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We describe 10 girls (mean age 3.7 years, range 1.9-6.9) with precocious sexual maturation and clinical findings intermediate between those of premature thelarche and central precocious puberty. Studies of spontaneous gonadotropin secretion and ovarian ultrasound morphology also revealed findings intermediate between those of isolated premature thelarche and central precocious puberty. There was no response in 6 of the girls treated with GnRH analogue, whether administered intranasally, sc, or by monthly depot injections. We have called this condition thelarche variant because the gonadotropin independence and cyclical nature of breast development may well be due to a lesion of folliculogenesis.

The existence of a short-loop feedback inhibition of pituitary ACTH release by administration of β-endorphin was postulated. However, data on the effect of peripherally administered β-endorphin in humans are highly controversial. We infused human synthetic β-endorphin at a constant rate of 1 μg · kg−1 · min−1 or normal saline to 7 normal volunteers for 90 min. Thirty min after starting the β-endorphin or placebo infusion, releasing hormones were injected as a bolus iv (oCRH and GHRH 1 μg/kg, GnRH 100 μg, TRH 200 μg) and blood was drawn for measurements of β-endorphin immunoreactivity, all other pituitary hormones, and cortisol. Infusion of β-endorphin resulted in high β-endorphin plasma levels with a rapid decrease after the infusion was stopped. During the control infusion, β-endorphin plasma levels rose in response to CRH. Plasma ACTH and serum cortisol levels in response to the releasing hormone were not different in subjects infused with β-endorphin or placebo. The PRL response to TRH was significantly higher after β-endorphin than after placebo (area under the stimulation curve 1209±183 vs 834±104 μg · 1−1 · h). There was no difference in the response of all other hormones measured. Our data on ACTH and cortisol secretion do not support the concept of a shortloop negative feedback of β-endorphin acting at the site of the pituitary.

The effect of anaesthesia with atropine and pentobarbital (PB) in three groups of volunteers has been tested. The dose used for the anaesthesia was 20 mg/kg body weight of PB. The experiment was performed with and without stimulation test with GnRH. Anaesthesia was shown to increase serum levels of LH and to increase the response to stimulation with GnRH. Atropine and mental stress seem to have no effect on the hormones. No variation in serum levels of FSH in the anaesthesized volunteers was noted. The data obtained have been statistically controlled.

A group of 15 patients with secondary amenorrhoea were studied. In 8 of these patients (hypothalamic-pituitary dysfunction, HPD) the baseline plasma Oe2 concentration (47.7 ± 6.0 pg ml) was similar to the early follicular phase levels of the 9 control subjects (48.8 ± 3.3 pg/ml). The remaining 7 patients (hypothalamic-pituitary failure, HPF) had significantly lower plasma Oe2 values (19.8 ± 2.6 pg/ml). Plasma LH and FSH levels were similar in the two groups of patients as well as the controls. All patients and control subjects were sequentially stimulated with insulin induced hypoglycaemia, GnRH-TRH and then GnRH alone. Although abnormal GH and PRL responses to hypoglycaemia and blunted PRL responses to TRH were obtained in the two groups of patients the results could not be correlated with plasma Oe2 baseline values. In both groups of patients, cortisol and TSH responses were comparable to the control subjects. Following the administration of GnRH the maximum LH response in patients of the HPF group was significantly lower (P < 0.025) than the LH response in patients of the HPD group. No statistically significant difference was found in the FSH response between the two groups. In conclusion, most of the patients with secondary amenorrhoea due to HPD have been shown to have most probably a hypothalamic derangement. Patients with HPF may have either a more profound hypothalamic involvement or an intrinsic pituitary alteration.

To investigate whether short-term fasting affects the pituitary-testicular axis in obese subjects, 9 massively obese men (Body Mass Index 39.0 ± 1.3, mean± sem) were given two identical iv GnRH tests, the first (control) after an overnight fast, the second after 56 h of food deprivation. Short-term fasting augmented the GnRH-induced LH incremental area by 26%(1317±251 vs 1661 ± 297 U · 1−1 · min−1, p −1 · min−1, p

Biosynthesis and secretion of LH were investigated using primary cultures of rat gonadotropes following exposure to swainsonine, an inhibitor of N-linked oligosaccharide processing. The inhibition of terminal glycosylation by swainsonine was assessed by the increase in incorporation of [3H]mannose and the decrease of incorporation of [3H]galactose into the trichloroacetic acid-insoluble and anti-ovine LHβ immunoprecipitable fractions. This inhibitor led to an increase in cellular LH with high affinity for concanavalin A and a decrease of LH with less affinity for the lectin, although the cellular LH was not decreased. The swainsonine-treated cells accumulated highly basic LH (pI, 9.6, 10.0) in a time-dependent manner. Although the treated cells showed a decrease in the responsiveness to hormonal stimulation, there were no differenes in the rate and extent of LH secretion induced by maximal stimulation with 100 nmol/I GnRH between the control and treated cells over 24 h. The accumulation of highly basic LH molecules did not result from a decrease in their secretion, since they were secreted in the presence and absence of GnRH. It is concluded that significant changes in N-linked oligosaccharide structure of LH have no effect upon its secretion.

Twenty patients with hypogonadism (19 women with amenorrhoea and 1 man with impotence and infertility), galactorrhoea and hyper-prolactinaemia (range: 36 to 344 ng/ml) were studied. The radiological study of the sella turcica, including in all cases hypocycloidal tomograms, allowed classification of the patients into 3 groups: group I (n = 4) had a grossly enlarged sella turcica, group II (n = 12) had localized alterations indicating the probable existence of a prolactin-secreting microadenoma ("microdeformation") while group III patients presented no radiological abnormality. Before treatment, all the patients were submitted to a complete evaluation of the function of their anterior pituitary, including the LH and FSH responses to iv administration of Gn-RH. All the group I patients had low basal LH levels and a blunted response to Gn-RH. The basal LH and in response to Gn-RH were normal in most of the group II patients and in all of the group III patients. An exaggerated FSH response to Gn-RH was observed in 6/12 patients with microdeformation (group II) but not in groups I and III patients. A low LH and a blunted LH response to Gn-RH is highly suggestive of the existence of a pituitary prolactin-secreting adenoma in case of amenorrhoea and hyper-prolactinaemia patients; a normal response does not however rule out such a diagnosis. The reasons for a exaggerated FSH response to Gn-RH in patients with suspected prolactin-secreting microadenoma remain to be investigated though this pattern can also occur in other cases of amenorrhoea. Hence the Gn-RH test might contribute to the assessment of the hypothalamo-pituitary axis of patients with hyper-prolactinaemia. Six patients treated for 4 months with bromocriptine (CB-154) were submitted to re-evaluation of their pituitary gonadotrophins reserve. All the women experienced restoration of menses with 39 days of treatment and the male patient regained potency. It was observed that bromocriptine treatment and subsequent normalized prolactin levels in the 4 group II women tested were associated with normalization of their previously exaggerated FSH response to Gn-RH; LH responses were also diminished in these cases. These data are compatible with the hypothesis that hyper-prolactinaemia per se could interfere with the endogenous secretion of Gn-RH at the hypothalamic level. In one patient with grossly enlarged sella turcica and a previous lack of an LH and FSH response to Gn-RH, bromocriptine treatment restored a normal gonadotrophins response, confirming that, in this case, the alteration of this response was indeed due to a prolonged lack of endogenous Gn-RH secretion.Gonadotropin levels in basal conditions and after gonadotropin-releasing hormone (Gn-RH) stimulation in 20 patients with hypogonadism, galactorrhea, and hyperprolactinemia are reported and, in some cases, results after treatment with bromocriptine are included. The radiological study of the sella turcica allowed classification of the patients in 3 groups: 1) grossly enlarged sella turcica, 2) localized alterations indicating the probable existence of a prolactin-secreting microadenoma, and 3) no radiological abnormality. All the Group 1 patients had low basal luteinizing hormone (LH) levels and a blunted response to Gn-RH. Basal LH and the response to Gn-RH were normal in most of Group 2 and in all of Group 3. An exaggerated follicle stimulating hormone (FSH) response to Gn-RH was observed in 6 of 12 patients in Group 2 but not in the other groups. It is suggested that the Gn-RH test might contribute to the assessment of the hypothalamo-pituitary axis of patients with hyperprolactinemia. 6 patients teated for 4 months with bromocriptine experienced restoration of menses or regained potency. These results are compatible with the hypothesis that hyperprolactinemia per se could interfere with the endogenous secretion of Gn-RH at the hypothalamic level.

The effects of an increase in endogenous gonadotrophin secretion on the production of oestradiol, progesterone, androstenedione and testosterone by the ovaries of anaesthetized anoestrous and oestrous ferrets were followed. Gonadotrophin secretion was enhanced by the injection of gonadotrophin releasing factor (GnRF), and serial blood samples were collected over 9 h for hormone assay. Thyrotrophic hormone releasing factor (TRF) or acetic acid were injected for control purposes. The plasma content of oestradiol in oestrous females was significantly higher than during anoestrus, but secretion of this steroid was not increased by any means. The plasma concentration of progesterone in anoestrous females was significantly higher than during oestrus. It was increased by GnRF in anoestrous ferrets and less markedly in oestrous females. The plasma concentration of androstenedione was raised by GnRF to a greater extent during anoestrus than during oestrus. Testosterone was present in higher concentration in the plasma during anoestrus than during oestrus, and the level was increased by GnRF administration. These findings indicate that the ovaries of the anoestrous ferret secrete significant quantities of steroid hormones, and that they respond readily to gonadotrophic hormone.

The responses of serum prolactin (Prl) to metoclopramide and LH and FSH to GnRH were studied simultaneously in 9 boys with hypogonadotrophic hypogonadism (HH), 7 boys with constitutional delay of puberty (D) and 15 controls. Metoclopramide increased the Prl levels in all groups. The boys with HH had lower Prl responses than the controls, whereas the boys with D had similar responses to the controls. Of the 9 boys with HH, 8 had subnormal Prl responses, 3 subnormal LH and none subnormal FSH. A metoclopramide test is clearly more sensitive than a GnRH test in differentiating HH and D and appears to make the differentiation between these two conditions clearer.

Estradiol is known to stimulate gonadotropin-releasing hormone release from the rat mediobasal hypothalamus. Studies were made in an in vitro perifusion system on whether catecholamine and/or histamine was involved in estradiol-induced GnRH release. Normal cycling female rats were decapitated in diestrus II and their medio-basal hypothalami were combined and, perifused with Earl's balanced salt solution containing 0.01% bovine serum albumin bubbled with 95% O2 and 5% CO2. The levels of norepinephrine, dopamine, and histamine and of GnRH in the effluent were measured by HPLC and radioimmunoassay, respectively. Administration of 10−6 mol/l estradiol resulted in releases of norepinephrine, dopamine, histamine and GnRH at levels of 98, 70, 91 and 288%, respectively, of initial values. Administration of 10−6 mol/l norepinephrine or dopamine resulted in no increase in histamine release, and administration of 10−6 mol/l histamine did not increase release of norepinephrine or dopamine. These data suggest that estradiol stimulates the releases of GnRH, catecholamine and histamine from the rat medio-basal hypothalamus, and that it increases GnRH release independently through catecholamine and histamine. As we found previously that norepinephrine or histamine stimulates GnRH release from the mediobasal hypothalamus, we conclude that estradiol stimulates releases of norepinephrine and histamine, resulting in GnRH release from the medio-basal hypothalamus.

In six healthy subjects serum oestradiol was selectively decreased by administering an aromatase activity inhibitor, hydrotestolactone (HT). After HT administration serum oestradiol (Oe2) decreased from 18.7 ± 2.3 (sem) to 6.7 ± 0.6 pg/ml whereas testosterone (T) and dihydrotestosterone (DHT) blood levels were not modified. These oestradiol changes were associated with a significant increase in serum LH and FSH concentrations (P < 0.001). The administration of tamoxifen, an oestrogen antagonist, to 5 subjects caused a sharp increase in LH and FSH levels (P < 0.001). Oe2 was unchanged after the treatment with tamoxifen, whereas T levels were significantly higher. The sum of these data suggests that oestradiol under physiological conditions plays a specific role in the feedback mechanism of gonadotrophin release.

In acromegalics, the percent peak GH responses to TRH (pTRH) and bromocriptine (pBr) are inversely related with those to GHRH, favouring the hypothesis that in the adenomas of some patients there is a preponderance of GH-producing cells with lactotrope-like characteristics, whereas in others pure somatotropes predominate. The aim of the present study was to investigate whether patients responsive to TRH with allegedly lactotrope-like tumours differ from those patients not responding to TRH with more somatotrope-like adenomas in their answer to the GH inhibitors Br and SRIH and the releasing hormones GHRH and GnRH. The present study demonstrates that the observed reciprocal relations between the GH responses to GHRH, TRH and Br are only present in acromegalics paradoxically responding to TRH (pGHRH vs pTRH −0.73, pGHRH vs pBr −0.60, pTRH vs pBr +0.54, P < 0.0002–P < 0.02, N = 20), not in TRH-non-responders (N = 10). In contrast, in these latter patients, not in the former, close relations were found between the percent peak GH responses to GnRH (pGnRH) and pGHRH (r = +0.81, P < 0.005) and between pGnRH and the percent maximum GH decrements in response to SRIH (pSRIH) (r = +0.64, P < 0.05). Expectedly, the GH response to Br in the TRH-responders was significantly higher than in the non-responders (75 ± 4% vs 54 ± 3%, P < 0.02), although it was also substantial in the latter. The GH response to SRIH was remarkedly similar in both groups (64 ± 5 vs 57 ± 9%, P > 0.10). Although the close relations between the GH responses to releasing and inhibiting agents favour the presence of GH-secreting tumours with more somatotrope or more lactotrope characteristics in subsets of acromegalics, they do not effectively predict the therapeutic response to these suppressive agents.

To study the acute and direct effects of GnRH agonists preovulatory follicles were isolated from PMSG-treated immature rats and incubated for 15–360 min in modified Kreb's bicarbonate buffer. The levels of cAMP, prostaglandin E, and progesterone were analysed in the tissue and/or incubation media. GnRH and two GnRH agonists produced a dose-dependent stimulation of progesterone production with maximal levels 5–6-fold higher than the control group. As compared to LH the magnitude of this effect was small and was detected only after 240–360 min of incubation. GnRH also stimulated prostaglandin E accumulation and this effect was as pronounced as for LH. There were no detectable changes in cAMP levels for any concentration of GnRH when the incubation time varied between 15 and 120 min whether or not a phosphodiesterase inhibitor was present, but after 240 min of incubation a 2-fold incease in cAMP was found. Consistent with previous results, LH caused a pronouced (40–50-fold) increase in follicular cAMP which was already detectable after 15 min of incubation. Indomethacin abolished the rise in prostaglandin E induced either by GnRH or LH but did not affect the response in terms of cAMP or progesterone, and did not affect the stimulation of meiotic maturation of the follicle-enclosed oocytes caused by the hormones. It is concluded that GnRH can exert acute and LH-like stimulatory effects on the preovulatory rat follicle but that the mechanism of GnRH action is different from that of LH.

Cultured pituitary cells from adult female rats were treated for 4 or 24 h in the absence or presence of E2 (10−9 mol/1) with increasing concentrations of the transcription inhibitor actinomycin-D or the translation inhibitor puromycin. During the last 4 h of incubation, LH release was stimulated with 5 × 10−10 mol/1 GnRH. The positive E2 effect observed after 24 h treatment with the steroid was clearly abolished by actinomycin-D at concentrations ≥10−10 mol/1 and by puromycin at concentrations ≥ 10−5 mol/1. These findings indicate that the positive E2 effect on GnRH-induced LH release is fully dependent on intact mRNA and protein synthesis. The negative E2 effect observed after 4 h treatment with the steroid was not affected by actinomycin-D and abolished by puromycin only at concentrations ≥ 10−4 mol/1. Similar results were obtained, when cells had been treated for 4 h with actinomycin-D or puromycin before the 4 h E2 treatment started. Thus, the negative E2 effect seems to be independent of mRNA synthesis and dependent on protein synthesis to a lesser extent than the positive E2 effect. In an attempt to identify positively the subcellular mechanism via which E2 exerts its negative effect, several steps in the GnRH stimulus secretion coupling mechanism were checked whether or not they are modulated by E2. The negative effects of E2 (10−9 mol/1) on LH release induced by GnRH (10−10, 10−9 mol/1) and by the activators of voltage dependent Ca2+ channels K+ (64 mmol/1) or veratridine (3.3 × 10−5 mol/1) were comparable to those of the calcium antagonist verapamil (10−6 mol/1). These findings supported the speculation that E2 might act on the Ca2+ channels. The LH release induced by the Ca2+ ionophores A 23 187 (10 −4 mol/1) or ionomycin (6.6 × 10−5 mol/1), however, was also significantly reduced by 10−9 mol/1 E2, indicating that the steroid modulated a mechanism secondary to the increase of intracellular Ca2+. Also GnRH (10−9, 10−8 mol/1) induced accumulation of [3H]inositol phosphates was not influenced by E2 (10−9 mol/1) treatment, though the steroid exerted a significant negative effect on the LH release by these cells, indicating that phosphatidylinositol-4,5-biphosphate breakdown is not the point of attack for the estrogen. The LH release induced by the activation of protein kinase C with the phorbol ester 12-O-tetradecanoylphorbol-13-acetate (TPA, 10−7 mol/1) or with the synthetic diacylglycerol 1,2-dioleyl-sn-glycerol (DOG, 5 × 10−5 mol/1, 10−4 mol/1) was significantly reduced by E2 treatment. These findings give rise to the speculation that E2 might exert its negative effect on GnRH-induced LH release in cultured pituitary cells at the level of protein kinase C or a mechanism secondary to the activation of this enzyme.

The effects of prolonged gonadotrophin-releasing hormone (GnRH) infusions on LH, FSH, oestrogens and progesterone secretion were studied in female rhesus monkeys at various times of the menstrual cycle and after castration. GnRH was infused at the rate of 15 μg/h for 48 h. This resulted in mean peripheral GnRH levels of 398 ± 31.5 pg/ml (± se) as measured by radioimmunoassay. As expected the pattern of gonadotrophin responses to GnRH varied considerably with the phase of the menstrual cycle. The largest LH increase was seen during the late follicular phase (6-fold over baseline), with a 3-fold increase during the luteal phase and a 2-fold one during the early follicular phase and in the period following the LH surge. Significant FSH increases (4-fold) were seen only during the follicular phase. Oestrogens increased about 2-fold within 4 h of the start of the infusion during the early follicular phase. In the late follicular phase and during the LH surge, they declined within 24 and 1 h, respectively. Large progesterone increases were seen only during the luteal phase. Of special interest is the fact that the increase in gonadotrophin secretion could not be maintained for the entire duration of the experiment even though GnRH continued to be infused at rates sufficient to elicit initial increases of several fold over baseline. Gonadotrophin release declined 4–28 h after the initial stimulation. A further decrease below pre-infusion control levels was particularly evident during the midcycle surge and, for FSH, after ovariectomy. These results indicate that a continuous mode of administration may rapidly induce a desensitization phenomenon at the level of the gonadotroph.

The pituitary reserve of GH, prolactin, TSH, LH, and FSH has been studied in a group of 13 acromegalic patients with the aim of evaluating the pituitary function and the activity of the disease. Plasma GH, TSH and prolactin were determined after thyrotrophin releasing hormone (TRH) administration, plasma gonadotrophins and GH after luteinizing hormone releasing hormone (LH-RH) administration. The plasma TSH response to TRH was generally blunted in the patients treated with pituitary irradiation; however, none of the patients with diminished TSH reserve had signs of hypothyroidism. Six acromegalics showed prolactin basal levels higher than controls; none had galactorrhoea; 4 of them complained of impairment of the gonadal function. The prolactin response to TRH was variable and not related to prolactin basal levels. A subnormal LH reserve after LH-RH stimulation was observed in 5 out of 10 patients; 4 of them had also clinical signs of hypogonadism. A normal FSH response to LH-RH was present in all patients. A non-specific GH response to TRH and/or LH-RH was obtained in about half of the acromegalics studied. The GH responsiveness to TRH and/or LH-RH was not related to the activity of the disease or to a specific derangement of the hypothalamo-pituitary function. A concordant response was observed only between GH and prolactin response to TRH; the highest prolactin responses to TRH were obtained in the GH responsive patients. Each patient showed a constant GH pattern of response on repeated testing. Even after pituitary irradiation the pattern of GH response was unchanged in spite of lowered GH plasma levels.

The aim of the present study was to determine the effects of exogenous GnRH pulsatile infusions on the diurnal variations of LH and testosterone secretion which occur in late pre-puberty and early puberty. GnRH infusions were administered to 12 short stature males in pre-puberty or early puberty, over 6-day periods. In 6 patients, GnRH doses of 2.5, 7.5 and 15 μg/pulse were used and 24-h profiles of serum LH and testosterone were measured before and at the end of the infusions. In the remaining 6 patients GnRH was administered at a dose of 7.5 μg/pulse and profiles between 21.00 and 06.00 h the following day were determined. Pre-infusion profiles demonstrated nocturnal LH and testosterone rises in all patients. Median pre-infusion serum LH prior to midnight was 2.2 U/l (range 1.0–5.4) rising to 3.7 U/l (range 1.9–10.7) during GnRH administration (p < 0.005). After midnight, median pre-treatment serum LH concentration was 4.3 U/l (range 2.7–7.5) which remained unaltered by GnRH administration (median 4.8 U/l, range 2.9–7.9, p > 0.05). Median pre-therapy serum testosterone before midnight was 0.8 nmol/l (range 0.1−7.1) rising significantly (p < 0.05) to 4.1 nmol/l (range 0.2–8.0). Following therapy, post-midnight median serum testosterone rose from 4.8 (range 0.4–9.4) to 7.0 nmol/l (range 0.5–13.9, p > 0.05). Diurnal variation in LH and testosterone secretion, therefore, is maintained during exogenous GnRH administration to pre-pubertal and pubertal boys. Response to exogenous GnRH pulses may be significantly influenced by endogenous GnRH.

Dispersed pituitary cells from adult female rats were preincubated for different time periods (0– 12 h) in the absence or presence of 10−9 moestradiol (E2) or 4-hydroxyoestradiol (4-OHE2). Then the media were changed and the cells incubated for 4 h with either vehicle, or E2, or 4-OHE2 and additionally with different concentrations (10−11– 10−7 m) of gonadotrophin-releasing hormone (GnRH). Treatment of pituitary cells with E2 for 4 h (i.e. no preincubation with E2) significantly decreased the LH-response to GnRH at concentrations ≥ 10−10 m of the decapeptide. During a transition time of approximately 10 h (i.e. in cultures preincubated with E2 or vehicle for 2, 4, 6 or 8 h and then coincubated with E2 or vehicle and GnRH for 4 h) no differences between E2-and vehicle-treated cultures were observed. After 14 and 16 h of E2-treatment (i.e. 10 or 12 h preincubation and 4 h coincubation with GnRH) the LH-responses to GnRH in these cultures were significantly higher than in the respective controls. A nearly identical reaction pattern was observed when 4-OHE2 was used instead of E2. In a second series of experiments dispersed rat pituitary cells were suspended in a carrier gel and continuously perifused with medium, using small chromatography columns. When these cells were exposed for 4 min to 10−9 m GnRH at 60 or 48 min intervals, they reacted with reproducible pulsatile LH-discharges during at least 6 subsequent stimuli with the decapeptide. When E2 (10−9 m) was added to the perifusion medium, the LH-responses to GnRH were significantly reduced, starting 36 min after the onset of E2-treatment. These data indicate: 1) In the rat, the negative oestrogen effect is at least in part directly mediated at the pituitary level. 2) The sensitizing effect of oestrogens on rat gonadotrophs to GnRH is significant already after 14 to 16 h. 3) E2 and the catecholoestrogen 4-OHE2 have the same effects in this system. 4) The negative E2-effect on GnRH-induced LH-release is significant after only 36 min, a finding bringing up the question of a non-genomic mechanism.

The in vivo release rates of GABA from the preoptic/anterior hypothalamic area (PO/AH) of ovariectomized rats were assessed by means of a focal perfusion cannula system. Seven days after surgery all animals received a sc silastic capsule implant containing either estradiol-17β (E2) or corn oil, and they were perfused 3 days later. Perfusate fractions were sampled at 5-min intervals and blood was collected every 10 min over a period of 5 h. In ovariectomized animals PO/AH GABA release was pulsatile without any diurnal rhythm. Prior to frequence analysis by means of the pulsar-programme, LH and GABA values were z-transformed. Significant LH peaks of all examined ovariectomized rats were superimposed and GABA data were arranged accordingly. It became evident that LH episodes are preceded by a significant reduction of preoptic anterior hypothalamic GABA release. The secretion patterns of GABA and LH were profoundly affected by E2 replacement. During early noon when LH levels were low, constantly elevated hypothalamic GABA release rates were observed in E2-substituted rats in comparison to ovariectomized rats. GABA release rates fell significantly during the E2-induced LH surge. Our previous demonstration of the existence of a large number of estrogen-respective GABAergic neurons in the PO/AH is suggestive of these neurons changing their activity in response to estrogen treatment. We conclude that these estrogen-respective GABAergic neurons are involved in the generation of GnRH pulses as well as in the generation of the so-called positive feedback effect of E2 on LH release. Whether the same GABAergic neurons have those effects or whether two populations of GABAergic neurons exist, one responsive for generation of pulsatile LH release, the other responsive for the estrogen-induced LH surge, remains to be elucidated.

The present study was undertaken in order to establish whether muscarinic cholinergic receptors are involved in the anomalous GH response to GnRH in men with insulin-dependent diabetes mellitus and in male patients with major depression. For this purpose, 16 male diabetics, 18 depressed men and 9 normal controls were tested with GnRH (25 μg iv) with and without previous treatment with the muscarinic cholinergic receptor blocker pirenzepine (40 mg iv 10 min before GnRH). Additional experiments with TRH (200 μg iv 10 min after pirenzepine) were performed in the same subjects and used for comparison between responders to TRH and GnRH. The administration of GnRH stimulated GH release in 12 out of the 16 diabetics and in 8 out of the 18 depressed patients, but not in the normal controls. Control and diabetic non-responders to GnRH did not respond to TRH. In contrast, all diabetic responders to GnRH, except 2, showed paradoxical GH responses to TRH. All depressed responders to GnRH and 3 of the non-responders, were responsive to TRH. The pattern and magnitude of the secretory responses to TRH and GnRH were similar in depressed and diabetic patients. When the effects of GnRH and TRH were restudied in the presence of pirenzepine, neither GnRH nor TRH enhanced the serum concentrations of GH in any patient. These data indicate that a muscarinic cholinergic mechanism is involved in the anomalous responses of GH to GnRH and TRH in diabetic men and in male patients affected by major depression.

The purpose of this study was to investigate the kinetics and estrogen dependence of the facilitory progesterone action on LH and FSH secretion from pituitary cells under dynamic culture conditions. Anterior pituitary cells obtained from ovariectomized or intact adult Wistar rats were cultivated on Cytodex 1 microcarrier beads. The perifusion experiments were performed with four separate perifusion chambers. The cells of chambers I+II had been pretreated with medium containing vehicle (0.1% ethanol), those of chambers III + IV with medium containing 1 nmol/l estradiol for 48 h. After perifusion was started, each of the chambers was challenged with an initial 2-min GnRH (1 nmol/l) pulse. Chamber I was further perifused with medium containing vehicle, chamber II with medium containing vehicle + 100 nmol/l progesterone, chamber III with medium containing 1 nmol/l estradiol, and chamber IV with medium containing 1 nmol/l estradiol + 100 nmol/l progesterone. Three further GnRH pulses were administered at 50-min intervals to each of the chambers. In estradiolprimed cells from intact rats, progesterone induced a positive effect on LH and FSH secretion after 50 min of exposure to progesterone. After 100 min of progesterone treatment, LH and FSH release were enhanced to 420 and 500 per cent, respectively. When such cells were not primed with estradiol, a slight though insignificant positive action of progesterone on LH release was present after 50 and 100 min of treatment, whereas FSH secretion was not influenced. The facilitory effect of progesterone occurred only after 100 min when estradiol-primed cells from ovariectomized rats were used. LH and FSH secretion were only enhanced by up to 210 and 233 per cent, respectively. When these cells were not primed with estradiol the positive progesterone effect was absent. In conclusion, progesterone induces an acute facilitory effect on GnRH-induced LH and FSH secretion which is estrogen-dependent.

Primary monolayer cultures of adult male rat anterior pituitary cells secreted both LH and FSH in a biphasic manner when incubated with gonadotrophin releasing hormone (GnRH). The peaks of secretion of LH and FSH were coincident; the first occurred between 15 and 30 min and the second between 1 and 3 h after the addition of GnRH. Approximately 20% of the total amount of gonadotrophins secreted in the 6 h treatment with GnRH was contained in the first peak. Inhibitors of the secretion of gonadotrophins affected LH and FSH secretion differently. Inhibin suppressed the secretion of FSH to a greater extent than that of LH, whereas testosterone and cycloheximide had a greater effect on LH. Neither phase of secretion of LH or FSH was reduced preferentially by inhibin or testosterone but the greater effect of cycloheximide was on the second phase of secretion.

One hundred excessively tall boys with a height prediction of 205.32 ± 5.28 cm (mean ± sd) were treated with 500 mg testosterone oenanthate (TE) every 14 days for a period of 14.96 ± 5.29 months. Following therapy, the hypothalamo-pituitary-gonadal axis was evaluated, using a standardized GnRH-test at median time intervals of 14 days, 6 weeks, 13 weeks, 6 months and 16 months. Basal and stimulated LH- and FSH-values were not measurable or severely suppressed in all boys 14 days after termination of therapy. Starting at 6 weeks, normalization of pituitary-gonadal function was demonstrated in 93 boys (group 1) with follow-up periods of up to 48 months. Six boys (group 2) developed transitory hypergonadotrophic LH- and FSH-secretory patterns for up to 11 months after the last TE-injection. Testosterone and gonadotrophins were within the normal range in all 6 boys, when prospectively re-evaluated at 12 to 27 months after termination of therapy. During TE-administration, testicular volume was reduced in some, and in most boys did not show the normal enlargement occurring during puberty. However, return to normal testicular size was seen several months after treatment. The data suggest that no major functional alterations of the hypothalamo-pituitary-gonadal axis occur in pubertal boys after high-dose and long-term testosterone therapy for excessively tall stature.

Effect of intravenous administration of a synthetic gonadotrophin releasing factor (GnRF) on circulating LH and FSH concentrations was investigated in normal and gonadectomized, adult male rats injected subcutaneously each day during seven days with various doses of oestradiol-17β or 5α-dihydrotestosterone in sesame oil. Higher increase in serum LH and FSH levels subsequent to intravenously administered GnRF was observed in castrated control animals than in intact control animals, though this increment was not of significant nature for serum FSH. Pre-treatment of normal and gonadectomized rats with oestradiol-17β resulted in an augmented response of serum LH and FSH concentrations to GnRF iv. Pre-treatment of normal and gonadectomized rats with 5α-dihydrotestosterone diminished serum LH and FSH response following administration of the same amount of GnRF. For these steroids to affect the response pattern of serum LH and FSH to GnRF iv, higher doses were required in normal than in gonadectomized animals. The data indicate that oestrogen and androgen may act at different sites in the male hypothalamic-pituitary axis with respect to regulation of pituitary gonadotrophins.

Cranial irradiation is known to affect the neuroendocrine control of GH secretion. The aim of the present study was to clarify the effect of leukaemia treatment on neuroendocrine control of PRL secretion in long-term survivors of acute lymphoblastic leukaemia (ALL). Pituitary LH, FSH and PRL responses to GnRH and to the dopamine receptor antagonist metoclopramide were evaluated in 18 boys 1.6–9 years after discontinuation of medication for ALL. All the boys had been subjected to prophylactic cranial irradiation and 8 to testicular irradiation. Eight of the boys had supranormal gonadotropin levels either basally or after GnRH. All had normal basal PRL levels, but in 14 of the 18 boys the metoclopramide-releasable PRL levels were subnormal. The decreased PRL responses cannot be explained by the hypergonadotropism, since exaggerated rather than decreased responses are frequently found in patients with primary gonadal failure. We conclude that abnormal regulation of PRL secretion is a frequent consequence of treatment of ALL, probably being due to decreased pituitary lactotrop mass.

The value of pulsatile GnRH therapy for induction of ovulation in patients with polycystic ovarian disease remains unclear. Intravenous pulsatile GnRH therapy was administered to a defined group of 5 patients with polycystic ovarian disease; all were infertile, had an LH: FSH ratio of 2:1 on two or more occasions, and had multiple cysts on ovarian ultrasonography. All had failed to respond to clomiphene citrate. The 5 patients received increasing doses of GnRH (5–40 μg/pulse) continuously for up to 6 weeks. The response was evaluated by serial hormone levels and ovarian ultrasonography. During nine treatment periods no patient ovulated, and in only one did the LH:FSH ratio revert to normal. Four patients have subsequently had wedge resection of the ovaries and in each case the diagnosis of polycystic ovarian disease was confirmed. Pulsatile GnRH therapy was of no value in the management of this group of infertile patients with strictly defined polycystic ovarian disease.

By centrifugal elutriation and subsequent Percoll density gradient centrifugation, we obtained highly enriched cell populations of the anterior pituitary of ovariectomized adult rats. Enrichment of somatotrophs, gonadotrophs and mammotrophs was 90%, 80% and 77%, respectively (determined by immunocytochemistry). 41% of the GH-cells showed a medium cell size and the highest density. 52% of the LH-cells and 44% of the FSH-cells were found within the population of the large and medium-dense cells. 69% of the prolactin-cells belonged to the small-sized cells with low density. Thyrotrophs were found within all of the size ranges. The majority of the corticotrophs (82%) showed a small cell size. The enriched somatotrophs, LH-gonadotrophs, and mammotrophs responded to GRF, LRH and TRH. Before and after enrichment, somatotrophs and mammotrophs showed the same responsiveness to releasing hormones. In contrast, enriched LH-gonadotrophs exhibited a higher responsiveness to LRH than unfractionated LH-gonadotrophs.

During late gestation in the ewe, the pituitary content of LH is reduced by about 95%, presumably due to the presence of high concentrations of ovarian steroids. The aim of this study was to determine whether the pituitary content of LH in the ewe can increase after long-term administration of ovarian steroids, when only estradiol (E) is removed or if both E and progesterone (P) must be withdrawn to allow synthesis of LH to occur. Ten ovariectomized ewes were treated with implants containing E and P. After 3 weeks of treatment, the E implants were removed from 5 ewes (–E+P) and both steroid implants were removed from the remaining 5 ewes (−E−P). Five ovariectomized ewes received P implants at the beginning of the experiment and these implants were left in place for the duration of the study; 5 ovariectomized ewes served as controls (C). All animals were injected with 100 μg GnRH iv 3, 6 and 9 weeks after the initiation of treatment. The area under the LH-response curve was used as an indication of the pituitary content of LH. All steroid treatments markedly reduced basal levels of LH. LH levels increased only in −E−P ewes, beginning 6 weeks after initiation of the study. After 3 weeks, −E+P and −E−P ewes released less LH (P < 0.05) in response to GnRH than did C ewes, whereas P animals did not differ from controls. LH release in response to GnRH in −E+P and −E−P groups had increased by 6 and 9 weeks and was not different from that of C ewes. After 9 weeks, LH release in P ewes was reduced (P < 0.05) compared with C ewes. These data suggest that, after the pituitary content of LH has been suppressed by ovarian steroids, the presence of P alone does not inhibit replenishment of the pituitary content of LH.

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.

A comparison was made of the efficacy of intravenously (iv) and intraventricularly administered synthetic Gn-RH in inducing ovulation in conscious and urethane anaesthetized rabbits. Via the iv route of administration, the minimum dose of Gn-RH required to induce ovulation in all animals of the conscious group was 300 ng/kg body weight. This is in contrast to the larger systemic dose (400 ng/kg body weight) of Gn-RH required to achieve the same results in urethane anaesthetized but otherwise identically treated rabbits. This suggests that although ovulation is not blocked by an anaesthetic dose of urethane (1.25 g/kg), the ovulatory response is altered in some way by the anaesthetic. When administered intraventricularly, the minimum dose of Gn-RH required to induce ovulation in anaesthetized animals was only 100 ng/kg body weight or 25% of the equivalent systemic dose. Systemically ineffective doses of Gn-RH (≤ 200 ng/kg body weight) also induced ovulation in the anaesthetized group when administered via the third cerebral ventricle. All animals that ovulated did so between 13 and 16 h following Gn-RH administration and the number of ova recovered paralleled the number of ruptered follicles. A graded increase in the number of ova released with dosage of Gn-RH was not observed. In this study, the smaller effective dose requirement for intraventricularly administered decapeptide supports the view that Gn-RH in the CSF circumvents the general circulation and reaches the portal vasculature directly via the median eminence.

Intravenous metoclopramide (MET) (10 mg) induced a brisk PRL response with a mean ± sem peak of 85.3 ± 7.7 ng/ml maximal at 30 min. l-Dopa, but not atropine pre-treatment, attenuated the prolactin (PRL) response to MET. This indicates that the antidopaminergic properties of MET mediate PRL secretion. MET did not influence basal levels of TSH, LH or FSH. Neither did it affect their response to the respective releasing hormones. Our results indicate that dopaminergic blockade induced by iv MET, does not influence the secretion of the pituitary glycoprotein hormones.

Columns of enzymatically dispersed rat pituitary cells have been used to examine the effects of estradiol and the estrogen receptor antagonist, tamoxifen, on the responsiveness of the gonadotropes to repeated challenges with GnRH. Synthetic GnRH stimulated readily the release of LH from the pituitary cells. When challenged repeatedly with the same submaximal dose (8 nmol/1) of the releasing hormone, cells from animals in diestrus 2 or pro-estrus, like those from juvenile rats, showed a marked progressive increase in their sensitivity, but those from rats in estrus or diestrus 1 did not. Adenohypophysial cells removed from adult rats ovariectomized 14 days previously also failed to exhibit sensitization even when estradiol-17β (1 and 10 nmol/1) was included in the perifusion fluid. However, those from ovariectomized rats treated with appropriate doses of the steroid (1.5 μg/day sc for 14 days or 5 μg sc 22 h before decapitation) did. The capacity of pituitary cells from intact rats to exhibit priming on the anticipated day of pro-estrus was unaffected by the administration of the estrogen receptor antagonist, tamoxifen (700 μg/100 g sc on the morning of diestrus 1 and 24 h later). Similarly, addition of tamoxifen (1 μmol/1) to the incubation medium failed to influence the capacity of pituitary cells from juvenile rats to exhibit priming in vitro. The results support the concept that the ability of GnRH to 'prime' the gonadotropes is dependent on estradiol and indicate that the exposure of the hypothalamo-pituitary complex to critical levels of estradiol prior to autopsy is necessary for the gonadotropes to retain their capacity to exhibit priming in vitro.

The aim of this study was to investigate effects of androgens on gonadotrophin release in response to gonadotrophin-releasing hormone (Gn-RH) stimulation in vitro. Hemipituitaries of mature male rats were pre-incubated for 90 min with T, DHT, 3α- or 3β-diol (4 ng or 4 μg/ml medium), and the incubation continued for 240 min after adding Gn-RH (1 ng/ml medium). Gn-RH caused a 4-5-fold rise in the secretion of LH and a 2-fold rise in FSH secretion. The effect of the androgens was dose-dependent. At low levels, T and DHT exerted no effect on Gn-RH-stimulated gonadotrophin release, whereas the two androstanediols (3α- and 3β-diol) augmented the Gn-RH stimulation of both gonadotrophins, though preferentially LH. With high doses of androgens, the results obtained showed: a) no effect of T; b) DHT suppression of the Gn-RH-stimulated FSH release; c) suppression of Gn-RH stimulation by 3α- and 3β-diol regarding both LH and FSH. It is concluded that T exerts through its reduced metabolites a feedback effect on the pituitary gland responsiveness to Gn-RH stimulation.

The effects of substance P on the release of LH and GnRH were examined in a sequential double-chamber perifusion system by perifusing the medio-basal hypothalamus and/or pituitary excised from normal female rats in dioestrus or ovariectomized rats. When the medio-basal hypothalamus and pituitary from normal rats were perifused in series with substance P (10−6 mol/l), the concentration of LH in the efflux was significantly (P < 0.05) increased by 70– 120% compared with that before the injection, but substance P had no effect on LH release from the pituitary perifused alone. This LH release by substance P increased in a dose-dependent manner and was blocked by substance P antagonist. Administration of 10−6 mol/l substance P induced a significant release (40–80% increase, P < 0.05) of GnRH from the medio-basal hypothalamus. Infusion of 10−6 mol/l substance P induced significant release (50–100% increase, P < 0.05) of LH and GnRH in ovariectomized rats with an implanted oestradiol capsule, but caused no significant increase in LH release in ovariectomized rats without an oestradiol capsule. Progesterone injection to both ovariectomized rats and ovariectomized rats with an implanted oestradiol capsule had no significant effect on the response of LH to substance P. These findings suggest that substance P induces GnRH release from the medio-basal hypothalamus, resulting in LH release from the pituitary, and that oestrogen may be involved in these processes.

Four normal male subjects received iv injections of synthetic luteinizing hormone- and follicle stimulating hormone-releasing hormone (LH/FSH-RH) in doses of 12.5, 25, 100, 200 and 400 μg, respectively. A dose of 12.5 μg of LH/FSH-RH caused a significant increase in serum FSH, and 25 μg significantly increased the serum LH. The peak responses occurred 15 to 30 min after the LH/FSH-RH injections in most of the experiments. The increase in the mean maximum serum LH and FSH levels was 2 to 4 fold. There was great variation in response between the subjects, but when tested repeatedly with the same dose of LH/FSH-RH a given individual responded in a consistent manner. The log dose-response curve between LH/FSH-RH and serum LH, and between LH/FSH-RH and serum FSH was approximately linear. A small but significant (P < 0.05) rise in serum thyrotrophin (TSH) was found after LH/FSH-RH in doses ranging from 25 to 400 μg. There was no significant rise in serum growth hormone (HGH). On the basis of the present study a standard 100 μg iv LH/FSH-RH test is suggested.

Plasma immunoreactive insulin and growth hormone of 20–24 weeks' gestation human foetuses were assayed in serial samples following delivery by hysterotomy. The mean umbilical cord plasma growth hormone concentration was 71 ng/ml (range 13–120 ng/ml) and the mean plasma insulin was 5 μU/ml (range 2–8 μU/ml). Following delivery the growth hormone levels increased, but there was no significant change in plasma insulin concentration. The hypothalamic-hypophyseal axis controlling growth hormone secretion appears to be developed by 20 weeks' gestation, and »stress« appears to be a provocative stimulus.

Polyacrylamide gel electrophoresis at neutral pH, 0°C, was used to test the identity of prolactin derived from serum and pituitary extracts of rats. Animals were studied after treatment with perphenazine, a prolactin releasing agent, and during post partum lactation. After gel electrophoresis, the position on the gel of prolactin activity was determined by the pigeon crop-sac bioassay. The biologically active prolactin exhibited identical electrophoretic mobility, whether derived from serum or pituitary, of either normal lactating or perphenazine treated rats.

The uptake of radioactivity by various tissues of the domestic fowl has been studied following a single intravenous dose, at different times during the ovulatory cycle. It has been found that the pituitary gland showed significant cyclical accumulations of radioactivity, when calculated in terms of dpm/mg wet weight, at different stages in the ovulatory cycle, the greatest accumulation being in the period 12–16 hours before the expected ovulation. The results are discussed in relation to the role of oestrogens in the timing of ovulation. It is proposed that oestradiol acts as a major regulator of the timing of ovulation in the domestic fowl.

There are several in vitro methods to analyse the function of the adenohypophysis or the mechanisms of its regulation. The present paper deals with single cell culture, organ culture and short term incubation techniques by which the morphology and gonadotrophin-secreting function of the adenohypophysis were studied. In trypsin-dispersed cell culture, the adenohypophysial cells showed extensive propagation to form numerous cell colonies and finally develop into a confluent monolayer cell sheet covering completely the surface of culture vessels. Almost all of the cultured cells, however, became chromophobic, at least at the end of the first week of cultivation, when gonadotrophin was detectable neither in the culture medium nor in the cells themselves. After the addition of the hypothalamic extract, gonadotrophin became detectable again, and basophilic or PAS-positive granules also reappeared within the cells, suggesting that the gonadotrophs were stimulated by the extract to produce gonadotrophin. In organ culture and short term incubation, the incorporation of [3H] leucine into the adenohypophysial cells in relation to the addition of hypothalamic extract was examined. It was obvious that the ability to incorporate [3H] leucine into the gonadotrophs in vitro was highly dependent upon the presence of the hypothalamic extract.

Only partial studies evaluating the endocrine profile in molar pregnancy have been performed. In order to characterize the neuro-endocrine pattern during and after molar pregnancy, we studied the basal hormonal levels of hCG, human placental lactogen (hPL), FSH, GH, TSH, free thyroxine index (FTI), oestradiol-17β (E2), and progesterone (PG), as well as the anterior pituitary response to TRH, GnRH, and hypoglycaemia induced by insulin in 7 patients during molar pregnancy and one week after molar abortion. hCG showed significantly higher serum levels during rather than after molar pregnancy and hPL was detectable in only 4 patients during, but in none after molar pregnancy. FSH values were in the follicular phase range before and after molar abortion (12.7 ± 0.8 and 12.7 ± 3.5 IU/l). PRL had elevated basal levels before and after molar abortion; 103.0 ± 16.5 and 43 ± 10.6 μg/l, respectively (P < 0.05). GH levels were distinctly elevated in 3 patients during molar pregnancy; after molar abortion, the basal GH values were normal in all patients < 10 μg/l. Basal cortisol and TSH levels were in the normal range before and after molar abortion. The FTI was above the normal range in 3 patients during molar pregnancy, whereas after molar abortion the values were normalized. E2 levels were elevated before and after molar abortion, 1881 ± 477 and 96.5 ± 39.2 ng/l, respectively (P < 0.01). PG levels before and after molar abortion were 30.9 ± 5.4 and 10 ± 6.7 μg/l, respectively (P < 0.05). Comparison of women with molar pregnancy vs women with normal pregnancy showed a significant difference only as to β hCG (P < 0.05), whereas comparison of women in molar puerperium vs women in normal puerperium after term pregnancy showed a significant difference in β hCG (P < 0.01), PRL (P < 0.01), and PG (P < 0.01). Six of the 7 patients had a PRL increase of over 100% of the basal level in the first 15 min after TRH administration; one week after molar abortion, all patients had a normal PRL response to TRH. During molar pregnancy, only 3 patients showed a normal TSH response to TRH, 4 patients did not respond; 3 of them had an FTI above normal range. During the second challenge one week after molar abortion, 5 patients showed a normal TSH response to TRH. The hypoglycaemia stimulus induced a normal cortisol and GH response in 6 and in 5 patients, respectively, during molar pregnancy; after molar abortion, all patients had a normal cortisol and GH response to hypoglycaemia. FSH had no response to GnRH during or after molar pregnancy. These results establish the neuro-endocrine pattern in patients with molar pregnancy showing a clear disturbance in the neuro-endocrine regulation of the thyroid axis as well as in the GH release.

Long-term treatment with the potent and long-acting stimulatory luteinizing hormone-releasing hormone (LRH) analogue D-Ser(TBU)6-EA10-LRH was given to 4 healthy men to study its effects on pituitary gonadotrophin secretion and gonadal function. Five μg of the LRH agonist was self-administered sc once daily over 17 weeks. Weekly basal blood samples were obtained for determination of follicle-stimulating hormone (FSH), luteinizing hormone (LH), prolactin (PRL) and testosterone. The gonadotrophin responses to the LRH analogue were also determined during the treatment period. LRH tests were performed after treatment. Seminal fluid specimens were collected during and after treatment. A reduction of the basal serum gonadotrophin and testosterone levels were observed during the treatment period. The FSH and LH responses to the analogue were also diminished. After discontinuation of treatment the gonadotrophin and testosterone concentrations returned to pre-treatment levels within a week. The PRL levels and the seminal fluid specimens did not show any significant changes during the study period. The results suggest that chronic treatment with D-Ser(TBU)6-EA10-LRH has an inhibitory effect on the pituitary gonadotrophin secretion in healthy men. It seems likely that the reduced testosterone level is secondary to the diminished gonadotrophin secretion.

In 10 hyperthyroid women studied in the follicular phase of the menstrual cycle, basal plasma PRL was normal, but PRL release after TRH was significantly suppressed compared with that in 11 control women. The suppressed PRL response to TRH was not explained by changes in serum estradiol or sex hormone-binding globulin. It recovered after treatment of hyperthyroidism. When normal women were treated with T4 (0.5 mg daily for 6 to 10 days), their mean serum free T4 level increased to about 70% of that in the hyperthyroid patients, whereas their serum free T3 levels increased to a lesser degree. During T4 administration, these women had PRL changes similar to those of the hyperthyroid patients. When the normal women took T3 (60–120) μg for 6 to 8 days), their serum free T3 increased to almost the level of the hyperthyroid patients, but the TRH stimulated PRL release remained close to the control level. The PRL increase after dopaminergic blockade with metoclopramide was significantly suppressed in hyperthyroid patients, and they had no PRL response to TRH after pretreatment with metoclopramide. In conclusion, the PRL changes in hyperthyroidism were reproduced by administration of T4, but not by administration of T3 to healthy women. The site of action is suggested to be pituitary, but additional hypothalamic effects cannot be excluded.

LH release from the anterior pituitary was studied by the method of an in vitro superfusion of bovine anterior pituitary tissue slices. LH release was stimulated by increased potassium concentration (23 and 59 mm) and by synthetic GnRH (1 and 4 ng/ml). While the potassium effect was completely dependent on extracellular calcium, that of GnRH was only partially dependent. Additive effect was observed when GnRH followed enhanced potassium infusion but not vice versa. This suggested that the mechanism responding to potassium may be contained within the stimulatory pathway of GnRH. There was a difference in the dynamics of the LH response: maximum response was attained in about 10 min of potassium infusion while infusion of GnRH resulted mostly in multiphasic stepwise release of LH reaching a plateau in 60 to 90 min only. It is speculated that the potassium effect involves the K+, Na+ dependent ATPase.

The present experiments were undertaken to throw light on the physiological role played by alpha-MSH in the advent of female puberty. The serum and pituitary alpha-MSH concentrations were determined in prepubertal rats 5, 15, 25, 30, 33 days old and at vaginal opening. The serum levels showed a significant increase on day 30 and the pituitary alpha-MSH levels increased steadily from day 5 to day 33. To determine if a relation exists between the increase in alpha-MSH levels and the peak of LH prior to vaginal opening, prepubertal rats of the same age range as above received an injection of alpha-MSH or saline solution and were sacrificed 30 min later. The peptide increased LH serum levels only at 30 days of age. To examine the hypothesis that alpha-MSH was involved in determining the timing of puberty, prepubertal rats pretreated with estradiol benzoate received an injection of alpha-MSH or saline solution on day 28. This treatment advanced the time of vaginal opening by 2 days in the experimental animals. With the same experimental procedure, an increase in serum levels of LH and progesterone was also observed at 29 days of age. The action of alpha-MSH on GnRH was analysed by incubating median eminence from prepubertal rats 20, 25 and 30 days of age, with the peptide, alpha-MSH increased GnRH release only in 25-day-old rats. The above results show that acute treatment with alpha-MSH induces modifications in the hormones related to puberty and hence allows us to include alpha-MSH in the chain of neuroendocrine events involved in reproductive maturation.

Rat pituitary cells were dispersed with trypsin and separated by sedimentation at unit gravity. The distributions of prolactin (Prl), luteinizing hormone (LH), and follicle-stimulating hormone (FSH) were determined, and two enriched cell populations (mammotrophs and gonadotrophs) were subsequently cultured. During a 4 h incubation, gonadotrophin-releasing hormone (GnRH) stimulated the release of LH and of FSH by both the unfractionated population and the enriched gonadotrophs; the magnitude of this stimulation increased with the length of the pre-culture periods, and the amount of LH released into the medium correlated strongly with the amount of FSH, whatever the length of the pre-culture period. The cellular cAMP content was also enhanced during the 4 h incubation, but no correlation was found between the hormone release and the cAMP accumulation. Furthermore, during the first 30 min of incubation with GnRH there was no increase of cellular cAMP, whatever cell population used. We conclude that the gonadotrophin release was independent of the cAMP accumulation observed in pituitary cells several hours after stimulation by GnRH; consequently, the late increase in the nucleotide is suggested to be a non-specific secondary process.

The effect of two synthetic injectable progestins, norethisterone oenanthate3 (NET-e) and medroxyprogesterone acetate (MPA) upon the hypothalamic pituitary unit was evaluated in post-menopausal women (PMW) and in castrated individuals with androgen un-responsiveness in order to gain insight into their mechanism of gonadotrophin inhibition. Continuous administration of natural progesterone (P4) to PMW by medicated vaginal rings was used as the experimental control. Elevated base line levels of serum immunoreactive gonadotrophins and normal pituitary LRH responses were found in PMW and pseudohermaphrodites. Progesterone did not affect circulating gonadotrophin levels or the pituitary LRH response in PMW. Administration of MPA 150 mg induced a clear decrease in serum gonadotrophins and a significant decrease in LRH pituitary responsiveness in a post-menopausal woman whereas no effect was observed on serum gonadotrophins and pituitary response to LRH in a patient with androgen unresponsiveness. Administration of NET-e 200 mg resulted in a significant decrease in circulating gonadotrophins in PMW and pseudohermaphrodites, while LRH pituitary sensitivity was diminished in all subjects except one post-menopausal woman. These results demonstrated that two structurally different synthetic progestins (NET-e and MPA) had a potent gonadotrophin inhibitory activity in PMW while natural progesterone did not, thus indicating a different mode of action. Furthermore their differing effect on individuals with complete androgen unresponsiveness suggested that antigonadotrophic activity of NET-e is mediated by its oestrogenic-progestational effect, while MPA acts through its androgenic potency and therefore requires the presence of androgen receptors.

The effects of chronic renal failure on the hypothalamic-pituitary-ovarian axis in 25 girls, aged 9.1 to 20.9 years (mean 13.8) were studied. Twelve patients on conservative treatment (group A) had serum creatinine values between 176.8 and 1502.8 μmol/l; 9 patients were on haemodialysis (group B); and 12 patients had received a renal transplant (group C). Tanner stage of breast development was delayed relative to chronological age in 5 out of 18 patients. Serum oestradiol was normal or low when related to pubertal stages in all groups. Serum LH was elevated in group A and B patients, but normal in group C patients. Serum FSH was elevated in 6 out of the 21 patients in group A plus B, and in 2 out of the 12 patients in group C. Serum PRL was elevated in 12/12, 6/8, and 4/11 patients in group A, B, and C respectively. After GnRH administration to 4 patients in group A, 3 showed delayed or absent gonadotropin response; all 4 patients studied in group C showed normal gonadotropin response. The data indicate a decreased E2 secretion, abnormal gonadotropin and PRL levels and a blunted gonadotropin response to GnRH in girls with chronic renal failure. These results seem to indicate an alteration of the hypothalamic-pituitary unit that can be reversed after successful renal transplantation.

The effects of vasoactive intestinal peptide (VIP) on the releases of LH and GnRH were examined in a sequential double chamber perifusion system by perifusing the medio-basal hypothalamus and/or pituitary excised from normal female rats in diestrus or ovariectomized rats. When the medio-basal hypothalamus and pituitary from normal rats in series were perifused with VIP (10−6 mol/l), the concentration of LH in the efflux was increased by 59–181% above that before the injection (P < 0.05), VIP having a dosedependent effect. VIP had no effect on LH release from the pituitary perifused alone. Infusion of VIP at 10−6 mol/l induced a significant release (84–159% increase, P < 0.05) of GnRH from the medio-basal hypothalamus. Infusion of 10−6 mol/l VIP induced a significant release (41–99% increase, P < 0.05) of LH in ovariectomized rats. These findings suggest that VIP induces GnRH release from the medio-basal hypothalamus, resulting in LH release from the pituitary, and that this process does not require ovarian estrogen.

The sensitizing effect of oestrogens on GnRH-stimulated LH release was evaluated in pituitary cells from adult female rats, cultured for 2 days in the presence of 10−13 to 10−6 m concentrations of oestradiol and selected A- and D-ring modified oestrogens. With almost all steroids tested, bell-shaped dose-response curves with comparable LH-maxima but different EDmax values were obtained for the LH response to a submaximal GnRH stimulus (5 × 10−10 m). Maximal LH response to 5 × 10−10 m GnRH were found at the following oestrogen concentrations: oestradiol and 4-hydroxyoestradiol = 10−11 m; 2-methyloestradiol = 10−9 m; 2-hydroxyoestradiol = 10−8 m; with 4-methyloestradiol no significant maximum was observed. When cells were pretreated with 10−13, 10−11 and 10−9 m oestradiol, or 4-hydroxyoestradiol, or 2-hydroxyoestradiol, and exposed to increasing concentrations of GnRH (10−11 to 10−7 m), an almost 10-fold decrease in the ED50 for GnRH was observed after pretreatment with 10−11 m oestradiol and 4-hydroxyoestradiol. With 2-hydroxyoestradiol at this concentration, the sensitizing effect was much less pronounced. Increasing the steroid concentration to 10−9 m slightly decreased the effect of oestradiol and 4-hydroxyoestradiol, whereas it increased the effect of 2-hydroxyoestradiol. Thus, at the target cell 4-hydroxyoestradiol has the same potency as oestradiol, while 2-hydroxyoestradiol is significantly less active. The sensitizing effect of oestradiol is only slightly decreased by the presence of a methyl group in position 2, but is markedly reduced by a methyl group in position 4. Our results also demonstrate the high sensitivity of the pituitary to oestrogen-induced enhancement of GnRH-stimulated gonadotrophin release, as well as the decrease of the positive effect at high oestrogen concentrations. The bell-shaped dose-response curves for oestrogen action should be taken into account when evaluating the effects of oestrogens and their derivatives upon gonadotrophin release from the pituitary gland.

The effect of chronic pulsatile low-dose GnRH treatment on the juvenile testis and associated structures was evaluated in relation to hormonal parameters in the peripheral blood in the pig. Starting at 8 weeks of age, male pigs (crossbreds of Dutch Landrace and Yorkshire breeds) were injected 6 times daily im with 0, 75 or 250 ng GnRH/kg body weight during 4 weeks. Immediately after the treatment period, a GnRH stimulation test with 750 ng GnRH/kg iv was carried out. Samples for plasma LH, FSH, testosterone and 5 alpha DHT measurement were obtained weekly (basal level) and after GnRH stimulation. The pigs were castrated at 12 weeks of age and the weights and lengths of the testis, epididymis and cremaster muscle were recorded. Intratesticular testosterone and 5 alpha DHT concentrations were determined, and the testis and epididymis were evaluated for histological changes. Basal plasma hormone concentrations, intratesticular androgen concentrations and the response of the pituitary gland to stimulation had not been affected by GnRH treatment. Pigs receiving the higher treatment dose of GnRH showed less increase in testosterone levels in response to the stimulation dose at 12 weeks of age than the other pigs. Morphologically, no changes were observed in the epididymis and cremaster muscle after GnRH treatment and no signs of reactivation of structures that can provoke testicular descent could be seen. The development of the seminiferous epithelium was more advanced in the GnRH-treated groups, apparently in a dose-dependent manner.