Silvia Carbone

Sexual maturation modifies the GABAergic control of gonadotrophin secretion in female rats

Administration of aminooxyacetic acid, (an inhibitor of gamma-aminobutyric acid (GABA)-transaminase which enhances the hypothalamic GABA content) increased luteinizing hormone (LH) and follicle stimulating hormone (FSH) levels in prepubertal rats (16 days of age), and decreased the levels of these pituitary hormones in peripubertal rats (30 days of age). Prepubertal and peripubertal female rats were administered with muscimol, a GABAA-antagonist, with baclofen, a GABAB agonist, and with bicuculline, a GABA-antagonist, and the serum concentrations of LH and FSH were determined. In prepubertal rats, muscimol increased both LH and FSH levels, while in peripubertals the GABAA agonist showed the opposite effect, i.e. both gonadotrophins were decreased after its administration. Baclofen lowered serum concentrations of LH and FSH at the different ages studied. Administration of bicuculline produced a decrease in LH and FSH concentrations in prepubertal rats, and an increase of these values in peripubertal rats. These results indicate that GABA exerts a stimulatory tone on gonadotrophin secretion in prepubertal rats and an inhibitory one in peripubertal animals. This effect is most probably mediated by GABAA receptors. It is suggested that the change in the effect on gonadotrophin secretion of the activation of GABAA receptors which takes place during sexual maturation in the female rats is related to the central mechanisms involved in the onset of puberty.

In vivo and in vitro studies on the effect of the serotorinergic system on luteinizing hormone and luteinizing hormone-releasing hormone secretion in prepubertal and peripubertal female rats

The present investigations were designed to assess the effect of the serotoninergic system on luteinizing hormone (LH) and LH-releasing hormone (LH-RH) secretion in female rats aged 14 and 30 days. The administration of 5-hydroxytryptophan (5-HTP; 75 mg/kg i.p.) increased hypothalamic serotonin (5-HT) concentrations in both age groups, and did not affect hypothalamic norepinephrine (NE) concentrations or release. Serum LH levels were raised by 5-HTP in 14-day-old, but not in 30-day-old rats. Basal and KCl- (28 mM) stimulated LH-RH release by incubated hypothalamic fragments was significantly enhanced when 5-HTP was injected previously to 14-day-old animals. In 30-day-old rats, 5-HTP treatment did not modify basal LH-RH release, and decreased the KCl-stimulated LH-RH output. Similarly, the addition of 5-HT (10(-7) M) to superfused hypothalamic fragments enhanced basal LH-RH release in 14-day-old rats and blocked the increment in LH-RH release evoked by KCl in 30-day-old rats. The present results show that in 14-day-old female rats, the serotoninergic system (activated in vivo by 5-HTP treatment, or in vitro by 5-HT addition) exerts a stimulatory effect on LH-RH, and thus, on LH release. On the contrary, in 30-day-old animals, stimulated LH-RH secretion was inhibited by 5-HT. Apparently, the hypothalamic NE system is not implicated in this response. The participation of this changing effect of 5-HT on LH-RH/LH release at the onset of puberty is postulated.

Sexual differences in the effect of the GABAergic system on LH secretion and in the hypothalamic ontogenesis of GABAA receptors in prepubertal rats

The administration of aminooxyacetic acid (AOAA), which increases hypothalamic GABA concentrations, induced a significant increase in LH levels in female rats of 12, 16 and 18 days of age. This stimulatory effect of AOAA on serum LH levels was not observed at 21 and 25 days of age whereas a significant decrease in the LH concentrations by AOAA was found at 30 days of age. The neonatal androgenization of female rats abolished the effects of AOAA and no effects of AOAA were found in male rats at the different ages studied. The muscimol binding sites in medial basal and preoptic anterior hypothalamic areas showed a significant different sexual ontogenic pattern. Males rats castrated at birth showed a very similar ontogenic pattern of [3H]muscimol binding sites to female rats and significantly different to male control rats. It is concluded that there are sexual differences in the effect of GABAergic system on LHRH-LH secretion and in the development of hypothalamic GABAA receptors during sexual maturation. These differences are probably connected with the neonatal exposure to androgens that would induced changes in the composition and probably in the properties of GABAA receptor that in turn modified its effects on LHRH neurons.

Differential effects of the N-methyl-D-aspartate and non-N-methyl-D-aspartate receptors of the excitatory amino acids system on LH and FSH secretion. Its effects on the hypothalamic luteinizing hormone releasing hormone during maturation in male rats.

The present experiments describe the effect of NMDA and kainate agonists of the NMDA and non-NMDA subtype of receptors respectively of the excitatory amino acids (EAAs) system in prepubertal (16 days of age) and peripubertal (30-day-old rats) male rats on the in vitro hypothalamic release of GnRH, and on the in vivo LH and FSH levels as well as the effect of testosterone on these effects. The addition of NMDA or kainate to the medium containing APOA-MBH areas significantly increased (P < 0.01) the GnRH release as compared with the respective controls. The increase in GnRH release observed with kainate was significantly higher (P < 0.01) than those observed with NMDA. NMDA administration increased significantly (P < 0.01) serum LH levels at both ages of sexual maturation while no effect was observed by kainate administration. MK 801, an antagonist of NMDA neurotransmission, and testosterone abolished the LH release response to NMDA. Contrary to that observed on LH, while NMDA did not modify serum FSH concentrations a significant increase (P < 0.01) was observed with kainate administration in prepubertal and peripubertal rats on this pituitary hormone, and CNQX, an antagonist of non-NMDA neurotransmission, and testosterone administrations blocked this FSH release effect of kainate. The NMDA and kainate release effect on LH and FSH respectively was significantly higher in prepubertal than in peripubertal rats. At both ages NMDA released more LH than kainate FSH. In conclusion, our experiments demonstrated that both subtypes of glutamate receptors NMDA and non-NMDA subtypes of EAAs increased GnRH release by APOA-MBH in vitro during sexual maturation. Nevertheless, while NMDA administration only increased serum LH levels, kainate showed only an effect on increasing FSH concentrations. These differential effects of NMDA and non-NMDA subtypes of EAA receptors on LH and FSH could probably explain some aspects of the differential modifications of LH and FSH observed in different physiological circumstances.

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)

Interactions between GABAergic and serotoninergic systems with excitatory amino acid neurotransmission in the hypothalamic control of gonadotropin secretion in prepubertal female rats

The present studies were designed to study the interrelationships between GABAergic, serotoninergic and excitatory amino acids systems (EAAs) in the control of gonadotropin secretion in prepubertal female rats. For this purpose we determined the effects of N-methyl-d-aspartate (NMDA), an exogenous agonist of EAAs receptors, on LH and FSH secretion in 16-day-old female rats in which the GABA-A and GABA-B receptors were blocked by bicuculline and baclofen or serotonin (5-HT) depleted by p-choloroamphetamine (PCA). In addition the effects of the GABAergic and serotoninergic systems on LH and FSH secretion were evaluated in animals treated with dibenzocycloalkenimine (diocilpine MK-801), an antagonist of NMDA neurotransmission. While muscimol, a GABA- A agonist, induced a significant increase in LH and FSH levels (P<0.01), baclofen, a GABA-B agonist, had an inhibitory effect on these hormones (P<0.01). MK 801, a NMDA receptor antagonist, not only suppressed the stimulatory effect of NMDA on LH and FSH but also blocked the stimulatory effect of muscimol without modifying the inhibitory action of baclofen on both gonadotropins. Bicuculline, a GABA-A receptor antagonist, did not modify the release effect of NMDA on LH and FSH. 5-HTP, a precursor of 5-HT that increases the levels of this neurotransmitter in the central nervous system significantly increased (P<0.01) the plasma levels of LH and FSH, and this effect was blocked by the NMDA receptor antagonist MK-801. We conclude that the stimulatory effects of GABAergic and serotoninergic systems in prepubertal female rats are connected with the activation of EAA neurotransmission, while the stimulatory effects of NMDA appear to be independent of serotoninergic and GABAergic actions on LH and FSH secretion. Since both GABA and serotonin systems change their effects on LH and FSH during sexual maturation from a stimulatory action in prepubertal to an inhibitory action in adult rats and since NMDA neurotransmission has a stimulatory effect on gonadotropin secretion both in prepubertal and adult rats, it is clear that the interrelationships between GABAergic and serotoninergic systems with EAAs in the gonadotropin control are different in prepubertal and in adult rats.

Effect of Ovarian Hormones on the Hypothalamic Excitatory Amino Acids System during Sexual Maturation in Female Rats

The present experiments were designed to study in female rats during sexual maturation: (1) the hypothalamic release of aspartate (Asp), glutamate (Glu) and glycine (Gly) which are the excitatory amino acids (EAAs) involved in NMDA neurotransmission and of taurine (Tau), a putative inhibitory amino acid of GnRH secretion; (2) the relationships between the effect of estrogen-progesterone (EP) on the release of these EAAs and the secretion of gonadotropins, and (3) the effect of hypothalamic NMDA receptor stimulation on EAAs release by the hypothalamus as well as the effect of EP on this release. The release of EAAs by the anterior preoptic and medial-basal hypothalamic areas (APOA-MBH) is significantly higher in peripubertal than in prepubertal rats (p < 0.01). EP treatment in prepubertal rats (16 days of age) decreased LH and FSH plasmatic levels and also the in vitro release of Asp, Glu, Gly and Tau. Contrary to the observations in prepubertal rats, in 30-day-old peripubertal rats the ovarian hormones significantly (p < 0.01) increased the levels of LH and FSH as well as the release to the medium of these amino acids.(ABSTRACT TRUNCATED AT 250 WORDS)

Hypothalamic excitatory amino acid system during sexual maturation in female rats

The present results indicate that during sexual maturation the APOA-MBH from rats of 30 days of age released significantly higher quantities of GnRH than the tissue from 16-day-old rats (P < 0.01). The addition of NMDA, an agonist of the excitatory amino acids system (EAAs), to the medium after 30 min of incubation significantly increased (P < 0.01) the GnRH release in normal rats of both ages and this increase was significantly (P < 0.01) higher in 30-day-old rats (to 661%) than in rats of 16 days of age (to 273%). The administration of estrogen-progesterone (EP) to rats of 16 days of age did not modify the GnRH release response to NMDA. On the contrary, at 30 days of age EP administration significantly potentiated the GnRH release response to NMDA since while in the control group NMDA increased the GnRH release to 630%, in the EP-pretreated group this was to around 4700% (P < 0.01). EP pretreatment of prepubertal rats decreases the hypothalamic release of aspartate and glutamate, the excitatory amino acids involved in NMDA neurotransmission and glycine but increases EAAs release in peripubertal rats. On the basis of these results it is proposed that the increase in EAAs release by the hypothalamus is directly connected with the onset of puberty and that the maturation of the positive feedback effect of ovarian hormones on gonadotropin secretion is related to the maturation of the capacity of EP to increase hypothalamic EAAs. Before this maturational event EP inhibits EAAs release as well as gonadotropin release (prepubertal rats). NMDA receptor stimulation leads to a positive mechanism which increases the release of Asp and Glu from APOA-MBH both in prepubertal and peripubertal rats, but EP potentiates this mechanism only in peripubertal rats. This could be an additional neuroendocrine mechanism involved in the increase of gonadotropin during sexual maturation which induces the onset of puberty and the preovulatory discharge of these pituitary hormones.

Different effects by sex on hypothalamic–pituitary axis of prepubertal offspring rats produced by in utero and lactational exposure to di-(2-ethylhexyl) phthalate (DEHP)

This study investigated the effect of pre and perinatal exposure to di-(2-ethylhexyl) phthalate (DEHP) on the neuroendocrine parameters that regulate reproduction in prepubertal male and female rats. DEHP at doses of 3 and 30mg/kgbw/day was administered orally in the drinking water to dam rats since pregnancy onset until the moment of pups sacrifice at 15 days of age. In these animals gonadotropin serum level and the hypothalamic contents of the amino acids aspartate, glutamate and gamma-aminobutyric acid were determined. No changes in gonadotropin levels and amino acid neurotransmitters were detected at the low dose in both sexes. However, DEHP administered at high dose (30mg/kgbw/day) to dams produced a significant decrease in the inhibitory neurotransmitter GABA and an increase in the stimulatory neurotransmitter aspartate in prepubertal male offspring rats. These modifications were accompanied by gonadotropin serum levels increase. On the contrary, in treated female rats this chemical increased both, aspartate and GABA, which exert a characteristic stimulatory action on gonadotropin in 15-day-old normal females. This study provides new data about changes produced by DEHP on the hypothalamic amino acid neurotransmitters involved in the neuroendocrine reproductive regulation, in prepubertal male and female rat offspring from dams exposed during gestational and lactational periods. These alterations induced by DEHP exposure could be related to the gonadotropin modifications also described in this work, and with changes in the production of sexual hormones previously reported by other authors.

Exposure to a low dose of bisphenol A impairs pituitary-ovarian axis in prepubertal rats: Effects on early folliculogenesis

The research work studies the effect of providing a low dose of bisphenol A (BPA), on the reproductive axis of prepubertal female rats. Wistar mated rats were treated with either 0.1% ethanol or BPA in their drinking water until their offspring were weaned on the 21 day of birth. The estimated average dose of exposure to dams was approximately 3μg/kg/day. The pups were sacrificed at the 30th day of life. Body weight at the moment of the sacrifice was significantly higher in the group exposed to BPA; ovarian weight and its relative weight were not modified. LH and estradiol levels increased significantly, meanwhile FSH ones showed no significant changes. The number of primary, secondary and atretic follicles increased and antral ones was decreased. Our results demonstrated that early exposure to a low dose of BPA disrupts the normal function of the reproductive axis in prepubertal female rats.