The Effect Of Prolactin Releasing Peptide In Brain On Electroacupuncture Normalizing The Dysfunction Of Hypothalamus-pituitary-ovary Axis Of Ovariectomized Rats

Aim: Prolactin releasing peptide (PrRP) is a novel 31 amino acid neuropeptide reported as a potent and specific stimulator of PRL secretion. It is the endogenous hypothalamic ligand for the GPR10 orphan receptor. The first biological function identified for the PrRP peptide was the prolactin release from pituitary cells. More recent reports have suggested new potential actions of PrRP in the regulation of cardiovascular system, stress, diet and sleep. In addition to these functions, the morphological and functional studies suggested that PrRP may participate in the regulation of reproductive endocrine system.There are two important functional axises regulating the female reproductive endocrine system: hypothalamus-pituitary-ovary axis (HPOA) and hypothalamus-pituitary prolactin axis. HPOA plays an important role in regulating the reproductive function. It is reported that GnRH is the initial factor of HPOA and the final common pathway of reproductive regulating system. There is a close relationship between HPOA and hypothalamus-pituitary prolactin axis. ICV injection of PrRP can stimulate the release of LH and FSH. The expression of PrRP mRNA was regulated by the level of gonadal steroid. The level of estrogen can also affect the synthesis and secretion of PRL. But it is not clear whether the stimulating effect of PrRP on LH and FSH is through indirect way or direct way-GnRH and what the relationship is between HPOA and hypothalamus-pituitary prolactin axis.The origin of menopause syndrome is the disfunction of HPOA due to the decrease of estrogen. Clinical researches have proved that acupuncture have positive effects on the disease. Previous researches done by our lab have suggested that electroacupuncture (EA) can regulate the disfunction of HPOA. But it is still unknown whether PrRP is involved in the regulatory effect of EA on HPOA.PRL and E2 can stimulate their receptors to affect the synthesis and secretion of PrRP, GnRH and other neurotransmitters or modulators. The effects of EA on the expression of PRL, E2 and their receptors are also under our investigation.Methods: Female Sprague-Dawley rats were purchased from Medical Experimental Animals Center of Fudan University. Half of them underwent ovariectomy with ether anesthesia, which were then divided randomly into two groups: ovariectomized (OVX) and ovariectomized with EA (OVX+EA). The rest were treated as controls which were divided into two groups: intact (INT) and intact with EA (INT+EA). The stimulation was generated by an EA apparatus and lasted for 30 min (08:00-10:00 h), Q.D, for 3 days altogether.Push-Pull perfusion technique was use to detect the release of GnRH in MPOA. The effects of ovariectomy and EA on the number of PrRP-ir and GnRH-ir cells were observed by immunohistochemistry. In situ hybridization combined with immunohistochemistry were employed to explore whether the PrRP receptor mRNA co-localize with GnRH. Dot blot and RT-PCR were used to observe the effects of EA and ovariectomy on the expression of PrRP-R mRNA , GnRH receptor mRNA , PRL receptor mRNA, ER-a and ER-P mRNA. The levels of E2 and PRL in serum were detected by RIA.Results: The expression of PrRP mRNA and the number of PrRP cells in medulla oblongata were decreased in OVX compared with INT. The expression of PrRP receptor mRNA in medulla oblongata, hypothalamus, and pituitary of OVX was increased compared with those of INT; The expression of GnRH mRNA in hypothalamus and the release of GnRH in MPOA were increased in OVX compared with those of INT. The number of GnRH-ir cells in hypothalamus and the expression of GnRH receptor mRNA were decreased in OVX compared with those of INT.The results showed that the PrRP receptor mRNA and GnRH co-localize by immunohistochemistry combined in situ hybridization.EA can increase the expression of PrPP mRNA and the number of PrRP cells in medulla oblongata of ovariectomized rats. The expression of PrRP receptor mRNA of ovariectomized rats was decreased after EA treatment. The expression of GnRH mRNA and the release of GnRH i...