Pulsatile gonadotropin-releasing hormone treatment in idiopathic delayed puberty

Idiopathic delayed male puberty is defined as a delay of puberty beyond the age of 16, with prepubertal testosterone levels, normal gonadotropin responses to GnRH (excluding pituitary failure), and normal androgen responses to a single hCG injection (excluding testicular Leydig cell dysfunction), in absence of serious disease. Ten boys with this condition were evaluated as to their spontaneous LH, FSH, and PRL secretory patterns during a 24-h sampling period (20-min intervals). After this all patients were treated with pulsatile infusions of GnRH (25 ng/kg . pulse every 90 min for 10 days. Two groups could be distinguished by means of their pretreatment LH secretory pattern. Five patients had nighttime pulsatile elevation of LH levels, as usually occurs in early puberty. The other five patients did not have such a pattern (prepubertal type). The GnRH treatment resulted in increased LH and testosterone levels in both groups. All patients with pretreatment nighttime pulsatile LH secretion had steady pubertal development during the post-GnRH treatment observation period, whereas the other patients did not. In conclusion, among a number of tests, including chronic pulsatile GnRH treatment for 10 days, only the nocturnal LH secretory pattern differentiated delayed puberty from permanent hypothalamic hypogonadism in boys.     

Dose response effects of pulsatile GnRH administration on restoration of pituitary GnRH receptors and pulsatile LH secretion during lactation

Ovariectomized (OVX) rats suckling 8 pups have a complete suppression of pulsatile LH secretion and a decrease in pituitary GnRH receptor (GnRH-R) content. Removing the suckling stimulus for 24 h results in a sharp increase in GnRH-R and a restoration of pulsatile LH secretion. These findings suggest that the suckling stimulus induces a suppression of GnRH secretion, and removal of the suckling stimulus permits the restoration of GnRH secretion. Indeed, if GnRH antiserum is injected at the time of pup removal, the restoration of pituitary GnRH-R and LH secretion is prevented. The present studies were designed to test our hypothesis that the deficits in pituitary gonadotroph function observed during lactation are due to suckling-induced suppression of GnRH. Exogenous GnRH was administered in a pulsatile regimen to OVX lactating rats on days 10 and 11 postpartum, and the effects on pituitary GnRH-R levels, pituitary sensitivity to GnRH, and pulsatile LH secretion were assessed. GnRH doses of 0, 0.5, 2.0 or 5.0 ng/pulse were administered every 50 min for 24 h beginning on day 10. Administration of 0.5 ng GnRH/pulse for 24 h increased GnRH-R from 35 +/- 3 to 63 +/- 8 fmol/pituitary. There was a clear GnRH dose-related upregulation of GnRH-R to approach nonsuckling levels (140-160 fmol/pituitary) with the 5 ng GnRH dose. At the beginning of GnRH administration, the pituitary was very unresponsive to GnRH. Consistent LH pulses were only observed with 5 ng GnRH/pulse.(ABSTRACT TRUNCATED AT 250 WORDS)     

A new compound heterozygous mutation of the gonadotropin-releasing hormone receptor (L314X, Q106R) in a woman with complete hypogonadotropic hypogonadism: chronic estrogen administration amplifies the gonadotropin defect

We describe a woman with complete hypogonadotropic hypogonadism and a new compound heterozygous mutation of the GnRH receptor (GnRHR) gene. A null mutation L314X leading to a partial deletion of the seventh transmembrane domain of the GnRHR is associated with a Q106R mutation previously described. L314X mutant receptor shows neither measurable binding nor inositol phosphate production when transfected in CHO-K1 cells compared to the wild-type receptor. The disease is transmitted as an autosomal recessive trait, as shown by pedigree analysis. Heterozygous patients with GnRHR mutations had normal pubertal development and fertility. The present study shows an absence of LH and FSH response to pulsatile GnRH administration (20 microg/pulse, sc, every 90 min). However, GnRH triggered free alpha-subunit (FAS) pulses of small amplitude, demonstrating partial resistance to pharmacological doses of GnRH. FSH, LH, and FAS concentrations were evaluated under chronic estrogen treatment and repeat administration of GnRH. Not only were plasma FSH, LH, and FAS concentrations decreased, but FAS responsiveness was reduced. This new case emphasizes the implication of the GnRH receptor mutations in the etiology of idiopathic hypogonadotropic hypogonadism. We also have evidence for a direct negative estrogen effect on gonadotropin secretion at the pituitary level, dependent on the GnRHR signaling pathway.     

Increase of serum leptin after short-term pulsatile GnRH administration in children with delayed puberty

OBJECTIVE: Leptin is known to play an important role in pubertal development in humans, probably acting as one permissive factor for the onset of puberty. Leptin serum concentrations change during pubertal development and an initial increase before the onset of puberty has been reported. The underlying mechanism for this increase in leptin levels is unknown. We hypothesized that the pulsatile release of GnRH stimulates leptin metabolism. In this study, the effect of short-term pulsatile GnRH administration on leptin levels in children with delayed onset of puberty was investigated. METHODS: Nineteen children (15 males and four females, mean age 15.5 years, range 13.1-20.5 years), who underwent evaluation for delayed sexual maturation, were included in the study. Sixteen subjects received 36 h of pulsatile intravenous GnRH, using an infusion pump that released 5 microg GnRH every 90 min. Serum concentrations of LH, FSH, testosterone, estradiol and leptin were analysed before and up to 36 h after GnRH administration. Eight patients received a single dose GnRH-agonist stimulation test (buserelin acetate test, 10 microg/kg body weight) with a 24-h follow-up (five patients underwent both tests). RESULTS: Mean (+/-s.e.m.) serum leptin increased significantly (P<0.01) after 36 h of pulsatile GnRH administration (7.26+/-1.35 vs 9.75+/-1.76 ng/ml). In contrast, no increase in leptin concentrations was observed after administration of a single dose of buserelin. CONCLUSIONS: These findings suggested that the increase in serum leptin at the onset of puberty is triggered by the pulsatile release of GnRH.     

Pulsatile GnRH treatment of the ovariectomized rat and release of LH and FSH

The effect of pulsatile GnRH administration on the levels of LH and FSH was investigated in rats that had been ovariectomized 2 weeks earlier. Also the asynchronous occurrence of endogenous and GnRH-induced LH and FSH pulses was analysed. A small pulse dose of GnRH (1.25 ng/100 g) was given iv at a frequency of once every 60 min or once every 120 min during 24 h. A larger dose of 5 ng/100 g was given once every 60 or 120 min during either 24 h or 96 h. Blood was sampled arterially every 5 min around the two first and last GnRH injections and LH and FSH were measured. Only the treatment with the larger GnRH pulse dose resulted in a change of LH and FSH plasma levels. LH levels declined under all circumstances, whereas FSH was found to be increased temporarily after 24 h of treatment. The pituitary LH response to pulses of GnRH (5 ng/100 g) decreased irrespective of the frequency or duration with which GnRH was administered. There was a marked asynchronicity between LH and FSH pulses and almost every injection of GnRH (5 ng/100 g) resulted in clear LH pulses but not in FSH pulses.     

Gonadotropin-releasing hormone (GnRH) agonists and GnRH antagonists do not alter endogenous GnRH secretion in short-term castrated rams

The administration of GnRH agonists and antagonists suppresses pituitary LH secretion. However little is known about their effects on endogenous GnRH secretion. To determine if GnRH analogs act on GnRH secretion through a short or ultrashort loop feedback mechanism, experiments were performed to analyze GnRH secretion in hypophyseal portal blood of conscious short-term castrated rams under both agonist or antagonist treatment. In Study 1, six rams were castrated and surgically prepared for portal blood collection on day -7. Portal and peripheral blood were collected simultaneously every 10 min for 14-15 h on day 0. Five hours after the beginning of the portal blood collection, animals were injected im with 5 mg potent GnRH antagonist (Nal-Glu). In Study 2, six rams were treated daily from day -11 to day 0 with the GnRH agonist D-Trp6 GnRH (0.5 mg im). Castration and surgical preparation for portal blood collection were performed on day -7. On day 0 portal and peripheral blood were collected simultaneously every 10 min for 10-11 h. In both studies, to determine whether an increase in GnRH concentration in hypophyseal portal blood can overcome the inhibitory effect of the GnRH analogs, between 5 and 5.5 h after the injection of the analogs, endogenous GnRH secretion was stimulated by Naloxone administration (3 x 100 mg, iv, at 30-min intervals) followed by a bolus of exogenous GnRH (2 x 10 micrograms, iv at 30-min intervals). In Study 1, Nal-Glu administration led to a rapid cessation of pulsatile LH secretion for the duration of blood collection while GnRH pulse frequency and amplitude were not affected. GnRH and LH pulse frequency before and after Nal-Glu administration were, 6.2 +/- 0.6 vs. 5.7 +/- 0.8 (NS) and 5.3 +/- 0.3 vs. 0.3 +/- 0.2 pulses/6 h (P less than 0.001) respectively. In Study 2, peripheral LH secretion was completely suppressed while GnRH secretion (portal blood) remained pulsatile. GnRH pulses frequency and pulse amplitude were 4.3 +/- 0.3 pulses/6 h and 43.0 +/- 4.7 pg/ml, respectively. In both experiments, neither stimulation of endogenous GnRH secretion by naloxone nor administration of exogenous GnRH allowed reinitiation of LH secretion. However, additional studies in two animals of each treatment group (study-III) showed that this was clearly a dose related effect in antagonist treated but not in agonist-treated animals since higher doses of exogenous GnRH (i.e. 100 micrograms or 1000 micrograms) can increase significantly LH levels.(ABSTRACT TRUNCATED AT 400 WORDS)     

Absence of regular pulsatile LH secretion during pre- and post-implantation periods in sheep

Two experiments were conducted to determine the patterns of LH secretion and to evaluate the LH responses to pulsatile administration of GnRH during early pregnancy in ewes. In experiment 1, pregnant ewes (n=16) were used to determine the concentration of LH in plasma of jugular blood samples collected every 15 min for 6h before (day 10 post-mating) and after (days 20 and 30 post-mating) implantation. In experiment 2, the pituitary LH responses to exogenous pulsatile administration of GnRH were examined on day 10 post-mating in 4 pregnant ewes. A small dose of GnRH (200 ng/ml saline) was injected (i.v.) every 3h and jugular blood samples were collected every 15 min for 12h beginning at the onset of GnRH administration and continuing through the 4th GnRH pulse. During the frequent-sample bleeding at any of the stages of pregnancy examined, LH concentrations oscillated in a pulsatile manner. However, pulsatile LH release occurred irregularly and infrequently. Overall mean LH concentrations, frequency and amplitude of LH pulses were not significantly different between any of the stages of pregnancy examined. Pulsatile administration of GnRH on day 10 post-mating induced regular pulses of LH. In conclusion, these data demonstrate that: (i) pulsatile LH secretion occurs irregularly during early pregnancy, and (ii) the absence of regular pulsatile LH release during early pregnancy is not attributed to a lack of pituitary responsiveness to GnRH.     

Combined hypothalamic-pituitary-gonadal defect in a hypogonadic man with a novel mutation in the DAX-1 gene.

We have studied a 20-yr-old male patient with adrenal hypoplasia congenita and hypogonadotropic hypogonadism (HH) due to a C to A transversion at nucleotide 825 in the DAX-1 gene, resulting in a stop codon at position 197. The same mutation was detected in his affected first cousin (adrenal hypoplasia congenita and HH) and in a heterozygous state in their carrier mothers. The patient had had acute adrenal insufficiency at the age of 2 yr and 6 months, bilateral cryptorchidism corrected surgically at the age of 12 yr, and failure of spontaneous puberty. Plasma testostereone (T) was undetectable (<0.30 nmol/L), gonadotropin levels were low (LH, <0.4 IU/L; FSH, 1.5 IU/L) and not stimulated after i.v. injection of 100 microg GnRH. The endogenous LH secretory pattern was apulsatile, whereas free alpha-subunit (FAS) levels depicted erratic pulses, suggesting an incomplete deficiency of hypothalamic GnRH secretion. During i.v. pulsatile GnRH administration (10 microg/pulse every 90 min for 40 h), each GnRH pulse induced a LH response of low amplitude (0.54 +/- 0.05 UI/L), whereas mean LH (0.45 +/- 0.01 IU/L) and FAS (63 +/- 8 mU/L) levels remained low. Amplitude of LH peaks (0.83 +/- 0.09 IU/L), mean LH (0.53 +/- 0.02 IU/L), and FAS (161 +/- 18 mU/L) levels increased (P < 0.01), whereas the T concentration remained low (0.75 nmol/L) when the pulsatile GnRH regimen was raised to 20 microg/pulse for a 40-h period, suggesting a partial pituitary resistance to GnRH. Thereafter, plasma T levels remained in prepubertal value after three daily im injections of 5000 IU hCG (3.6 nmol/L) and after 1-yr treatment with weekly i.m. injections of 1500 IU hCG (1.2 nmol/L), implying Leydig cell resistance to hCG. The patient had a growth spurt, bone maturation, progression of genital and pubic hair stages, and normalization of plasma T level (15.8 nmol/L) after a 12-month treatment with twice weekly injections of hCG and human menopausal gonadotropin (75 IU International Reference Preparation 2) preparations, suggesting that, in presence of FSH, a Sertoli cell-secreted factor stimulated Leydig cell production of T. In conclusion, we report a novel mutation in the DAX-1 gene in patients with AHC and HH. Our results suggest that the hypogonadism is due to a combined hypothalamic-pituitary-gonadal defect and imply that the DAX-1 gene may play a critical role in human testicular function.     

Idiopathic hypogonadotropic hypogonadism in men: dependence of the hormone responses to gonadotropin-releasing hormone (GnRH) on the magnitude of the endogenous GnRH secretory defect

Idiopathic isolated gonadotropin deficiency (IGD) is associated with a spectrum of clinical findings as well as variable gonadotropin responses to GnRH. In this study we investigated whether patterns of gonadotropin and testosterone responses to pulsatile GnRH therapy (25 ng/kg, iv, every 2 h for 4 days) were related to the magnitude of the GnRH secretory defect in patients with IGD. Eight men with IGD were studied. Patients with partial IGD (p-IGD) and those who had no evidence of GnRH secretion (n-IGD) were differentiated by the presence or absence of spontaneous LH secretory pulses during 24 h of every 20-min blood sampling. In response to the first GnRH injection, no LH rise occurred in the n-IGD patients, while LH increases in the p-IGD patients were similar to those in normal men. Continuation of GnRH therapy in patients with n-IGD resulted in predominant FSH secretion and absent or minimal augmentation of LH and T secretion. In contrast, predominant LH secretion occurred in the p-IGD patients and resulted in a significant increase in serum testosterone. A bolus dose of GnRH 2 days after the termination of GnRH therapy caused significant augmentation of gonadotropin responses in the n-IGD, while in the p-IGD group, both LH and FSH responses were unchanged compared to those after the first GnRH pulse. These results indicate that IGD is characterized by variable degrees of endogenous GnRH deficiency. Moreover, the hormone responses to GnRH in IGD patients depend on the magnitude of the underlying GnRH secretory defect.     

X-linked adrenal hypoplasia congenita: a mutation in DAX1 expands the phenotypic spectrum in males and females

X-linked adrenal hypoplasia congenita (AHC) is a disorder associated with primary adrenal insufficiency and hypogonadotropic hypogonadism (HH). The gene responsible for X-linked AHC, DAX1, encodes a member of the nuclear hormone receptor superfamily. We studied an extended kindred with AHC and HH in which two males (the proband and his nephew) were affected with a nucleotide deletion (501delA). The proband's mother, sister, and niece were heterozygous for this frameshift mutation. At age 27 yr, after 7 yr of low dose hCG therapy, the proband underwent a testicular biopsy revealing rare spermatogonia and Leydig cell hyperplasia. Despite steadily progressive doses of hCG and Pergonal administered over a 3-yr period, the proband remained azoospermic. The proband's mother, sister (obligate carrier), and niece all had a history of delayed puberty, with menarche occurring at ages 17-18 yr. Baseline patterns of pulsatile gonadotropin secretion and gonadotropin responsiveness to exogenous pulsatile GnRH were examined in the affected males. LH, FSH, and free alpha-subunit were determined during 12.5-24 h of frequent blood sampling (every 10 min). Both patients then received pulsatile GnRH (25 ng/kg) sc every 2 h for 6-7 days. Gonadotropin responses to a single GnRH pulse iv were monitored daily to assess the pituitary responsiveness to exogenous GnRH. In the proband, FSH and LH levels demonstrated a subtle, but significant, response to GnRH over the week of pulsatile GnRH therapy. Free alpha-subunit levels demonstrated an erratic pattern of secretion at baseline and no significant response to pulsatile GnRH. We conclude that 1) affected males with AHC/HH may have an intrinsic defect in spermatogenesis that is not responsive to gonadotropin therapy; 2) female carriers of DAX1 mutations may express the phenotype of delayed puberty; and 3) although affected individuals display minimal responses to pulsatile GnRH, as observed in other AHC kindreds, subtle differences in gonadotropin patterns may nevertheless exist between affected individuals within a kindred.     

Hormonal responses in pubertal males to pulsatile gonadotropin releasing hormone (GnRH) administration

Twenty-two boys (9 with delayed puberty and 13 with short stature) ages 12.3 - 17.8 yr, and 10 adult males with idiopathic hypogonadotropic hypogonadism (ages 17.3 - 41.1 yr) have been studied following pulsatile, sc GnRH therapy (240 ng/kg/pulse) over 6 days. Mean pre- and post-therapy LH and FSH concentrations were estimated by 15 min blood sampling over 3-h periods immediately before and at the end of the treatment period. There were significant correlations between the mean pre- and posttreatment LH and FSH concentrations (r = 0.82, p less than 0.001 and r = 0.51, p less than 0.02, respectively) for the 2 groups of peripubertal boys when assessed together. Nine of the 10 adults with hypogonadism showed proportionately greater gonadotropin increments following pulsatile therapy when compared with the peripubertal boys. Standard bolus GnRH tests (100 micrograms iv) did not differentiate between the three groups of patients before pulsatile GnRH therapy. Bolus GnRH tests could predict the subsequent response to pulsatile therapy in the peripubertal boys only. There was no significant change in LH increments following the GnRH bolus tests in either group, after pulsatile GnRH administration (p greater than 0.1). Early response to pulsatile GnRH administration is dependent upon the maturity of the hypothalamic-pituitary-testicular axis in males with delayed puberty or short stature. Patients with hypogonadotropic hypogonadism do not show this relationship.     

Serum leptin levels in males with delayed puberty during short-term pulsatile GnRH administration

Leptin may be a possible trigger for puberty. In normal males, it has been shown that leptin increases from the pre-pubertal to the early pubertal stage, and then declines in the late pubertal stage. We examined leptin levels in six male adolescents (mean age 16.3+/-0.6 yr; range 14.2-17.6 yr) with delayed puberty (constitutional delay of puberty no.=2; idiopathic hypogonadotropic hypogonadism no.=4) during 120 days of subcutaneous pulsatile GnRH administration. A group of subjects in pre-puberty (no.=11), early-puberty (n=10) and mid-puberty (no.=7) were evaluated as controls. Morning blood samples were taken for determination of leptin, testosterone, LH and FSH levels. In delayed puberty subjects blood samples were taken every 30 days after the start of GnRH administration. At each examination BMI and testicular volume were evaluated. A follow-up examination was performed in the 6 patients 1.3-7.5 yr after the end of the 120 days of GnRH therapy. At baseline evaluation in delayed puberty mean leptin levels were 11.3+/-2.0 microg/l (median 11.3 microg/l; range 4.7-17.3 microg/l) and were higher than those found in pre-puberty (p=0.04) and mid-puberty (p=0.001). During GnRH administration there was no change in BMI and leptin levels but there was an increase in gonadotrophin levels, testosterone and testicular volume. One hundred and twenty days after, mean serum leptin were 10.1+/-2.1 microg/l (median 9.1 microg/l; range 3.4-16.8 microg/l). At the end of the study, leptin levels were higher in delayed puberty than in mid-puberty (p=0.002). At the follow-up examination leptin levels were 4.3+/-1.3 microg/l (median 3.4 microg/l; range 1.4-9.1 microg/l) (p=0.03 vs end of 120 days GnRH therapy) while testosterone and BMI were not changed. In conclusion 120-day pulsatile GnRH administration induced in males with delayed puberty physiological-like pubertal changes but not the decline in leptin levels reported during the progression of puberty. Therefore, in males with delayed puberty an impairment in the phenomenon of leptin decline associated with progression of puberty could be suggested. However after retrospective diagnosis of pubertal delay and long-term therapy in subjects with idiopathic hypogonadotropic hypogonadism leptin levels declined. These data seem to indicate that time more than increase in testosterone levels and testicular volume is the determinant of leptin decline at puberty.     

The inhibitory effects of neurokinin B on GnRH pulse generator frequency in the female rat

Neurokinin B (NKB) and its receptor (neurokinin-3 receptor) are coexpressed with kisspeptin and dynorphin A (Dyn) within neurons of the hypothalamic arcuate nucleus, the suggested site of the GnRH pulse generator. It is thought that these neuropeptides interact to regulate gonadotropin secretion. Using the ovariectomized (OVX) and OVX 17β-estradiol-replaced rat models, we have carried out a series of in vivo neuropharmacological and electrophysiological experiments to elucidate the hierarchy between the kisspeptin, NKB, and Dyn signaling systems. Rats were implanted with intracerebroventricular cannulae and cardiac catheters for frequent (every 5 min) automated serial blood sampling. Freely moving rats were bled for 6 h, with intracerebroventricular injections taking place after a 2-h control bleeding period. A further group of OVX rats was implanted with intra-arcuate electrodes for the recording of multiunit activity volleys, which coincide invariably with LH pulses. Intracerebroventricular administration of the selective neurokinin-3 receptor agonist, senktide (100-600 pmol), caused a dose-dependent suppression of LH pulses and multiunit activity volleys. The effects of senktide did not differ between OVX and 17β-estradiol-replaced OVX animals. Pretreatment with a selective Dyn receptor (κ opioid receptor) antagonist, norbinaltorphimine (6.8 nmol), blocked the senktide-induced inhibition of pulsatile LH secretion. Intracerebroventricular injection of senktide did not affect the rise in LH concentrations after administration of kisspeptin (1 nmol), and neither did kisspeptin preclude the senktide-induced suppression of LH pulses. These data show that NKB suppresses the frequency of the GnRH pulse generator in a Dyn/κ opioid receptor-dependent fashion.     

Graded inhibition of pulsatile luteinizing hormone secretion by a selective gonadotropin-releasing hormone (GnRH)-receptor antagonist in healthy men: evidence that age attenuates hypothalamic GnRH outflow

Healthy older men manifest concomitant hypoandrogenemia and attenuation of LH pulse size. Because exogenous GnRH remains effective, a plausible intuition is that aging reduces hypothalamic GnRH secretion, thus mediating relative hypogonadotropic hypogonadism. To assess the impact of age on central GnRH outflow indirectly, we quantitated graded suppression of pulsatile LH secretion by saline and escalating doses of a potent and selective GnRH-receptor antagonist, ganirelix, in 18 healthy men ages 23-72 yr. The rationale is that ganirelix should reduce the amplitude of LH pulses in proportion to both drug concentration and endogenous GnRH feedforward. To this end, blood was sampled every 10 min for 2 h before and 16 h after sc administration of saline or ganirelix and for 3 additional hours after iv injection of a fixed dose of GnRH (100 ng/kg); concentrations of LH and ganirelix were measured by immunochemiluminometry and RIA, respectively; and pulsatile LH secretion was quantitated by a deconvolution procedure. Log-linear regression analysis was used to estimate the sensitivity of pulsatile LH secretion to inhibition by a unit increase in serum ganirelix concentrations in each subject. Statistical analyses revealed that increasing age markedly attenuated the capability of ganirelix to decrease LH pulse size (viz., r = -0.648; P = 0.004). In contrast, age did not modify the competitive interaction between injected GnRH and ganirelix. These joint outcomes support the clinical hypothesis that age diminishes hypothalamic GnRH outflow without impairing GnRH action in healthy men.     

The same molecular defects of the gonadotropin-releasing hormone receptor determine a variable degree of hypogonadism in affected kindred

Detailed endocrinological studies were performed in the three affected kindred of a family carrying mutations of the GnRH receptor gene. All three were compound heterozygotes carrying on one allele the Arg262Gln mutation and on the other allele two mutations (Gln106Arg and Ser217Arg). When expressed in heterologous cells, both Gln106Arg and Ser217Arg mutations altered hormone binding, whereas the Arg262Gln mutation altered activation of phospholipase C. The propositus, a 30-yr-old man, displayed complete idiopathic hypogonadotropic hypogonadism with extremely low plasma levels of gonadotropins, absence of pulsatility of endogenous LH and alpha-subunit, absence of response to GnRH and GnRH agonist (triptorelin), and absence of effect of pulsatile administration of GnRH. The two sisters, 24 and 18 yr old, of the propositus displayed, on the contrary, only partial idiopathic hypogonadotropic hypogonadism. They both had primary amenorrhea, and the younger sister displayed retarded bone maturation and uterus development, but both sisters had normal breast development. Gonadotropin concentrations were normal or low, but in both cases were restored to normal levels by a single injection of GnRH. In the two sisters, there were no spontaneous pulses of LH, but pulsatile administration of GnRH provoked a pulsatile secretion of LH in the younger sister. The same mutations of the GnRH receptor gene may thus determine different degrees of alteration of gonadotropin function in affected kindred of the same family.     

Lack of gonadotropic response to pulsatile gonadotropin-releasing hormone in isolated hypogonadotropic hypogonadism associated to congenital adrenal hypoplasia

Congenital adrenal hypoplasia (AH) is a rare condition, known to be associated with isolated hypogonadotropic hypogonadism (IHH). Three studies have reported attempts to stimulate gonadotropin secretion with pulsatile gonadotropin-releasing hormone (GnRH) in a total of 4 patients presenting such a syndrome, with conflicting results. In the present study, one patient with idiopathic IHH and AH was treated with pulsatile sc GnRH--doses ranging from 2.5 to 10.0 micrograms/pulse, every 90 min--during 8 weeks in an attempt to induce puberty. The prepubertal basal plasma levels of LH, FSH and testosterone, and saliva testosterone levels remained unaltered throughout treatment, at all doses of GnRH tested. The gonadotropin response to an acute iv GnRH administration (0.1 mg) also remained at the prepubertal level after pulsatile GnRH treatment. No circulating anti-GnRH antibodies were detected. The absence of gonadotropic response to exogenous pulsatile GnRH suggests that the IHH of patients with AH is due to an abnormal pituitary function rather than to a lack of endogenous GnRH.     

Utility of free alpha-subunit as an alternative neuroendocrine marker of gonadotropin-releasing hormone (GnRH) stimulation of the gonadotroph in the human: evidence from normal and GnRH-deficient men.

To examine the hypothesis that the secretion of free alpha-subunit (FAS) can serve as an alternative to LH as a neuroendocrine marker of gonadotroph stimulation by GnRH in euthyroid humans, we have investigated the relationship of pulsatile FAS secretion in euthyroid GnRH-deficient men (n = 10) before and after exogenous GnRH stimulation and in normal men under the influence of endogenous GnRH secretion (n = 18). Before GnRH exposure, the GnRH-deficient men showed a complete absence of both LH and FAS pulses. During the initial 7 days of GnRH exposure, all GnRH-deficient men exhibited pulsatile release of FAS by the third day, whereas the appearance of pulsatile release of LH and FSH was more variable. Long term administration of GnRH led to pulses of LH and FAS that were 100% concordant with a demonstrable dose-response relationship between GnRH and FAS, which was quantitatively similar to but more exuberant than that for LH. All doses of GnRH that produced LH pulses within the normal adult range yielded supraphysiological FAS pulses. Analysis of distribution histograms of interpulse intervals and pulse amplitudes of LH and FAS in both normal and GnRH-deficient subjects demonstrated no significant difference between these glycoproteins in interpulse intervals in either the normal or GnRH-deficient groups or in the pulse amplitudes in the GnRH-deficient subjects. There was, however, a significant difference (P less than 0.01) between the distribution histogram of LH and FAS pulse amplitudes in normal men. We conclude that the pulsatile secretion of FAS in euthyroid men 1) is determined by GnRH secretion, 2) is the initial glycoprotein to be secreted in a pulsatile fashion from the gonadotroph during early GnRH exposure in GnRH-deficient men, 3) demonstrates a dose-response relationship to exogenous GnRH which is more robust than that of LH in GnRH-deficient men receiving GnRH, and 4) can, therefore, serve as a complementary and powerful tool with LH for the study of GnRH neurosecretory dynamics.     

The influence of pulsatile GnRH administration to the ovariectomized rat on the pituitary response to GnRH and the occurrence of spontaneous LH pulses

Otherwise untreated adult ovariectomized rats were given pulses of GnRH (5 ng/100 g body weight iv) once every 60 or 120 min for 24 or 96 h. On the first and last day of the experiment plasma LH was estimated during the administration of GnRH pulses. Endogenous LH pulses between exogenously generated LH pulses were observed in nearly all animals on both the first and the last day, without any change in nadir and amplitude values. Shortly after an injection of GnRH, the spontaneous LH pulses were fewer than expected. The number of these pulses, however, increased again with time after the injections. The response to exogenous GnRH was reduced on the last day of the experiment. However, not all GnRH injections led to LH pulses. Most injections which did not result in an LH pulse appeared to be given within 15 min after a preceding endogenous LH pulse. The results obtained are in agreement with the hypothesis of an acute short-lasting desensitization of the pituitary gland caused by exogenous as well as endogenous pulses of GnRH.     

Disruption of GnRH pulses by anti-GnRH serum and phentolamine obliterates pulsatile LH but not FSH secretion in ovariectomized rabbits

It has been hypothesized that the secretion of gonadotropins, i.e. luteinizing hormone (LH) and follicle-stimulating hormone (FSH), is driven by a synchronized neural network ('pulse generator'). This network, regulated in part by alpha-adrenergic activity, ultimately generates bursts of hypothalamic gonadotropin-releasing hormone (GnRH) release. In this study, we used the push-pull (PP) perfusion technique in ovariectomized rabbits to investigate three aspects of the ('GnRH/gonadotropin pulse generator') hypothesis. The objectives were to determine: (1) if plasma LH and FSH pulses occur concomitantly with mediobasal hypothalamic (MBH-) GnRH pulses, (2) changes in the patterns of pulsatile LH and FSH secretion when pulsatile MBH GnRH signals are interrupted by either local immunoneutralization of GnRH or intravenous infusion of the alpha-adrenergic antagonist phentolamine (PHEN, 4 mg/kg BW), and (3) whether third cerebroventricular (3VT-) GnRH patterns reflect neuronal GnRH release from the MBH. We found that while both plasma LH and FSH patterns were pulsatile, MBH GnRH pulses were significantly coupled only with LH pulses (94% coincidence). Both the local immunoneutralization of MBH GnRH pulses and the PHEN-induced suppression of MBH GnRH pulses obliterated the pulsatile secretion of LH, but not FSH. Neither MBH GnRH nor plasma LH or plasma FSH pulses were concurrent with 3VT GnRH pulses. However, the PP perfusion of the 3VT appeared to alter the pulsatile release of MBH GnRH and pituitary LH. The results support the hypothesis that in the absence of ovarian signals, the 'pulse generator' is maintained by tonic alpha-adrenergic input and that a 'cellular unity' of MBH GnRH release (GnRH pulses) drives the gonadotrophs to secrete LH in pulses. In contrast, the pulsatile release of FSH appears to involve additional nonovarian regulatory events to those controlling LH secretion.