[Effects of Nourishing Yin Removing Fire Chinese Herbs on Gene Expression of Hypothalamic Ghrelin and its Receptor in Female Precocious Rats].

OBJECTIVE: To observe the effect of nourishing yin removing fire Chinese herbs (NYRF-CH) on the gene expression of hypothalamic growth hormone secretion peptide (Ghrelin) and its receptor growth hormone secretion peptide receptor 1alpha (GHSR1-alpha) at the puberty onset of danazol induced female precocious rats. METHODS: Forty female SD rats were randomly divided into 4 groups, i.e., the normal group (N), the model group (M), the normal saline intervention group (NS), and the NYRFCH intervention group (NI), 10 in each group. 300 microg danazol was subcutaneously injected to all rats except those in the N group to prepare precocious rat model. NYRFCH and normal saline was respectively administered to rats in the NI and the NS group from the 15th day old for 7-10 days. No treatment was given to rats in the N group. Time of rats' vulva opening was recorded. Ovary index and uterus index were calculated. Peripheral blood levels of estradiol (E2), follicle stimulating hormone (FSH), and luteinizing hormone (LH), and hypothalamic contents of gonadotropin releasing hormone (GnRH) as well as the gene expression of hypothalamic Ghrelin and GHSR1-alpha were determined. Results Compared with the N group, the vulva opening time was advanced in the model group; peripheral blood levels of E2 and LH, uterus index, hypothalamic contents of GnRH increased; peripheral blood FSH levels and mRNA levels of hypothalamic Ghrelin and GHSR1-alpha decreased (P < 0.05, P < 0.01). Compared with the M group and the NS group, the vulva opening time was not advanced in the NI group; peripheral blood levels of E2 and LH, uterus index and hypothalamic contents of GnRH obviously decreased (P < 0.05, P < 0.01); mRNA levels of hypothalamic Ghrelin and GHSR1-alpha increased (all P < 0.01). But there was no statistical difference in the hypothalamic contents of Ghrelin, or the number and activity of GHSR1-alpha (P > 0.05). CONCLUSION: NYRFCH had regulatory effect on regulating hypothalamic Ghrelin and GHSR1-alpha at gene transcription levels.

[Effect of electroacupuncture intervention on expression of corticotropin-releasing factor protein in hypothalamus and hippocampus of traumatized rats].

OBJECTIVE: To observe changes of anesthesia analepsia time and hypothalamic and hippocampal corticotropin-releasing factor (CRF) protein expression after electroacupuncture (EA) intervention in acute traumatic rats, in order to explore its mechanism underlying relief of post-operative stress. METHODS: Twenty male Sprague-Dawley rats were randomly divided into normal control group, model group, sham EA group and EA group (5 rats/group). Acute traumatic model was established by left lobectomy. For rats of the sham EA group, two filiform needles were inserted into the right "Zusanli" (ST 36) and "Sanyinjiao" (SP 6) respectively, without needle-twirling and without electrical stimulation. For rats of the EA group, the right ST 36 and SP 6 were electrically stimulated at 2 Hz/100 Hz and 2 mA for 30 minutes after needle insertion. Anesthesia analepsia time was recorded and righting reflex was tested for the acute traumatized rats. The expression of CRF protein in the hypothalamus and hippocampus was analyzed with Western blot. RESULTS: The anesthesia analepsia time and righting reflex recovery time were significantly shorter in the EA group than in the model group and sham EA group (P < 0.01). There was a close correlation between the anesthesia analepsia time and righting reflex recovery time (P < 0.05). In comparison with the normal group, the expression level of hippocampal CRF protein was significantly decreased whereas that of hypothalamic CRF protein evidently increased in the model group (P < 0.05). Compared with the model group, hippocampal CRF protein expression was considerably upregulated and hypothalamic CRF protein remarkably down-regulated in the EA group (P < 0.05). A markedly negative correlation was found between hypothalamic CRF and hippocampal CRF protein expression, and between hippocampal CRF expression and the stimulating mode (P < 0.001), and a positive correlation was found between hypothalamic CRF expression and righting reflex, and between hippocampal CRF expression and EA intervention (P < 0.01). CONCLUSION: EA intervention can shorten anesthesia analepsia time and righting reflex recovery time in liver-traumatized rats, which is closely associated with its effects in upregulating hippocampal CRF protein expression and down-regulating hypothalamic CRF expression. It suggests that CRF mediated hippocampus-hypothalamus-pituitary-axis is involved in EA induced reduction of post-surgical stress.

[Neurokinkin B and it's function on reproductive endocrine].

Neurokinkin B (NKB) is a member of tachykinin family and plays a role mainly through its receptor NK3R. NKB and NK3R are wide spread through the neural system. Studies revealed that NKB has kinds of biological functions such as constringing the smooth muscle of hollow viscus, relaxing blood vessels, reducing mean arterial pressure, slowing heart rate, exciting in vitro spinal cord neurons of rats and astringing sphincter pupillae. For the past few years, people pay more and more attentions to the regulating action of NKB on reproductive endocrine and a lot of research are made to discuss the function of NKB in HPGA. This article summarizes the distribution and physiological function of NKB and NK3R, discusses their functions in reproductive endocrine. Future studies will be needed to determine the precise mechanism of NKB.

[Effects of electroacupuncture on the expression of corticotropin-releasing factor (CRF) family and CRF receptor 1 in traumatic injury rats].

OBJECTIVE: To observe the effects of electroacupunture (EA) on the abnormal hypothalamus-pituitary-adrenal (HPA) axis and the expression of hypothalamic corticotropin-releasing factor (CRF) peptides family and CRF receptor (R)1 mRNA in traumatized rats, so as to study its underlying mechanism in improving traumatic injury. METHODS: Forty male Sprague-Dawley rats were randomly divided into normal control group (n=10), normal+ EA group (n=10), surgical trauma (model) group (n=10), and model+ EA group (n=10). Traumatic injury model was established by performing an exploratory laparotomy in the rats under anesthesia. EA (2 Hz/15 Hz, 1-2 mA) was administered for 30 min via two stainless steel needles inserted in "Zusanli" (ST 36) and "Sanyinjiao" (SP 6). Serum adrenocorticotropic hormone (ACTH) , corticosterone (Cort), luteinizing hormone (LH) and testosterone (T) contents were detected by double-antibody radioimmnoassay. Hypothalamic CRF and ORF R1 mRNA expression was assayed by reversed transcription polymerase chain reaction (RT-PCR) technique. RESULTS: The serum ACTH concentration decreased significantly in model group (P < 0.05) compared with those in normal control and normal+ EA groups, and was significantly higher in model+ EA group than that in model group (P < 0.05). No significant difference was found between normal control and normal+ EA groups in serum ACTH levels (P > 0.05). The serum Cort level increased significantly in model group (P < 0.05) compared with those in normal control and normal + EA groups, and was obviously lower in model+EA group than that in model group (P < 0.05). While serum LH and T contents had no apparent changes in the four groups (P > 0.05). The expression of hypothalamic CRF mRNA decreased significantly in model group (P < 0.05) compared with that in normal control group, but increased significantly in model + EA group (P < 0.05) compared with that in model group. However, no significant difference was found between normal control and normal + EA groups in the CRF mRNA expression level. The expression of hypothalamic Ucn 1 mRNA increased significantly in model group than that in normal control group (P < 0.05), and decreased considerably in model+ EA group than that in model group (P < 0.05). There were no significant changes in the expression of hyp, thalamic Ucn 2 mRNA, Ucn 3 mRNA and CRF R1 mRNA in the four groups (P > 0.05). CONCLUSION: EA of "Zusanli" (ST 36) and "Sanyinjiao"(SP 6) can improve surgical trauma-induced HPA disorders, which is probably related to its effects in upregulating CRF mRNA expression and downregulating Ucn 1 mRNA expression in the hypothalamus.

Electroacupuncture stimulates hypothalamic aromatization.

We have previously reported that the repeated electroacupuncture (EA) stimulation significantly increased the concentrations of circulating estradiol and restored the depressed function of the hypothalamus-pituitary-ovary axis (HPOA) in ovariectomized (OVX) rats. We hypothesize that extragonadal aromatization in specific brain areas might be responsible for these changes. Thus, various assays, including radiometric assay, Western blot, and reverse transcriptase polymerase chain reaction (RT-PCR), were employed to determine the aromatization in the hypothalamus of rats that received both OVX and electroacupuncture (OVX + EA). The results showed that EA significantly increased the aromatase activity as well as the expressions of its mRNA and protein (P < 0.05) in the OVX rats. These results suggest that EA enhances brain aromatization, which might contribute to influence the function of gonadotropin-releasing hormone (GnRH) neurons and promote the hypofunction of the HPOA in the ovariectomized rats.

Decreased hypothalamic aromatization in female rats of true precocious puberty.

The true precocious puberty animal model induced by the single dose of danazol was used for investigating the expressions of hypothalamic aromatase in the advanced onset of puberty in rats. The day of vaginal opening and first estrus showed significant advancement in the model rats compared with the normal and vehicle rats (P < 0.01, respectively). The hypothalamic gonadotropin-releasing hormone (GnRH) mRNA expression increased significantly in the model rats compared with that in the normal and vehicle ones (P < 0.01). The levels of aromatase mRNA and protein expressions detected by RT-PCR and Western blot both decreased in the model rats compared with those in the normal and vehicle groups (P < 0.05). The results suggested that the hypothalamic aromatization might diminish in the onset of true precocious puberty of female rats.

Increased corticortropin-releasing hormone release in ovariectomized rats' paraventricular nucleus: effects of electroacupuncture.

Utilizing push-pull perfusion, we examined secretary profiles of corticotropin-releasing hormone (CRH) in the nucleus paraventricularis (PVN) of freely moving intact (INT) and ovariectomized (OVX) rats, and in the meanwhile the effects of electroacupuncture (EA) on the release patterns of CRH were observed. The PVN was perfused with artificial cerebrospinal fluid between 08:00 and 12:00 h, and perfusates were collected every 10 min. The average CRH output was significantly larger in OVX rats than that in INT and INT with EA (INT+EA) groups. Interestingly enough, the CRH output showed a significant elevation in OVX with EA (OVX+EA) group during the EA procedure and further increase immediately after the EA. It is the first time to present the temporal profiles of CRH secretion in the PVN of OVX and OVX+EA rats.

[Effect of Chinese herbal medicine for nourishing yin and purging fire on mRNA expressions of gonadotropin-releasing hormone and its receptor in precocious puberty model rats].

OBJECTIVE: To probe the mechanism of Chinese herbal medicine (CHM) for nourishing Yin and purging Fire on the expressions of gonadotropin-release hormone (GnRH) and its mRNA expression in hypothalamus and GnRH receptor mRNA in pituitary in danazol induced precocious puberty model rats. METHODS: Rats were divided into the normal group, the model group, the blank control group and the CHM group. Rats, except that in the normal group, were subcutaneously administered danazol 300 micrograms at 5 days of age individually and CHM was fed to rats in the CHM group from 15 days of age, in the meantime, normal saline was fed to rats in the blank control group. Expression of GnRH in hypothalamus was observed by immunohistochemical method and expressions of GnRH mRNA in hypothalamus and GnRH receptor mRNA in pituitary were determined by RT-PCR. RESULTS: Compared with rats in the normal groups, the vaginal opening and the onset of first estrus were ahead of time, the number of GnRH immunoreactive positive cells decreased and the expressions of GnRH mRNA in hypothalamus and GnRH receptor mRNA in pituitary up-regulated in the model rats and blank control rats. Compared with the model and the blank control groups, in CHM group, all the above-mentioned abnormally changed parameters improved significantly after treatment. CONCLUSION: CHM for nourishing Yin and purging Fire may inhibit the abnormal hyperfunction of hypothalamus-pituitary-ovary axis in precocious puberty rat induced by danazol via reducing the synthesis and release of GnRH, and lowering the responsibility of pituitary cells to GnRH. This may be the primary mechanism of CHM in effectively treating the true precocious puberty.

An important role of corticotropin-releasing hormone in electroacupuncture normalizing the subnormal function of hypothalamus-pituitary-ovary axis in ovariectomized rats.

In the present study, the effects of electroacupuncture (EA) at a group of specific acupoints on corticotropin-releasing hormone (CRH) and gonadotropin-releasing hormone (GnRH) immunoreactivity (ir) in the hypothalamuses of ovariectomized rats were observed. Meanwhile the blood E2 level was detected. The results showed that EA might significantly increase the blood level of E2 and GnRH cell number in the ovariectomized rats. The number of CRH neurons was higher in the group ovariectomized with EA than that in the ovariectomized and intact groups. Interestingly enough, only in the group ovariectomized with EA was observed the co-localization of CRH-ir and GnRH-ir substances in one cell of hypothalamic nucleus paraventricularis by immunofluorescent double-labeling histochemistry combining laser con-focal scanning microscope. The results suggest that CRH might be an important factor in EA normalizing the subnormal function of hypothalamus-pituitary-ovary axis.