My 28 Days - a global digital women's health initiative for evaluation and management of secondary amenorrhea: case report and literature review.

There is a need to close the gap between knowledge and action in health care. Effective care requires a convenient and reliable distribution process. As global internet and mobile communication increase capacity, innovative approaches to digital health education platforms and care delivery are feasible. We report the case of a young African woman who developed acute secondary amenorrhea at age 18. Subsequently, she experienced a 10-year delay in the diagnosis of the underlying cause. A global digital medical hub focused on women's health and secondary amenorrhea could reduce the chance of such mismanagement. Such a hub would establish more efficient information integration and exchange processes to better serve patients, family caregivers, health care providers, and investigators. Here, we show proof of concept for a global digital medical hub for women's health. First, we describe the physiological control systems that govern the normal menstrual cycle, and review the pathophysiology and management of secondary amenorrhea. The symptom may lead to broad and profound health implications for the patient and extended family members. In specific situations, there may be significant morbidity related to estradiol deficiency: (1) reduced bone mineral density, 2) cardiovascular disease, and 3) cognitive decline. Using primary ovarian insufficiency (POI) as the paradigm condition, the Mary Elizabeth Conover Foundation has been able to address the specific global educational needs of these women. The Foundation did this by creating a professionally managed Facebook group specifically for these women. POI most commonly presents with secondary amenorrhea. Here we demonstrate the feasibility of conducting a natural history study on secondary amenorrhea with international reach to be coordinated by a global digital medical hub. Such an approach takes full advantage of internet and mobile device communication systems. We refer to this global digital women's health initiative as My 28 Days®.

The fifty years following the discovery of gonadotropin-releasing hormone.

This article highlights key milestones in GnRH research that have occurred in the 50 plus years since the discovery of the decapeptide. It is by no means exhaustive and inevitably reflects our limitations and idiosyncratic perspectives.

Neurobiology of puberty and its disorders.

Puberty, which in humans is considered to include both gonadarche and adrenarche, is the period of becoming capable of reproducing sexually and is recognized by maturation of the gonads and development of secondary sex characteristics. Gonadarche referring to growth and maturation of the gonads is fundamental to puberty since it encompasses increased gonadal steroid secretion and initiation of gametogenesis resulting from enhanced pituitary gonadotropin secretion, triggered in turn by robust pulsatile GnRH release from the hypothalamus. This chapter reviews the development of GnRH pulsatility from before birth until the onset of puberty. In humans, GnRH pulse generation is restrained during childhood and juvenile development. This prepubertal hiatus in hypothalamic activity is considered to result from a neurobiological brake imposed upon the GnRH pulse generator resident in the infundibular nucleus. Reactivation of the GnRH pulse generator initiates pubertal development. Current understanding of the genetics and physiology of the brake will be discussed, as will hypotheses proposed to account for timing the resurgence in pulsatile GnRH and initiation of puberty. The chapter ends with a discussion of disorders associated with precocious or delayed puberty with a focus on those with etiologies attributed to aberrant GnRH neuron anatomy or function. A pediatric approach to patients with pubertal disorders is provided and contemporary treatments for both precocious and delayed puberty outlined.

Intertwined reproductive endocrinology: Puberty and polycystic ovary syndrome.

Polycystic ovary syndrome (PCOS) is a heterogeneous familial disorder often emerging during the peri-pubertal years concomitantly with the onset of gonadarche and adrenarche. Both gonadarche and PCOS reflect functional changes in the hypothalamic-pituitary-ovarian axis. During this transition, normal girls manifest features consistent with PCOS such as irregular menses, mild hyperandrogenism, and multi-follicular ovary morphology. Themes common to puberty and PCOS, neuroendocrine features, androgen exposure, and insulin sensitivity, will be considered to address the possibility that PCOS interferes with the normal pubertal transition.

The neurobiological mechanism underlying hypothalamic GnRH pulse generation: the role of kisspeptin neurons in the arcuate nucleus.

This review recounts the origins and development of the concept of the hypothalamic gonadotropin-releasing hormone (GnRH) pulse generator. It starts in the late 1960s when striking rhythmic episodes of luteinizing hormone secretion, as reflected by circulating concentrations of this gonadotropin, were first observed in monkeys and ends in the present day. It is currently an exciting time witnessing the application, primarily to the mouse, of contemporary neurobiological approaches to delineate the mechanisms whereby Kiss1/NKB/Dyn (KNDy) neurons in the arcuate nucleus of the hypothalamus generate and time the pulsatile output of kisspeptin from their terminals in the median eminence that in turn dictates intermittent GnRH release and entry of this decapeptide into the primary plexus of the hypophysial portal circulation. The review concludes with an examination of questions that remain to be addressed.

Gonadotrophin-mediated miRNA expression in testis at onset of puberty in rhesus monkey: predictions on regulation of thyroid hormone activity and DLK1-DIO3 locus.

Molecular mechanisms responsible for the initiation of primate spermatogenesis remain poorly characterized. Previously, 48 h stimulation of the testes of three juvenile rhesus monkeys with pulsatile LH and FSH resulted in down-regulation of a cohort of genes recognized to favor spermatogonia stem cell renewal. This change in genetic landscape occurred in concert with amplification of Sertoli cell proliferation and the commitment of undifferentiated spermatogonia to differentiate. In this report, the non-protein coding small RNA transcriptomes of the same testes were characterized using RNA sequencing: 537 mature micro-RNAs (miRNAs), 322 small nucleolar RNAs (snoRNAs) and 49 small nuclear RNAs (snRNAs) were identified. Pathway analysis of the 20 most highly expressed miRNAs suggested that these transcripts contribute to limiting the proliferation of the primate Sertoli cell during juvenile development. Gonadotrophin treatment resulted in differential expression of 35 miRNAs, 12 snoRNAs and four snRNA transcripts. Ten differentially expressed miRNAs were derived from the imprinted delta-like homolog 1-iodothyronine deiodinase 3 (DLK1-DIO3) locus that is linked to stem cell fate decisions. Four gonadotrophin-regulated expressed miRNAs were predicted to trigger a local increase in thyroid hormone activity within the juvenile testis. The latter finding leads us to predict that, in primates, a gonadotrophin-induced selective increase in testicular thyroid hormone activity, together with the established increase in androgen levels, at the onset of puberty is necessary for the normal timing of Sertoli cell maturation, and therefore initiation of spermatogenesis. Further examination of this hypothesis requires that peripubertal changes in thyroid hormone activity of the testis of a representative higher primate be determined empirically.

The testicular transcriptome associated with spermatogonia differentiation initiated by gonadotrophin stimulation in the juvenile rhesus monkey (Macaca mulatta).

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.

Estradiol Upregulates Kisspeptin Expression in the Preoptic Area of both the Male and Female Rhesus Monkey (Macaca mulatta): Implications for the Hypothalamic Control of Ovulation in Highly Evolved Primates.

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.

The Increase in Signaling by Kisspeptin Neurons in the Preoptic Area and Associated Changes in Clock Gene Expression That Trigger the LH Surge in Female Rats Are Dependent on the Facilitatory Action of a Noradrenaline Input.

In rodents, kisspeptin neurons in the rostral periventricular area of the third ventricle (RP3V) of the preoptic area are considered to provide a major stimulatory input to the GnRH neuronal network that is responsible for triggering the preovulatory LH surge. Noradrenaline (NA) is one of the main modulators of GnRH release, and NA fibers are found in close apposition to kisspeptin neurons in the RP3V. Our objective was to interrogate the role of NA signaling in the kisspeptin control of GnRH secretion during the estradiol induced LH surge in ovariectomized rats, using prazosin, an α1-adrenergic receptor antagonist. In control rats, the estradiol-induced LH surge at 17 hours was associated with a significant increase in GnRH and kisspeptin content in the median eminence with the increase in kisspeptin preceding that of GnRH and LH. Prazosin, administered 5 and 3 hours prior to the predicted time of the LH surge truncated the LH surge and abolished the rise in GnRH and kisspeptin in the median eminence. In the preoptic area, prazosin blocked the increases in Kiss1 gene expression and kisspeptin content in association with a disruption in the expression of the clock genes, Per1 and Bmal1. Together these findings demonstrate for the first time that NA modulates kisspeptin synthesis in the RP3V through the activation of α1-adrenergic receptors prior to the initiation of the LH surge and indicate a potential role of α1-adrenergic signaling in the circadian-controlled pathway timing of the preovulatory LH surge.

The Distribution of Substance P and Kisspeptin in the Mediobasal Hypothalamus of the Male Rhesus Monkey and a Comparison of Intravenous Administration of These Peptides to Release GnRH as Reflected by LH Secretion.

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.

Epigenetic regulation of puberty via Zinc finger protein-mediated transcriptional repression.

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.

The NK3 Receptor Antagonist ESN364 Interrupts Pulsatile LH Secretion and Moderates Levels of Ovarian Hormones Throughout the Menstrual Cycle.

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.

Application of three-dimensional culture systems to study mammalian spermatogenesis, with an emphasis on the rhesus monkey (Macaca mulatta).

In vitro culture of spermatogonial stem cells (SSCs) has generally been performed using two-dimensional (2D) culture systems; however, such cultures have not led to the development of complete spermatogenesis. It seems that 2D systems do not replicate optimal conditions of the seminiferous tubules (including those generated by the SSC niche) and necessary for spermatogenesis. Recently, one of our laboratories has been able to induce proliferation and differentiation of mouse testicular germ cells to meiotic and postmeiotic stages including generation of sperm in a 3D soft agar culture system (SACS) and a 3D methylcellulose culture system (MCS). It was suggested that SACS and MCS form a special 3D microenvironment that mimics germ cell niche formation in the seminiferous tubules, and thus permits mouse spermatogenesis in vitro. In this review, we (1) provide a brief overview of the differences in spermatogenesis in rodents and primates, (2) summarize data related to attempts to generate sperm in vitro, (3) report for the first time formation of colonies/clusters of cells and differentiation of meiotic (expression of CREM-1) and postmeiotic (expression of acrosin) germ cells from undifferentiated spermatogonia isolated from the testis of prepubertal rhesus monkeys and cultured in SACS and MCS, and (4) indicate research needed to optimize 3D systems for in vitro primate spermatogenesis and for possible future application to man.

60 YEARS OF NEUROENDOCRINOLOGY: The hypothalamo-pituitary-gonadal axis.

This review provides an outline of how our understanding of the neuroendocrine control of the hypothalamo-pituitary-gonadal axis has evolved since the publication of Geoffrey Harris' renowned monograph in 1955. Particular attention is directed to the neurobiology underlying pulsatile GnRH release from the hypothalamus, the neuroendocrine control of ovarian cycles, puberty and seasonality of gonadal function, and to ideas that have emerged as a result of examining the relationship between growth and the reproductive axis. The review closes with i) a brief discussion of how knowledge gained as a result of pursuing the early hypotheses of Harris has led to major clinical and therapeutic applications, and ii) a personal glimpse into the future of research in this fascinating area of biology.

Neuroendocrine control of the onset of puberty.

This chapter is based on the Geoffrey Harris Memorial Lecture presented at the 8th International Congress of Neuroendocrinology, which was held in Sydney, August 2014. It provides the development of our understanding of the neuroendocrine control of puberty since Harris proposed in his 1955 monograph (Harris, 1955) that "a major factor responsible for puberty is an increased rate of release of pituitary gonadotrophin" and posited "that a neural (hypothalamic) stimulus, via the hypophysial portal vessels, may be involved." Emphasis is placed on the neurobiological mechanisms governing puberty in highly evolved primates, although an attempt is made to reverse translate a model for the timing of puberty in man and monkey to non-primate species.

Spermatogonial SOHLH1 nucleocytoplasmic shuttling associates with initiation of spermatogenesis in the rhesus monkey (Macaca mulatta).

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.

Kisspeptin and puberty in mammals.

Since the discovery of the G-protein coupled receptor 54 (kisspeptin receptor) and its ligand, kisspeptin, our understanding of the neurobiological mechanisms that govern the pituitary-gonadal axis has evolved dramatically. In this chapter, we have reviewed progress regarding the relationship between kisspeptin and puberty, and have proposed a novel hypothesis for the role of kisspeptin signaling in the onset of this crucial developmental event. According to this hypothesis, although kisspeptin neurons in the arcuate nucleus (ARC) are critical for puberty, this is simply because these cells are an integral component of the hypothalamic GnRH pulse generating mechanism that drives intermittent release of the decapeptide, as an increase in GnRH is obligatory for the onset of puberty. In our model, ARC kisspeptin neurons play no "regulatory" role in controlling the timing of puberty. Rather, as a component of the neural network responsible for GnRH pulse generation, they subserve upstream regulatory mechanisms that are responsible for the timing of puberty.

The decline in pulsatile GnRH release, as reflected by circulating LH concentrations, during the infant-juvenile transition in the agonadal male rhesus monkey (Macaca mulatta) is associated with a reduction in kisspeptin content of KNDy neurons of the arcuate nucleus in the hypothalamus.

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.

Insufficient androgen and FSH signaling may be responsible for the azoospermia of the infantile primate testes despite exposure to an adult-like hormonal milieu.

BACKGROUND: In humans, as well as in other higher primates, the infantile testis is exposed to an adult-like hormonal milieu, but spermatogenesis is not initiated at this stage of primate development. In the present study, we examined the molecular basis of this intriguing infertile state of the primate testis. METHODS: The integrity of androgen receptor (AR) and FSH receptor (FSHR) signaling pathways in primary cultures of Sertoli cells (Scs) harvested from azoospermic infant and spermatogenic pubertal monkey testes were investigated under identical in vitro hormonal conditions. In order to synchronously harvest Scs from early pubertal testis, the activation of testicular puberty was timed experimentally by prematurely initiating gonadotrophin secretion in juvenile animals with an intermittent infusion of gonadotrophin-releasing hormone. RESULTS: While qRT-PCR demonstrated that AR and FSHR mRNA expression in Scs from infant and pubertal testes were comparable, androgen-binding and FSH-mediated cAMP production by infant Scs was extremely low. Compromised AR and FSHR signaling in infant Scs was further supported by the finding that testosterone (T) and FSH failed to augment the expression of the T responsive gene, claudin 11, and the FSH responsive genes, inhibin-ßB, stem cell factor (SCF) and glial cell line-derived neurotrophic factor (GDNF) in Scs harvested at this stage of development. CONCLUSION: These results indicate that compromised AR and FSHR signaling pathways in Scs underlie the inability of the infant primate testis to respond to an endogenous hormonal milieu that later in development, at the time puberty, stimulates the initiation of spermatogenesis. This finding may have relevance to some forms of idiopathic infertility in men.