RESUMO
Immune challenge inhibits reproductive function and endocannabinoids (eCB) modulate sexual hormones. However, no studies have been performed to assess whether the eCB system mediates the inhibition of hormones that control reproduction as a result of immune system activation during systemic infections. For that reason, we evaluated the participation of the hypothalamic cannabinoid receptor CB1 on the hypothalamic-pituitary-gonadal (HPG) axis activity in rats submitted to immune challenge. Male adult rats were treated i.c.v. administration with a CB1 antagonist/inverse agonist (AM251) (500 ng/5 µL), followed by an i.p. injection of lipopolysaccharide (LPS) (5 mg/kg) 15 minutes later. Plasmatic, hypothalamic and adenohypophyseal pro-inflammatory cytokines, hormones and neuropeptides were assessed 90 or 180 minutes post-LPS. The plasma concentration of tumour necrosis factor α and adenohypophyseal mRNA expression of Tnfα and Il1ß increased 90 and 180 minutes post i.p. administration of LPS. However, cytokine mRNA expression in the hypothalamus increased only 180 minutes post-LPS, suggesting an inflammatory delay in this organ. CB1 receptor blockade with AM251 increased LPS inflammatory effects, particularly in the hypothalamus. LPS also inhibited the HPG axis by decreasing gonadotrophin-releasing hormone hypothalamic content and plasma levels of luteinising hormone and testosterone. These disruptor effects were accompanied by decreased hypothalamic Kiss1 mRNA expression and prostaglandin E2 content, as well as by increased gonadotrophin-inhibitory hormone (Rfrp3) mRNA expression. All these disruptive effects were prevented by the presence of AM251. In summary, our results suggest that, in male rats, eCB mediate immune challenge-inhibitory effects on reproductive axis at least partially via hypothalamic CB1 activation. In addition, this receptor also participates in homeostasis recovery by modulating the inflammatory process taking place after LPS administration.
Assuntos
Encefalite/imunologia , Sistema Hipotálamo-Hipofisário/imunologia , Receptor CB1 de Canabinoide/imunologia , Reprodução , Animais , Corticosterona/sangue , Citocinas/sangue , Dinoprostona/metabolismo , Encefalite/induzido quimicamente , Encefalite/metabolismo , Hormônios Hipotalâmicos/metabolismo , Sistema Hipotálamo-Hipofisário/metabolismo , Mediadores da Inflamação/sangue , Mediadores da Inflamação/imunologia , Kisspeptinas/metabolismo , Lipopolissacarídeos , Hormônio Luteinizante/sangue , Masculino , Ratos Sprague-Dawley , Canais de Cátion TRPV/metabolismo , Testosterona/sangue , Fator de Necrose Tumoral alfa/sangueRESUMO
All forms of stress, including restraint stress (RS) and lipopolysaccharide (LPS) administration, activate the hypothalamic-pituitary-adrenal (HPA) axis. LPS binds to a recognition protein (CD14) and toll-like receptor 2/4 in different cells and tissues, including the adrenal gland, to induce the production of cytokines and cause upregulation of cyclooxygenase and nitric oxide synthase (NOS) enzymes. Acute ethanol exposure activates the HPA axis, but in some conditions prolonged administration can dampen this activation as well as decrease the inflammatory responses to LPS. Therefore, this study was designed to evaluate the adrenal response to a challenge dose of LPS (50 µg/kg) injected i.p., after submitting male rats to RS, twice a day (2 h each time) for 5 days and/or ethanol administration (3 g/kg) by gavage also for 5 days, twice daily. At the end of the experiment, plasma corticosterone concentrations and adrenal gland content of prostaglandin E (PGE) and NOS activity were measured as stress mediators. The results showed that repetitive ethanol administration attenuated the adrenal stress response to LPS challenge alone and after RS, by preventing the increase in plasma corticosterone concentrations and by decreasing the PGE content and NOS activity in the adrenal gland. Therefore, we conclude that moderate alcohol consumption could attenuate the effects of psychophysical stress and impair an inflammatory response.
Assuntos
Glândulas Suprarrenais/efeitos dos fármacos , Glândulas Suprarrenais/fisiologia , Etanol/farmacologia , Lipopolissacarídeos/farmacologia , Animais , Corticosterona/sangue , Ciclo-Oxigenase 1/biossíntese , Ciclo-Oxigenase 2/biossíntese , Inflamação/prevenção & controle , Receptores de Lipopolissacarídeos/metabolismo , Lipopolissacarídeos/antagonistas & inibidores , Masculino , Proteínas de Membrana/biossíntese , Óxido Nítrico Sintase/metabolismo , Prostaglandinas E/metabolismo , Ratos , Ratos Sprague-Dawley , Restrição Física , Estresse Psicológico/tratamento farmacológico , Receptor 4 Toll-Like/metabolismoRESUMO
Bacterial and viral products, such as bacterial lipopolysaccharide (LPS), cause inducible (i) NO synthase (NOS) synthesis, which in turn produces massive amounts of nitric oxide (NO). NO, by inactivating enzymes and leading to cell death, is toxic not only to invading viruses and bacteria, but also to host cells. Injection of LPS induces interleukin (IL)-1beta, IL-1alpha, and iNOS synthesis in the anterior pituitary and pineal glands, meninges, and choroid plexus, regions outside the blood-brain barrier. Thereafter, this induction occurs in the hypothalamic regions (such as the temperature-regulating centers), paraventricular nucleus (releasing and inhibiting hormone neurons), and the arcuate nucleus (a region containing these neurons and axons bound for the median eminence). Aging of the anterior pituitary and pineal with resultant decreased secretion of pituitary hormones and the pineal hormone melatonin, respectively, may be caused by NO. The induction of iNOS in the temperature-regulating centers by infections may cause the decreased febrile response in the aged by loss of thermosensitive neurons. NO may play a role in the progression of Alzheimer's disease and parkinsonism. LPS similarly activates cytokine and iNOS production in the cardiovascular system leading to coronary heart disease. Fat is a major source of NO stimulated by leptin. As fat stores increase, leptin and NO release increases in parallel in a circadian rhythm with maxima at night. NO could be responsible for increased coronary heart disease as obesity supervenes. Antioxidants, such as melatonin, vitamin C, and vitamin E, probably play important roles in reducing or eliminating the oxidant damage produced by NO.
Assuntos
Envelhecimento/fisiologia , Óxido Nítrico/metabolismo , Animais , Aterosclerose/metabolismo , Sistema Nervoso Central/fisiologia , Corticosterona/metabolismo , Hormônio Liberador de Gonadotropina/metabolismo , Humanos , Hipotálamo/anatomia & histologia , Hipotálamo/metabolismo , Isoenzimas/metabolismo , Leptina/metabolismo , Lipopolissacarídeos/metabolismo , Modelos Biológicos , Doenças Neurodegenerativas/metabolismo , Óxido Nítrico Sintase/metabolismo , Glândula Pineal/metabolismo , Fator de Necrose Tumoral alfa/metabolismoRESUMO
TNF-alpha is involved in the regulation of normal tissue homeostasis affecting cell proliferation, differentiation, and death. We previously reported that TNF-alpha reduces anterior pituitary cell proliferation and PRL release in an estrogen-dependent manner. In the present project we studied the induction of apoptosis by TNF-alpha in anterior pituitary cells from female rats. TNF-alpha (50 ng/ml) decreased the viability of anterior pituitary cells. Incubation with TNF-alpha for 24 h increased the percentage of terminal deoxynucleotidyltransferase-mediated deoxyuridine triphosphate nick end labeling-positive cells. TNF-alpha increased the percentage of somatotropes and lactotropes with apoptotic nuclear morphology without affecting the proportion of apoptotic corticotropes or gonadotropes. TNF-alpha increased the percentage of apoptotic lactotropes in cultured cells from rats killed in proestrus and estrus, but not in diestrus. This effect was significantly higher in cells from rats in proestrus than in estrus. In anterior pituitary cells from ovariectomized rats, TNF-alpha significantly increased the percentage of apoptotic lactotropes only when the cells were incubated in the presence of 17beta-estradiol. These results indicate that TNF-alpha induces apoptosis in somatotropes and lactotropes from female rats. The apoptotic effect of TNF-alpha on lactotropes is dependent on estrogens and could be involved in the regulation of anterior pituitary cell renewal during the estrous cycle.
Assuntos
Apoptose , Adeno-Hipófise/citologia , Prolactina/metabolismo , Fator de Necrose Tumoral alfa/farmacologia , Animais , Células Cultivadas , Eletroforese em Gel de Ágar , Estradiol/farmacologia , Ciclo Estral , Estro , Feminino , Hormônio do Crescimento/metabolismo , Marcação In Situ das Extremidades Cortadas , Ovariectomia , Adeno-Hipófise/metabolismo , Proestro , Ratos , Ratos WistarRESUMO
Tumor necrosis factor-alpha (TNF-alpha) is a pleiotropic cytokine that markedly affects neuroendocrine functions. This cytokine is expressed in the anterior pituitary where its receptors are also present. Nitric oxide (NO) is synthesized in gonadotropes and folliculo-stellate cells of the anterior pituitary. Since NO directly inhibits prolactin secretion, we investigated the involvement of NO in the inhibitory effect of TNF-alpha on prolactin release from anterior pituitary cells of female rats. The presence of L-NAME (1 mM), an inhibitor of NO synthase (NOS), in the incubation medium significantly blunted the inhibition of prolactin release produced by TNF-alpha (50 ng/ml). TNF-alpha increased nitrite release to the incubation medium. The activity of NOS as measured by [(14)C]citrulline production was significantly enhanced when anterior pituitary cells were incubated with TNF-alpha for 8 h or more. Also, TNF-alpha induced iNOS gene expression in anterior pituitary cells as assessed by reverse transcriptase-polymerase chain reaction. The current results indicate that NO is involved in the inhibitory effect of TNF-alpha on prolactin secretion and that TNF-alpha induces iNOS transcription and stimulates NO synthesis in anterior pituitary cells.
Assuntos
Expressão Gênica/efeitos dos fármacos , Óxido Nítrico Sintase/metabolismo , Óxido Nítrico/metabolismo , Prolactina/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , Animais , Células Cultivadas , Inibidores Enzimáticos/metabolismo , Inibidores Enzimáticos/farmacologia , Feminino , Expressão Gênica/genética , Óxido Nítrico/agonistas , Óxido Nítrico Sintase/antagonistas & inibidores , Óxido Nítrico Sintase/genética , Óxido Nítrico Sintase Tipo II , Hipófise/citologia , Prolactina/antagonistas & inibidores , Ratos , Ratos Wistar , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Fator de Necrose Tumoral alfa/efeitos dos fármacos , Fator de Necrose Tumoral alfa/farmacologiaRESUMO
Neurokinin A (NKA) is a tachykinin that participates in the control of neuroendocrine functions. The posterior pituitary lobe (PP) contains abundant nitric oxide synthase (NOS), suggesting that nitric oxide (NO) may play a role in controlling the release of neuropeptides and neurotransmitters. In the present project, we investigated the in vitro effect of NKA on oxytocin release from hypothalamic explants and PP of male rats and the possible involvement of NO in the action of NKA. Since NKA inhibits gamma-aminobutyric acid (GABA) release from PP, we also examined the role of NO in the effect of NKA on basal and K(+)-evoked GABA release. NKA (10(-7)-10(-5) M) significantly decreased oxytocin release from PP, whereas it did not affect its release from hypothalamic explants. The inhibitory effect of NKA on oxytocin release from PP was completely blocked by the NOS inhibitors N(G)-monomethyl-L-arginine (L-NMMA, 0.5 mM) or N(G)-nitro-L-arginine-methyl-ester (L-NAME, 1 mM). Sodium nitroprusside (0.5 mM), an NO releaser, had no effect on basal GABA release but significantly decreased K(+)-evoked GABA release. L-NMMA (0.3 mM) and L-NAME (0.5 mM) increased K(+)-evoked GABA release, indicating that NO plays an inhibitory role in GABA release from PP. The inhibition in both basal and K(+)-evoked GABA release induced by NKA (10(-7) M) was reduced by L-NAME (1 mM). Also, NKA (10(-7) M) increased NO synthesis as measured by [(14)C] citrulline production. Considered all together, our data indicate that NO may mediate the inhibitory effect of NKA on the release of both oxytocin and GABA from PP.
Assuntos
GMP Cíclico/análogos & derivados , Neurocinina A/farmacologia , Óxido Nítrico Sintase/efeitos dos fármacos , Ocitocina/efeitos dos fármacos , Neuro-Hipófise/efeitos dos fármacos , Ácido gama-Aminobutírico/efeitos dos fármacos , Animais , GMP Cíclico/farmacologia , Relação Dose-Resposta a Droga , Inibidores Enzimáticos/farmacologia , Hipotálamo/efeitos dos fármacos , Hipotálamo/metabolismo , Técnicas In Vitro , Masculino , NG-Nitroarginina Metil Éster/farmacologia , Óxido Nítrico/fisiologia , Óxido Nítrico Sintase/metabolismo , Ocitocina/metabolismo , Neuro-Hipófise/metabolismo , Potássio/farmacologia , Ratos , Ratos Wistar , Tionucleotídeos/farmacologia , Ácido gama-Aminobutírico/metabolismo , ômega-N-Metilarginina/farmacologiaRESUMO
Considering that tumor necrosis factor-alpha (TNF-alpha) is involved in normal tissue homeostasis and that its receptors are expressed in the anterior pituitary, we examined the effect of this cytokine on pituitary cell growth. Because anterior pituitary function depends on hormonal environment, we also investigated the influence of gonadal steroids in the effects of TNF-alpha on cell proliferation and the release of PRL from anterior pituitary cells. In addition, the release of TNF-alpha and its action on the release of PRL from anterior pituitary cells of rats at different stages of the estrous cycle was evaluated. In minimum essential medium D-valine, a medium that restricts fibroblastic proliferation, TNF-alpha (10 and 50 ng/mL) reduced 3H-Thymidine incorporation, DNA content, and active cell number. TNF-alpha failed to affect proliferation of cells from ovariectomized (OVX) rats. However, it significantly inhibited growth of cells from OVX rats cultured with 17beta-estradiol (E2) (10(-9) M) and from chronically estrogenized rats. TNF-alpha decreased the release of PRL from cells of intact rats, especially in proestrous, OVX rats cultured with E2 and chronically estrogenized rats. The release of anterior pituitary TNF-alpha was higher in proestrous rats. These results indicate that TNF-alpha plays an inhibitory role in anterior pituitary cell growth and the release of PRL in an estrogen-dependent manner.
Assuntos
Divisão Celular/efeitos dos fármacos , Estrogênios/farmacologia , Adeno-Hipófise/citologia , Adeno-Hipófise/metabolismo , Prolactina/metabolismo , Fator de Necrose Tumoral alfa/farmacologia , Animais , Células Cultivadas , DNA/biossíntese , Estradiol/farmacologia , Estrogênios/fisiologia , Estro , Feminino , Fibroblastos/citologia , Interleucina-6/farmacologia , Ovariectomia , Adeno-Hipófise/efeitos dos fármacos , Ratos , Ratos Wistar , Fator de Necrose Tumoral alfa/metabolismoRESUMO
OBJECTIVE: In order to determine the mechanism by which nitric oxide (NO) inhibits prolactin release, we investigated the participation of cGMP-dependent cAMP-phosphodiesterases (PDEs) and protein kinase G (PKG) in this effect of NO. METHODS: Anterior pituitary glands of male rats were incubated with inhibitors of PDE and PKG with or without sodium nitroprusside (NP). Prolactin release, and cAMP and cGMP concentrations were determined by RIA. RESULTS AND CONCLUSIONS: The inhibitory effect of NP (0.5 mmol/l) on prolactin release and cAMP concentration was blocked by EHNA (10(-4)mol/l) and HL-725 (10(-4)mol/l), inhibitors of cGMP-stimulated cAMP-PDE (PDE2). 8-Br-cGMP (10(-4) and 10(-3)mol/l), which mimics cGMP as a mediator of NP effects on prolactin release, also decreased cAMP concentration. Zaprinast (10(-4)mol/l), a selective inhibitor of specific cGMP-PDE (PDE5), potentiated the NP effect on cAMP concentration. Rp-8-[(4-chlorophenyl)thio]-cGMP triethylamine (Rp-8-cGMP, 10(-7)-10(-6)mol/l), an inhibitor of PKG, reversed the effect of NP on prolactin release. The present study suggests that several mechanisms are involved in the inhibitory effect of NO on prolactin release. The activation of PDE2 by cGMP may mediate the inhibitory effect of NO on cAMP concentration and therefore on prolactin release. NO-activated PKG may also be participating in the inhibitory effect of NO on prolactin release.
Assuntos
Óxido Nítrico/farmacologia , Diester Fosfórico Hidrolases/metabolismo , Adeno-Hipófise/efeitos dos fármacos , Adeno-Hipófise/metabolismo , Prolactina/metabolismo , Proteínas Quinases/metabolismo , 3',5'-AMP Cíclico Fosfodiesterases/metabolismo , 3',5'-GMP Cíclico Fosfodiesterases/antagonistas & inibidores , 3',5'-GMP Cíclico Fosfodiesterases/metabolismo , Animais , AMP Cíclico/metabolismo , GMP Cíclico/análogos & derivados , GMP Cíclico/metabolismo , GMP Cíclico/farmacologia , Proteínas Quinases Dependentes de GMP Cíclico , Sinergismo Farmacológico , Ativação Enzimática/efeitos dos fármacos , Inibidores Enzimáticos/farmacologia , Masculino , Nitroprussiato/farmacologia , Inibidores de Fosfodiesterase/farmacologia , Purinonas/farmacologia , Ratos , Ratos WistarRESUMO
The release of cytokines during infection, inflammation and stress induces brain-mediated responses, including alterations of neuroendocrine functions. We examined the effect of interleukin-6 (IL-6) and tumor necrosis factor-alpha (TNF-alpha) on release of gamma-aminobutyric acid (GABA) from mediobasal hypothalamic (MBH) explants and posterior pituitaries (PP) of male rats. IL-6 (10 ng/ml) did not modify basal GABA release from MBH and PP, but significantly increased GABA release under depolarizing conditions (40 mM K(+)). This effect was abolished by incubation of the tissue with indomethacin, an inhibitor of cyclooxygenase activity, indicating that prostaglandins could mediate the stimulation of GABA release induced by IL-6. On the contrary, TNF-alpha (50 ng/ml) significantly decreased K(+)-evoked GABA release from both MBH and PP. This inhibitory effect was not modified by indomethacin. Neither IL-6 nor TNF-alpha affected nitric oxide synthesis, as measured by [(14)C]citrulline production. The current results indicate that IL-6 stimulates GABA release from both hypothalamus and posterior pituitary by a mechanism mediated by prostaglandins. On the contrary, TNF-alpha inhibits GABA release from both tissues. These results suggest the possibility that GABAergic activity in the hypothalamic-pituitary axis could be involved in neuroendocrine responses to cytokines.
Assuntos
Hipotálamo Médio/metabolismo , Interleucina-6/farmacologia , Neuro-Hipófise/metabolismo , Fator de Necrose Tumoral alfa/farmacologia , Ácido gama-Aminobutírico/metabolismo , Animais , Inibidores de Ciclo-Oxigenase/farmacologia , Hipotálamo Médio/efeitos dos fármacos , Hipotálamo Médio/enzimologia , Técnicas In Vitro , Indometacina/farmacologia , Interleucina-6/antagonistas & inibidores , Masculino , Potenciais da Membrana/efeitos dos fármacos , Óxido Nítrico/metabolismo , Óxido Nítrico Sintase/metabolismo , Neuro-Hipófise/efeitos dos fármacos , Neuro-Hipófise/enzimologia , Potássio/agonistas , Potássio/antagonistas & inibidores , Potássio/farmacologia , Ratos , Ratos WistarRESUMO
In this research we examined the mechanisms by which ethanol (EtOH) inhibits luteinizing hormone-releasing hormone (LHRH) release from incubated medial basal hypothalamic explants. EtOH (100 mM) stimulated the release of two inhibitory neurotransmitters: gamma-aminobutyric acid (GABA) and beta-endorphin. EtOH also inhibited NO production, indicative of a suppression of nitric oxide synthase (NOS) activity. This inhibition was reversed by naltroxone (10(-8) M), a micro-opioid receptor blocker, indicating that the inhibition of NOS by EtOH is mediated by beta-endorphin. EtOH also blocked N-methyl-d-aspartic acid-induced LHRH release, but the blockade could not be reversed by either the GABA receptor blocker, bicuculline (10(-5) M), naltroxone (10(-8) M), or both inhibitors added together. However, increasing the concentration of naltrexone (10(-6) M) but not bicuculline (10(-4) M) reversed the inhibition. When we lowered the concentration of EtOH (50 mM), the EtOH-induced blockade of LHRH release could be reversed by either bicuculline (10(-5) M), naltroxone (10(-8) M), or the combination of the two blockers. Therefore, GABA is partially responsible for the blockade of N-methyl-d-aspartic acid-induced LHRH release. The block by GABA was exerted by inhibiting the activation of cyclooxygenase by NO, because it was reversed by prostaglandin E(2), the product of activation of cyclooxygenase. Because the inhibition caused by the higher concentration of EtOH could not be reduced by bicuculline (10(-4) M) but was blocked by naltroxone (10(-6) M), the action of alcohol can be accounted for by stimulation of beta-endorphin neurons that inhibit LHRH release by inhibition of activation of NOS and stimulation of GABA release.