The Effects Of Peripheral Sex Hormone Levels On Hypothalamus During Estrus Cycle And Following Gonadectomy In Rats

Background and Objective:Sex hormones are a kind of steroid mainly synthesized by sexual gonads. The major sex hormones in body are estrogens and androgen. The ovary in a female and the testis in a male are the most important organs where secrete sex hormones are produced. The hypothalamus is the centre of neuroendocrine system, which regulates the secretion of sex hormones through the hypothalamo-pituitary-gonad (HPG) axis. It is clear that sex hormones are also produced in the brain. Since sex hormones are small hydrophobic steroids that can diffusion through the blood-brain barrier (BBB) freely, the contents of sex hormones in the brain are, at least partly, determined by both sex steroids derived from the peripheral and those produced de novo in the brain. The question how the peripheral sex hormone levels contribute to the levels of sex hormones in the brain, however, remains so far as unclear. In addition, it is well known that there is a close interaction between the HPG and the hypothalamo-pituitary-adrenal (HPA) axis that is the key system involved in the stress response of body. Changes of sex hormones can influence the activity of the HPA-axis, participating in the mechanism of sex differences in the stress responses and a lot of neuropsychiatric disorders. However, so far it is lack of study on the effect of the fluctuating sex hormones in the periphery on the activity of the HPA axis, thus on the vulnerability to the neuropsychiatric disorders. The present study thus aimed to investigate the influence of changes in circulating sex hormones during normal estrous cycle or after gonadectomy upon the hypothalamic sex hormone levels, the activity of the HPA axis, and the transcriptional levels of relevant hormone receptors and neuropeptides in the hypothalamus, and whether there are sex differences.Methods:The estrous cycle stages of female Sprague-Dawley (SD) rats (weigh 220-250g) were monitored by daily vaginal smears. After at least three consecutive regular cycles, rats were sacrificed in proestrus, estrus and diestrus stage respectively, with 12 rats each stage(=group). Another 30 female SD rats (weigh 220-250g) and 20 male SD rats (weigh 290-320g) were recruited and divided at random into sham-operated and gonadectomy groups, respectively. Under 10% chloral hydrate anesthesia, the rats in sham groups were only treated with excising a part of the fat tissue next to the ovary or testis, while the rats in ovariectomy (OVX) or in castration (CAS) group, the ovaries or the testes were removed respectively under the same operation condition. After one week of recovery, female rats of the sham-operation-group were monitored for three consecutive regular cycles then sacrificed in the proestrus stage, the same period as the OVX, CAS and the sham-operation male groups were sacrificed. Animals were decapitated between 9:00-11:00hr. Trunk blood and hypothalamus were collected. Estradiol (E2) and testosterone (T) levels in plasma, the hypothalamus, and CRH levels in plasma were determined by radioimmunoassay. Coricosterone (CORT) levels in plasma were measured by enzyme-linked immunosorbent assay (ELISA). Expression levels of estrogen receptor-a (ER-a)-, ER-β, androgen receptor (AR)-, aromatase-, mineralcorticoid receptor (MR)-, glucocorticoid receptor (GR)-, CRH-, Vasopressin (AVP)-and oxytocin (OXT)-mRNA in the hypothalamus were tested by means of real-time quantitative PCR (QPCR).Results:1. During estrus cycle, E2 and T levels in plasma were dramatically higher in proestrus than in estrus (p<0.0001,p<0.0001) or diestrus (p<0.0001,p<0.0001). No significant changes were found in plasma CORT and CRH (p>0.05). E2 and T levels in the hypothalamus did not show significant changes (p>0.05). There were on significant correlations between plasma-and hypothalamus-sex hormone levels (p>0.05). The mRNA levels of the receptors and neuropeptides detected in the the present study did not show any significant changes (p>0.05) in hypothalamus, except for the expression of aromatase-mRNA showed a trend for increase in the proestrus compared with the estrus stage (p=0.06).2. After OVX, E2 and T levels in plasma significantly declined (p<0.0001, p<0.01),while there was a trend for increase of E2 in the hypothalamic (p=0.055). The hypothalamic T levels did not change significantly in the OVX group compared with the sham-operation group. In addition, the expression of ER-a-mRNA significantly increased (p<0.01), while aromatase-, CRH-and OXT-mRNA significantly decreased (p<0.01, p<0.01, p<0.05 respectively) in hypothalamus of OVX females compared to sham-operation group. Other relevant receptors and neuropeptides detected in the hypothalamus showed no significant changes (p>0.05).3. After castration, T levels in plasma and hypothalamus were both significantly decreased compared to the sham-operation group (p<0.01, p<0.05), and they showed a significant positive correlation in between (r=0.774, p=0.014). In addition, CORT level in plasma increased in CAS group (p<0.05). The receptors and neuropeptides detected did not show significant changes in hypothalamus except that the aromatase-mRNA expression was dramatically decreased in the CAS group (p<0.001).4. Females had a higher CORT level in plasma than males in sham-operation groups (p<0.0001). After gonadectomy, CORT level was down-regulated in female OVX rats but increased in male CAS subjects, while the CORT levels in OVX females showed no significant differences from the CAS males. In addition, OVX female rats showed decreased levels of neuropeptides(CRH and OXT) in the hypothalamus while CAS male rats did not show any significant changes in these neuropeptides, compared with the sham-operation control subjects.Conclusion:The significant fluctuation of circulating sex hormone levels during the estrus cycle does not remarkably affect the homeostasis in the hypothalamus, in the sense of the levels of sex hormones, and HPA activity. The long-lasting dramatic changes of sex hormone levels in the periphery by for instance gonadectomy may break down such homeostasis leading to significant changes in levels of sex hormones, transcriptional levels of relevant receptors and neuropeptides in the hypothalamus. There is a sex difference in the influence of changed peripheral sex hormones on the HPA axis and related neuropeptides in the hypothalamus.