RESUMO
There is considerable evidence of the neuroendocrine control involved in luteal regression in the rat. In addition, circulating prolactin (PRL), which increases during the night before parturition, may gain access to the coeliac ganglion (CG), indirectly impacting the physiology of the ovary because of the known connection between the CG and the ovary via the superior ovarian nerve (SON). In this work we investigated in the CG-SON-ovary system and whether PRL added to the CG has an impact, indirectly via the SON, on luteal regression on Day 21 of pregnancy. The system was incubated without (control) or with PRL added to the CG. We measured the ovarian release of progesterone (P), oestradiol and prostaglandin F2 alpha (PGF2α) by radioimmunoassay, and nitrites (NO) by the Griess method. Luteal mRNA expression of 3ß-hydroxysteroid dehydrogenase (3ß-HSD), 20α-HSD, aromatase, inducible nitric oxide synthase (iNOS) and apoptosis regulatory factors was analysed by reverse transcription-polymerase chain reaction. P release, the expression of Bcl-2 and the Bcl-2:Bax ratio was lower than control preparations, while the expression of 20α-HSD and the release of NO and PGF2α were higher in the experimental group. In conclusion, PRL acts at the CG and, by a neural pathway, modulates luteal function at the end of pregnancy.
Assuntos
Corpo Lúteo/inervação , Gânglios Simpáticos/efeitos dos fármacos , Luteólise/efeitos dos fármacos , Ovário/inervação , Prolactina/farmacologia , 20-alfa-Hidroxiesteroide Desidrogenase/genética , 20-alfa-Hidroxiesteroide Desidrogenase/metabolismo , 3-Hidroxiesteroide Desidrogenases/genética , 3-Hidroxiesteroide Desidrogenases/metabolismo , Animais , Aromatase/genética , Aromatase/metabolismo , Corpo Lúteo/enzimologia , Corpo Lúteo/patologia , Dinoprosta/metabolismo , Estradiol/metabolismo , Feminino , Gânglios Simpáticos/fisiologia , Idade Gestacional , Óxido Nítrico Sintase Tipo II/genética , Óxido Nítrico Sintase Tipo II/metabolismo , Nitritos/metabolismo , Ovário/metabolismo , Gravidez , Progesterona/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/genética , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Ratos Sprague-Dawley , Fatores de Tempo , Proteína X Associada a bcl-2/genética , Proteína X Associada a bcl-2/metabolismoRESUMO
OBJECTIVE: To investigate whether cholinergic ganglionic stimulus modifies the release of gonadotropin-releasing hormone (GnRH), catecholamines, and progesterone at the ovarian level. DESIGN: Animal study. SETTING: University animal laboratory. ANIMAL(S): Six to eight virgin adult Holtzman rats. INTERVENTION(S): Superior mesenteric ganglion-ovarian nerve plexus-ovary system removed and placed in one cuvette with two compartments, with acetylcholine added to the ganglion in the experimental group. MAIN OUTCOME MEASURE(S): Measurement of ovarian liquid obtained from catecholamines by high-performance liquid chromatography; measurement of progesterone (P(4)), GnRH, and luteinizing hormone (LH) by radioimmunoassay; and measurement of gene expression of 3ß-hydroxysteroid dehydrogenase (3ß-HSD) and 20α-hydroxysteroid dehydrogenase (20α-HSD) by reverse-transcriptase polymerase chain reaction (RT-PCR). RESULT(S): The study focused on the estrus and diestrus II (DII) stages. On the estrus days, the release of GnRH, NA, and 20α-HSD increased, while P(4) and 3ß-HSD decreased. On the DII days, GnRH, P(4), and 3ß-HSD increased, while 20α-HSD and NA decreased. The ovarian liquid with GnRH showed biologic activity, namely, an increase in LH release during the DII stage and a decrease during the estrus stage. CONCLUSION(S): Neural stimulus from the superior mesenteric ganglion influences the release of NA, adrenaline, and GnRH. We also have demonstrated that these neurotransmitters participate in the atretogenic processes of the ovary, thus providing evidence of the necessity of the sympathetic neural pathway.
Assuntos
Catecolaminas/metabolismo , Gânglios Simpáticos/metabolismo , Hormônio Liberador de Gonadotropina/metabolismo , Ovário/inervação , Ovário/metabolismo , Progesterona/metabolismo , Receptores Colinérgicos/metabolismo , 20-alfa-Hidroxiesteroide Desidrogenase/genética , 20-alfa-Hidroxiesteroide Desidrogenase/metabolismo , 3-Hidroxiesteroide Desidrogenases/genética , 3-Hidroxiesteroide Desidrogenases/metabolismo , Acetilcolina/metabolismo , Animais , Cromatografia Líquida de Alta Pressão , Diestro/metabolismo , Estro/metabolismo , Feminino , Ovário/enzimologia , RNA Mensageiro/metabolismo , Radioimunoensaio , Ratos , Ratos Sprague-Dawley , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Fatores de TempoRESUMO
The aims of this work were to investigate if oestradiol 10(-8)M in the incubation media of either the ovary alone (OV) or the ganglion compartment of an ex vivo coeliac ganglion-superior ovarian nerve-ovary system (a) modifies the release of ovarian progesterone (P4) and oestradiol (E2) on dioestrus II, and (b) modifies the ovarian gene expression of 3ß-HSD and 20α-HSD enzymes and markers of apoptosis. The concentration of ovarian P4 release was measured in both experimental schemes, and ovarian P4 and E2 in the ex vivo system by RIA at different times. The expression of 3ß-hydroxysteroid dehydrogenase, 20α-hydroxysteroid dehydrogenase and antiapoptotic bcl-2 and proapoptotic bax by RT-PCR were determined. E2 added in the coeliac ganglion caused an increase in the ovarian release of the P4, E2 and 3ß-HSD, while in the ovary incubation alone it decreased P4 and 3ß-HSD but increased and 20α-HSD and bax/bcl-2 ratio. It is concluded that through a direct effect on the ovary, E2 promotes luteal regression in DII rats, but the addition of E2 in the coeliac ganglion does not have the same effect. The peripheral nervous system, through the superior ovarian nerve, has a protective effect against the apoptotic mechanism on DII.
Assuntos
Apoptose/efeitos dos fármacos , Diestro , Estradiol/farmacologia , Gânglios Simpáticos/efeitos dos fármacos , Ovário , 20-alfa-Hidroxiesteroide Desidrogenase/genética , 20-alfa-Hidroxiesteroide Desidrogenase/metabolismo , 3-Hidroxiesteroide Desidrogenases/genética , 3-Hidroxiesteroide Desidrogenases/metabolismo , Animais , Estradiol/biossíntese , Estradiol/metabolismo , Feminino , Gânglios Simpáticos/metabolismo , Ovário/efeitos dos fármacos , Ovário/inervação , Ovário/metabolismo , Progesterona/metabolismo , Ratos , Proteína X Associada a bcl-2/metabolismo , Proteína de Morte Celular Associada a bcl/biossíntese , Proteína de Morte Celular Associada a bcl/metabolismoRESUMO
Whether prolactin (PRL) has a luteotrophic or luteolytic effect in the rat ovary depends on the nature of the corpora lutea present in the ovaries and the hormonal environment to which they are exposed. The aim was to investigate the effect of PRL acting on the coeliac ganglion (CG) on the function of the corpora lutea on day 4 postpartum under either lactating or non-lactating conditions, using the CG-superior ovarian nerve-ovary system. The ovarian release of progesterone (P), estradiol, PGF2α, and nitrites was assessed in the ovarian compartment at different incubation times. Luteal mRNA expression of 3ß-HSD, 20α-HSD, aromatase, PGF2α receptor, iNOS, Bcl-2, Bax, Fas and FasL was analysed in the corpus luteum of pregnancy at the end of the experiments. Comparative analysis of control groups showed that the ovarian release of P, nitrites, and PGF2α, the expression of PGF2α receptor, and the Bcl-2/Bax ratio were lower in non-lactating rats, with increased release of estradiol, and higher expression of aromatase, Fas and FasL, demonstrating the higher luteal functionality in ovaries of lactating animals. PRL added to the CG compartment increased the ovarian release of P, estradiol, nitrites and PGF2α, and decreased the Bcl-2/Bax ratio in non-lactating rats; yet, with the exception of a reduction in the release of nitrites, such parameters were not modified in lactating animals. Together, these data suggest that the CG is able to respond to the effect of PRL and, via a neural pathway, fine-tune the physiology of the ovary under different hormonal conditions.
Assuntos
Gânglios Simpáticos/efeitos dos fármacos , Gânglios Simpáticos/metabolismo , Lactação/efeitos dos fármacos , Lactação/metabolismo , Ovário/inervação , Ovário/metabolismo , Período Pós-Parto/metabolismo , Prolactina/farmacologia , 20-alfa-Hidroxiesteroide Desidrogenase/genética , 3-Hidroxiesteroide Desidrogenases/genética , Animais , Aromatase/genética , Estradiol/metabolismo , Proteína Ligante Fas/genética , Feminino , Nitritos/metabolismo , Ovário/efeitos dos fármacos , Período Pós-Parto/efeitos dos fármacos , Gravidez , Progesterona/metabolismo , Prostaglandinas/metabolismo , Radioimunoensaio , Ratos , Receptores de Prostaglandina/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Proteína X Associada a bcl-2/genéticaRESUMO
There is evidence suggesting that estradiol (E(2)) regulates the physiology of the ovary and the sympathetic neurons associated with the reproductive function. The objective of this study was to investigate the effect of E(2) on the function of late pregnant rat ovaries, acting either directly on the ovarian tissue or indirectly via the superior ovarian nerve (SON) from the celiac ganglion (CG). We used in vitro ovary (OV) or ex vivo CG-SON-OV incubation systems from day 21 pregnant rats. Various concentrations of E(2 )were added to the incubation media of either the OV alone or the ganglion compartment of the CG-SON-OV system. In both experimental schemes, we measured the concentration of progesterone in the OV incubation media by radioimmunoassay at different times. Luteal messenger RNA (mRNA) expression of 3ß-hydroxysteroid dehydrogenase (3ß-HSD) and 20α-hydroxysteroid dehydrogenase (20α-HSD) enzymes, respectively, involved in progesterone synthesis and catabolism, and of antiapoptotic B-cell lymphoma 2 (Bcl-2) and proapoptotic Bcl-2-associated X protein (Bax), were measured by reverse transcriptase-polymerase chain reaction (RT-PCR) at the end of the incubation period. Estradiol added directly to the OV incubation or to the CG of the CG-SON-OV system caused a decline in the concentration of progesterone accumulated in the incubation media. In addition, E(2), when added to the OV incubation, decreased the expression of 3ß-HSD and the ratio of Bcl-2/Bax. We conclude that through a direct effect on the OV, E(2) favors luteal regression at the end of pregnancy in rats, in association with neural modulation from the CG via the SON.
Assuntos
Corpo Lúteo/efeitos dos fármacos , Estradiol/farmacologia , Gânglios Simpáticos/efeitos dos fármacos , Luteólise/efeitos dos fármacos , Ovário/efeitos dos fármacos , Progesterona/metabolismo , 20-alfa-Hidroxiesteroide Desidrogenase/genética , 20-alfa-Hidroxiesteroide Desidrogenase/metabolismo , 3-Hidroxiesteroide Desidrogenases/genética , 3-Hidroxiesteroide Desidrogenases/metabolismo , Animais , Corpo Lúteo/enzimologia , Corpo Lúteo/inervação , Corpo Lúteo/fisiologia , Feminino , Gânglios Simpáticos/enzimologia , Gânglios Simpáticos/fisiologia , Técnicas In Vitro , Luteólise/fisiologia , Ovário/enzimologia , Ovário/inervação , Ovário/fisiologia , Gravidez , Proteínas Proto-Oncogênicas c-bcl-2/genética , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , RNA Mensageiro/química , RNA Mensageiro/genética , Ratos , Ratos Sprague-Dawley , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Proteína X Associada a bcl-2/genética , Proteína X Associada a bcl-2/metabolismoRESUMO
The objective of the present study was to examine some factors involved in follicular development of women with polycystic ovary syndrome (PCOS). Women with PCOS showed increased levels of serum luteinizing hormone (LH) but decreased follicular production of progesterone and estradiol by pre-ovulatory follicles. The mRNA expression corresponding to steroidogenic acute regulatory protein (StAR), and 20alpha-hydroxysteroid dehydrogenase (20α-HSD) was increased, while that corresponding to cytochrome P450 aromatase (P450arom) was decreased in PCOS follicles as compared to controls. No changes in the mRNA expression for 3beta-hydroxysteroid dehydrogenase 2 (3ß-HSD2), cytochrome P450 side-chain cleavage (P450scc), cytochrome P450 17 alpha hydroxylase/lyase (P450c17), cyclooxygenase 2 (COX2), and transcription factors (GATA-4 and GATA-6) were found. We conclude that despite the hyper-luteinized environment of PCOS follicles, these follicles produce lower levels of progesterone and estradiol, and that this is characterized by increased degradation of progesterone and decreased estradiol synthesis. Our data demonstrate that the synthesis of prostaglandin F2α (PGF2α) may be affected in PCOS-follicles and that the transcription factors GATA-4 and GATA-6 are present in PCOS-follicles but they are not involved in the abnormal transcription observed in the steroidogenic enzymes.
Assuntos
Folículo Ovariano/patologia , Síndrome do Ovário Policístico/patologia , 20-alfa-Hidroxiesteroide Desidrogenase/biossíntese , 20-alfa-Hidroxiesteroide Desidrogenase/genética , 3-Hidroxiesteroide Desidrogenases/biossíntese , 3-Hidroxiesteroide Desidrogenases/genética , Aromatase/biossíntese , Aromatase/genética , Ciclo-Oxigenase 2/biossíntese , Ciclo-Oxigenase 2/genética , Estradiol/biossíntese , Feminino , Fator de Transcrição GATA4/genética , Fator de Transcrição GATA4/fisiologia , Fator de Transcrição GATA6/genética , Fator de Transcrição GATA6/fisiologia , Humanos , Hormônio Luteinizante/sangue , Folículo Ovariano/metabolismo , Fosfoproteínas/biossíntese , Fosfoproteínas/genética , Síndrome do Ovário Policístico/sangue , Síndrome do Ovário Policístico/enzimologia , Síndrome do Ovário Policístico/metabolismo , Progesterona/biossíntese , RNA Mensageiro/biossíntese , RNA Mensageiro/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Esteroide 17-alfa-Hidroxilase/biossíntese , Esteroide 17-alfa-Hidroxilase/genéticaRESUMO
The ovarian function is controlled by endocrine factors and neural influence. In late pregnant rat, androstenedione, from the coeliac ganglion, has a luteotrophic effect in the ex vivo coeliac ganglion-superior ovarian nerve-ovary system. In this work we investigate the presence of androgen receptors in the coeliac ganglion of late pregnant rats by immunohistochemistry. We also explore, from a physiological point of view, the potential participation of these receptors in the androstenedione ganglionic action on progesterone release and metabolism, as well as on nitrites release in the ovary compartment. The coeliac ganglion was isolated after being fixed in situ and immunohistochemistry was performed. In the system, three experimental groups were used with the addition of (a) androstenedione, (b) flutamide, and (c) androstenedione plus flutamide in the ganglion compartment. Progesterone and nitrite concentrations were determined in the ovary compartment at different incubation times. Corpora lutea samples isolated at the end of incubation were used to determine the expressions and activities of the progesterone synthesis (3beta-hydroxysteroid-dehydrogenase, 3beta-HSD) and degradation (20alpha-hydroxysteroid-dehydrogenase, 20alpha-HSD) enzymes. Immunohistochemistry revealed cytoplasmatic androgen receptor immunoreactivity in neural somas in the coeliac ganglion. In the coeliac ganglion-superior ovarian nerve-ovary system, androstenedione addition increased 3beta-HSD and decreased 20alpha-HSD, showed a tendency to decrease 20alpha-HSD expression, and increased nitrites release in relation to control. Androstenedione plus flutamide decreased progesterone and nitrites release in relation to the androstenedione group. This work demonstrates the presence of androgen receptors in neurons of celiac ganglion and provides evidence for the luteotrophic action of androstenedione via a neural pathway that may be mediated by these receptors.
Assuntos
Gânglios Simpáticos/metabolismo , Receptores Androgênicos/metabolismo , 20-alfa-Hidroxiesteroide Desidrogenase/genética , 20-alfa-Hidroxiesteroide Desidrogenase/metabolismo , 3-Hidroxiesteroide Desidrogenases/genética , 3-Hidroxiesteroide Desidrogenases/metabolismo , Antagonistas de Androgênios/farmacologia , Androstenodiona/farmacologia , Animais , Corpo Lúteo/efeitos dos fármacos , Corpo Lúteo/metabolismo , Feminino , Flutamida/farmacologia , Gânglios Simpáticos/citologia , Gânglios Simpáticos/efeitos dos fármacos , Imuno-Histoquímica , Masculino , Nitrilas/metabolismo , Ovário/efeitos dos fármacos , Ovário/metabolismo , Gravidez , Progesterona/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Ratos , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Fatores de TempoRESUMO
A decrease in serum progesterone at the end of pregnancy is essential for the induction of parturition in rats. We have previously demonstrated that LH participates in this process through: 1) inhibiting 3beta-hydroxysteroid dehydrogenase (3beta-HSD) activity and 2) stimulating progesterone catabolism by inducing 20alpha-hydroxysteroid dehydrogenase (20alpha-HSD) activity. The objective of this investigation was to determine the effect of LH and progesterone on the luteal expression of the steroidogenic acute regulatory protein (StAR), cytochrome P450 side-chain cleavage (P450(scc)), 3beta-HSD, and 20alpha-HSD genes. Gene expression was analyzed by Northern blot analysis 24 and 48 h after administration of LH or vehicle on Day 19 of pregnancy. StAR and 3beta-HSD mRNA levels were lower in LH-treated rats than in rats administered with vehicle at both time points studied. P450(scc) mRNA levels were unaffected by LH. The 20alpha-HSD mRNA levels were not different between LH and control rats 24 h after treatment; however, greater expression of 20alpha-HSD, with respect to controls, was observed in LH-treated rats 48 h after treatment. Luteal progesterone content dropped in LH-treated rats at both time points studied, whereas serum progesterone decreased after 48 h only. In a second set of experiments, the anti-progesterone RU486 was injected intrabursally on Day 20 of pregnancy. RU486 had no effect on 3beta-HSD or P450(scc) expression but increased 20alpha-HSD mRNA levels after 8 h treatment. In conclusion, the luteolytic effect of LH is mediated by a drop in StAR and 3beta-HSD expression without effect on P450(scc) expression. We also provide the first in vivo evidence indicating that a decrease in luteal progesterone content may be an essential step toward the induction of 20alpha-HSD expression at the end of pregnancy in rats.
Assuntos
20-Hidroxiesteroide Desidrogenases/genética , 3-Hidroxiesteroide Desidrogenases/genética , Enzima de Clivagem da Cadeia Lateral do Colesterol/genética , Corpo Lúteo/metabolismo , Expressão Gênica/efeitos dos fármacos , Fosfoproteínas/genética , 20-alfa-Hidroxiesteroide Desidrogenase , Animais , Northern Blotting , Feminino , Idade Gestacional , Antagonistas de Hormônios/farmacologia , Hormônio Luteinizante/farmacologia , Mifepristona/farmacologia , Gravidez , Progesterona/antagonistas & inibidores , RNA Mensageiro/análise , Ratos , Ratos WistarRESUMO
In this study, we investigated whether progesterone exerts a local action regulating the function of the corpus luteum of pregnancy in rats. The luteal activities of the enzymes 3beta-hydroxysteroid dehydrogenase (3beta-HSD), involved in progesterone biosynthesis, and 20alpha-hydroxysteroid dehydrogenase (20alpha-HSD), that catabolizes progesterone and reduces progesterone secretion by the corpus luteum, were evaluated after intrabursal ovarian administration of progesterone in pregnant rats that had received a luteolytic dose of prostaglandin F2alpha (PGF2alpha). Luteal 3beta-HSD activity decreased and 20alpha-HSD activity increased after PGF2alpha treatment (100 microg x 2 intraperitoneally on Day 19 of pregnancy at 12:00 p.m. and 4:00 p.m.) when compared with controls sacrificed at 8:00 p.m. on Day 20 of pregnancy. This effect of PGF2alpha on the luteal 3beta-HSD and 20alpha-HSD activities was abolished in animals that also received an intraovarian dose of progesterone (3 microg/ovary on Day 19 of pregnancy at 8:00-9:00 a.m.). In a second functional study, luteal cells obtained from 19-day pregnant rats responded to the synthetic progestin promegestone (R5020) in a dose-dependent manner, with an increase in the progesterone output. In addition, the glucocorticoid agent hydrocortisone did not affect progesterone accumulation in the same luteal cell culture. We also examined by immunocytochemistry the expression of progesterone receptors (PR) in the corpora lutea during pregnancy and demonstrated the absence of PR in this endocrine gland in all the days of pregnancy studied. In the same pregnant rats, positive staining for PR was observed in cells within the uteroplacental unit, such as cells of the decidua basalis and trophoblast giant cells of the junctional zone. In addition, positive PR staining was observed in the ovarian granulosa and theca cells of growing follicles, but not in corpora lutea of ovaries obtained from cycling rats at proestrus. In summary, this report provides further evidence of a local action of progesterone regulating luteal function in the rat despite the absence of a classic PR.
Assuntos
Corpo Lúteo/fisiologia , Prenhez/fisiologia , Progesterona/fisiologia , Receptores de Progesterona/fisiologia , 20-Hidroxiesteroide Desidrogenases/metabolismo , 20-alfa-Hidroxiesteroide Desidrogenase , 3-Hidroxiesteroide Desidrogenases/metabolismo , Animais , Corpo Lúteo/efeitos dos fármacos , Corpo Lúteo/enzimologia , Feminino , Hidroxicorticosteroides/farmacologia , Ovário/metabolismo , Placenta/metabolismo , Gravidez , Promegestona/farmacologia , Ratos , Ratos Wistar , Receptores de Progesterona/metabolismoRESUMO
We examined the participation of the intraluteal levels of progesterone (P4) and prostaglandin F2 alpha (PGF2 alpha) in the induction of luteolysis by LH and its relationship with the induction of the 20 alpha-hydroxysteroid dehydrogenase activity (20 alpha-HSD). Subcutaneous administration of four doses of 10 microgram ovine LH (oLH) at 0800, 0900, 1000 and 1100 h on day 19 of pregnancy induced a decrease in the activity of the enzyme 3 beta-HSD 24 and 48 h after treatment and an increase in luteal 20 alpha-HSD activity 48 h after oLH treatment when compared with control rats. Intraluteal and serum P4 levels were lower than control values 24 and 48 h after oLH treatment, with a significant increase in luteal PGF2 alpha content and a decrease in corpus luteum (CL) weight 48 h after oLH treatment. Intrabursal ovarian (i.b.) treatment with an inhibitor of PG's biosynthesis (diclofenac) (70 microgram/ovary) or P4 (3 microgram/ovary) on day 20 of pregnancy, prevented the increase in 20 alpha-HSD activity observed 48 h after oLH treatment, without any effect on 3 beta-HSD activity. The i.b. administration of P4 prevented the increase in intraluteal PGF2 alpha content induced by oLH treatment and the increases in 20 alpha-HSD activity and intraluteal PGF2 alpha content observed in control animals on day 21 of pregnancy. The inhibition of PG biosynthesis also prevents the decrease in intraluteal and serum P4 level induced by oLH. These results provide good evidence of the important participation of intraluteal P4 and PGF2 alpha on the oLH-induced luteolysis in pregnant rats. We also found the P4 produced by the CL is involved, in part, in the regulation of luteal PG synthesis. Thus, the early decline in 3 beta-HSD activity and the consequent fall in intraluteal P4 content, may trigger the synthesis of PGs and thereafter the increase in luteal 20 alpha-HSD activity to establish luteolysis.