RESUMO
The GnRH decapeptide controls reproductive function through its release from neuroendocrine terminals in the median eminence, a site where there is a convergence of numerous nerve terminals and glial cells. Previous work showed dynamic changes in the GnRH-glial-capillary network in the median eminence under different physiological conditions. Because aging in rats is associated with a diminution of GnRH release and responsiveness to estradiol feedback, we examined effects of age and estradiol treatment on these anatomical interactions. Rats were ovariectomized at young (4 months), middle-aged (11 months), or old (22-23 months) ages, allowed 4 wk to recover, and then treated with vehicle or estradiol for 72 h followed by perfusion. Immunofluorescence of GnRH was measured, and immunogold electron microscopic analyses were performed to study the ultrastructural properties of GnRH neuroterminals and their microenvironment. Although the GnRH immunofluorescent signal showed no significant changes with age and estradiol treatment, we found that the median eminence underwent both qualitative and quantitative structural changes with age, including a disorganization of cytoarchitecture with aging and a decrease in the apposition of GnRH neuroterminals to glia with age and estradiol treatment. Thus, although GnRH neurons can continue to synthesize and transport peptide, changes in the GnRH neuroterminal-glial-capillary machinery occur during reproductive senescence in a manner consistent with a disconnection of these elements and a potential dysregulation of GnRH neurosecretion.
Assuntos
Envelhecimento , Estradiol/farmacologia , Hormônio Liberador de Gonadotropina/metabolismo , Eminência Mediana/metabolismo , Animais , Estrogênios/farmacologia , Feminino , Imuno-Histoquímica , Eminência Mediana/inervação , Eminência Mediana/ultraestrutura , Microscopia de Fluorescência , Microscopia Imunoeletrônica , Neuroglia/efeitos dos fármacos , Neuroglia/metabolismo , Neuroglia/ultraestrutura , Ovariectomia , Terminações Pré-Sinápticas/efeitos dos fármacos , Terminações Pré-Sinápticas/metabolismo , Terminações Pré-Sinápticas/ultraestrutura , Ratos , Ratos Sprague-DawleyRESUMO
Pyroglutamyl peptidase II (PPII), a highly specific membrane-bound metallopeptidase that inactivates TRH in the extracellular space, is tightly regulated by thyroid hormone in cells of the anterior pituitary. Whether PPII has any role in the region where axons containing hypophysiotropic TRH terminate, the median eminence, is unknown. For this purpose, we analyzed the cellular localization and regulation of PPII mRNA in the mediobasal hypothalamus in adult, male rats. PPII mRNA was localized in cells lining the floor and infralateral walls of the third ventricle and coexpressed with vimentin, establishing these cells as tanycytes. PPII mRNA extended in a linear fashion from the tanycyte cell bodies in the base of the third ventricle to its cytoplasmic and end-feet processes in the external zone of the median eminence in close apposition to pro-TRH-containing axon terminals. Compared with vehicle-treated, euthyroid controls, animals made thyrotoxic by the i.p. administration of 10 microg L-T(4) daily for 1-3 d, showed dramatically increased accumulation of silver grains in the mediobasal hypothalamus and an approximately 80% increase in enzymatic activity. PPII inhibition in mediobasal hypothalamic explants increased TRH secretion, whereas i.p. injection of a specific PPII inhibitor increased cold stress- and TRH-induced TSH levels in plasma. We propose that an increase in circulating thyroid hormone up-regulates PPII activity in tanycytes and enhances degradation of extracellular TRH in the median eminence through glial-axonal associations, contributing to the feedback regulation of thyroid hormone on anterior pituitary TSH secretion.
Assuntos
Aminopeptidases/fisiologia , Axônios/fisiologia , Sistema Hipotálamo-Hipofisário/fisiologia , Eminência Mediana/inervação , Neuroglia/fisiologia , Ácido Pirrolidonocarboxílico/análogos & derivados , Glândula Tireoide/fisiologia , Aminopeptidases/genética , Aminopeptidases/metabolismo , Animais , Axônios/metabolismo , Comunicação Celular/fisiologia , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Sistema Hipotálamo-Hipofisário/metabolismo , Masculino , Eminência Mediana/citologia , Eminência Mediana/enzimologia , Modelos Biológicos , Neuroglia/citologia , Neuroglia/efeitos dos fármacos , Neuroglia/metabolismo , Precursores de Proteínas/metabolismo , Ácido Pirrolidonocarboxílico/metabolismo , RNA Mensageiro/metabolismo , Ratos , Ratos Sprague-Dawley , Glândula Tireoide/metabolismo , Tireotropina/sangue , Hormônio Liberador de Tireotropina/metabolismo , Tiroxina/farmacologia , Distribuição TecidualRESUMO
Activation of the G-protein-coupled receptor GPR54 by kisspeptins during normal puberty promotes the central release of gonadotropin-releasing hormone (GnRH) that, in turn, leads to reproductive maturation. In humans and mice, a loss of function mutations of GPR54 prevents the onset of puberty and leads to hypogonadotropic hypogonadism and infertility. Using electrophysiological, morphological, molecular, and retrograde-labeling techniques in brain slices prepared from vGluT2-GFP and GnRH-GFP mice, we demonstrate the existence of two physiologically distinct subpopulations of GnRH neurons. The first subpopulation is comprised of septal GnRH neurons that colocalize vesicular glutamate transporter 2 and green fluorescent protein and is insensitive to metabotropic glutamate receptor agonists, but is exquisitely sensitive to kisspeptin which closes potassium channels to dramatically initiate a long-lasting activation in neurons from prepubertal and postpubertal mice of both sexes. A second subpopulation is insensitive to kisspeptin but is uniquely activated by group I metabotropic glutamate receptor agonists. These two physiologically distinct classes of GnRH cells may subserve different functions in the central control of reproduction and fertility.
Assuntos
Hormônio Liberador de Gonadotropina/metabolismo , Neurônios/classificação , Neurônios/fisiologia , Receptores de Glutamato Metabotrópico/agonistas , Proteínas Supressoras de Tumor/farmacologia , Animais , Compostos de Bário/farmacologia , Capilares/inervação , Cloretos/farmacologia , Feixe Diagonal de Broca/citologia , Feixe Diagonal de Broca/metabolismo , Feixe Diagonal de Broca/fisiologia , Resistência a Medicamentos , Eletrofisiologia , Feminino , Hormônio Liberador de Gonadotropina/genética , Proteínas de Fluorescência Verde/genética , Técnicas In Vitro , Masculino , Eminência Mediana/irrigação sanguínea , Eminência Mediana/inervação , Metoxi-Hidroxifenilglicol/análogos & derivados , Metoxi-Hidroxifenilglicol/farmacologia , Camundongos , Camundongos Transgênicos , Neurônios/efeitos dos fármacos , Neurônios/metabolismo , Canais de Potássio/efeitos dos fármacos , Canais de Potássio/metabolismo , RNA Mensageiro/metabolismo , Ratos , Ratos Sprague-Dawley , Sinapses/fisiologia , Proteína Vesicular 2 de Transporte de Glutamato/genéticaRESUMO
Recent studies have localized the glutamatergic cell marker type-2 vesicular glutamate transporter (VGLUT2) to distinct peptidergic neurosecretory systems that regulate hypophysial functions in rats. The present studies were aimed to map the neuronal sources of VGLUT2 in the median eminence and the posterior pituitary, the main terminal fields of hypothalamic neurosecretory neurons. Neurons innervating these regions were identified by the uptake of the retrograde tract-tracer Fluoro-Gold (FG) from the systemic circulation, whereas glutamatergic perikarya of the hypothalamus were visualized via the radioisotopic in situ hybridization detection of VGLUT2 mRNA. The results of dual-labeling studies established that the majority of neurons accumulating FG and also expressing VGLUT2 mRNA were located within the paraventricular, periventricular and supraoptic nuclei and around the organum vasculosum of the lamina terminalis and the preoptic area. In contrast, only few FG-accumulating cells exhibited VGLUT2 mRNA signal in the arcuate nucleus. Dual-label immunofluorescent studies of the median eminence and posterior pituitary to determine the subcellular location of VGLUT2, revealed the association of VGLUT2 immunoreactivity with SV2 protein, a marker for small clear vesicles in neurosecretory endings. Electron microscopic studies using pre-embedding colloidal gold labeling confirmed the localization of VGLUT2 in small clear synaptic vesicles. These data suggest that neurosecretory neurons located mainly within the paraventricular, anterior periventricular and supraoptic nuclei and around the organum vasculosum of the lamina terminalis and the preoptic area secrete glutamate into the fenestrated vessels of the median eminence and posterior pituitary. The functional aspects of the putative neuropeptide/glutamate co-release from neuroendocrine terminals remain to be elucidated.
Assuntos
Ácido Glutâmico/metabolismo , Hipotálamo/metabolismo , Eminência Mediana/inervação , Vias Neurais/metabolismo , Neuro-Hipófise/inervação , Proteína Vesicular 2 de Transporte de Glutamato/metabolismo , Animais , Biomarcadores/metabolismo , Hipotálamo/ultraestrutura , Hibridização In Situ , Masculino , Eminência Mediana/irrigação sanguínea , Eminência Mediana/ultraestrutura , Glicoproteínas de Membrana/metabolismo , Microcirculação/citologia , Microcirculação/fisiologia , Microscopia Imunoeletrônica , Proteínas do Tecido Nervoso/metabolismo , Vias Neurais/ultraestrutura , Neurônios/citologia , Neurônios/metabolismo , Sistemas Neurossecretores/metabolismo , Sistemas Neurossecretores/ultraestrutura , Hipófise/irrigação sanguínea , Hipófise/inervação , Hipófise/fisiologia , Neuro-Hipófise/irrigação sanguínea , Neuro-Hipófise/ultraestrutura , Terminações Pré-Sinápticas/metabolismo , Terminações Pré-Sinápticas/ultraestrutura , RNA Mensageiro/metabolismo , Ratos , Ratos Wistar , Estilbamidinas , Vesículas Sinápticas/metabolismo , Vesículas Sinápticas/ultraestrutura , Proteína Vesicular 2 de Transporte de Glutamato/genéticaRESUMO
Reproduction in Japanese quail is primarily regulated by photoperiod. Vasoactive intestinal peptide (VIP) has been suggested as a transducer of environmental information, especially photoperiodic cues, to the hypothalamo-pituitary-gonadal axis. To investigate the possible interaction of VIP and the reproductive (gonadotropin-releasing hormone, GnRH) system, double-immunocytochemical staining for VIP and cGnRH-I was conducted in sexually mature male quail held under a long-day photoperiod (16L:8D; LD) and in sexually quiescent males held under a short-day photoperiod (8L:16D; SD). VIP-immunoreactive (ir) cells were found primarily in three locations: lateral septal organ (LSO) in nucleus accumbens (Ac), ventral hypothalamus, and infundibular area. VIP-ir cells in LSO displayed characteristics typical of cerebrospinal fluid (CSF)-contacting cells, and co-existed with cGnRH-I-ir cells and beaded fibers. In contrast, VIP-ir cells in the infundibular area did not co-exist with cGnRH-I-ir structures. The number of visible VIP-ir cells in the infundibular area of SD males was significantly lower than that of LD males, while the number of visible VIP-ir cells in Ac/LSO was not altered by photoperiod. A cluster of cGnRH-I-ir cells in the caudalmost septal area was heavily innervated by VIP-ir fibers, which appeared to contact cGnRH-I-ir cells directly at this location. Both VIP- and cGnRH-I-ir fibers heavily innervated the external layer of the median eminence (ME). These data suggest that Ac/LSO, the caudalmost septal area, and ME are possible sites of interaction between the VIP and the GnRH systems.