Sexual maturation modifies the catecholaminergic control of gonadotrophin secretion and the effect of ovarian hormones on hypothalamic neurotransmitters in female rats.

alpha-Methyl-p-tyrosine (alpha-MT), a competitive inhibitor of tyrosine hydroxylase, was used to block the synthesis of hypothalamic catecholamines in immature female rats of 14, 16 and 30 days of age and in castrated adults. The administration of alpha-MT (300 mg/kg body weight, free base) induced a significant decay in the hypothalamic content of norepinephrine (NE) and dopamine (DA) within the first 120 min. A second dose (150 mg/kg body weight), given 2 h after the first injection, did not further modify the low catecholamine levels observed 120 min after the first alpha-MT administration. The administration of 300 mg/kg body weight of alpha-MT induced a significant increase in LH concentrations in rats aged 14 and 16 days. On the contrary, after an alpha-MT injection, a significant LH decrease was observed in 30-day-old and in adult castrated rats. alpha-MT also increased FSH levels in prepubertal rats of 16 days of age, but no change occurred in 30-day-old and in adult rats. The administration of estrogen-progesterone (EP) to prepubertal rats of 16 days of age induced a significant decrease in serum LH levels as well as in the serotonin (5-HT) and 5-hydroxyindole-acetic acid (5-HIAA) concentrations in the anterior-preoptic hypothalamic area (AH-POA), but not in the medial basal hypothalamus. No modifications in the catecholamine content of these hypothalamic areas were observed in this age group after EP administration. On the contrary, in 30-day-old rats, EP induced a significant LH release as well as an increase in AH-POA concentrations of 5-HT, 5-HIAA and catecholamines.(ABSTRACT TRUNCATED AT 250 WORDS)     

Changes in gonadotrophin binding to rat ovaries during sexual maturation

Changes in LH and FSH binding to rat ovaries during sexual maturation were measured by in vitro binding assay and autoradiography. Ovaries were obtained from rats at various ages ranging from 5 till 35 days of age. Animals older than 35 days of age were classified in three groups: anoestrus (uterine weight below 60 mg), early pro-oestrus (uterine weight 60-120 mg) and pro-oestrus (uterus distended with fluid). Measurements were made of specific binding of radioiodinated hCG and rFSH to ovarian homogenates (day 5-35) and to granulosa cells isolated from the largest follicles (day 35-pro-oestrus). The earliest age at which specific hCG and FSH binding could be demonstrated was day 11. Thereafter hCG binding to ovarian homogenates, expressed in CPM/microgram DNA, increased with age. Specific binding of hCG to granulosa cells increased from 1000 CPM/microgram DNA at day 35 to 4000 CPM/microgram DNA at pro-oestrus. FSH binding to ovarian homogenates increased till day 21 and remained fairly constant thereafter. Autoradiography revealed the presence of hCG binding to interstitial tissue and thecal cells. hCG binding to granulosa cells was only found in large pre-ovulatory follicles present at early pro-oestrus and pro-oestrus. FSH binding was observed to granulosa cells of all types of follicles, but not to interstitial and thecal cells. The results demonstrate that during sexual maturation FSH binding to granulosa cells remains constant, whereas LH binding to interstitium and theca and ultimately to granulosa cells increases with age suggesting an increased responsiveness of the ovaries to LH.     

Changes in the effect of gamma-aminobutyric acid on prolactin secretion during sexual maturation in female rats.

To evaluate the effects of the gamma-aminobutyric acid (GABA)ergic system on PRL secretion during sexual development in female rats, aminooxyacetic acid (an inhibitor of GABA-transaminase which increases the hypothalamic GABA concentration) was administered to 12- to 16- and 30-day-old female rats. The increased GABA level in the hypothalamus was accompanied by a significant increase in the serum PRL concentration in rats 16 days of age and a decrease in the serum concentration of the hormone in 30-day-old rats. No changes in PRL levels were observed at 12 days of age. Muscimol, a GABA-A receptor agonist, increased serum PRL concentrations in 16-day-old rats. By contrast, in rats 30 days of age, muscimol induced a significant decrease in PRL levels. On the other hand, the administration of baclofen, a GABA-B receptor agonist, induced a pattern of PRL modification similar to that observed with muscimol, i.e. stimulation in rats 16 days of age and inhibition in 30-day-old rats. These findings indicate that both the stimulatory and inhibitory effects of the GABAergic system on PRL secretion during sexual maturation are mediated by GABA-A and GABA-B receptors. The present results show that during sexual maturation there is a qualitative modification of the effect of the GABAergic system on PRL secretion. These findings could be related to the complex neuroendocrine mechanisms involved in the onset of puberty in the female rat.     

Influence of anti-oestrogens on gonadotrophin secretion in control and ACTH-infused immature rats

The objective of this study was to determine whether anti-oestrogens (nafoxidine, MER-25) would block the suppressive effects of ACTH on gonadotrophin secretion in immature rats. Female rats were castrated at 25-26 days of age, and an Alzet osmotic minipump containing ACTH (1-24) or saline was implanted in each animal. ACTH was administered at a rate of 1 IU/day by constant infusion. Beginning on the day of surgery, animals were injected daily for 5 days with 0.25, 5 or 25 micrograms/100 g body weight of nafoxidine or 5 mg MER-25 and sacrificed on the sixth day following castration. ACTH lowered serum LH concentrations and increased pituitary LH levels. Serum androstenedione concentrations were more than two times greater in ACTH-infused than in control rats, but serum oestrone levels were not affected. Serum testosterone and oestradiol concentrations in ACTH-infused rats remained below levels of detection. Administration of 0.25 micrograms of nafoxidine prevented the suppressive effects of ACTH on serum LH. Serum levels of LH in these animals were comparable to saline-treated controls (418 +/- 94 vs 443 +/- 73 ng/ml). The two higher doses of nafoxidine and MER-25 were ineffective in suppressing the actions of ACTH on serum LH. MER-25 reduced serum LH values in both controls and ACTH-infused rats. Serum FSH concentrations were not altered by ACTH or nafoxidine treatment. MER-25 elevated pituitary FSH concentrations in both control and ACTH-infused rats.(ABSTRACT TRUNCATED AT 250 WORDS)     

Role of the organum vasculosum laminae terminalis in the control of gonadotrophin secretion in rats

The organum vasculosum laminae terminalis (OVLT) was destroyed by radiofrequency lesions in regularly cycling and in long-term ovariectomized adult rats. After OVLT lesion practically all cyclic females (16 out of 22) became dioestrous, as indicated by vaginal smears. At the time of killing these animals (8 days after the lesion) serum LH levels were undetectable, while serum FSH was as low as in cyclic animals in dioestrus. In the few OVLT-lesioned animals which exhibited some sort of oestrous cyclicity, serum LH showed a small subphysiological increase at pro-oestrus; this was not accompanied by a parallel increase in serum FSH and in these animals a delayed peak of FSH occurred on the day of oestrus. Ovariectomized rats bearing OVLT lesions had serum titres of Lh and FSH as high as those of ovariectomized control rats. It is suggested that the OVLT may play a role in the control of the cyclic release of gonadotrophins but is not involved in the tonic regulation of gonadotrophin secretion.     

Pituitary gonadotrophin secretion during the first weeks of pregnancy.

A longitudinal study of basal plasma LH and FSH and their responses to 25 microng LRH iv as well as basal levels of oestradiol, progesterone, prolatin and HCG was performed every week in 3 women, pregnant after heterologous insemination, from conception until the 6th week of gestation. A comparative study was carried out in 7 women in cycles in which no conception occurred after insemination. All hormones were assayed with radioimmunoassay. LH was measured with a specific assay for native HL, which did not cross-react with HCG. A decrease in basal levels of LH and FSH as well as decreasing responses to LRH was found during the first 2 weeks of gestation. These changes did not differ from what was observed during the luteal phase in the non-conception cycles. One week later the basal FSH levels and the FSH response in the pregnant women showed a further decrease, while in the non-pregnant women, now reaching the early follicular phase, a rise in FSH basal levels occurred. The basal levels of LH and the LH response, however, did not differ from that found in the non-pregnant woment at this time. FSH basal levels remained below the lower normal limit in eumenorrhoic women from the 3rd week of gestation. By this time the FSH response was almost completely inhibited. The LH basal levels, however, remained above the lower normal limit in eumenorrhoic women, but the LH response to LRH progressively decrease and was completely inhibited by the 5th week of gestation. In the non-conception cycles the LH response varied with the levels of oestradiol in plasma. This was not found in the pregnant women as the decrease in gonadotrophin response occurred while oestradiol remained at mid-cycle levels during the first 4 weeks of gestation. Rather it seems that the increasing and continuously elevated level of progesterone, in the presence of appropriate levels of oestradiol, might be the main go nadal steroid responsible for the diminishing pituitary secretion. The contribution of HCG to the further decrease in gonadotrophin secretion after the 2nd week of pregnancy cannot be answered by the present studies. Prolactin remained at non-pregnant levels until the 6th week of gestation, and appeared to have no influence on the secretion of gonadotrophins during early pregnancy.     

Ovarian control of gonadotropin secretion during induced precocious sexual maturation in the rat.

In sexually immature female rats 'primed' with a unilateral basal hypothalamic lesion on day 23 of life, a gonadotropin surge was triggered by progesterone (P) on day 26. The influence of sequential ovariectomy (OVX) on this response was studied in an attempt to elucidate the mode of action of brain lesions inducing precocious sexual maturation in this species. OVX, carried out on days 23-26, abolished or greatly reduced the LH surge, but had little effect on basal LH concentrations. In contrast, basal FSH concentrations rose rapidly after OVX. In lesioned rats, the post-castration rise of FSH could be prevented by leaving the ovaries in place during the 1st 24 or 48 h. It is concluded that the lesion-induced maturation of the positive feedback mechanism underlying the pubertal preovulatory gonadotropin surge is mediated by the ovaries. In prepubertal rats at this age, tonic FSH release, but not tonic LH release, is under tight negative feedback control.     

Changes in oxytocin and vasopressin secretion during sexual activity in men

We measured plasma oxytocin (OT) and arginine vasopressin (AVP) concentrations in 13 normal men during sexual arousal and ejaculation. Mean plasma AVP increased from 1.4 +/- 0.2 (+/- SE) to 5.3 +/- 1.7 pmol/L (P less than 0.05) during arousal, but there was no significant change in OT. In contrast, at ejaculation mean plasma OT rose from a basal value of 1.4 +/- 0.3 to 7.3 +/- 0.6 pmol/L (P less than 0.01) and then fell to basal concentrations in 30 min. AVP, however, had returned to basal levels at the time of ejaculation and remained stable thereafter. We conclude that in man AVP is secreted during sexual arousal, and there is, subsequently, a selective release of OT at the time of ejaculation.     

Serotoninergic control of gonadotrophin and prolactin secretion in women

OBJECTIVE Due to conflicting observations from previous investigations, the role of serotonin (5-HT) in the regulation of the human menstrual cycle has not been clearly established. We have therefore investigated the possible participation of 5-HR in the control of gonadotrophin and PRL secretion in women, using the potent 5-HT₃ receptor antagonist ondansetron as a pharmacological probe. DESIGN Serum profiles of LH, FSH and PRL were obtained in 9 normally cycling women during a control and a treatment cycle, during which ondansetron (8 mg orally) was administered daily. On day 10 of both cycles, the serum pulsatility of LH, FSH and PRL was assessed by frequent blood sampling (at 10-minute intervals for 10 hours). Pituitary responsiveness was tested by administration of a GnRH bolus (25 μg I.v. after 8 hours). MEASUREMENTS LH, FSH and PRL were serially determined in all blood samples by immunofluorescence assays. The resulting hormone data arrays were searched for significant fluctuations by the Cluster pulse algorithm. RESULTS Compared with control cycles, the temporal organization and the endocrine characteristics of the treatment cycles remained virtually unaltered. Serotonin antagonism did not noticeably affect the LH pulse attributes (frequencies, interpulse intervals, amplitudes). Although FSH amplitudes declined markedly (P < 0·05), the remaining pulse attributes were unchanged. A clear increase (P < 0·05) in the PRL pulse frequency was noted, while PRL pulse amplitudes tended to increase (P= 0·1). Gonadotrophin and PRL release in response to GnRH administration was unaltered by ondansetron treatment. CONCLUSIONS Serotoninergic blockade by a selective 5-HT₃ receptor antagonist failed to modify pulsatile LH secretion, but induced distinct changes in episodic FSH and PRL secretion. Since the pituitary gonadotrophin and PRL responsiveness remained unaltered during 5-HT₃ receptor blockade, the observed alterations in the FSH and PRL secretion presumably relate to altered hypo-thalamic regulation of these pituitary hormones. Thus, the central regulation of pulsatile FSH and PRL release in women appears to involve 5-HT₃ receptor-mediated processes.     

Changes in insulin, somatostatin, and glucagon secretion during the development of obesity in ventromedial hypothalamic-lesioned rats

Changes in insulin, somatostatin, and glucagon secretion during the development of obesity in rats after ventromedial hypothalamic (VMH) lesions were evaluated by measuring fasting hormone levels and their secretion from the isolated perfused pancreas. Fasting peripheral insulin levels were not altered 1 week after the VMH lesions but became progressively elevated at 3-4, 8-9, and 11-12 weeks compared to the values in sham-operated and age-matched control rats. In the portal vein, insulin levels also progressively increased in VMH-lesioned rats, but the portal-peripheral gradient of insulin in the later phase of VMH obesity was significantly lower than in the early phase after VMH lesions. On the contrary, the arginine-induced insulin release from the perfused pancreas was highest at 1 week and gradually decreased thereafter, although it continued to remain higher than that of controls. The perfusate somatostatin response to arginine also was exaggerated in the VMH-lesioned rats. However, both the peripheral glucagon level and the glucagon secretion from the perfused pancreas of the VMH-lesioned rats were not significantly different from the controls. These results show that VMH lesions result in an increased insulin and somatostatin secretion. Using the cyclically perfused liver in situ, we have found that the hepatic extraction rate of insulin is indeed reduced in rats 8-9 weeks after VMH lesioning, and so have at least partly accounted for the decreased portal-peripheral gradient of insulin in the later VMH postoperative phase.     

Gene regulation by NMDA receptor activation in the SDN-POA neurons of male rats during sexual development

The present study was designed to identify possible signaling pathways, which may play a role in prevention of neuronal apoptosis in the sexually dimorphic nucleus of the preoptic area (SDN-POA) after physiological activation of the N-methyl-D-aspartate (NMDA) receptor. Gene response to the blockage of the NMDA receptor by an antagonist (dizocilpine hydrogen maleate; MK-801) was screened after suppression subtractive hybridization (SSH). The results showed that differential screening after SSH detected the presence of some neurotrophic genes (RNA binding motif protein 3 (RBM3), alpha-tubulin) as well as apoptosis-related genes (Bcl-2, cytochrome oxidase subunit II, cytochrome oxidase subunit III) in the SDN-POA of male rats, which were down-regulated by blocking the NMDA receptor. The RT-PCR products of the aforementioned genes in MK-801-treated males were significantly less than that in untreated males. In particular, the expression of Bcl-2 mRNA, including Bcl-2 protein, in male rats were significantly suppressed by MK-801 treatment. Moreover, the binding activity of nuclear factor kappaB (NFkappaB) was significantly higher in male rats than in females, but significantly diminished by blocking the NMDA receptor with MK-801 in male rats. No significant difference in cAMP response element-binding protein (CREB) binding activity was observed among untreated male, MK-801-treated male, untreated female and MK-801-treated female groups. These results suggest that genes regulated by NMDA receptor activation might participate in neuronal growth and/or anti-apoptosis, and support an important signaling pathway of NFkappaB activation and its target gene, Bcl-2, in preventing neuronal apoptosis in the SDN-POA of male rats during sexual development.     

Development and control of the opioid inhibition of gonadotropin secretion during the sexual maturation of the male rat.

The endogenous opioid system tonically inhibits the LH-releasing apparatus in adult male rats but not in immature animals. To determine the relationship between the onset of this effect and the peripubertal testosterone rise, male rats were examined at 5-day intervals from day 25 through day 65. They were injected subcutaneously with saline, 0.25 or 1.0 mg/kg body weight naloxone and sacrificed 20 min later. Another group of immature males was castrated, implanted with testosterone-filled capsules and tested with naloxone 4 days later. The peripubertal increase in testosterone and the ability of naloxone to increase serum LH concentrations were both first statistically significant on day 45. Testosterone treatment of immature rats did not induce a naloxone effect. The ability of hypothalamic fragments to release LHRH in vitro in response to naloxone also appeared to occur at the same time as the peripubertal testosterone rise. Hypothalamic fragments obtained from immature male rats treated in vivo with testosterone were capable of responding to naloxone with LHRH release in vitro. These data suggest that in the rat the maturation of the endogenous opioid system is a component of male puberty that is induced by the peripubertal testosterone rise.