Lateral ventricle injection of orexin-A ameliorates central precocious puberty in rat via inhibiting the expression of MEG3

Background: Central precocious puberty (CPP) is characterized as increasing gonadotropin-releasing hormone (GnRH) release. Orexin-A has also been shown to affect GnRH release. However, there are few reports about the effect of orexin A on the treatment of CPP. Methods: After establishing the precocious puberty model, the rats were divided into four groups: normal control, precocious puberty rats, precocious puberty rats treated with normal saline and precocious puberty rats treated with orexin-A. The vaginal opening time, second estrus cycle, ovarian index and uterus index of rats in each group were detected. qRT-PCR was performed to examine the expression of MEG3 and kisspeptin in rats. HT22 cells were transfected with pcDNA-MEG3 to detect the expression of Kisspeptin. Results: In this study, we found that orexin-A not only delayed the day of vaginal opening and regular estrus cycle days but also decreased the ovarian index and uterus index in rats with CPP. In addition, orexin-A reversed the up-regulation of MEG3 and kisspeptin in rats with CPP. In HT22 cells, the mRNA and protein level of kisspeptin were enhanced by pcDNA-MEG3. Conclusion: Our results suggest that orexin-A ameliorates central precocious puberty in rat and MEG3 might be involved in this effect, suggesting that MEG3 might be a novel target in treating central precocious puberty.     

Sex differences in circadian timing systems: Implications for disease

Virtually every eukaryotic cell has an endogenous circadian clock and a biological sex. These cell-based clocks have been conceptualized as oscillators whose phase can be reset by internal signals such as hormones, and external cues such as light. The present review highlights the inter-relationship between circadian clocks and sex differences. In mammals, the suprachiasmatic nucleus (SCN) serves as a master clock synchronizing the phase of clocks throughout the body. Gonadal steroid receptors are expressed in almost every site that receives direct SCN input. Here we review sex differences in the circadian timing system in the hypothalamic–pituitary–gonadal axis (HPG), the hypothalamic–adrenal–pituitary (HPA) axis, and sleep–arousal systems. We also point to ways in which disruption of circadian rhythms within these systems differs in the sexes and is associated with dysfunction and disease. Understanding sex differentiated circadian timing systems can lead to improved treatment strategies for these conditions.     

Arg-Phe-amide-related peptides influence gonadotropin-releasing hormone neurons

The hypothalamic Arg-Phe-amide-related peptides, gonadotropin-inhibitory hormone and orthologous mammalian peptides of Arg-Phe-amide, may be important regulators of the hypothalamus-pituitary-gonadal reproductive axis. These peptides may modulate the effects of kisspeptins because they are presently recognized as the most potent activators of the hypothalamus-pituitary-gonadal axis. However, their effects on gonadotropin-releasing hormone neurons have not been investigated. In the current study, the GT1-7 cell line-expressing gonadotropin-releasing hormone was used as a model to explore the effects of Arg-Phe- amide-related peptides on kisspeptin activation. Intracellular calcium concentration was quantified using the calcium-sensitive dye, fura-2 acetoxymethyl ester. Gonadotropin-releasing hormone released into the medium was detected via enzyme-linked immunosorbent assay. Results showed that 100 nmol/L kisspeptin-10 significantly increased gonadotropin-releasing hormone levels (at 120 minutes of exposure) and intracellular calcium concentrations. Co-treatment of kisspeptin with 1 μmol/L gonadotropin-inhibitory hormone or 1 μmol/L Arg-Phe-amide-related peptide-1 significantly attenuated levels of kisspeptin-induced gonadotropin-releasing hormone but did not affect kisspeptin-induced elevations of intracellular calcium concentration. Overall, the results suggest that gonadotropin-inhibitory hormone and Arg-Phe-amide-related peptide-1 may have inhibitory effects on kisspeptin-activated gonadotropin-releasing hormone neurons independent of the calcium signaling pathway.     

健康男性血浆kisspeptin与迟发性性腺功能减退关系研究

目的探讨健康男性血浆kisspeptin与更年期发生的关系。方法对上海市宝山区99名健康志愿者进行PADAM问卷调查、外周血激素测定,分析血浆kisspeptin与年龄、更年期发生及激素间的关系。结果成年男性99名血浆kisspeptin随年龄增加而增长,PADAM症状阳性率也随年龄增高。PADAM症状阳性组与阴性组间年龄差异有统计学意义(P<0.01),而kisspeptin差异无统计学意义(P>0.05)。血浆kisspeptin与年龄(P<0.01)、雌二醇(E2)(P=0.039)密切相关,年龄是血浆kisspeptin变化的重要影响因素(P=0.004)。结论血浆kisspeptin与更年期发生无明显关系,提示成年后kisspeptin在HPG轴中的作用可能不及青春期启动时重要。     

Ovarian Regulation of Kisspeptin Neurones in the Arcuate Nucleus of the Rhesus Monkey (Macaca mulatta)

Tonic gonadotrophin secretion throughout the menstrual cycle is regulated by the negative‐feedback actions of ovarian oestradiol (E₂) and progesterone. Although kisspeptin neurones in the arcuate nucleus (ARC) of the hypothalamus appear to play a major role in mediating these feedback actions of the steroids in nonprimate species, this issue has been less well studied in the monkey. In the present study, we used immunohistochemistry and in situ hybridisation to examine kisspeptin and KISS1 expression, respectively, in the mediobasal hypothalamus (MBH) of adult ovariectomised (OVX) rhesus monkeys. We also examined kisspeptin expression in the MBH of ovarian intact females, and the effect of E₂, progesterone and E₂ + progesterone replacement on KISS1 expression in OVX animals. Kisspeptin or KISS1 expressing neurones and pronounced kisspeptin fibres were readily identified throughout the ARC of ovariectomised monkeys but, on the other hand, in intact animals, kisspeptin cell bodies were small in size and number and only fine fibres were observed. Replacement of OVX monkeys with physiological levels of E₂, either alone or with luteal phase levels of progesterone, abolished KISS1 expression in the ARC. Interestingly, progesterone replacement alone for 14 days also resulted in a significant down‐regulation of KISS1 expression. These findings support the view that, in primates, as in rodents and sheep, kisspeptin signalling in ARC neurones appears to play an important role in mediating the negative‐feedback action of E₂ on gonadotrophin secretion, and also indicate the need to study further their regulation by progesterone.     

Kisspeptin对女性生殖功能调控机制的研究进展

在女性生殖活动中,kisspeptin参与调控下丘脑-垂体-性腺(HPG)轴的功能,并通过介导雌激素的正负反馈调节促性腺激素释放激素(Gn RH)的分泌,解除促性腺激素抑制激素(Gn IH)对HPG轴的负性调控作用,继而调节促性腺激素(Gn)的分泌及类固醇甾体激素的分泌,并与多种卵巢功能异常疾病的病理过程密切相关。Kisspeptin可调控卵泡发育和排卵发生,并影响子宫内膜容受性的形成。此外,在辅助生殖技术(ART)过程中kisspeptin可代替传统h CG扳机,并降低接受ART治疗的不孕症女性卵巢过度刺激综合征(OHSS)的发生。     

Kisspeptin levels in idiopathic hypogonadotropic hypogonadism diagnosed male patients and its relation with glucose-insulin dynamic

Male hypogonadism is defined as the deficiency of testosterone or sperm production synthesized by testicles or the deficiency of both. The reasons for hypogonadism may be primary, meaning testicular or secondary, meaning hypothalamohypophyseal. In hypogonadotropic hypogonadism (HH), there is indeficiency in gonadotropic hormones due to hypothalamic or hypophyseal reasons. Gonadotropin-releasing hormone (GnRH) is an important stimulant in releasing follicular stimulant hormone (FSH), mainly luteinizing hormone (LH). GnRH omitted is under the effect of many hormonal or stimulating factors. Kisspeptin is present in many places of the body, mostly in hypothalamic anteroventral periventricular nucleus and arcuate nucleus. Kisspeptin has a suppressor effect on the metastasis of many tumors such as breast cancer and malign melanoma metastases, and is called “metastin” for this reason. Kisspeptin is a strong stimulant of GnRH. In idiopathic hypogonadotropic hypogonadism (IHH) etiology, there is gonadotropic hormone release indeficiency which cannot be clearly described.

A total of 30 male hypogonatropic hypogonadism diagnosed patients over 30 years of age who have applied to Haydarpasa Education Hospital Endocrinology and Metabolic Diseases Service were included in the study. Compared to the control group, the effect of kisspeptin on male patients with hypogonatropic hypogonadism and on insulin resistance developing in hypogonadism patients was investigated in our study.

A statistically significant difference was detected between average kisspeptin measurements of the groups (p?kisspeptin and LH/FSH levels. Although a positive low relation was detected between kisspeptin measurements of patient group cases and homeostasis model assessment of insulin resistance (HOMA-IR) measurements, this relation was statistically insignificant. When the patient and control groups were compared for HOMA-IR, no statistically significant difference was detected.

The reason for high kisspeptin levels in the patient group compared to the control group makes us consider that there may be a GPR54 resistance or GnRH neuronal transfer pathway defect. When patients and control groups were compared for HOMA-IR, the difference was not statistically significant. It is considered that kisspeptin is one of the reasons for hypogonatropic hypogonadism and has less effect on insulin resistance.     

Role for the membrane estrogen receptor alpha in the sexual differentiation of the brain

Estrogens exert pleiotropic effects on multiple physiological and behavioral responses. Male and female sexual behavior in rodents constitutes some of the best‐characterized responses activated by estrogens in adulthood and largely depend on ERα. Evidence exists that nucleus‐ and membrane‐initiated estrogen signaling cooperate to orchestrate the activation of these behaviors both in short‐ and long‐term. However, questions remain regarding the mechanism(s) and receptor(s) involved in the early brain programming during development to organize the circuits underlying sexually differentiated responses. Taking advantage of a mouse model harboring a mutation of the ERα palmitoylation site, which prevents membrane ERα signaling (mERα; ERα‐C451A), this study investigated the role of mERα on the expression of male and female sexual behavior and neuronal populations that differ between sexes. The results revealed no genotype effect on the expression of female sexual behavior, while male sexual behavior was significantly reduced, but not abolished, in males homozygous for the mutation. Similarly, the number of kisspeptin‐ (Kp‐ir) and calbindin‐immunoreactive (Cb‐ir) neurons in the anteroventral periventricular nucleus (AVPv) and the sexually dimorphic nucleus of the preoptic area (SDN‐POA), respectively, were not different between genotypes in females. In contrast, homozygous males showed increased numbers of Kp‐ir and decreased numbers of Cb‐ir neurons compared to wild‐types, thus leading to an intermediate phenotype between females and wild‐type males. Importantly, females neonatally treated with estrogens exhibited the same neurochemical phenotype as their corresponding genotype among males. Together, these data provide evidence that mERα is involved in the perinatal programming of the male brain.     

Direct inhibition of arcuate kisspeptin neurones by neuropeptide Y in the male and female mouse

Adverse energy states exert a potent suppressive influence on the reproductive axis by inhibiting the pulsatile release of gonadotrophin‐releasing hormone and luteinising hormone. One potential mechanism underlying this involves the metabolic‐sensing pro‐opiomelanocortin and agouti‐related peptide/neuropeptide Y (AgRP/NPY) neuronal populations directly controlling the activity of the arcuate nucleus kisspeptin neurones comprising the gonadotrophin‐releasing hormone pulse generator. Using acute brain slice electrophysiology and calcium imaging approaches in Kiss1‐GFP and Kiss1‐GCaMP6 mice, we investigated whether NPY and α‐melanocyte‐stimulating hormone provide a direct modulatory influence on the activity of arcuate kisspeptin neurones in the adult mouse. NPY was found to exert a potent suppressive influence upon the neurokinin B‐evoked firing of approximately one‐half of arcuate kisspeptin neurones in both sexes. This effect was blocked partially by the NPY1R antagonist BIBO 3304, whereas the NPY5R antagonist L152,804 was ineffective. NPY also suppressed the neurokinin B‐evoked increase in intracellular calcium levels in the presence of tetrodotoxin and amino acid receptor antagonists, indicating that the inhibitory effects of NPY are direct on kisspeptin neurones. By contrast, no effects of α‐melanocyte‐stimulating hormone were found on the excitability of arcuate kisspeptin neurones. These studies provide further evidence supporting the hypothesis that AgRP/NPY neurones link energy status and luteinising hormone pulsatility by demonstrating that NPY has a direct suppressive influence upon the activity of a subpopulation of arcuate kisspeptin neurones.     

Neuroanatomical connections between kisspeptin neurones and somatostatin neurones in the female and male rat hypothalamus: A possible involvement of SSTR1 in kisspeptin release

Somatostatin (SST), a neuropeptide involved in the central modulation of several physiological functions, is co‐distributed in the same hypothalamic areas as kisspeptin (KP), the most potent secretagogue of the gonadotrophin‐releasing hormone (GnRH) secretion known to date. Because SST infused i.c.v. evoked a potent inhibition of GnRH release, we explored the neuroanatomical relationships between KP and SST populations in male and female rats. Accordingly, intact males and ovariectomised oestradiol‐replaced females were killed and their brains processed aiming to simultaneously detect KP, SST and synapsin, a marker for synapses. We observed numerous appositions of KP on SST neurones both in the female and male arcuate nucleus (ARC) and ventromedial hypothalamus. A large association between SST terminals and KP neurones at the level of the pre‐optic area was also observed in female rats and in a more limited frame in males. Finally, most KP neurones from the ARC showed SST appositions in both sexes. To determine whether SST could affect KP cell activity, we assessed whether SST receptors (SSTR) were present on KP neurones in the ARC. We also looked for the presence of SSTR1 and SSTR2A in the brain of male rats. Brains were processed using a sequential double immunocytochemistry aiming to detect KP and SSTR1 or KP and SSTR2A. We observed overlapping distributions of immunoreactive neurones for SSTR1 and KP and counted approximately one‐third of KP neurones with SSTR1. By contrast, neurones labelled for SSTR2A or KP were often juxtaposed in the ARC and the occurrence of double‐labelled neurones was sporadic (< 5%). These results suggest that the action of SST on KP neurones would pass mainly through SSTR1 at the level of the ARC.