Biomedical subjects

C Libertun

Publications and source records attributed to C Libertun.

At least 19 recordsLinked to original sources

PubMed2006

Experimental data supporting the expression of the highly conserved GnRH-II in the brain and pituitary gland of rats.

The second GnRH form, originally identified in chickens (cGnRH-II or GnRH-II), is the most ubiquitous peptide of the GnRH neuropeptide family, being present from jawed fish to human beings. However, the presence of GnRH-II in such an important experimental model as the rat is still an object of discussion. Here we present chromatographic, immunologic and biologic activity evidence supporting the expression of GnRH-II in the rat. Olfactory bulb, hypothalamus, remnant brain and anterior pituitary from a pool of 50 female adult rats were extracted and subjected to RP-HPLC on a C-18 column. The fractions were collected and evaluated by using two different RIA systems, specific for GnRH-I and GnRH-II respectively. Under these conditions the GnRH-I standard eluted in fraction 21 (f21) was only detected with the GnRH-I RIA system, whereas the GnRH-II standard was only detected in the fraction 27 (f27) by using a GnRH-II RIA system. In the olfactory bulbs extract, the fractions analyzed by the GnRH-I RIA systems showed a single peak in f21, whereas by using the GnRH-II RIA system a single peak at f27 was observed. In the hypothalamus GnRH-I was detected in f21 meanwhile GnRH-II could not be detected. When the remnant brain and pituitary gland extracts were analyzed, both GnRH forms were detected. To the best of our knowledge, this is the first report concerning GnRH-II detection in a mammalian pituitary. Serial dilutions of f27 and GnRH-II presented similar displacement of radioiodinated-GnRH-II, demonstrating that both molecules share immunological properties. Moreover, after 60 min stimulation, both f27 and GnRH-II had similar LH and FSH releasing activity in 12 day-old rat pituitary primary cell cultures. However, we failed to characterize the GnRH-II gene in this model. These results provide strong evidence for the expression of GnRH-II in the rat brain and pituitary gland.

Animals↗

PubMed2005

Differential gonadotropin releasing hormone (GnRH) expression, autoregulation and effects in two models of rat luteinized ovarian cells.

GnRH has been suggested to participate in corpus luteum function. Here we studied the expression of GnRH mRNA and peptide in two models of rat luteinized tissues: ovarian cells from PMSG-hCG treated prepubertal rats (SPO) and from intrasplenic ovarian tumors (Luteoma). A GnRH autoregulatory effect was evaluated as well as its action on cell proliferation and apoptosis. GnRH mRNA was present in SPO, isolated corpora lutea from SPO and Luteoma from 1 week to 7 months of development. In vitro cultures of Luteoma cells expressed 2-fold higher GnRH mRNA and 10-fold higher GnRH peptide than SPO cells. Buserelin (GnRH analog) increased GnRH mRNA and peptide expression in SPO but not in Luteoma cells. While basal proliferation was very low in Luteoma cells, SPO cells showed a significant increase in cell number by both the thymidine and the MTS methods after 72 h in culture. Buserelin induced a decrease in cell number in both cell types to a similar degree. Although basal apoptosis levels were higher in SPO than in Luteoma cells, Buserelin-induced apoptosis was only detected in Luteoma cells after 48 h treatment. These results show that the two types of rat, luteinized tissues, Luteoma and SPO, markedly differed in some intrinsic properties and in their local GnRH systems. Luteoma cells proliferate very weakly, express and secrete high amounts of GnRH, do not show an autoregulatory effect and respond to the decapeptide with apoptosis stimulation. In contrast SPO cells proliferate significantly, secrete low levels of GnRH but possess a positive, autoregulatory mechanism and respond to GnRH stimulation with impairment of proliferation.

Animals↗

PubMed2002

Actions of immunosuppressor drugs on the development of an experimental ovarian tumor.

Immunosuppression has been related to the incidence of tumor apparition, including endocrine tumors. The intrasplenic ovarian tumor (luteoma) is a typical benign endocrine tumor that develops under high gonadotropin stimulation and, from the immunological perspective, is located in a critical organ involved in immune response. To establish if immunosuppression could alter the development of this experimental tumor, the effects of cyclosporin A (CsA) and dexamethasone (Dex) were evaluated. After surgery, tumor-bearing and sham animals were kept without treatment for 4 weeks; thereafter, they were distributed into CsA (25 mg/kg), Dex (0.1 mg/kg), or vehicle (75:25 castor oil:ethanol) groups and were injected on alternate days for 50 days. Body weight was evaluated weekly. Animals were sacrificed after a jugular vein blood sample was obtained. Thymi were weighed. Tumors were measured and placed in formaline for histological studies. Serum luteinizing hormone (LH), follicle-stimulating hormone (FSH), prolactin (PRL), and estradiol were measured by radioimmunoassay. Hematological parameters were determined. CsA induced a significant decrease in survival rates both in tumor-bearing and sham animals (P < 0.01). Dex significantly impaired weight increase in both groups of animals. CsA induced a significant weight loss in sham animals, not observed in tumor-bearing animals. Dex induced thymus weight loss in both groups, whereas CsA induced thymus weight loss only in sham animals. Only Dex induced a decrease in lymphocyte number in both groups. CsA induced an increase in monocyte number only in sham animals. Treatments did not alter LH, FSH, or estradiol, whereas PRL was increased by CsA only in sham rats. Neither Dex nor CsA induced any significant variations in tumor volume, nor did they alter tumor histology. In addition, no visible metastases or alterations in other organs were observed. We conclude that, though immunological parameters were altered by the treatments, immunosuppressor drugs did not condition tumor development. In addition, tumors secrete one or more factor/s that counteract CsA effect.

Animals↗

PubMed2002

Disruption of the D2 dopamine receptor alters GH and IGF-I secretion and causes dwarfism in male mice.

We determined the consequences of the loss of D2 receptors (D2R) on the GH-IGF-I axis using mice deficient in functional dopamine D2 receptors by targeted mutagenesis (D2R(-/-)). Body weights were similar at birth, but somatic growth was less in male D2R(-/-) mice from 1-8 months of age and in D2R(-/-) females during the first 2 months. The rate of skeletal maturation, as indexed by femur length, and the weight of the liver and white adipose tissue were decreased in knockout male mice even though food intake was not altered. The serum GH concentration was significantly decreased during the first 2 months in knockout female and male mice, and IGF-I and IGF-binding protein-3 levels were lower in knockout mice. PRL was significantly higher in knockout mice, and females attained higher levels than males. Pituitaries from adult knockout mice had impaired basal GH release and a lower response to GHRH in vitro. We propose that the D2R participates in GHRH/GH release in the first month of life. In accordance, the D2R antagonist sulpiride lowered GH levels in 1-month-old wild-type mice. Our results indicate that lack of D2R alters the GHRH-GH-IGF-I axis, and impairs body growth and the somatotrope population.

Animals↗

PubMed2002

Development of an experimental ovarian tumor: immunocytochemical analysis.

OBJECTIVE: The aim of the present work was to study whether immunocytochemical parameters present in the normal ovary were altered after tumor development under high gonadotropin levels. METHODS: Ovarian tumors (luteoma): castrated female rats had an ovary grafted into the spleen; tumors were left to develop for 1, 2, 3 or 7 months. The presence of apoptotic cells (TUNEL method) and the expression of proliferating cell nuclear antigen (PCNA), gap junction protein (Cx43), steroidogenic acute regulatory protein (StAR), aromatase and synaptosome-associated protein of 25 kDa (SNAP-25) were determined by immunocytochemistry. Some of these findings were confirmed by RT-PCR (Cx43, StAR, SNAP-25). Inhibin subunit mRNAs were investigated by Northern blot. RESULTS: PCNA staining of tumors was mainly found in granulosa cells of transforming follicles and was absent from luteinized follicles. A nearly complete absence of apoptosis was observed. Cx43 was mainly found in follicles, while it was very weakly expressed or absent in luteinized follicles. StAR protein expression, indicating active steroidogenesis, was demonstrated only in luteinized follicles and in thecal cells, but was absent from granulosa cells. Aromatase immunoreactivity was very intense in granulosa and also present in luteal cells. Membrane-associated and cytoplasmic SNAP-25 immunostaining was determined in patches of endocrine cells in the follicles, as well as in the luteinized follicles. The expression of mRNAs for Cx43, StAR and SNAP-25 (RT-PCR) and inhibin subunits (Northern blots) were confirmed in 1-, 3- and 7-month-old tumors. CONCLUSIONS: These results indicated that luteoma most likely develop from unruptured follicles by hypertrophy and proliferation of follicular cells. Circulating gonadotropins seem to play a fundamental role in maintaining the expression of proteins typically expressed in normal ovary, while avoiding apoptosis in this tissue.

Animals↗

PubMed2001

Ontogenic expression of anterior pituitary GABA(B) receptor subunits.

gamma-Aminobutyric acid (GABA) is involved in the neuroendocrine control of hypophyseal secretion, acting both in the central nervous system and directly at the pituitary. We have characterized the properties of anterior pituitary GABA(B) receptors. In this work the ontogeny of rat anterior pituitary GABA(B) receptors and the pattern of subunit expression in rats of both sexes were determined. Western blot analysis showed a temporal decrease in GABA(B) subunits GABA(B(1a)) and GABA(B(1b)) expression in female anterior pituitary membranes from day 4 to adulthood, with GABA(B(1a)) being significantly more abundant than GABA(B(1b)) at early stages of development; the GABA(B(2)) subunit was barely detectable. In the male, GABA(B(1a)) followed a similar pattern and appeared to be significantly less abundant than in 4- and 12-day-old females; GABA(B(1b)) and GABA(B(2)) expression in the male was barely detectable. Scatchard plot analysis showed a temporal decrease in binding sites in female anterior pituitary membranes, in agreement with the western blot results. The number of binding sites was significantly higher in female than in male 4-day-old membranes. Dissociation constant values were similar for both sexes at all ages studied. This study reports for the first time the ontogeny of anterior pituitary GABA(B) receptors, showing a particular developmental pattern of subunit expression and a clear sexual dimorphism.

Animals↗

PubMed2001

GABA(B) receptors in anterior pituitary cells. Mechanism of action coupled to endocrine effects.

The activation of pituitary GABA(B) receptors by the specific agonist baclofen inhibits pituitary hormone secretion in vitro. Here we studied the mechanism of action of GABA(B) receptors in rat adenohypophysis. Anterior pituitary cells were obtained by trypsinization and were either plated for hormonal studies and cAMP determination or incubated in FURA 2AM for calcium measurements. Baclofen (BACL: 1 x 10(-5) M) significantly inhibited basal and thyrotropic releasing hormone (TRH)-stimulated (1 x 10(-7) M) PRL secretion in anterior pituitary cells from proestrous rats. In the presence of pertussis toxin (PTX: 150 ng/ml, 20 h), which leads to the uncoupling of the G(i/o)-protein from the receptor, both effects of BACL were abolished while the effect of dopamine (DA: 1 x 10(-8) M), used as an inhibitory control, was reduced from 70 to 25%. PTX also reversed BACL-induced inhibition of gonadotropin-releasing hormone (GnRH)-elicited luteinizing hormone (LH) secretion in anterior pituitary cells from 15-day-old female rats. In addition, though working in a pituitary mixed cell population, in which only some cell types possess GABA(B) receptors, BACL (1 x 10(-5) M) attenuated the forskolin-induced (0.5 microM) increase in cAMP. This effect was prevented by co-incubation with the antagonist 2 hydroxysaclofen and by preincubation with PTX. BACL (5 x 10(-5) M) and DA (5 x 10(-7) M) inhibited basal intracellular calcium concentrations ([Ca(2+)](i)) in pituitary cells and the effect of the latter was significantly stronger. The effect of BACL on [Ca(2+)](i) was abolished after preincubation with PTX. In the presence of the potassium channel blocking agents barium (200 microM and 1 mM) and tetraethylammonium (10 mM), BACL was still able to inhibit [Ca(2+)](i). Blockade of voltage-sensitive calcium channels (VSCC) with either verapamil (5 x 10(-6) M) or nifedipine (1 x 10(-6) M) completely abolished the effect of BACL on [Ca(2+)](i). In the presence of 12.5 mM potassium concentration baclofen significantly inhibited [Ca(2+)](i). In conclusion, our results describe the negative coupling of adenohypophyseal GABA(B) receptors to VSCC through PTX-sensitive G-proteins. These characteristics suggest a resemblance of these receptors to the typical presynaptic GABA(B) sites described in the central nervous system.

Animals↗

PubMed2001

Structure and biological activity of gonadotropin-releasing hormone isoforms isolated from rat and hamster brains.

Rat and hamster brain tissues were used to investigate the possible existence of a follicle stimulating hormone (FSH)-releasing factor with similar characteristics to the lamprey gonadotropin-releasing hormone III (lGnRH-III) form proposed in previous reports. The present studies involved isolation and purification of the molecule by high-performance liquid chromatography (HPLC), identification by radioimmunoassay, sequence analysis by automated Edman degradation, mass spectrometry and examination of biological activity. Hypothalamic extracts from both species contained an HPLC fraction that was immunoreactive to GnRH and coeluted with lGnRH-III and 9-hydroxyproline mGnRH ([Hyp(9)]GnRH). Determination of primary structure from purified total brain material demonstrated that the isolated molecule was [Hyp(9)]GnRH. This is the first report showing the presence of the posttranslationally modified form already known as [Hyp(9)]GnRH by primary sequence analysis. The biological activity of distinct GnRH peptides was also tested in vitro for gonadotropin release using rat pituitary primary cell cultures. The results showed that [Hyp(9)]GnRH stimulated both luteinizing hormone and FSH release, as already reported, whereas lGnRH-III had no action on the secretion of either gonadotropin.

Amino Acid Sequence↗

PubMed2000

Bromocriptine restores angiotensin II response in pituitary hyperplasia.

In estrogen-induced pituitary hyperplasia AII-evoked prolactin release is decreased and the octapeptide does not generate a spike elevation in [Ca(2+)](i) in vitro. We studied whether or not bromocriptine could restore AII response in diethylstilbestrol treated rats. Co-administration of bromocriptine resulted in involution of pituitary size and lowering of serum prolactin. In vitro, prolactin release per cell was reduced in the hyperplastic group, and levels were not significantly increased by in vivo bromocriptine treatment. Immunocytochemical analysis revealed that hyperplastic pituitaries contained fewer prolactin granules than control pituitaries, and that bromocriptine, did not increase prolactin storage. Nevertheless, in this group, prolactin response to AII increased, and AII evoked a consistent spike in [Ca(2+)](i), albeit lower than in the control group. Such spike was abolished by thapsigargin, and not by removal of extracellular calcium or by K(+), indicating that it was mainly dependent on intracellular calcium stores, as in normal cells. We conclude that bromocriptine treatment partially restores AII response in the hyperplastic pituitary.

Angiotensin II↗

PubMed2000

Endocrine studies in ivermectin-treated heifers from birth to puberty.

Continuous treatment with ivermectin from birth to puberty advanced sexual maturation by 3.7 wk in Holstein heifers grazing pastures naturally infected with nematodes. Every 14 d jugular blood samples were taken from birth to 45 wk of age from all heifers. No differences in serum FSH, estradiol, or thyroxine levels were observed during the trial between the treated and untreated group. Mean LH levels were diminished in untreated heifers 4 wk before the first estrus and the amplitude of LH pulses was augmented in treated heifers when puberty was reached. Serum IGF-I levels increased from birth to 22 wk of age and then reached a plateau in both groups, but levels were consistently higher in treated heifers from 26 wk of age onward. Body weight gain was retarded in parasitized heifers and IGF-I values were positively correlated with body weight only during the first 20 wk of life. We suggest that enhanced prepubertal IGF-I levels in conjunction with increased prepubertal LH levels and pubertal LH pulse amplitude might be involved in the accelerated somatic maturation and in puberty advancement observed in ivermectin-treated heifers.

Animals↗

PubMed1999

Development of an experimental ovarian tumor over a year in the rat.

Tumor growth, possible malignant transformation or metastatic propagation and hormonal patterns were evaluated over a year in luteoma induced by introducing an ovary into the spleen of ovariectomized 60 day-old rats. Sham castrated animals had a piece of muscle inserted into the spleen. Jugular blood samples were taken monthly. After a year animals were cycled and decapitated. Troncal blood was collected, autopsies were performed and luteoma were measured and fixed in 10% buffered formalin. Serum LH, FSH, PRL, estradiol and progesterone were measured. Serum inhibin content was determined in one month-old tumors-bearing animals and estrous rats as controls. After one year no external changes in tumor-bearing rats were observed, nor differences in body weight or mortality rates compared to Sham animals. Metastatic propagation was absent. Routine histological examination showed two types of tumors according to either granulosa or luteal cell predomination, tumor type did not determine hormonal patterns. However, a clear relationship between gonadotropin levels and tumor size was established. Low gonadotropins: Small tumors, 18.7% of cases and high gonadotropins: Large tumors, 81.3%. In Sham animals gonadotropins attained castrate levels and remained elevated until the end of the experiment. In the Small group no increases in gonadotropins or estradiol were detected, progesterone and PRL fluctuated. In the Large tumor group LH increased to Sham titers until month 7, then fell to initial levels, FSH augmented significantly as from month three and remained high up to month 5. No variations in either estradiol, progesterone or PRL were observed. Serum inhibin of one month-old tumor-bearing rats was significantly elevated, justifying the lack of FSH increase at this time point. We conclude that these luteoma do not suffer malignant transformation or induce metastases. They appear in two histological types. Tumor size depends on hormonal patterns. The delay in the initial increase and the sharp decrease observed in FSH in animals bearing Large tumors suggest a possible role for inhibin in this regulation.

Animals↗

PubMed1999

Calcium influx and intracellular stores in angiotensin II stimulation of normal and hyperplastic pituitary cells.

In rat pituitary cells from estrogen-induced hyperplasia, angiotensin II (ANG II) does not evoke a clear spike elevation of intracellular Ca2+ concentration ([Ca2+]i) but induces a plateau increase. The present work was undertaken to establish whether this difference was related to a differential participation of intracellular and/or plasma membrane Ca2+ channels. We first tested the effect of 10 nM ANG II on [Ca2+]i in the absence of extracellular Ca2+ in cells depolarized with 25 mM K+ or in the presence of blockers of L-type voltage-sensitive Ca2+ channels (VSCC). These treatments did not alter spike elevation in [Ca2+]i in controls but reduced plateau levels in hyperplastic cells. Intracellular Ca2+ stores were similar in both groups, as assessed by thapsigargin treatment, but this drug abolished spike increase in controls and scarcely modified plateau levels in hyperplastic cells. Finally, inositol trisphosphate (InsP3) production in response to ANG II was significantly higher in control cells. We conclude that the observed plateau rise in hyperplastic cells results mainly from Ca2+ influx through VSCC. In contrast, in control cells, the ANG II-induced spike increase in [Ca2+]i results from mobilization of Ca2+ from thapsigargin-sensitive internal channels, activated by higher inositol 1,4,5-trisphosphate generation.

Angiotensin II↗

PubMed1999

Alterations in intracellular messengers mobilized by gonadotropin-releasing hormone in an experimental ovarian tumor.

Cells derived from an experimental luteinized ovarian tumor are more sensitive to GnRH endocrine action than control luteal cells. In an attempt to understand the possible causes of the differential sensibility to GnRH action, we examined the number and affinity of GnRH receptors and the second messenger response to GnRH stimulation in both tissues. For GnRH receptor studies membranes were obtained from 4- to 6-week-old ovarian tumors (luteoma) and ovaries from prepubertal rats treated with 25 IU PMSG and 25 IU hCG (SPO) and were incubated with [125I]Buserelin. The number of GnRH receptors were increased in luteoma compared with that in SPO ovaries; dissociation constants were similar in both tissues. GnRH stimulation of second messenger release was assessed in cells obtained from luteoma and SPO ovaries by collagenase treatment. Buserelin (100 ng/ml) induced a significant 35% calcium increase in SPO cells, as determined by the fura-2 method; in luteoma cells no response was observed after buserelin stimulation, although a calcium transient was induced by thapsigargin (0.5 microM), an inhibitor of Ca2+-adenosine triphosphatase associated with the endoplasmic reticulum. The effect of buserelin on inositol phosphates was evaluated after incubation of luteoma and SPO cells with [3H]myoinositol for 48 h. Buserelin induced a 400% increase in inositol trisphosphate in SPO cells. Again, luteoma cells did not respond to buserelin stimulation, although NaF (10 mM), an activator of G proteins coupled to phospholipase C, induced an 800% increase in inositol trisphosphate. Although the number of GnRH receptors is augmented in luteoma cells, justifying an increased endocrine response, neither inositol phosphates nor intracellular calcium were released by a GnRH analog, indicating the uncoupling of GnRH receptors from phospholipase C. These data provide evidence that the transformation of the ovary into a luteoma implies the acquisition of novel characteristics in the GnRH receptor second messenger-generating system.

Animals↗

PubMed1999

Effects of continuous ivermectin treatment from birth to puberty on growth and reproduction in dairy heifers.

The effect of continuous ivermectin treatment from birth to puberty on growth and reproductive performance was studied in Holstein heifer calves grown on pastures in comparison to naturally nematode-infected, untreated animals. Ivermectin effectively abated the presence of nematode eggs in feces. Eggs per gram (EPG) in parasitized animals increased rapidly from wk 12 to 18 of age and then decreased. Animals treated with ivermectin grew faster than untreated ones, and differences in body weight became significant at 6 wk of life, even before eggs appeared in the feces of either treatment group. Ivermectin-treated heifers reached puberty 3 wk earlier than infected ones as assessed with serum progesterone concentrations (ivermectin, 30.4 +/- .8 vs untreated, 33.7 +/- 1.3 wk of age). This delay was not directly related to body weight. In addition, pelvic area at 39 wk and at 15 mo of age was increased in treated heifers (8 and 11%, respectively) compared with parasitized animals. No differences in the wither heights were observed. We conclude that ivermectin treatment in dairy heifers may increase growth rate during development, advance the onset of ovarian function, and positively affect yearling pelvic area.

Animals↗

PubMed1998

Angiotensin II-induced Ca2+ mobilization and prolactin release in normal and hyperplastic pituitary cells.

We evaluated the effects of angiotensin II (ANG II) and its antagonists on prolactin release, intracellular calcium ([Ca2+]i) mobilization, and [3H]thymidine uptake in cells from normal rat pituitaries and from estrogen-induced pituitary tumors. ANG II (10(-7) to 10(-9) M) increased prolactin release significantly in control and not in tumoral cells. In control cells, ANG II (10(-6) to 10(-9) M) produced an immediate spike of [Ca2+]i followed by a plateau. Spike levels rose significantly between 10(-10) and 10(-8) M ANG II, whereas the onset of the spike was retarded with decreasing concentrations. In tumoral cells, ANG II did not produce a spike phase even at 10(-6) M. ANG II-induced prolactin release and calcium mobilization were blocked by losartan (AT1 receptor antagonist) and not by PD-123319 (AT2 antagonist). Finally, [3H]thymidine uptake was not modified by ANG II (10(-7) to 10(-10) M) or its antagonists in either group. Our results suggest that chronic in vivo estrogenic treatment alters in vitro pituitary response to ANG II. Alterations might function to limit excessive prolactin secretion of hypersecreting tumors. Besides, ANG II does not modify DNA synthesis in vitro of cells from normal or tumor-derived hypophyses.

Angiotensin II↗

PubMed1998

Ontogenic and sexual differences in pituitary GnRH receptors and intracellular Ca2+ mobilization induced by GnRH.

The present experiments were designed in order to elucidate the participation of the developing hypophysis in determining the changing sensitivity of gonadotrophins to gonadotropin-releasing hormone (GnRH) during ontogeny in the rat. To that end, we chose two well defined developmental ages that differ markedly in sexual and ontogenic characteristics of hypophyseal sensitivity to GnRH, 15 and 30 d. In order to study sex differences and the role of early sexual organization of the hypothalamus, experiments were carried out in males, females, and neonatally androgenized females (TP females). We evaluated (1) the characteristics of pituitary GnRH receptors, and (2) associated changes in GnRH-induced mobilization of intracellular Ca2+ (a second messenger involved in gonadotropins exocytosis). We measured binding characteristics of the GnRH analog D-Ser(TBu)6-des-Gly10-GnRH ethylamide in pituitary homogenates. We found that Kds did not vary among the different sex groups. Total number and concentration of receptors decreased in the female rat from 15-30 d of age, whereas in the male and TP female, receptors/pituitary increased, and the concentration/mg tissue did not change. Also, at 30 days of age, males presented higher content and concentration of receptors than females, and higher content than TP females. In order to evaluate if developmental and sexual differences in pituitary sensitivity to GnRH might be expressed through variations in the intracellular Ca2+ signal, we studied the mobilization of intracellular Ca2+ induced by GnRH (1 x 10(-8) to 1 x 10(-11) M) in a suspension of dispersed pituitary cells in the six groups. In cells from 15-d-old females, Ca2+ response was greater than in 30-d-old females at the doses of 10(-8) to 10(-10) M, indicating that in the infantile female rat activation of highly concentrated GnRH receptors is reflected in an increase in signal transduction mediated by Ca2+. In males and in female rats androgenized at birth, there was also a decrease in the magnitude of intracellular, Ca2+ mobilization induced by GnRH (10(-8) to 10(-10) M) from 15-30 d of age, even though the concentration of GnRH receptors did not change in the same period. In conclusion, the present results suggest that high sensitivity to GnRH, which has been described in the female infantile rat, may be related to elevated concentration of hypophyseal receptors coupled to an increase of intracellular calcium response to GnRH, both parameters decreasing as the rat matures. In males, the greater sensitivity that has been described for GnRH at 30 d in comparison to 15 d is paralleled by an increase in the total number of GnRH receptors per pituitary (and not in their concentration), but not in an increase in the magnitude of Ca2+ mobilization induced by GnRH. On the other hand, neonatal sexual organization of the hypothalamus is involved in the differential expression of GnRH receptors, but does not modulate mobilization of intracellular Ca2+ induced by the decapeptide.

Aging↗

PubMed1998

Effect of stage of development and sex on gonadotropin-releasing hormone secretion in in vitro hypothalamic perifusion.

Marked sexual and ontogenic differences have been described in gonadotropin regulation in the rat. These could arise from events occurring both at the hypothalamic or hypophyseal levels. The present experiments were designed to evaluate the capacity of the hypothalamus in releasing GnRH in vitro, basally and in response to depolarization with KCl, during ontogeny in the rat. To that end we chose two well-defined developmental ages that differ markedly in sexual and ontogenic characteristics of gonadotropin regulation, 15 and 30 days. We compared GnRH release from hypothalami of females, neonatal androgenized females and males. Mediobasal hypothalami were perifused in vitro, and GnRH measured in the effluent. Basal secretion of the decapeptide increased with age in the three groups with no sexual differences encountered. When studying GnRH release induced by membrane depolarization, no differences within sex or age were encountered. On the other hand FSH serum levels decreased with age in females and increased in males, and in neonatal androgenized females followed a similar pattern to that of females. LH levels were higher in infantile females than in age-matched males or androgenized females. Such patterns of gonadotropin release were therefore not correlated to either basal or K+-induced GnRH release from the hypothalamus. We conclude that sexual and ontogenic differences in gonadotropin secretion in the developing rat are not dependent on the intrinsic capability of the hypothalamus to release GnRH in response to membrane depolarization. The hormonal differences observed during development and between sexes are probably related to differences in the sensitivity of the GnRH neuron to specific secretagogue and neurotransmitter regulation, and/or to differences in hypophyseal GnRH receptors and gonadotrope sensitivity.

Aging↗

PubMed1997

GnRH receptors and GnRH endocrine effects on luteoma cells.

An ovary implanted into the spleen of an ovariectomized rat develops into a luteinized tumor, growing in response to gonadotrophins. Previously, it was shown that in vivo Buserelin, a gonadotrophin-releasing hormone (GnRH) analog, inhibited tumor growth. To determine if GnRH had a direct effect on tumor cells, the presence of GnRH receptors as well as the endocrine effects of buserelin were studied on tumoral tissue. GnRH receptors were present in luteoma in similar concentrations and dissociation constant (Kd) to control estrous ovaries. In vivo treatment with buserelin did not modify luteoma GnRH receptors. In organ incubations, luteoma secreted significantly higher estradiol and lower progesterone than estrous ovaries; addition of buserelin did not modify steroid secretion. The same difference in basal steroid secretion between luteoma cells and luteal cells superovulated prepubertal ovaries was observed in cell cultures. Although luteinizing-hormone (LH)-stimulated progesterone in both kinds of cells, buserelin significantly inhibited LH-stimulated progesterone only in luteoma cells. These results describe clear differences in basal steroid secretion between tumoral and normal tissue. Furthermore, they show that luteoma possess GnRH receptors similar to those in normal ovarian tissue, and that GnRH analogs have endocrine effects on these cells. Therefore, a direct effect of buserelin on luteoma cells can be postulated.

Animals↗