[Show abstract][Hide abstract] ABSTRACT: During reproductive aging female rats enter an anovulatory state of persistent estrus (PE). In an animal model of reinstatement of estrus cyclicity in middle-aged PE rats we injected the animals with progesterone (0.5 mg progesterone/kg body weight) at 12:00 for 4 days whereas control animals received corn oil injections. After the last injection animals were analyzed at 13:00 and 17:00. Young regular cycling rats served as positive controls and were assessed at 13:00 and 17:00 on proestrus. Progesterone treatment of middle-aged PE rats led to occurrence of luteinizing hormone (LH), follicle stimulating hormone (FSH), and prolactin surges in a subset of animals that were denoted as responders. Responding middle-aged rats displayed a reduction of ER-beta mRNA in the preoptic area which was similar to the effect in young rats. Within the mediobasal hypothalamus, only young rats showed a decline of ER-alpha mRNA expression. A decrease of ER-alpha mRNA levels in the pituitary was observed in progesterone-responsive rats and in young animals. ER-beta mRNA expression was reduced in young regular cycling rats. ER-beta mRNA levels in the ovary were reduced following progesterone treatment in PE rats and in young rats. Taken together our data show that cyclic administration of progesterone reinstates ovulatory cycles in intact aging females which have already lost their ability to display spontaneous cyclicity. This treatment leads to the occurrence of preovulatory LH, FSH and prolactin surges which are accompanied by differential modulation of ERs in the hypothalamus, the pituitary and the ovary.
[Show abstract][Hide abstract] ABSTRACT: In females, it is well established that changes in the expression of neurotransmitters and peptides regulating the activity of the gonadotrophin-releasing hormone (GnRH) pulse generator are altered during ageing. By contrast, little is known about whether those age-related changes also occur in males. Therefore, we designed an animal study with orchidectomised young and middle-aged male rats to investigate changes in luteinising hormone (LH) secretion profiles and changes in the mRNA expression of genes regulating the activity of the GnRH pulse generator. Our results demonstrate that middle-aged rats exhibit lower serum LH levels and relatively fewer LH pulses with attenuated amplitude compared to young animals. Furthermore, upon ageing, GnRH mRNA expression is up-regulated in the preoptic area and the septum where GnRH neurones reside. Analysis of mRNA levels of glutamate decarboxylase (GAD) enzymes revealed that GAD(65) and GAD(67) mRNA expression increased in the mediobasal hypothalamus (MBH) and that GAD(67) mRNA levels decreased in the suprachiasmatic nucleus. In addition, we observed an age-related increase of oestrogen receptor (ER)alpha mRNA in the MBH, and both ERalpha and ERbeta mRNA expression was up-regulated in the pituitary of middle-aged rats compared to young animals. Taken together, our data support the existence of a male 'andropause' that is, like the menopause in females, accompanied by changes in neurotransmitter and hormone receptor expression that are involved in regulating the function of the GnRH pulse generator.
Journal of Neuroendocrinology 11/2007; 19(10):779-87. DOI:10.1111/j.1365-2826.2007.01589.x · 3.14 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: GnRH agonists are the established treatment of precocious puberty caused by premature stimulation of gonadotropin secretion. It has been reported that after an initial stimulation ("flare-up") they reduce LH secretion by desensitization of pituitary GnRH receptors. Little has been published about the use of GnRH antagonists such as cetrorelix to control the onset of puberty and whether they are potentially advantageous compared with GnRH agonists. We conducted two multigroup experiments (12 and 10 d, respectively) treating prepubertal/peripubertal female rats with either the GnRH agonist triptorelin or buserelin and compared them with rats treated with the GnRH antagonist cetrorelix and controls to assess the effects on pubertal progress and serum hormones. In the second experiment, the effects of buserelin and cetrorelix on gene expression of the GnRH receptor, LH-beta, FSH-beta, and the alpha subunit genes in the pituitary were also investigated. Cetrorelix, triptorelin, and buserelin retarded the onset of puberty as determined by delayed vaginal opening, lower ovarian weights, and lower serum estradiol levels. However, although LH and FSH levels were stimulated by both agonists, they were inhibited by cetrorelix. In the cetrorelix versus buserelin experiment, pituitary gene expression of the GnRH receptor and LH-beta subunit were significantly lower in cetrorelix treated rats compared with controls whereas buserelin had little effect. Expression of FSH-beta and alpha subunit were stimulated by buserelin but not by cetrorelix. Even though all three of these GnRH analogues inhibited gonadal development and delayed the onset of puberty, the GnRH agonists had stimulating and inhibiting effects on the pituitary-gonadal axis whereas cetrorelix exerted only inhibiting effects. We conclude from this female rat model that cetrorelix may offer advantages for a more controlled medical treatment of precocious puberty compared with GnRH agonist treatment.
Pediatric Research 11/2000; 48(4):468-74. DOI:10.1203/00006450-200010000-00009 · 2.31 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Increasing evidence suggests that interleukin-1β (IL-1β) regulates luteinizing hormone (LH) release primarily through modulation of the gonadotropin-releasing hormone (GnRH) neuronal activity. This study was undertaken to elucidate the effect of IL-1β on GnRH as well as GnRH receptor (GnRHR) gene expression in the preoptic area. IL-1β (100 ng/rat) or saline was administered into the lateral ventricle of castrated rats. RNA samples were isolated from micropunches of the preoptic area and mediobasal hypothalamus from individual brain slices and GnRH mRNA levels in the preoptic area and GnRHR mRNA levels in the mediobasal hypothalamus were determined by competitive reverse transcription-polymerase chain reaction (RT-PGR) protocols. Serum LH concentrations were decreased from 1 h to 3 h after IL-1β treatment, but rebounded at 5 h, while serum concentrations of follicle-stimulating hormone (FSH) and prolactin were not altered. There were no significant changes in GnRH mRNA levels from the micropunched preoptic area, while GnRHR mRNA levels from the preoptic area and mediobasal hypothalamus micropunch samples, but not in the anterior pituitary, showed a pattern similar to the serum LH profile following i.c.v. administration of IL-1β. We then examined the effect of IL-1β on the translational efficiency of the GnRH mRNA. After the separation and fractionation of polyribosome-associated cytoplasmic RNA from the hypothalamic fragments containing the preoptic area-anterior hypothalamic area of control (saline-treated) and IL-1β-treated group 3 h after administration, GnRH transcript levels were examined from the each fraction. IL-1β decreased the translational efficiency of the transcribed GnRH mRNA. These results clearly demonstrate that central administration of IL-1β suppresses the translational activity of GnRH mRNA. Moreover, GnRHR may play an important role in the modulation of GnRH neuronal activity through GnRHR-expressing neurones (or glia) in the hypothalamus.
Journal of Neuroendocrinology 06/2000; 12(5):421-9. DOI:10.1046/j.1365-2826.2000.00466.x · 3.14 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Cranial irradiation in prepubertal children with leukemia or brain tumors can lead to precocious or in high doses to late puberty. To unravel the underlying mechanisms, we developed a rat model with selective cranial Co60-irradiation technique. Infantile (12-16 d old) or juvenile (21-23 d old) female Sprague-Dawley rats received a single dose of 4, 5, 6, 9 or 2 x 9 Gy (at days 21 and 23). Each group consisted of 7-20 animals. High radiation doses (9 Gy and more) caused retardation of sexual development, whereas low radiation doses (5 or 6 Gy) led to accelerated onset of puberty in 20% of infantile irradiated rats animals as determined by vaginal opening. Interestingly, at peripubertal age (postnatal day 32-34), 5 or 6 Gy infantile irradiated rats had significantly higher serum LH levels stimulated by GnRH and estradiol levels (p < 0.05). 2 x 9 Gy irradiated rats had at the age of 3 mo a marked growth retardation and significantly lower GH levels than the controls (p < 0.05) whereas prolactin, FSH, TSH, T4, and corticosterone levels were comparable with controls. These studies demonstrate that the GnRH-pulse generator is very radiosensitive as precocious activation occurred after low dose irradiation (5 or 6 Gy) of infantile rats without any other endocrine disorder. High radiation doses (9 or 2 x 9 Gy) induced retardation of sexual maturation and later on growth hormone deficiency. Moreover this model of cranial irradiation seems to be suitable to study the molecular mechanisms of radiation induced pubertal changes.
Pediatric Research 06/2000; 47(5):586-91. DOI:10.1203/00006450-200005000-00005 · 2.31 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: There is increasing evidence that in the rat prior to and during the preovulatory LH surge, release rates of GABA in the preoptic area (POA) are decreased while no such changes occurred in the mediobasal hypothalamus (MBH). In addition, GnRH release appears to be facilitated by an increased preoptic excitation of glutamate (GLU). To investigate whether such changes of secretory activity of intrahypothalamic GABA or GLU neurons are associated with altered gene expression of biosynthetic enzymes or transporter proteins characteristic for either neuronal system, we determined mRNA levels of the two forms of the GABA-synthesizing enzyme glutamate decarboxylase (GAD65 and GAD67), the glutamate-synthesizing enzyme glutaminase (GLS), the GABA transporter type 1 (GAT-1) and the glutamate-aspartate transporter type 1 (GLAST). Competitive RT-PCRs using mutant cRNAs as internal standards were conducted with mRNA extracted from microdissected tissue of POA and MBH from diestrous, proestrous, and estrous rats. Proestrous animals were subgrouped according to their endocrine status as follows: 'prior to', on the 'ascending' or on the 'descending' limb of the LH peak, and 'after the LH surge (post)'. During the preovulatory LH surge, mRNA concentrations of GAD67 and GAT-1 in the POA were significantly increased compared to those observed on diestrous (2.8-fold for GAD67 and 2.5-fold for GAT-1, p < 0.01), while in the MBH the amount of both mRNAs remained constant. The expression levels of GAD65, GLS and GLAST were without any changes in the POA as well as in the MBH. These findings support the hypothesis that in rats induction of the preovulatory LH surge is controlled at the level of GnRH perikarya, and suggest that altered activities of intrapreoptic GABA neurons at both transcriptional and secretory levels are pivotal for the preovulatory activation of GnRH neurons.
[Show abstract][Hide abstract] ABSTRACT: Gonadotropin releasing-hormone (GnRH) analogues contain amino acid substitutions of the native decapeptide. Depending on the substitutions, the analogues have GnRH agonistic or antagonistic properties. GnRH agonists are the established treatment in cases of central precocious puberty caused by premature activation of the hypothalamic GnRH pulse generator. Much less data exist on the use of GnRH antagonists to influence the onset of puberty. Using the GnRH antagonist cetrorelix we conducted a 5 day treatment of peripubertal male rats (cetrorelix group n=12, 100 microg/d intraperitoneally injected; placebo n=10, NaCl 0.9% intraperitoneally injected) from postnatal day 32 to 36 and decapitated on postnatal day 37 to investigate the effects on pubertal development, serum gonadotropin and testosterone levels as well as the GnRH release from explanted hypothalami. A control group of 5 male rats was added for hypothalamus superfusion experiments on day 25. We observed no progress of testicular development in the cetrorelix group. Cetrorelix injected rats had lower testicular weights (531+/-13 versus controls 819+/-25 mg, mean+/-SEM, p<0.0001). 12 h after the last injection testosterone levels were in the castrate range (serum testosterone, median controls: 1.7 ng/ml, median cetrorelix <0.30 ng/ml, p<0.001), and they showed lower serum LH and FSH compared to the same age placebo group. After decapitation the preoptic mediobasal hypothalamic area (POA/MBH) was dissected from 5 randomly selected rats from each treatment group and the release rates of GnRH were determined in superfusion experiments: The hypothalamic GnRH secretion was comparable in the CET and the same age placebo rats but significantly higher than in the 25 day old control group. Conclusion: The GnRH antagonist cetrorelix inhibits the pituitary-gonadal axis in peripubertal male rats and may be effective in treating central precocious puberty in males.
[Show abstract][Hide abstract] ABSTRACT: There is increasing evidence that in the rat prior to and during the preovulatory LH surge, release rates of GABA in the preoptic area (POA) are decreased while no such changes occurred in the mediobasal hypothalamus (MBH). In addition, GnRH release appears to be facilitated by an increased preoptic excitation of glutamate (GLU). To investigate whether such changes of secretory activity of intrahypothalamic GABA or GLU neurons are associated with altered gene expression of biosynthetic enzymes or transporter proteins characteristic for either neuronal system, we determined mRNA levels of the two forms of the GABA-synthesizing enzyme glutamate decarboxylase (GAD65 and GAD67), the glutamate-synthesizing enzyme glutaminase (GLS), the GABA transporter type 1 (GAT-1) and the glutamate-aspartate transporter type 1 (GLAST). Competitive RT-PCRs using mutant cRNAs as internal standards were conducted with mRNA extracted from microdissected tissue of POA and MBH from diestrous, proestrous, and estrous rats. Proestrous animals were subgrouped according to their endocrine status as follows: ‘prior to’, on the ‘ascending’ or on the ‘descending’ limb of the LH peak, and ‘after the LH surge (post)’. During the preovulatory LH surge, mRNA concentrations of GAD67 and GAT-1 in the POA were significantly increased compared to those observed on diestrous (2.8-fold for GAD67 and 2.5-fold for GAT-1, p < 0.01), while in the MBH the amount of both mRNAs remained constant. The expression levels of GAD65, GLS and GLAST were without any changes in the POA as well as in the MBH. These findings support the hypothesis that in rats induction of the preovulatory LH surge is controlled at the level of GnRH perikarya, and suggest that altered activities of intrapreoptic GABA neurons at both transcriptional and secretory levels are pivotal for the preovulatory activation of GnRH neurons.
[Show abstract][Hide abstract] ABSTRACT: In rats, fasting or restriction of feed intake impairs the activity of the hypothalamic gonadotropin-releasing hormone (GnRH) pulse generator which results in reduced luteinizing hormone (LH) secretion. It is still unknown which neurotransmitters are involved in this phenomenon. However, it is known that increased GABA concentrations in the hypothalamus reduce GnRH biosynthesis and release. Therefore, we examined whether 17 days of feed restriction in male rats affected the hypothalamic gene expression of GnRH and the GABA-synthesizing enzymes glutaminase (GLS) and glutamic acid decarboxylase-which exists in two forms, GAD67 and GAD65-in the mammalian brain. Furthermore, the expression of the GnRH receptor (GnRH-R) and the GABA transporter 1 (GAT-1) were investigated. Feed restriction resulted in a 75% reduction in body weight (b.w.) compared to rats fed ad libitum. Serum concentrations of LH and testosterone in the feed restricted group were significantly reduced to approximately 15% of that of rats fed ad libitum, while the FSH concentration remained unchanged. In the mediobasal hypothalamus (MBH) where GnRH is released into the portal vessels, mRNA levels of GAD67 and GLS were increased twofold compared to rats fed ad libitum while no changes were observed in the preoptic area of the hypothalamus (POA) where GnRH is biosynthesised. Neither the expression of preoptic GnRH mRNA nor the expression of GAD65 and of GnRH-R mRNA in both hypothalamic structures was affected by feed restriction. In the anterior pituitary, a significant reduction of the expression of GnRH-R, LH-beta and the alpha subunit was observed in the feed restricted rats, whereas FSH-beta mRNA levels remained constant. Thus, feed restriction selectively increased the expression of GABA-synthesizing enzymes in the MBH but did not modify GnRH expression in the POA. However, the reduced expression of the LH-beta- and alpha-subunit and of the GnRH-R in the anterior pituitary indicates that pulsatile GnRH release may have been attenuated or even abolished. We suggest, that enhanced expression of GABA-synthesizing enzymes reflects increased GABAergic neurotransmission and thereby reducing GnRH release from the MBH.
Journal of Neuroendocrinology 09/1999; 11(8):613-9. · 3.14 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Since structural luteolysis involves deterioration of tissue, the gene expression of matrix-metalloproteinase-1 (MMP-1) and the respective tissue inhibitor of this metalloprotease (TIMP-1) were measured at various times on the day of pro-oestrus and in animals in which the preovulatory prolactin surge was blocked for the duration of 3 cycles by bromocriptine. An additional group of prolactin-blocked rats received a prolactin replacement injection on the afternoon of pro-oestrus. In spontaneously pro-oestrous rats, MMP-1 and TIMP-1 gene expression increased significantly (P<0.01) prior to the occurrence of the preovulatory LH surge but simultaneously with the onset of the preovulatory prolactin surge. When prolactin release was blocked by bromocriptine for 3 cycles, no such changes were observed during the afternoon of pro-oestrus. However, an intraperitoneal injection of bovine prolactin at the time when the preovulatory prolactin surge occurs normally, increased MMP-1 and TIMP-1 gene expression (P<0.01). These results indicate that MMP-1 and TIMP-1 gene expression are stimulated by the preovulatory prolactin surge. Previous work has shown that the preovulatory LH surge activates the enzymatic cascade which leads to increased collagenase activity.
European Journal of Endocrinology 06/1999; 140(6):583-9. DOI:10.1530/eje.0.1400583 · 4.07 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: During aging in female rats the estrous cycle ceases and the animals develop phases of constant estrous (CE) or constant diestrous (CD) prior to the irreversible transition into anestrous. In young rats, gamma-aminobutyric acid (GABA) is of pivotal importance for the release of GnRH. In the medial-preoptic area (MPO) where the majority of the GnRH perikarya are located in the rat, GABA release decreases at the time of the preovulatory LH surge. The suprachiasmatic nucleus (SCN) contains numerous GABA neurons. Neurochemical signals from this hypothalamic nucleus provide temporal information to GnRH neurons and thereby influence the preovulatory LH surge and the length of estrous cycles. To investigate aging-related changes of the activity of hypothalamic GABAergic neurons, we determined GABA turnover rates in various hypothalamic nuclei of CE and CD rats and compared them to those determined in young estrous (E) or diestrous rats (D1). In old female rats, GABA activity in the MPO was significantly decreased compared to E and D1 rats. A selective increase of GABA turnover rates was observed in the SCN of CE animals. No age-related changes were observed in the other examined brain areas. These data provide the first evidence for alterations in GABAergic activity in specific hypothalamic areas that depend on age and reproductive status. These may cause changes in ability to induce preovulatory LH surges and to maintain regular estrous cyclicity.
[Show abstract][Hide abstract] ABSTRACT: In summary, PACAP actions on gonadotrope cells are schematically outlined in FIGURE 5. PACAP is produced by and secreted from gonadotropes. The peptide increases LH release in an autocrine manner by a direct stimulatory action via the PACAP receptor and, as suggested by Culler and Paschall, by amplification of the LHRH stimulus. Hence, our data support the hypothesis that PACAP acts as an autocrine regulator on gonadotrope cell function.
Annals of the New York Academy of Sciences 01/1999; 865:486-91. · 4.38 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Amino acid neurotransmitters like gamma-aminobutyric acid (GABA) and glutamate (GLU) are involved in the regulation of hypothalamic gonadotropin releasing hormone (GnRH) release. We investigated, whether there are changes of gene expression in the rat hypothalamus for GnRH, GnRH receptor, as well as glutaminase and glutamate decarboxylase, two enzymes regulating neurotransmitter concentrations of GLU and GABA in the brain during the ontogeny. After reverse transcription-polymerase chain reaction (RT-PCR) we used an ELISA method to quantify PCR products. In 15-day old animals high plasma luteinizing hormone (LH) levels with pronounced variations were found. In 25-day old animals LH values were low, whereas in 35-day old rats LH levels increased significantly indicating the reactivation of the GnRH-pulse generator at the beginning of puberty. In parallel to these changes, the mRNA levels of the GnRH receptor in the mediobasal hypothalamus were high at day 15, significantly lower at day 25 and again high at day 35 after birth (ELISA O.D. GnRH-R day 15: 0.46+/-0.07, day 25: 0.16+/-0.04, day 35: 0.36+/-0.04; p<0.01), but no changes of GnRH receptor gene expression were found in the preoptic area. The mRNA of GnRH in the preoptic area as well as mRNA levels of glutaminase and glutamate decarboxylase in the mediobasal hypothalamus and the preoptic area did not change during ontogeny. We conclude that hypothalamic GnRH receptors are involved in the characteristic changes of LH secretion patterns during sexual maturation. Major changes of GnRH receptor gene expression occurred in the mediobasal hypothalamus and correlated well with plasma LH levels, whereas hypothalamic mRNA levels of GnRH, glutaminase and glutamate decarboxylase did not change within the different age groups. Thus the activity of the GABA- and glutamatergic system during ontogeny may be regulated at the receptor or postreceptor level.
Developmental Brain Research 09/1998; 110(1):105-14. DOI:10.1016/S0165-3806(98)00102-3 · 1.78 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: There is ample evidence that prostaglandin F2alpha (PGF2alpha) is a luteolytic substance in sows, however, there is also some evidence that it may stimulate progesterone (P4) secretion in young corpora lutea (CL). In vitro studies also suggested that tumor necrosis factor alpha (TNF) is inhibitory to luteal cell P4 and estradiol-17beta (E2) release. Since E2 is a strong luteotropic substance in porcine CL, we studied the effects of intraluteal application of PGF2alpha and TNF alone and in combination on the secretion of P4 and E2 in freely moving sows. Furthermore, the effects of intraluteal infusion of E2 and its stereoisomer, estradiol-17alpha, on luteal function, were also determined. Microdialysis systems were implanted into CL at Day 10 of the estrous cycle. After a 24-h recovery period, PGF2alpha (10(-6) M) or E2 (10(-6) M) was applied daily for 6 h into the CL. PGF2alpha caused a stimulation of E2 and P4, and E2 also stimulated P4 secretion at Days 11 and 12, but the stimulatory effect of both substances diminished as the CL approached luteolysis. Intraluteal TNF application resulted in a transient increase of P4 secretion, which was followed by a dramatic reduction of P4 release. When TNF-pretreated CL were exposed to PGF2alpha at Day 11 of the estrous cycle, the prostaglandin was no longer able to stimulate but rather inhibited E2 and P4 secretion. Intraluteal application of estradiol-17alpha had no effect on P4 secretion. These results are suggestive that the PGF2alpha-induced E2 secretion in young and middle-aged CL is stimulatory to P4 secretion. Under the influence of macrophage-derived TNF production, E2 secretion is inhibited, and thereby PGF2alpha and TNF cause functional luteolysis.
Biology of Reproduction 06/1998; 58(5):1310-5. · 3.32 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: In many species the GnRH pulse generator functions early postnatally to become arrested during infancy. In rats highly variable LH levels in 15-day-old animals are suggestive that LH is being released by the pituitary in pulses whereas between day 20 after birth and puberty LH levels are low indicating that the GnRH pulse generator is arrested. In the present study we show on the basis of consecutively withdrawn blood samples in 15-day-old animals that LH pulses are indeed present at that age. The proper function of GnRH receptors in the pituitary is crucially dependent on pulsatile GnRH release from the hypothalamus. In addition, GnRH receptors have been demonstrated in the medial preoptic area and in the mediobasal hypothalamus of adult rats. In 15-day-old animals the functional GnRH pulse generator results in upregulated GnRH receptor gene expression as demonstrated by quantitative RT-PCR. It is not known what neural mechanisms are involved in turning the GnRH pulse generator off during infancy and a GABAergic brake has been discussed. Indeed, when 30-day-old animals were injected with the GABA-A receptor blocking drug bicuculline, this resulted in increased serum LH levels indicating that a tonic GABAergic inhibition is indeed operative at this age.
[Show abstract][Hide abstract] ABSTRACT: The mediobasal hypothalamus of rats contains gonadotropin-releasing hormone (GnRH) receptors. These hypothalamic neurons also express the GnRH corresponding gene. Under these circumstances, the possibility exists that these GnRH receptors could be localized in other neurons, which are GnRH-receptive, unknowing the neurotransmitter quality. Therefore, we studied the in vitro effects of the GnRH agonist buserelin on GnRH, glutamate, gamma-amino-butyric acid (GABA) and taurine release from explanted superfused hypothalami of untreated and buserelin-pretreated (down-regulated) male rats. When buserelin was added to the superfusion medium it inhibited promptly the release of GnRH and the excitatory amino acid neurotransmitter glutamate, but stimulated the release of the inhibitory neurotransmitters, GABA and taurine. Hypothalamic release of GnRH from hypothalami collected from buserelin-treated (30 micrograms/100 g b.w. twice daily for 4 days) male rats released significantly less GnRH, glutamate and more GABA and taurine. The inhibitory effect of buserelin was maintained when the superfusion medium continuously contained the GnRH analog. When superfusion of hypothalami from buserelin-pretreated animals was performed in the absence of buserelin, GnRH and glutamate release increased significantly within 45-60 min, whereas GABA and taurine release decreased at this time point. When buserelin was added to the superfusion medium 2 h after buserelin-free superfusion, GnRH and glutamate release decreased whereas GABA and taurine release increased instantaneously. Buserelin-treated rats showed significantly low values of LH and testosterone than the untreated rats. These results suggest that GnRH receptors may not only be present in GnRH axon terminals in the median eminence, but also on glutamatergic, GABAergic and taurinergic neurons by which GnRH may exert an autoinhibitory ultrashort loop feedback on its own secretion. This effect appears to be connected with glutamatergic, GABAergic and taurinergic neurons.
[Show abstract][Hide abstract] ABSTRACT: In order to evaluate the possible participation of the hypothalamic excitatory and inhibitory amino acid neurotransmitter systems in the GnRH release response to GABAergic drugs, hypothalami (preoptic and mediobasal area) of immature (26 days of age) and adult male rats were perifused with GABA-A and -B agonists and antagonists. GnRH and amino acid neurotransmitter concentrations (glutamate, taurine, GABA) were measured in perfusate samples collected every 15 min during 150 min. In immature rats, muscimol and baclofen (GABA-A and GABA-B agonists, respectively) increased GnRH, glutamate and GABA release and decreased taurine output, while in adults these agonists showed opposite effects on GnRH and glutamate release, and increased GABA and taurine output. On the other hand, in immature rats bicuculline and phaclofen (GABA-A and GABA-B antagonists, respectively) decreased GnRH, glutamate and GABA release, increasing taurine outflow. In adult animals, these antagonists enhanced GnRH and glutamate release, decreasing taurine and GABA outflow. These results indicate that GABA stimulates GnRH release in immature male rats and confirm the inhibitory role of this amino acid neurotransmitter in adult animals. This effect might be associated, at least partially, with the modifications observed in the excitatory and inhibitory amino acid release. On the other hand, in immature rats, stimulation of GABA-A and GABA-B receptors increased GABA release. Although ultrastructural studies have not produced any evidence of GABA-GABA neurointeractions, our results suggest the existence of a positive feedback mechanism of GABA autoregulation active during the prepubertal stage. Participation of this mechanism in the onset of puberty cannot be discarded.
[Show abstract][Hide abstract] ABSTRACT: Alterations in the immune system are often accompanied by reproductive disorders. The bacterial endotoxin lipopolysaccharide (LPS) has central inflammatory effects, and activates the release of cytokines (immune system mediatory factors) in the hypothalamus, where the LHRH neurons are located. Therefore, these studies were designed to investigate a possible effect of the LPS and LHRH secretion and the possible interaction with excitatory and inhibitory amino acids. Animals were decapitated at 09.00 h, and the preoptic mediobasal hypothalamic area (PO/MBH) dissected and superfused. Superfusate fractions were collected at 15-min intervals. After a stabilization superfusion period of 60 min four fractions were collected; LPS (100 ng/ml) was then added to the superfusion medium for the duration of 1 h. This was followed by a washout period of 60 min. The PO/MBH fragments were then subjected to a 56 mM K+ stimulus. Control PO/MBH fragments were continuously superfused with Earle's balanced salt solution. LHRH release was significantly (p < 0.05) reduced during and following exposure to LPS. At this time GABA and taurine concentrations increased in the superfusion medium, while glutamate decreased significantly compared with the control group. These observations indicate that LPS inhibits LHRH and glutamate release, but stimulates taurine and GABA secretion. These effects may be explained by the stimulation of cytokines of neuronal and/or glial source which may interact with the excitatory and inhibitory amino acids.
[Show abstract][Hide abstract] ABSTRACT: We review the crucial role of the two neurotransmitters norepinephrine (NE) and GABA in eliciting GnRH pulse. NE acts via an alpha l-receptor mechanism and also GABA acts at the alpha-subtype of the GABA receptor. The function of NE appears to be induction of phasic activation of GnRH neurons and GABA inhibits GnRH neurons tonically until they are all ready for phasic activation. By an unknown mechanism preoptic GABA release in dramatically reduced which causes simultaneous desinhibition of the GnRH neurons. Hence they release their product into the portal vessels simultaneously which is the appropriate signal for the pituitary ganodotrophs. The action of norepinephrine and GABA is most likely exerted at the perikarya level of the GnRH neurons since the alpha l-adreno receptor blocker doxazosin and GABA inhibit GnRH secretion only when applied into the medial preoptic/anterior hypothalamic area (where in the rat brain the GnRH perikarya are located). Utilizing a quantitative reverse transcription polymerase chain reaction, we demonstrate furthermore that GnRH receptors are present in the mediobasal hypothalamus as well as in the preoptic area of rats. Their function appears to serve autoinhibitory purposes since Buserelin added to medium significantly decreased GnRH release. Simultaneously, the release of GABA was increased and that of glutamate decreased. We conclude from these experiments that GABAergic and glutamatergic neurons in the hypothalamus may also be GnRH-receptive.