian effect. It is suggested that in contrast to FSH, the secretion of LH in response to GnRH is controlled by different ovarian mechanisms during the two phases of the menstrual cycle.     

Recessive Robinow syndrome: with emphasis on endocrine functions

We present the characteristic features of 14 children with the recessive form of Robinow syndrome and the growth hormone (GH) response to provocation with clonidine and the serum insulin-like growth factor-I (IGF-I) concentration in 12 of these children. The gonadotropin (luteinizing hormone [LH] and follicle-stimulating hormone [FSH]) response to gonadotropin-releasing hormone (GnRH) was evaluated in early pubertal and pubertal patients, and the testosterone response to human chorionic gonadotropin (HCG) was evaluated in males. Children with Robinow syndrome, born at full-term, were short at birth (length, 41.4+/-2.1 cm) and had markedly slow growth velocity (GV) during the first year (13.1+/-2.1 cm/yr); consequently, they were significantly short at the end of the first year of life (length, 54.4+/-2.9 cm). This intrauterine and early extrauterine growth delay reflected low growth potential. During childhood, the GV standard deviation score (GVSDS) remained low (-2.17+/-0.83). Despite the presence of empty sella in all of the patients, they had an adequate GH response to clonidine provocation (peak, 19.3+/-5.8 microg/L) and a normal serum IGF-I concentration (309+/-142 ng/mL) for their age. During childhood and early adolescence, boys with Robinow syndrome had low basal testosterone and a low testosterone response to HCG stimulation (3,000 IU/m2/d intramuscularly [IM] for 3 days). However, their basal and GnRH-stimulated FSH concentrations were normal. Two girls (Tanner II breast development) had a normal serum estradiol (E2) concentration but high LH and FSH responses to GnRH stimulation. This suggested either defective feedback of E2 on the hypothalamic-pituitary axis or hyporesponsiveness of the ovaries to gonadotropin. Four weeks of HCG therapy (2,500 IU/m2 IM twice weekly) in three boys with Robinow syndrome increased the penile length and testicular volume, denoting a significant Leydig cell response to prolonged HCG stimulation and the presence of functioning androgen receptors. It is suggested that HCG and/or testosterone therapy during infancy may improve the severe micropenis in these patients.     

In pubertal girls, naloxone fails to reverse the suppression of luteinizing hormone secretion by estradiol

Estradiol (E2) negative feedback on LH secretion was examined in 10 pubertal girls, testing the hypothesis that E2 suppresses LH pulse frequency and amplitude through opioid pathways. At 1000 h, a 32-h saline infusion was given, followed 1 week later by an E2 infusion at 13.8 nmol/m2 x h. During both infusions, four iv boluses of saline were given hourly beginning at 1200 h, and four naloxone iv boluses (0.1 mg/kg each) were given hourly beginning at 1200 h on the following day. Blood was obtained every 15 min for LH determination and every 60 min for E2 determination from 1200 h to the end of the infusion. E2 infusion increased the mean serum E2 concentration from 44+/-17 to 112+/-26 pmol/L (P < 0.01). The mean LH concentration between 2200-1200 h decreased from 3.19+/-0.89 to 1.99+/-0.65 IU/L (P = 0.014), and LH pulse amplitude decreased from 3.4+/-0.6 to 2.6+/-0.5 IU/L (P = 0.0076). Although there were 1.2 fewer pulses during E2 infusion compared to saline infusion, differences did not reach significance (P = 0.1; 95% confidence interval for the difference, -3.5, 1.1). Pituitary responsiveness to GnRH, assessed at the end of the infusion by administering 250 ng/kg GnRH iv, did not change during E2 infusion. The effect of naloxone blockade of opioid activity on LH secretion was determined by assessing the area under the curve (AUC) from 1200-1600 h. During saline infusion, the LH AUC was 1122+/-375 IU/L during saline boluses and 1575+/-403 IU/L during naloxone boluses (P = 0.39). When E2 was infused, the LH AUCs during saline and naloxone boluses were 865+/-249 and 866+/-250 IU/L, respectively. Thus, in pubertal girls: 1) E2 decreases the LH concentration and LH pulse amplitude; 2) the main site of negative feedback effect of E2 appears to be at the level of the hypothalamus; 3) an increase in LH secretion after naloxone administration could not be demonstrated in these girls and may depend on the maturity of the hypothalamic-pituitary-gonadal axis; and 4) opioid receptor blockade does not reverse the E2 inhibition of LH secretion even in the most mature girls. Thus, E2 suppression of LH secretion in pubertal girls appears to be mediated by a decrease in hypothalamic GnRH secretion that is independent of opioid pathways.     

A bolus of recombinant human follicle stimulating hormone at midcycle induces periovulatory events following multiple follicular development in macaques

The efficacy of follicle stimulating hormone (FSH) as an alternative to luteinizing hormone (LH)/human chorionic gonadotrophin (HCG) for the initiation of periovulatory events in primate follicles is unknown. A single bolus of 2500 IU recombinant (r)-hFSH was compared to 1000 IU r-HCG for its ability to promote oocyte nuclear maturation and fertilization, granulosa cell luteinization and corpus luteum function following r-hFSH (60 IU/day) induction of multiple follicular development in rhesus monkeys. Following the r-hFSH bolus, bioactive luteinizing hormone concentrations were <3 ng/ml. Peak concentrations of serum FSH (1455+/-314 mIU/ml; mean+/-SEM) were attained 2-8 h after r-hFSH, and declined by 96 h. Bioactive HCG concentrations peaked between 2-8 h after r-HCG and remained > or = 100 ng/ml for >48 h, while immunoreactive FSH concentrations were at baseline. The proportion of oocytes resuming meiosis and undergoing in-vitro fertilization (IVF) were comparable for r-hFSH (89%; 47+/-19%) and r-HCG (88%; 50+/-17%). In-vitro progesterone production and expression of progesterone receptors in granulosa cells did not differ between groups. Peak concentrations of serum progesterone in the luteal phase were similar, but were lower 6-9 days post-FSH relative to HCG. Thus, a bolus of r-hFSH was equivalent to r-HCG for the reinitiation of oocyte meiosis, fertilization and granulosa cell luteinization, but a midcycle FSH surge did not sustain normal luteal function in primates.     

Characterization of reproductive hormonal dynamics in the perimenopause

Medical therapy for women in the perimenopausal period is controversial, in part due to varying degrees of ovarian hormone secretion characteristic of this time of life. To extend our understanding of the reproductive endocrine milieu of perimenopausal women, we studied 6 cycling women, aged 47 yr and older, for 6 months with daily collections of first morning voided urine. Five additional older reproductive aged (43-47 yr old) women were studied with daily urine and serum sampling for a single menstrual cycle; their urinary hormone data were combined with the former group for menstrual cycle comparisons. Urine was assayed for LH, FSH, estrone conjugates, and pregnanediol glucuronide and normalized for creatinine (Cr). Eleven midreproductive aged (19-38 yr old) normally cycling women, 5 women with well defined premature ovarian failure, and 5 women aged 54 yr and older who were at least 1 yr postmenopausal were used for comparison. Perimenopausal women had shorter follicular phases (11 +/- 2 days vs. 14 +/- 1 days; P = 0.031) and, hence, shorter menstrual cycles than midreproductive aged controls. FSH excretion in perimenopausal women was greater than that in younger women (range of means, 4-32 vs 3-7 IU/g Cr; P = 0.0005). LH secretion was overall greater than that in younger normal subjects (range of means, 1.4-6.8 vs. 1.1-4.2 IU/g Cr; P < 0.026). Overall mean estrone conjugate excretion was greater in the perimenopausal women compared to that in the younger women [76.9 ng/mg Cr (range, 13.1-135) vs. 40.7 ng/mg Cr (range, 22.8-60.3); P = 0.023] and was similarly elevated in both follicular and luteal phases. Luteal phase pregnanediol excretion was diminished in the perimenopausal women compared to that in younger normal subjects (range for integrated pregnanediol, 1.0-8.4 vs. 1.6-12.7 microg/mg Cr/luteal phase; P = 0.015). Compared to postmenopausal women, perimenopausal women had more overall estrone excretion (2.5-6.2 ng/mg Cr in postmenopausal women; P = 0.02) and lower mean FSH (range of means for postmenopause, 24-85 IU/g Cr; P = 0.017) and LH (range for postmenopause, 4.3-14.8 IU/g Cr; P = 0.041). Compared to women with premature menopause, perimenopausal women again had lower FSH (range of means for premature menopause, 36-82 IU/g Cr; P = 0.0022), lower LH (range of means for premature menopause, 5.5-23.8 IU/g Cr; P = 0.0092), borderline higher mean estrone conjugates (range of means for premature menopause, 4-44 ng/mg Cr; P = 0.064), and far longer periods of ovarian activity (one to two cycles in prematurely menopausal women vs. three to six cycles in perimenopausal women). We conclude that altered ovarian function in the perimenopause can be observed as early as age 43 yr and include hyperestrogenism, hypergonadotropism, and decreased luteal phase progesterone excretion. These hormonal alterations may well be responsible for the increased gynecological morbidity that characterizes this period of life.     

Modulation of the action of gonadotrophin surge-attenuating factor by gonadotrophin-releasing hormone

Gonadotrophin surge-attenuating factor (GnSAF) is a putative non-steroidal ovarian factor which attenuates the luteinizing hormone (LH) surge in superovulated women through the reduction of the pituitary response to gonadotrophin-releasing hormone (GnRH). The mechanism of action of GnSAF on gonadotrophin secretion was further studied by investigating six normally ovulating women in two cycles--a spontaneous and a follicle-stimulating hormone (FSH)-treated cycle. The response of the pituitary to five consecutive pulses of GnRH was investigated in late follicular phase (follicle size 15 mm) of both cycles. GnRH pulses, 10 micrograms each, were injected i.v. every 2 h and LH was measured in blood samples taken before and 30, 60 and 120 min after each pulse. FSH was injected daily at the fixed dose of 225 IU starting on cycle day 2. Peak values of LH increment occurred 30 min after each pulse. However, maximal LH increment occurred in both cycles after the second GnRH dose. In the FSH cycles the response of LH to the first three pulses was significantly attenuated compared with the spontaneous cycles, while the response to the fourth and fifth pulses was similar in the two cycles. In both cycles, LH increment 30 min post GnRH (net increase above the previous value) was similar after the fourth and fifth pulses. Serum concentrations of oestradiol and immunoreactive inhibin, although higher in the FSH cycles, remained stable throughout the GnRH experimental period in both cycles. These results demonstrate that multiple submaximal doses of GnRH can override the attenuating effect of GnSAF on LH secretion.(ABSTRACT TRUNCATED AT 250 WORDS)     

Favourable effect of olive oil in patients with non-insulin-dependent diabetes. The effect on blood pressure, blood glucose and lipid levels of a high-fat diet rich in monounsaturated fat compared with a carbohydrate-rich diet

The aim of the study was to elucidate the role of the neuropeptide galanin in the regulation of somatotropic and gonadotropic function in normal women. Thirteen normally ovulating (aged 28 to 40 years), non-obese (body mass index, 18.4 to 27.1 kg/m2) women with infertility due to a tubal or male factor were studied. Each woman underwent three tests: (1) bolus intravenous (IV) injection of growth hormone (GH)-releasing hormone (GHRH) (1-29)NH2 1 microgram/kg plus gonadotropin-releasing hormone (GnRH) 100 micrograms at time 0; (2) IV infusion of porcine galanin 500 micrograms in 100 mL saline from -10 minutes; and (3) bolus IV injection of GHRH(1-29)NH2 1 microgram/kg plus GnRH 100 micrograms at time 0 plus IV infusion of porcine galanin 500 micrograms in 100 mL saline from -10 to +30 minutes. All results are expressed as the mean +/- SEM. GH peak after GHRH was 14 +/- 5 micrograms/L; porcine galanin significantly increased serum GH (GH peak, 7.3 +/- 1.2) with respect to baseline levels. No significant differences were observed between either GH peak or GH absolute values after galanin as compared with GHRH alone. Porcine galanin significantly enhanced GH response to GHRH (peak, 31.4 +/- 4.4 micrograms/L) with respect to either GHRH or galanin alone. Luteinizing hormone (LH)/follicle-stimulating hormone (FSH) peaks after GnRH were 16.5 +/- 5.3 and 17.4 +/- 4 IU/L, respectively. Porcine galanin did not cause significant increases in serum LH and FSH levels with respect to baseline.(ABSTRACT TRUNCATED AT 250 WORDS)     

Gonadotrophin and prolactin secretory dynamics in girls with normal puberty, idiopathic precocious puberty and precocious puberty due to hypothalamic hamartoma

OBJECTIVE: This study was designed to test the hypothesis that hypothalamic hamartoma causes precocious puberty through a different neuroendocrine mechanism than that of normal puberty or of idiopathic precocious puberty. DESIGN AND PATIENTS: We compared the pattern of gonadotrophin secretion among 4 girls with precocious puberty due to hypothalamic hamartoma, 27 girls with idiopathic precocious puberty, and 14 girls with normal puberty. All subjects were breast stage 3 or 4. Blood samples were obtained every 20 min for 4 h during the day (1.000 hours to 1400 h) and night (22.00 hours to 0200 h). MEASUREMENTS: LH, FSH, and prolactin were measured in each blood sample. Girls also underwent LHRH-stimulation with measurement of LH and FSH before and after stimulation. RESULTS: There were no significant differences in mean LH level, LH peak amplitude, or LH or FSH peak frequency during either the day or the night among the three diagnostic groups. However, the mean +/- SD LHRH-stimulated peak LH levels were greater in girls with hypothalamic hamartoma than in girls with normal puberty or with idiopathic precocious puberty (194 +/- 142 vs 85 +/- 60 or 66 +/- 54 IU/l, respectively, P < 0.05). The LHRH-stimulated peak FSH level in girls with hypothalamic hamartoma exceeded the level for the normal pubertal girls (31 +/- 19 vs 17 +/- 7 IU/l, P < 0.05), but not the level for the girls with idiopathic precocious puberty (25 + 12 IU/l). The peak LH to peak FSH ratio in the girls with hypothalamic hamartoma exceeded the ratio for the girls with idiopathic precocious puberty (7.3 +/- 3.9 vs 2.6 +/- 3.0 IU/l, P < 0.05), but not the ratio for the normal pubertal girls (5.0 + 2.9). There were no significant differences in mean prolactin level, peak amplitude or frequency, or in the ratio of mean night to mean day prolactin, among the 3 diagnostic groups. CONCLUSIONS: We conclude that spontaneous gonadotrophin and prolactin secretion are similar among girls with hypothalamic hamartoma, idiopathic precocious puberty, or normal puberty. However, the increased LHRH-stimulated peak LH in the girls with hypothalamic hamartoma suggests subtle differences in neuroendocrine regulation that may underlie their more rapid pubertal maturation.     

Evidence for a role for the cyclic adenosine 3',5'-monophosphate/protein kinase-A pathway in regulation of the gonadotropin subunit messenger ribonucleic acids

cAMP regulation of gonadotropin secretion and subunit mRNA levels was studied in pituitary cells perifused with pulses of GnRH. Pituitary cells from 7-week-old male rats castrated at 5 weeks of age were stimulated hourly for 9-24 h with 1-min pulses of GnRH, the adenylate cyclase activator forskolin, the cell-permeable cAMP analog 8-bromo-cAMP (8Br-cAMP), or control medium. Cells were also treated with the nonsteroidal antiinflammatory drug flufenamic acid, which reduces pituitary cAMP levels. During perifusion, the effluent was collected in 10-min fractions for FSH and LH assay. At the completion of perifusion, total RNA was extracted, and gonadotropin subunit mRNA levels were quantitated by Northern analysis. Continuous administration of flufenamic acid gradually reduced the amplitude of GnRH-stimulated FSH and LH pulses to nadir values of 40 +/- 4.7% and 62 +/- 12% of the control value, respectively. Flufenamic acid decreased (P < 0.05) FSH beta and alpha-subunit mRNA levels and blocked the effect of GnRH to lengthen LH beta mRNA. Pulses of forskolin or 8Br-cAMP released LH and FSH, and continuous forskolin or 8Br-cAMP potentiated the gonadotropin stimulatory effect of GnRH. Forskolin or 8Br-cAMP increased (P < 0.05) FSH beta mRNA and alpha-subunit mRNA levels when administered in pulses, but not when administered continuously, and lengthened LH beta mRNA. The Nal-Glu GnRH antagonist blocked the effects of GnRH pulses, but not the effects of 8Br-cAMP or forskolin. In conclusion, lowering intracellular cAMP levels with flufenamic acid attenuated GnRH-stimulated gonadotropin secretion, decreased alpha-subunit and FSH beta mRNA levels, and blocked the effect of GnRH to lengthen LH beta mRNA, whereas 8Br-cAMP or forskolin produced the opposite effect. These data extend previous results which suggested that cAMP modulates gonadotropin secretion and indicate that the cAMP/A-kinase pathway regulates each of the gonadotropin subunit mRNAs.     

The midcycle gonadotropin surge in normal women occurs in the face of an unchanging gonadotropin-releasing hormone pulse frequency

The midcycle gonadotropin surge is a critical event in normal reproductive cycles and requires functional integration of the hypothalamus, pituitary, and ovary. To determine whether a change in GnRH frequency occurs coincident with the onset or termination of the surge in normal women, 20 studies were performed at a sampling interval of every 5 min for up to 36 h. The frequency of pulsatile GnRH secretion was assessed by the use of two surrogate markers of its secretion, LH and free alpha-subunit (FAS). The timing of the studies was prospectively determined by serial ultrasound and previous cycle history, whereas measurements of LH, FSH, estradiol, and progesterone in daily blood samples were used retrospectively to locate the frequent sampling study in relation to the day of ovulation in each individual. The frequent sampling studies were divided into late follicular phase (LFP; days -4 to -2) and early, mid-, and late portions of the midcycle surge (days -1 to 1) in relation to the 95% confidence limits of the LH peak derived from daily samples in 69 normal ovulatory women. The patterns of LH and FAS secretion were pulsatile at all times during the midcycle surge. The amplitude of LH pulsations increased from the LFP and early surge to the midportion of the midcycle surge (5.9 +/- 6 and 15.1 +/- 5 vs. 39.0 +/- 3 IU/L; P < 0.0001) and decreased from the mid- to the late portion of the surge (13.4 +/- 5 IU/L; P < 0.0001). Likewise, the amplitude of FAS pulse increased from the LFP and early surge to the midportion of the surge (82.4 +/- 59 and 153.1 +/- 50 vs. 421.4 +/- 35 ng/L; P < 0.0001) and decreased from the mid- to the late portion of the surge (190.8 +/- 49 ng/L; P < 0.0002). Although there was excellent concordance of pulsatile secretion of LH and FAS, significantly more pulses of FAS were detected than of LH (P < 0.0001). There was no change in frequency (expressed as interpulse interval) between the LFP and the early and midportions of the surge for LH (70.0 +/- 8, 67.5 +/- 7, and 65 +/- 5 min, respectively) or FAS (55.1 +/- 7, 54.6 +/- 6, and 60.0 +/- 4 min). However, there was an increase in LH interpulse interval (decrease in pulse frequency) in the late portion of the surge (87.0 +/- 6 min) compared to the early and midportions of the surge (P < 0.02 and P < 0.0005, respectively).(ABSTRACT TRUNCATED AT 400 WORDS)     

Ontogeny of gonadotropin, testosterone, and inhibin secretion in normal boys through puberty based on overnight serial sampling

To investigate hormonal changes occurring in male puberty, we measured LH, FSH, testosterone, and alpha-inhibin immunoactivity in serum samples drawn every 10 min for 8 h (2100-0500 h) from each of 50 normal prepubertal and pubertal boys, aged 8.4-18.8 yr. We measured gonadotropins with ultrasensitive immunofluorometric assays, and testosterone and alpha-inhibin with RIAs. Unlike previous studies, which indexed pubertal development with Tanner stages, we used testicular volume, a more finely graduated indicator of development, to reveal patterns that were obscured when subjects were grouped by Tanner stage. The overnight mean concentration of each hormone increased with testis volume, but the rate of increase on a logarithmic scale slowed as testes grew. Log LH rose precipitously in the late prepubertal and early pubertal periods and plateaued during mid- and late puberty. Based on fitted regression curves, LH increased about 20-fold (from 0.11 IU/L) between testis volumes of 1 and 10 mL, but only an additional 1.5-fold by 30 mL. The developmental trajectory of log testosterone was like that of log LH, but rose less steeply early in puberty. From 0.14 micrograms/L at a testis volume of 1 mL, testosterone increased about 8.5-fold by 10 mL and an additional 3-fold by 30 mL. In contrast, logarithms of overnight mean FSH and alpha-inhibin concentrations rose at a more nearly constant rate throughout puberty. From 0.62 IU/L at a testis volume of 1 mL, the FSH concentration doubled by 10 mL and increased an additional 1.7-fold by 30 mL. From 270 ng/L at a testis volume of 1 mL, inhibin increased 1.5-fold by 10 mL and an additional 1.3-fold by 30 mL. Overnight pulse amplitudes exhibited developmental trajectories similar to those of the corresponding overnight mean concentrations. The number of LH and testosterone pulses during the sampling period averaged 2.2 and 2.1, respectively, at Tanner stage 1 and increased to 4.5 and 3.2, respectively, at Tanner stage 5. The number of FSH and inhibin pulses remained constant throughout puberty, averaging 3.3 and 3.5, respectively. Pairwise correlations among hormone concentrations were strong, reflecting common increasing trends through puberty; however, after accounting for developmental trends, FSH, LH, and testosterone concentrations remained correlated, whereas inhibin was uncorrelated with each of the other three hormones. Measuring gonadotropins with ultrasensitive assays and analyzing the results on a logarithmic scale as a function of testis volume made clear the dramatic hormonal changes that begin before the clinical changes of puberty.     

Follicle stimulating hormone alone supports follicle growth and oocyte development in gonadotrophin-releasing hormone antagonist-treated monkeys

Both follicle stimulating hormone (FSH) and luteinizing hormone (LH) are proposed requirements for follicular growth and steroidogenesis; however, the role of LH in primate folliculogenesis is unclear. Follicular stimulation by recombinant human FSH (n = 5) with and without recombinant LH (1:1; n = 6) following 90 days of gonadotrophin-releasing hormone (GnRH) antagonist (Antide) treatment in macaques was evaluated. Human chorionic gonadotrophin (HCG) was administered when six follicles > or = 4 mm were observed. Oocytes were aspirated 27 h later and inseminated in vitro. Chronic Antide reduced serum oestradiol and bioactive LH to concentrations observed in hypophysectomized rhesus monkeys. Multiple follicular growth required a longer interval following recombinant FSH (12 +/- 1 days) than recombinant FSH+recombinant LH (9 +/- 0.2 days), but the total number of follicles/animal did not differ between groups. The day prior to HCG, oestradiol concentrations were 4-fold less following recombinant FSH compared to recombinant FSH+recombinant LH. With recombinant FSH, more oocytes completed meiosis to metaphase II (51%) and fertilized (89 +/- 5%) relative to recombinant FSH+recombinant LH (12 and 52 +/- 11% respectively). Follicular growth and maturation in LH-deficient macaques occurred with FSH alone. Thus, LH is not required for folliculogenesis in primates. Higher fertilization rates following follicular stimulation with FSH alone suggest that the presence of LH with FSH (1:1) during the pre-ovulatory interval impairs gametogenic events in the periovulatory period.     

Menstrual abnormalities in women with Cushing's disease are correlated with hypercortisolemia rather than raised circulating androgen levels

Menstrual irregularity is a common complaint at presentation in women with Cushing's syndrome, although the etiology has been little studied. We have assessed 45 female patients (median age, 32 yr; range, 16-41 yr) with newly diagnosed pituitary-dependent Cushing's syndrome. Patients were subdivided into 4 groups according to the duration of their menstrual cycle: normal cycles (NC; 26-30 days), oligomenorrhea (OL; 31-120 days), amenorrhea (AM; > 120 days), and polymenorrhea (PM; < 26 days). Blood was taken at 0900 h for measurement of LH, FSH, PRL, testosterone, androstenedione, dehydroepiandrosterone sulfate, estradiol (E2), sex hormone-binding globulin (SHBG), and ACTH; cortisol was sampled at 0900, 1800, and 2400 h. The LH and FSH responses to 100 micrograms GnRH were analyzed in 23 patients. Statistical analysis was performed using the nonparametric Mann-Whitney U and Spearman tests. Only 9 patients had NC (20%), 14 had OL (31.1%), 15 had AM (33.3%), and 4 had PM (8.8%), whereas 3 had variable cycles (6.7%). By group, AM patients had lower serum E2 levels (median, 110 pmol/L) than OL patients (225 pmol/L; P < 0.05) or NC patients (279 pmol/L; P < 0.05), and higher serum cortisol levels at 0900 h (800 vs. 602 and 580 nmol/L, respectively; P < 0.05) and 1800 h (816 vs. 557 and 523 nmol/L, respectively; P < 0.05) and higher mean values from 6 samples obtained through the day (753 vs. 491 and 459 nmol/L, respectively; P < 0.05). For the whole group of patients there was a negative correlation between serum E2 and cortisol at 0900 h (r = -0.50; P < 0.01) and 1800 h (r = -0.56; P < 0.01) and with mean cortisol (r = -0.46; P < 0.05). No significant correlation was found between any serum androgen and E2 or cortisol. The LH response to GnRH was normal in 43.5% of the patients, exaggerated in 52.1%, and decreased in 4.4%, but there were no significant differences among the menstrual groups. No differences were found in any other parameter. In summary, in our study 80% of patients with Cushing's syndrome had menstrual irregularity, and this was most closely related to serum cortisol rather than to circulating androgens. Patients with AM had higher levels of cortisol and lower levels of E2, while the GnRH response was either normal or exaggerated. Our data suggest that the menstrual irregularity in Cushing's disease appears to be the result of hypercortisolemic inhibition of gonadotropin release acting at a hypothalamic level, rather than raised circulating androgen levels.     

Charge interactions in sperm-egg recognition

The effect of androgens on changes in circulating LH and FSH during pubertal development was examined longitudinally in a 3 year study in male hamadryas baboons. Baboon LH and FSH were measured by a species-specific radioimmunoassay and bioactive LH (B-LH) was measured by the mouse in vitro Leydig cell bioassay. Control baboons (n = 5) progressed normally through puberty. Eight baboons were castrated prepubertally; of these four received testosterone implants at the chronological age (CA) of clinical puberty (4.0 +/- 0.1 yr, mean +/- SEM). The timing of the postcastration rise in B-LH levels ranged between 1 and 15 months later (median 3.5 months) (CA 3.5 +/- 0.2 yr) thus supporting the hypothesis that central activation of gonadotrophins occurs at the time of puberty, independent of gonadal influences. Similar results were seen for immunoreactive-LH (IR-LH) and IR-FSH levels. IR- and B-LH levels continued to rise with age (P < 0.0003) in the untreated castrated baboons, associated with an increased LH B/I ratio. Administration of testosterone resulted in temporary suppression of B-LH, IR-LH and IR-FSH levels; however gonadotrophin levels subsequently rose with age despite increased testosterone levels. Thus the mechanisms initiating puberty involve both gonad-independent events as well as alterations in negative androgenic feedback sensitivity on gonadotrophin secretion.     

On the dynamics between gonadotrophin surge-inhibiting factor and gonadotrophin releasing hormone (GnRH): role of self-priming and desensitization in the luteinizing hormone response to GnRH after follicle stimulating hormone treatment

The effects were studied of follicle stimulating hormone (FSH)-induced production of gonadotrophin surge-inhibiting factor (GnSIF) on three phases of the pituitary responsiveness to gonadotrophin releasing hormone (GnRH): the unprimed, primed and desensitized phases. Rats were injected with FSH on two occasions during the oestrous cycle. Spontaneous luteinizing hormone (LH) surges were measured as well as GnRH-induced LH surges on the day of pro-oestrus during infusions with 100-4000 pmol GnRH/rat/10 h, in phenobarbital blocked rats. The spontaneous LH surges were attenuated or completely inhibited by the FSH treatment. FSH suppresses and prolongs the unprimed LH response and delays GnRH self-priming, especially during infusions with low concentrations of GnRH. This treatment does not affect the total LH response (area under curve) to the highest concentrations of GnRH and after ovariectomy. On the other hand, this response is suppressed during infusions with the lower concentrations of GnRH. Hence, FSH, via GnSIF, delays maximal priming of the LH response to GnRH, whereas the suppression of LH release is a consequence of the GnRH-induced progressed state of desensitization. The inconsistent effects of FSH on the mid-cycle LH surges are explained as a result of the interaction between the relative strengths of GnRH and GnSIF.