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1.
Hum Reprod ; 32(10): 2088-2100, 2017 10 01.
Artículo en Inglés | MEDLINE | ID: mdl-28938749

RESUMEN

STUDY QUESTION: What is the genetic landscape within the testis of the juvenile rhesus monkey (Macaca mulatta) that underlies the decision of undifferentiated spermatogonia to commit to a pathway of differentiation when puberty is induced prematurely by exogenous LH and FSH stimulation? SUMMARY ANSWER: Forty-eight hours of gonadotrophin stimulation of the juvenile monkey testis resulted in the appearance of differentiating B spermatogonia and the emergence of 1362 up-regulated and 225 down-regulated testicular mRNAs encoding a complex network of proteins ranging from enzymes regulating Leydig cell steroidogenesis to membrane receptors, and from juxtacrine and paracrine factors to transcriptional factors governing spermatogonial stem cell fate. WHAT IS KNOWN ALREADY: Our understanding of the cell and molecular biology underlying the fate of undifferentiated spermatogonia is based largely on studies of rodents, particularly of mice, but in the case of primates very little is known. The present study represents the first attempt to comprehensively address this question in a highly evolved primate. STUDY DESIGN, SIZE, DURATION: Global gene expression in the testis from juvenile rhesus monkeys that had been stimulated with recombinant monkey LH and FSH for 48 h (N = 3) or 96 h (N = 4) was compared to that from vehicle treated animals (N = 3). Testicular cell types and testosterone secretion were also monitored. PARTICIPANTS/MATERIALS, SETTING, METHODS: Precocious testicular puberty was initiated in juvenile rhesus monkeys, 14-24 months of age, using a physiologic mode of intermittent stimulation with i.v. recombinant monkey LH and FSH that within 48 h produced 'adult' levels of circulating LH, FSH and testosterone. Mitotic activity was monitored by immunohistochemical assays of 5-bromo-2'-deoxyuridine and 5-ethynyl-2'-deoxyuridine incorporation. Animals were bilaterally castrated and RNA was extracted from the right testis. Global gene expression was determined using RNA-Seq. Differentially expressed genes (DEGs) were identified and evaluated by pathway analysis. mRNAs of particular interest were also quantitated using quantitative RT-PCR. Fractions of the left testis were used for histochemistry or immunoflouresence. MAIN RESULTS AND THE ROLE OF CHANCE: Differentiating type B spematogonia were observed after both 48 and 96 h of gonadotrophin stimulation. Pathway analysis identified five super categories of over-represented DEGs. Repression of GFRA1 (glial cell line-derived neurotrophic factor family receptor alpha 1) and NANOS2 (nanos C2HC-type zinc finger 2) that favor spermatogonial stem cell renewal was noted after 48 and 96 h of LH and FSH stimulation. Additionally, changes in expression of numerous genes involved in regulating the Notch pathway, cell adhesion, structural plasticity and modulating the immune system were observed. Induction of genes associated with the differentiation of spermatogonia stem cells (SOHLH1(spermatogenesis- and oogenesis-specific basic helix-loop-helix 1), SOHLH2 and KIT (V-Kit Hardy-Zuckerman 4 feline sarcoma viral oncogene homolog)) was not observed. Expression of the gene encoding STRA8 (stimulated by retinoic acid 8), a protein generally considered to mark activation of retinoic acid signaling, was below our limit of detection. LARGE SCALE DATA: The entire mRNA data set for vehicle and gonadotrophin treated animals (N = 10) has been deposited in the GEO-NCBI repository (GSE97786). LIMITATIONS REASONS FOR CAUTION: The limited number of monkeys per group and the dilution of low abundance germ cell transcripts by mRNAs contributed from somatic cells likely resulted in an underestimation of the number of differentially expressed germ cell genes. WIDER IMPLICATIONS OF THE FINDINGS: The findings that expression of GDNF (a major promoter of spermatogonial stem cell renewal) was not detected in the control juvenile testes, expression of SOHLH1, SOHLH2 and KIT, promoters of spermatogonial differentiation in mice, were not up-regulated in association with the gonadotrophin-induced generation of differentiating spermatogonia, and that robust activation of the retinoic acid signaling pathway was not observed, could not have been predicted. These unexpected results underline the importance of non-human primate models in translating data derived from animal research to the human situation. STUDY FUNDING/COMPETING INTEREST(S): The work described was funded by NIH grant R01 HD072189 to T.M.P. P.A. was supported by an Endocrine Society Summer Research Fellowship Award and CONICET (Argentine Research Council), S.N. by a grant from Vali-e-Asr Reproductive Health Research Center of Tehran University of Medical Sciences (grant #24335-39-92) to Dr Batool Hosseini Rashidi, and M.P.H. by grants from the National Health and Medical Research Council of Australia, and the Victorian State Government's Operational Infrastructure Support Program. The authors have nothing to disclose.


Asunto(s)
Gonadotropinas/metabolismo , Espermatogonias/metabolismo , Testículo/metabolismo , Transcriptoma , Animales , Hormona Folículo Estimulante/metabolismo , Macaca mulatta/genética , Macaca mulatta/metabolismo , Masculino , Modelos Animales , ARN Mensajero/metabolismo , Maduración Sexual/genética , Maduración Sexual/fisiología , Espermatogénesis/genética , Espermatogonias/citología , Testículo/citología , Testosterona/metabolismo
2.
Neuroendocrinology ; 105(1): 77-89, 2017.
Artículo en Inglés | MEDLINE | ID: mdl-27454155

RESUMEN

The aim of this immunohistochemical study was to evaluate the distribution of kisspeptin neurons in the preoptic area (POA) of gonadally intact adult male and female rhesus monkeys, and to determine whether imposition of an estradiol (E2)-positive feedback signal in the castrate male increased kisspeptin in the POA. Additionally, kisspeptin in the POA of the intact female was examined during an LH surge induced prematurely by E2 administered in the early follicular phase. The number of kisspeptin neurons in the POA of males and females was similar. Immunoactive kisspeptin perikarya were not observed in the POA of castrate adult males, but such neurons in these animals were present within 12 h of imposing an increment in circulating E2 concentrations that in a screening study conducted 4-6 weeks earlier had elicited an LH surge. As expected, premature induction of an LH surge by E2 early in the follicular phase was associated with upregulation of kisspeptin in the POA. These results represent the first description of immunoreactive kisspeptin cell bodies in the POA of the macaque brain and provide further support for the view that (1) kisspeptin neurons in the POA of the female monkey are a target for the positive feedback action of E2 and (2) the hypothalamic mechanism which mediates this action of E2 in primates is not subjected to perinatal programming by testicular testosterone. Moreover, our findings indicate that maintenance of the kisspeptin content in the POA of intact male monkeys requires the action of E2, presumably generated by aromatization of testicular testosterone at the hypothalamic level.


Asunto(s)
Estradiol/farmacología , Estrógenos/farmacología , Kisspeptinas/metabolismo , Área Preóptica/efectos de los fármacos , Caracteres Sexuales , Regulación hacia Arriba/efectos de los fármacos , Análisis de Varianza , Animales , Anticuerpos/farmacología , Castración , Recuento de Células , Estradiol/sangre , Estrógenos/sangre , Femenino , Fase Folicular/efectos de los fármacos , Humanos , Histerectomía , Kisspeptinas/inmunología , Hormona Luteinizante/sangre , Macaca mulatta , Masculino , Neuronas/efectos de los fármacos , Neuronas/metabolismo , Ovulación/efectos de los fármacos , Área Preóptica/citología , Área Preóptica/metabolismo , Vasopresinas/metabolismo
3.
Neuroendocrinology ; 103(6): 711-23, 2016.
Artículo en Inglés | MEDLINE | ID: mdl-26580201

RESUMEN

Substance P (SP) was recently reported to be expressed in human kisspeptin/neurokinin B/dynorphin (KNDy) neurons and to enhance KNDy neuron excitability in the mouse hypothalamus. We therefore examined (1) interactions of SP and kisspeptin in the mediobasal hypothalamus of adult male rhesus monkeys using immunofluorescence, and (2) the ability of SP to induce LH release in GnRH-primed, agonadal juvenile male monkeys. SP cell bodies were observed only occasionally in the arcuate nucleus (Arc), but more frequently dorsal to the Arc in the region of the premammillary nucleus. Castration resulted in an increase in the number of SP cell bodies in the Arc but not in the other regions. SP fibers innervated the Arc, where they were found in close apposition with kisspeptin perikarya in the periphery of this nucleus. Beaded SP axons projected to the median eminence, where they terminated in the external layer and intermingled with beaded kisspeptin axons. Colocalization of the two peptides, however, was not observed. Although close apposition between SP fibers and kisspeptin neurons suggest a role for SP in modulating GnRH pulse generator activity, i.v. injections of SP failed to elicit release of GnRH (as reflected by LH) in the juvenile monkey. Although the finding of structural interactions between SP and kisspeptin neurons is consistent with the notion that this tachykinin may be involved in regulating pulsatile GnRH release, the apparent absence of expression of SP in KNDy neurons suggests that this peptide is unlikely to be a fundamental component of the primate GnRH pulse generator.


Asunto(s)
Hormona Liberadora de Gonadotropina/metabolismo , Hipotálamo Medio , Kisspeptinas/metabolismo , Hormona Luteinizante/metabolismo , Péptidos/administración & dosificación , Sustancia P/metabolismo , Administración Intravenosa , Animales , Castración , Relación Dosis-Respuesta a Droga , Hipotálamo Medio/citología , Hipotálamo Medio/efectos de los fármacos , Hipotálamo Medio/metabolismo , Macaca mulatta , Masculino , Neuronas/efectos de los fármacos , Neuronas/metabolismo
4.
Mol Hum Reprod ; 20(4): 350-7, 2014 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-24324034

RESUMEN

As the spermatogenesis- and oogenesis-specific basic helix-loop-helix 1 (SOHLH1) transcription factor has been shown to be essential for spermatogonial differentiation in mice, we examined the immunoexpression of this protein in the testis of the rhesus monkey (Macaca mulatta) during puberty, the stage of development when spermatogonial differentiation is initiated in higher primates. Immunopositive SOHLH1 cells were observed only on the basement membrane of the seminiferous cords and tubules. Prior to puberty, essentially 100% of SOHLH1-positive spermatogonia co-expressed the glial cell line-derived neurotrophic factor family receptor alpha 1 (GFRα1), a marker for undifferentiated spermatogonia, and >80% of the immunopositive SOHLH1 cells exhibited only cytoplasmic staining of this transcription factor. Nuclear-only SOHLH1 was found in <10% of spermatogonia in testes from pre-pubertal animals. Puberty was associated with a dramatic and progressive increase in the percentage of immunopositive SOHLH1 cells with nuclear-only staining, and this was associated with (i) a marked reduction in the fraction (∼100-20%) of SOHLH1-positive germ cells co-expressing GFRα1 and (ii) a significant increase in the proportion of SOHLH1-positive spermatogonia that co-expressed the tyrosine kinase receptor (cKIT). Spermatogonia exhibiting nuclear SOHLH1 staining were found to be cKIT positive, but not all cKIT-positive spermatogonia exhibited nuclear SOHLH1 staining. Taken together, these results suggest that, in the monkey, nuclear location of SOHLH1 is closely associated with spermatogonial differentiation.


Asunto(s)
Transporte Activo de Núcleo Celular/genética , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/genética , Macaca mulatta/genética , Espermatogénesis/genética , Espermatogonias/metabolismo , Testículo/metabolismo , Animales , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico/metabolismo , Diferenciación Celular , Regulación del Desarrollo de la Expresión Génica , Receptores del Factor Neurotrófico Derivado de la Línea Celular Glial/genética , Receptores del Factor Neurotrófico Derivado de la Línea Celular Glial/metabolismo , Macaca mulatta/crecimiento & desarrollo , Macaca mulatta/metabolismo , Masculino , Proteínas Proto-Oncogénicas c-kit/genética , Proteínas Proto-Oncogénicas c-kit/metabolismo , Maduración Sexual/genética , Espermatogonias/citología , Espermatogonias/crecimiento & desarrollo , Testículo/citología , Testículo/crecimiento & desarrollo
5.
Neuroendocrinology ; 94(3): 237-45, 2011.
Artículo en Inglés | MEDLINE | ID: mdl-21832818

RESUMEN

Human genetics have revealed that kisspeptin signaling and neurokinin B (NKB) signaling are both required for robust pulsatile gonadotropin-releasing hormone (GnRH) release, and therefore for puberty and maintenance of adult gonadal function. How these two peptides interact to affect GnRH pulse generation remains a mystery. To address the hierarchy of the NKB and kisspeptin signaling pathways that are essential for GnRH release, two experiments were conducted using agonadal, juvenile male monkeys. Pituitary responsiveness to GnRH was first heightened by a pulsatile GnRH infusion to use the in situ pituitary as a bioassay for GnRH release. In the first experiment (n = 3), the kisspeptin receptor (KISS1R) was desensitized by a continuous 99-hour i.v. infusion of kisspeptin-10 (100 µg/h). During the last 4 h of continuous kisspeptin-10 infusion, desensitization of KISS1R was confirmed by failure of an i.v. bolus of kisspeptin-10 to elicit GnRH release. Desensitization of KISS1R was associated with a markedly blunted GnRH response to senktide. The response to senktide was progressively restored during the 72 h following termination of continuous kisspeptin-10. An analogous design was employed in the second experiment (n = 2) to desensitize the NKB receptor (neurokinin 3 receptor, NK3R) by administration of a continuous 48-hour i.v. infusion of senktide (200 µg/h). While a bolus of senktide during the last 3 h of continuous senktide administration failed to elicit GnRH release, thus confirming desensitization of NK3R, the ability of kisspeptin to stimulate GnRH was unimpaired. The foregoing findings support the view that NKB stimulation of GnRH release is upstream from KISS1R.


Asunto(s)
Hormona Liberadora de Gonadotropina/metabolismo , Macaca mulatta , Modelos Animales , Neuroquinina B/fisiología , Orquiectomía , Receptores Acoplados a Proteínas G/metabolismo , Factores de Edad , Animales , Genitales Masculinos/metabolismo , Genitales Masculinos/cirugía , Humanos , Kisspeptinas/administración & dosificación , Kisspeptinas/farmacología , Hormona Luteinizante/metabolismo , Masculino , Neuroquinina B/metabolismo , Orquiectomía/veterinaria , Receptores de Neuroquinina-3/metabolismo , Transducción de Señal/efectos de los fármacos , Transducción de Señal/fisiología
6.
Peptides ; 30(1): 67-75, 2009 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-18662732

RESUMEN

The present article reviews recent studies of monkeys and, in some cases, humans that have been conducted to examine the role of kisspeptin-GPR54 signaling in the regulation of the hypothalamic-pituitary-gonadal axis in higher primates. This area of peptide biology was initiated in 2003 by the discovery that loss of function mutations of GPR54 in man were associated with hypogonadotropic hypogonadism and absent or delayed puberty. Puberty in the monkey, an experimental model commonly used to study this fundamental developmental stage, is first described. This is followed by a review of the role of kisspeptin in the regulation of the postnatal ontogeny of GnRH pulsatility. The roles of kisspeptin in GnRH pulse generation and in the feedback loops governing gonadotropin secretion in primates are then discussed. A brief section on kisspeptin-GPR54 signaling at the pituitary and gonadal levels is also included. The review concludes with a discussion of the phenomenon of GPR54 downregulation by continuous exposure to kisspeptin and its therapeutic implications.


Asunto(s)
Gónadas/fisiología , Sistema Hipotálamo-Hipofisario/fisiología , Macaca mulatta/fisiología , Proteínas Supresoras de Tumor/metabolismo , Animales , Hormona Liberadora de Gonadotropina/metabolismo , Humanos , Hormona Luteinizante/metabolismo , Macaca mulatta/anatomía & histología , Neuronas/metabolismo , Periodicidad , Pubertad/fisiología , Receptores Acoplados a Proteínas G/metabolismo , Receptores de Kisspeptina-1 , Transducción de Señal/fisiología
7.
Endocrinology ; 149(9): 4387-95, 2008 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-18511511

RESUMEN

Kisspeptin is recognized to play a critical role in eliciting the pubertal resurgence of pulsatile GnRH release, the proximal trigger of puberty in higher primates. Expression of the kisspeptin receptor (GPR54) by GnRH neurons indicates a direct action of kisspeptin on the GnRH neuronal network. The purpose of the present study was to examine the distribution of kisspeptin cell bodies in the monkey hypothalamus and to assess the structural basis for the stimulatory action of kisspeptin on the GnRH neuronal network. Three castrated male rhesus monkeys, 39-51 months of age, were deeply anesthetized and their brains perfused transcardially with 4% paraformaldehyde in PBS. Serial 25-microm coronal sections throughout the hypothalamus were prepared, and immunopositive neurons identified using a cocktail of specific primary antibodies (sheep anti-kisspeptin at 1:120,000, and rabbit anti-GnRH at 1:100,000) detected with fluorescently tagged secondary antibodies (antisheep, Alexa Fluor 488; antirabbit, Cy3) in combination with confocal microscopy. Kisspeptin perikarya were found only in the mediobasal hypothalamus (MBH) almost exclusively in the posterior two-thirds of the arcuate nucleus. Surprisingly, kisspeptin-beaded axons made only infrequent contacts with GnRH neurons (kisspeptin and GnRH profiles abutting in a 0.5- to 1.0-mum optical section) in the MBH. In the median eminence, kisspeptin and GnRH axons were found in extensive and intimate association. GnRH contacts on kisspeptin perikarya and dendrites were observed. These findings indicate that nonsynaptic pathways of communication in the median eminence should be considered as a possible mechanism of kisspeptin regulation of GnRH release, and provide an anatomical basis for reciprocal control of kisspeptin neuronal activity by GnRH.


Asunto(s)
Hormona Liberadora de Gonadotropina/metabolismo , Hipotálamo/metabolismo , Neuronas/fisiología , Proteínas Supresoras de Tumor/metabolismo , Animales , Técnica del Anticuerpo Fluorescente , Macaca mulatta , Masculino , Microscopía Confocal , Neuronas/metabolismo , Transmisión Sináptica/fisiología
8.
Endocrinology ; 148(7): 3364-70, 2007 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-17412800

RESUMEN

In agonadal juvenile male monkeys, continuous administration of human metastin 45-54 (hu metastin 45-54) leads to desensitization of its receptor, G protein-coupled receptor 54 (GPR54), and decreased LH. The present study extended this observation to the adult male monkey, a more preclinically relevant model in which robust activity in the hypothalamic-pituitary-testicular axis is present. Continuous iv infusion of hu metastin 45-54 at either 200 or 400 microg/h elicited a marked rise in circulating LH that peaked 2-3 h after initiation of treatment. Thereafter, levels declined, and by 24 h, LH in metastin 45-54-infused animals was similar to control. LH release in response to an iv bolus of hu metastin 45-54 (10-30 microg) during the final 3 h of continuous infusion was truncated or abolished (low and high peptide dose, respectively). GPR54 desensitization by the high-dose metastin 45-54 infusion was associated with compromised pituitary response to a bolus GnRH injection (0.3 microg). LH pulse amplitude and pulse frequency were markedly suppressed during high-dose metastin 45-54 treatment. Surprisingly, the fidelity of the relationship between circulating testosterone (T) and LH was distorted during the high-dose peptide infusion. Thus, for a given concentration of LH, T levels were invariably higher during the high-dose metastin 45-54 infusion than during vehicle, suggesting that the peptide may exert direct actions on the testis to amplify T production. These findings support the notion that GPR54 is desensitized by continuous exposure to ligand, and they raise the possibility of an intratesticular role of GPR54.


Asunto(s)
Sistema Hipotálamo-Hipofisario/efectos de los fármacos , Sistemas Neurosecretores/efectos de los fármacos , Fragmentos de Péptidos/farmacología , Proteínas Supresoras de Tumor/farmacología , Animales , Humanos , Sistema Hipotálamo-Hipofisario/metabolismo , Infusiones Intravenosas , Kisspeptinas , Hormona Luteinizante/sangre , Hormona Luteinizante/metabolismo , Macaca mulatta , Masculino , Sistemas Neurosecretores/metabolismo , Fragmentos de Péptidos/administración & dosificación , Receptores Acoplados a Proteínas G/metabolismo , Testosterona/sangre , Testosterona/metabolismo , Factores de Tiempo , Proteínas Supresoras de Tumor/administración & dosificación , Proteínas Supresoras de Tumor/química
9.
Endocrinology ; 148(4): 1784-96, 2007 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-17218411

RESUMEN

The present study was undertaken to examine the antispermatogenic effect of l-CDB-4022 in the adult male cynomolgus monkey. Monkeys (four per group) were dosed via nasogastric tube for 7 d with l-CDB-4022 at 12.5 mg/kg.d or vehicle (d 0=first day of dosing). Plasma levels of l-CDB-4022 and its deesterified metabolite were nondetectable prior to treatment and in all vehicle-treated monkeys. Peak levels of l-CDB-4022 and its metabolite were observed at 4 h after dosing with steady-state levels apparent around d 4. Sperm concentration and total sperm per ejaculate were decreased to levels below 1x10(6) sperm/ml or sperm/ejaculate in l-CDB-4022-treated monkeys by d 17 and remained suppressed through wk 6. Sperm motility also declined to 0% for 6 wk. Testicular volume was reduced in l-CDB-4022-treated monkeys through d 21. The left testis and epididymis were removed from all monkeys on d 24. At this time, the most mature germ cells in the seminiferous tubules of testes from l-CDB-4022-treated monkeys were either spermatocytes or round spermatids. Immature germ cells, but not mature sperm, were found in the efferent ducts and collapsed epididymal lumen of l-CDB-4022-treated monkeys. A steady recovery in sperm motility, concentration, and total sperm per ejaculate was observed in l-CDB-4022-treated monkeys such that these parameters were not different from those of vehicle-treated monkeys by wk 16. Volume of the remaining testis increased in vehicle- and l-CDB-4022-treated monkeys after hemicastration; however, the increase in l-CDB-4022-treated monkeys was delayed compared with that observed in the vehicle-treated monkeys. The morphology of the remaining testis and epididymis, which were removed on wk 17, was normal. Serum inhibin B levels were increased in l-CDB-4022-treated monkeys during the dosing interval; thereafter serum inhibin B levels declined such that there was no difference between the groups by wk 3. l-CDB-4022 treatment did not affect circulating levels of testosterone, LH, FSH, or estradiol. In conclusion, these data indicate that in the cynomolgus monkey, a representative higher primate, l-CDB-4022 exerts a selective antispermatogenic action, which was reversible under the conditions of this study and thus has potential as a nonhormonal oral male contraceptive.


Asunto(s)
Indenos/administración & dosificación , Oligospermia/inducido químicamente , Oligospermia/rehabilitación , Piperidinas/administración & dosificación , Recuperación de la Función , Administración Oral , Animales , Anticonceptivos Masculinos/administración & dosificación , Evaluación Preclínica de Medicamentos , Epidídimo/anatomía & histología , Epidídimo/efectos de los fármacos , Hormona Folículo Estimulante/sangre , Indenos/farmacocinética , Hormona Luteinizante/sangre , Macaca fascicularis , Masculino , Modelos Biológicos , Piperidinas/farmacocinética , Testículo/anatomía & histología , Testículo/efectos de los fármacos , Testosterona/sangre
10.
Endocrinology ; 147(2): 1007-13, 2006 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-16282350

RESUMEN

The purpose of the present study was to further examine the hypothesis that activation of G protein-coupled receptor 54 (GPR54) signaling at the end of the juvenile phase of primate development is responsible for initiation of gonadarche and the onset of puberty. Accordingly, we determined whether repetitive iv administration of the GPR54 receptor agonist kisspeptin-10 (2 microg as a brief 1-min infusion once every hour for 48 h) to the juvenile male rhesus monkey would prematurely elicit sustained, pulsatile release of hypothalamic GnRH, the neuroendocrine trigger for gonadarche. GnRH release was monitored indirectly by measuring LH secretion from the in situ pituitary, the GnRH responsiveness of which had been heightened before the experiment with an intermittent iv infusion of synthetic GnRH. Agonadal animals (n = 4) were employed to eliminate any confounding and secondary effects of changing feedback signals from the testis. The first brief infusion of kisspeptin-10 evoked an LH discharge that mimicked those produced by GnRH priming, and this was followed by a train of similar LH discharges in response to hourly activation of GPR54 by repetitive kisspeptin-10 administration. Concomitant treatment with a GnRH receptor antagonist, acyline, abolished kisspeptin-10-induced LH release. Repetitive kisspeptin-10 administration also provided a GnRH-dependent signal to FSH secretion. These findings are consistent with the notion that, in primates, the transition from the juvenile (attenuated GnRH release) to pubertal (robust GnRH release) state is controlled by activation of GPR54 resulting from increased expression of hypothalamic KiSS-1 and release of kisspeptin in this region of the brain.


Asunto(s)
Hormona Liberadora de Gonadotropina/metabolismo , Hipotálamo/metabolismo , Oligopéptidos/administración & dosificación , Receptores Acoplados a Proteínas G/metabolismo , Maduración Sexual/fisiología , Animales , Esquema de Medicación , Infusiones Intravenosas , Kisspeptinas , Hormona Luteinizante/metabolismo , Macaca mulatta , Masculino , Flujo Pulsátil , Receptores Acoplados a Proteínas G/efectos de los fármacos , Transducción de Señal/fisiología , Estadísticas no Paramétricas
11.
Endocrinology ; 147(5): 2122-6, 2006 May.
Artículo en Inglés | MEDLINE | ID: mdl-16469799

RESUMEN

The effect of continuous administration of the C-terminal fragment of metastin, the ligand for the G protein-coupled receptor, GPR54, on GnRH-induced LH secretion was examined in three agonadal, juvenile male monkeys whose responsiveness to GnRH was heightened by pretreatment with a chronic pulsatile iv infusion of synthetic GnRH. After bolus injection of 10 microg human (hu) metastin 45-54 (equivalent to kisspeptin 112-121), the GPR54 agonist was infused continuously at a dose of 100 microg/h and elicited a brisk LH response for approximately 3 h. This rise was then followed by a precipitous drop in LH despite continuous exposure of GPR54 to metastin 45-54. On d 4, during the final 3 h of the infusion, single boluses of hu metastin 45-54 (10 microg), N-methyl-DL-aspartic acid (NMDA) (10 mg/kg) and GnRH (0.3 microg) were administered to interrogate each element of the metastin-GPR54-GnRH-GnRH receptor cascade. Although the NMDA and GnRH boluses were able to elicit LH pulses, that of hu metastin 45-54 was not, demonstrating functional integrity of GnRH neurons (NMDA) and GnRH receptors (NMDA and GnRH) but desensitization of GPR54. The desensitization of GPR54 by continuous hu metastin 45-54 administration has therapeutic implications for a variety of conditions currently being treated by GnRH and its analogs, including restoration of fertility in patients with abnormal GnRH secretion (i.e. idiopathic hypogonadotropic hypogonadism and hypothalamic amenorrhea) and selective, reversible suppression of the pituitary-gonadal axis to achieve suppression of gonadal steroids (i.e. precocious puberty, endometriosis, uterine fibroids, and prostate cancer).


Asunto(s)
Proteínas/administración & dosificación , Receptores de Galanina/metabolismo , Animales , Bioensayo , Peso Corporal , Humanos , Infusiones Intravenosas , Kisspeptinas , Ligandos , Hormona Luteinizante/metabolismo , Macaca mulatta , Masculino , Modelos Biológicos , N-Metilaspartato/química , Hipófisis/metabolismo , Neoplasias de la Próstata/metabolismo , Proteínas/química , Receptores Acoplados a Proteínas G , Receptores de Kisspeptina-1 , Receptores LHRH/metabolismo , Factores de Tiempo , Proteínas Supresoras de Tumor
13.
Nat Commun ; 6: 10195, 2015 Dec 16.
Artículo en Inglés | MEDLINE | ID: mdl-26671628

RESUMEN

In primates, puberty is unleashed by increased GnRH release from the hypothalamus following an interval of juvenile quiescence. GWAS implicates Zinc finger (ZNF) genes in timing human puberty. Here we show that hypothalamic expression of several ZNFs decreased in agonadal male monkeys in association with the pubertal reactivation of gonadotropin secretion. Expression of two of these ZNFs, GATAD1 and ZNF573, also decreases in peripubertal female monkeys. However, only GATAD1 abundance increases when gonadotropin secretion is suppressed during late infancy. Targeted delivery of GATAD1 or ZNF573 to the rat hypothalamus delays puberty by impairing the transition of a transcriptional network from an immature repressive epigenetic configuration to one of activation. GATAD1 represses transcription of two key puberty-related genes, KISS1 and TAC3, directly, and reduces the activating histone mark H3K4me2 at each promoter via recruitment of histone demethylase KDM1A. We conclude that GATAD1 epitomizes a subset of ZNFs involved in epigenetic repression of primate puberty.


Asunto(s)
Epigénesis Genética , Factores de Transcripción GATA/genética , Regulación del Desarrollo de la Expresión Génica , Hipotálamo/metabolismo , Pubertad/genética , ARN Mensajero/metabolismo , Animales , Western Blotting , Inmunoprecipitación de Cromatina , Femenino , Técnica del Anticuerpo Fluorescente , Hormona Folículo Estimulante/metabolismo , Factores de Transcripción GATA/metabolismo , Hormona Liberadora de Gonadotropina/metabolismo , Gonadotropinas/metabolismo , Histona Demetilasas/metabolismo , Hibridación Fluorescente in Situ , Kisspeptinas/genética , Kisspeptinas/metabolismo , Hormona Luteinizante/metabolismo , Macaca mulatta , Masculino , Neuroquinina B/genética , Neuroquinina B/metabolismo , Ratas , Ratas Sprague-Dawley , Reacción en Cadena en Tiempo Real de la Polimerasa , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Dedos de Zinc/genética
14.
Endocrinology ; 156(11): 4214-25, 2015 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-26305889

RESUMEN

Women's health disorders such as uterine fibroids and endometriosis are currently treated by GnRH modulators that effectively suppress the hypothalamic-pituitary-gonadal axis. The neurokinin-3 receptor (NK3R) is an alternative target with an important role in the modulation of this axis. In this report, we demonstrate that systemic administration of an NK3R antagonist (ESN364) prolongs the LH interpulse interval in ovarectomized ewes and significantly lowers plasma LH and FSH concentrations in castrated nonhuman primates (Macaca fascicularis). Moreover, daily oral dosing of ESN364 throughout the menstrual cycle in M fascicularis lowered plasma estradiol levels in a dose-dependent manner, although nadir levels of estradiol were maintained well above menopausal levels. Nevertheless, estradiol levels during the follicular phase were sufficiently inhibited at all doses to preclude the triggering of ovulation as evidenced by the absence of the LH surge and failure of a subsequent luteal phase rise in plasma progesterone concentrations, consistent with the absence of normal cycle changes in the uterus. Apart from the point at surge, FSH levels were not altered over the course of the menstrual cycle. These effects of ESN364 were reversible upon cessation of drug treatment. Together these data support the proposed role of neurokinin B-NK3R signaling in the control of pulsatile GnRH secretion. Furthermore, in contrast to GnRH antagonists, NK3R antagonists induce a partial suppression of estradiol and thereby offer a viable therapeutic approach to the treatment of ovarian sex hormone disorders with a mitigated risk of menopausal-like adverse events in response to long-term drug exposure.


Asunto(s)
Estradiol/sangre , Hormona Luteinizante/sangre , Ciclo Menstrual/efectos de los fármacos , Progesterona/sangre , Receptores de Neuroquinina-3/antagonistas & inhibidores , Animales , Castración , Femenino , Hormona Folículo Estimulante/sangre , Macaca fascicularis , Masculino , Ciclo Menstrual/sangre , Ovinos
15.
Endocrinology ; 145(11): 5115-20, 2004 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-15308611

RESUMEN

This study examined, in adult monkeys, the role that gonadotropin-independent mechanisms play in compensation of testosterone (T) secretion by the testis that remains after unilateral orchidectomy (UO). We employed a model (testicular clamp), in which endogenous gonadotropin secretion was abolished with a GnRH receptor antagonist, and the gonadotropin drive to the testes was concomitantly replaced with an invariant iv pulsatile infusion of recombinant human LH and FSH (1-min pulse every 2.5 h: LH, 0.08-0.12 IU/kg.pulse; FSH, 0.12-0.32 IU/kg.pulse) that provided the Leydig cells with a physiological stimulus. Within 5 h of UO (n = 5), circulating T concentrations had declined to 43% of pre-UO levels. By d 4, however, loss of the first testis was partially compensated, as reflected by the finding that circulating T had reached a plateau of 67% of the pre-UO level, where it remained for the duration of the study (39 d). That the recovery in circulating T was the result of increased T secretion by the remaining testis was suggested by the finding that the pulsatile pattern and decay of T during the intergonadotropin pulse interval before and after UO were indistinguishable. Interestingly, inhibin B production by the remaining testis also showed a delayed, albeit, minor, compensation (13% on d 10-11; P > 0.05) after loss of the first testis. These results suggest that compensation in T production by the remaining testis after UO in adult monkeys may be achieved in part by a gonadotropin-independent mechanism that probably involves direct neural inputs to the primate testis.


Asunto(s)
Adaptación Fisiológica/fisiología , Gonadotropinas/sangre , Orquiectomía , Testículo/metabolismo , Testosterona/sangre , Factores de Edad , Animales , Hormona Folículo Estimulante/sangre , Gonadotropinas/farmacología , Inhibinas/sangre , Hormona Luteinizante/sangre , Macaca mulatta , Masculino , Testículo/cirugía , Testosterona/metabolismo
16.
Endocrinology ; 144(4): 1175-85, 2003 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-12639898

RESUMEN

This study examined the relative role of FSH and LH in governing testicular inhibin B secretion in the rhesus monkey. Adult male monkeys, rendered hypogonadotropic and hypogonadal by administration of a GnRH receptor antagonist (acyline), were implanted with testosterone (T)-filled or empty capsules. Following T-induced restoration of spermatogenesis, both groups received recombinant human FSH and vehicle for 12 d. Juvenile male monkeys received an 11-d infusion of single-chain recombinant human LH and recombinant human FSH, either alone or in combination. In adults, chronic hypogonadotropism resulted in a modest reduction of circulating inhibin B levels, which was more than fully reversed by FSH. In the presence of T, which exerted a marked suppression in inhibin B secretion, FSH restored inhibin B levels only to those observed before acyline treatment. In juveniles, treatment with single-chain recombinant human LH led to a suppression of inhibin B secretion and curtailed the FSH-induced stimulation of this testicular hormone. The T-induced decrease in inhibin B secretion was associated with suppression in inhibin-beta(B) mRNA levels, but FSH stimulation of inhibin B secretion occurred in the absence of clear changes in expression of this subunit gene. These findings indicate that inhibin B secretion by the monkey testis is governed by the inhibitory and stimulatory action of LH and FSH, respectively. The action of LH is presumably indirect and likely mediated by T inhibition of inhibin-beta(B) gene expression. The molecular basis of the stimulatory action of FSH on inhibin B secretion requires further study.


Asunto(s)
Hormona Folículo Estimulante/farmacología , Inhibinas/metabolismo , Hormona Luteinizante/farmacología , Testículo/metabolismo , Factores de Edad , Animales , Expresión Génica/efectos de los fármacos , Hormonas Esteroides Gonadales/farmacología , Hipogonadismo/tratamiento farmacológico , Hipogonadismo/metabolismo , Inhibinas/genética , Macaca mulatta , Masculino , Oligopéptidos/farmacología , Orquiectomía , Testículo/efectos de los fármacos , Testosterona/farmacología
17.
Spermatogenesis ; 4(2): e996025, 2014.
Artículo en Inglés | MEDLINE | ID: mdl-26413400

RESUMEN

Evaluation of testicular functions (production of sperm and androgens) is an important aspect of preclinical safety assessment and testicular toxicity is comparatively far more common than ovarian toxicity. This chapter focuses (1) on the histological sequelae of disturbed reproductive endocrinology in rat, dog and nonhuman primates and (2) provides a review of our current understanding of the roles of gonadotropins and androgens. The response of the rodent testis to endocrine disturbances is clearly different from that of dog and primates with different germ cell types and spermatogenic stages being affected initially and also that the end-stage spermatogenic involution is more pronounced in dog and primates compared to rodents. Luteinizing hormone (LH)/testosterone and follicle-stimulating hormone (FSH) are the pivotal endocrine factors controlling testicular functions. The relative importance of either hormone is somewhat different between rodents and primates. Generally, however, both LH/testosterone and FSH are necessary for quantitatively normal spermatogenesis, at least in non-seasonal species.

18.
Endocrinology ; 154(5): 1845-53, 2013 May.
Artículo en Inglés | MEDLINE | ID: mdl-23525220

RESUMEN

Puberty in primates is timed by 2 hypothalamic events: during late infancy a decline in pulsatile GnRH release occurs, leading to a hypogonadotropic state that maintains quiescence of the prepubertal gonad; and in late juvenile development, pulsatile GnRH release is reactivated and puberty initiated, a phase of development that is dependent on kisspeptin signaling. In the present study, we determined whether the arrest of GnRH pulsatility in infancy was associated with a change in kisspeptin expression in the mediobasal hypothalamus (MBH). Kisspeptin was determined using immunohistochemistry in coronal hypothalamic sections from agonadal male rhesus monkeys during early infancy when GnRH release as reflected by circulating LH concentrations was robust and compared with that in juveniles in which GnRH pulsatility was arrested. The distribution of immunopositive kisspeptin neurons in the arcuate nucleus of the MBH of infants was similar to that previously reported for adults. Kisspeptin cell body number was greater in infants compared with juveniles, and at the middle to posterior level of the arcuate nucleus, this developmental difference was statistically significant. Neurokinin B in the MBH exhibited a similar distribution to that of kisspeptin and was colocalized with kisspeptin in approximately 60% of kisspeptin perikarya at both developmental stages. Intensity of GnRH fiber staining in the median eminence was robust at both stages. These findings indicate that the switch that shuts off pulsatile GnRH release during infancy and that guarantees the subsequent quiescence of the prepubertal gonad involves a reduction in a stimulatory kisspeptin tone to the GnRH neuronal network.


Asunto(s)
Núcleo Arqueado del Hipotálamo/metabolismo , Hormona Liberadora de Gonadotropina/metabolismo , Kisspeptinas/metabolismo , Hormona Luteinizante/sangre , Neuronas/metabolismo , Maduración Sexual/fisiología , Animales , Animales Recién Nacidos/sangre , Animales Recién Nacidos/crecimiento & desarrollo , Animales Recién Nacidos/metabolismo , Núcleo Arqueado del Hipotálamo/crecimiento & desarrollo , Regulación hacia Abajo , Hormona Liberadora de Gonadotropina/sangre , Hipotálamo/crecimiento & desarrollo , Hipotálamo/metabolismo , Hormona Luteinizante/análisis , Macaca mulatta , Masculino , Orquiectomía , Concentración Osmolar , Flujo Pulsátil
19.
Endocrinology ; 151(9): 4494-503, 2010 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-20573725

RESUMEN

Human genetics indicate that kisspeptin and neurokinin B (NKB) signaling are necessary for generating pulsatile LH release and therefore for initiation of puberty and maintaining gonadal function. In the present study, male monkeys were employed to examine 1) whether activation of the NKB receptor (NK3R) is associated with GnRH release, and 2) hypothalamic localization of these peptides using immunofluorescence histochemistry. Agonadal juveniles, in which pituitary responsiveness to GnRH was heightened by GnRH priming, were employed to indirectly examine GnRH-releasing actions of NK3R and kisspeptin receptor agonists by tracking LH after their i.v. injection. Castrated adults were used for immunohistochemistry. Single i.v. injections of NKB or senktide (an NK3R agonist) elicited robust LH discharges that were abolished by GnRH receptor antagonism (acyline) confirming the ligands' hypothalamic action. Intermittent infusion of senktide (1-min pulse every hour for 4 h), in contrast to that of kisspeptin, failed to sustain pulsatile GnRH release. Repetitive senktide injections did not compromise the GnRH-releasing action of kisspeptin. NKB and kisspeptin were colocalized in perikarya of the arcuate nucleus and in axonal projections to the median eminence, confirming earlier findings in sheep. These results are consistent with the human genetics, and indicate that although brief activation of NK3R stimulates GnRH release, repetitive stimulation of this pathway, in contrast to that of kisspeptin receptor, fails to sustain pulsatile GnRH release. In addition, the data provide a platform for future elucidation of the interactions between NKB and kisspeptin that are required for generating pulsatile GnRH release in primates.


Asunto(s)
Núcleo Arqueado del Hipotálamo/metabolismo , Hormona Liberadora de Gonadotropina/metabolismo , Neuroquinina B/metabolismo , Oligopéptidos/metabolismo , Animales , Núcleo Arqueado del Hipotálamo/efectos de los fármacos , Interacciones Farmacológicas , Humanos , Inmunohistoquímica , Inyecciones Intravenosas , Kisspeptinas , Hormona Luteinizante/sangre , Hormona Luteinizante/metabolismo , Macaca mulatta , Masculino , Microscopía Confocal , Neuroquinina B/farmacología , Oligopéptidos/farmacología , Orquiectomía , Fragmentos de Péptidos/farmacología , Quinolinas/farmacología , Receptores Acoplados a Proteínas G/agonistas , Receptores Acoplados a Proteínas G/antagonistas & inhibidores , Receptores Acoplados a Proteínas G/metabolismo , Receptores LHRH/antagonistas & inhibidores , Receptores de Neuroquinina-3/agonistas , Receptores de Neuroquinina-3/antagonistas & inhibidores , Receptores de Neuroquinina-3/metabolismo , Sustancia P/análogos & derivados , Sustancia P/farmacología
20.
Biol Reprod ; 79(1): 93-9, 2008 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-18367678

RESUMEN

Testosterone shows circadian rhythms in monkeys with low serum levels in the morning hours. The decline relies on a diminished frequency of LH pulses. Inhibin B shows no diurnal patterns. In elderly men, the diurnal rhythm of testosterone is blunted and inhibin levels fall. Here we explore whether aging exerts similar effects in the rhesus monkey. We collected blood samples from groups of young (6-9 yr) and old (12-16 yr) male rhesus monkeys at 20-min intervals for a period of 24 h under remote sampling via a venous catheter. We determined moment-to-moment changes in plasma levels of testosterone, FSH, and LH by RIA, and of inhibin B by ELISA. We found significant diurnal patterns of testosterone in both groups. The circadian rhythm in testosterone was enhanced in older monkeys. Testosterone levels and pulse frequencies dropped significantly below those of young monkeys during midday hours. Diminished pulse frequency of LH appeared to be responsible for the midday testosterone decrease in old monkeys, while LH and testosterone pulse frequency did not change in young monkeys at corresponding time points. Old monkeys showed extended periods of LH-pulse quiescence in the morning and midday hours. Inhibin B and FSH levels were generally lower in old monkeys compared with the young group, but neither inhibin B nor FSH showed circadian rhythms. We conclude from these data that old rhesus monkeys have a more prominent circadian rhythm of LH and testosterone resulting from an extended midday period of quiescence in the hypothalamus-pituitary-gonadal axis.


Asunto(s)
Envejecimiento/sangre , Ritmo Circadiano/fisiología , Gonadotropinas/sangre , Inhibinas/sangre , Testosterona/sangre , Factores de Edad , Animales , Macaca mulatta , Masculino , Flujo Pulsátil
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