Differential Gene Expression Response to Gonadal Hormones by Preoptic Regulatory Factors-1 and -2 in the Female Rat Brain

Abstract

Steroids and neuropeptides interact in the central nervous system (CNS) to regulate reproductive function and behavior. The preoptic regulatory factors, PORF-1 and PORF-2, are unique neuropeptides for which roles in gender-related brain development and function have been proposed. PORF-1 and PORF-2 expression in rat brain are age, region and gender dependent, and castration or hypophysectomy alter the metabolism of the PORF-1 and PORF-2 mRNAs in male rat brain and testes. If these two peptides have a role in gender-dependent brain function, then gonadal steroids might well affect their expression. The present study was designed to investigate the response of the PORF-1 and PORF-2 mRNAs to sex steroids in the female rat brain and to compare this response to that of two peptides whose roles in the neuroendocrinology of reproduction are well established, gonadotropin-releasing hormone (GnRH) and neuropeptide Y (NPY). Rats were ovariectomized and treated with placebo, estradiol (E₂), progesterone (P₄) or a combination of the two (E₂/P₄) and NPY, PORF-2, GnRH and PORF-1 mRNAs were quantified by nuclease protection assays. PORF-1, PORF-2 and GnRH mRNAs were also measured in intact rats during estrus and proestrus. Responses were compared in the preoptic anterior hypothalamus (POA), medial basal hypothalamus (MBH), cerebral cortex (CC) and hippocampus (HIPP). Expression of PORF-1 and PORF-2 was also confirmed in the female rat hypothalamus by in situ hybridization analysis. PORF-1 and PORF-2 mRNAs were detected in the adult female rat brain by both in situ hybridization and ribonuclease protection analyses. In situ hybridization analysis demonstrated that PORF-1 and PORF-2 mRNAs are expressed in hypothalamic neurons. RNase protection analysis showed that PORF-1, PORF-2 and NPY mRNAs were present in all four brain regions examined while GnRH expression was detected only in the MBH and POA. Estradiol alone upregulated expression of the PORF-1 and PORF-2 mRNAs in the ovariectomized rat in the POA and HIPP, and of NPY mRNA in the MBH and HIPP. Progesterone alone had a stimulatory effect on NPY mRNA in the MBH and HIPP. Treatment with a combination of E₂/P₄ downregulated PORF-2 mRNA in the POA as well as PORF-1, PORF-2 and NPY mRNAs in the CC. In contrast, E₂/P₄ upregulated the PORF-2 and NPY mRNAs in the HIPP and NPY mRNA in the MBH. In the cycling rat, PORF-1 mRNA levels were higher during proestrus than estrus in both the MBH and POA, while PORF-2 mRNA levels did not change. In contrast GnRH mRNA was lower in the POA and higher in the MBH during proestrus compared with estrus. Thus, intrinsic factors, most likely both ovarian and neuroendocrine, regulate PORF-1 and GnRH expression in the intact cycling rat CNS in a region-dependent manner. In the ovariectomized rat, PORF-1, PORF-2, NPY and GnRH mRNAs all respond in a region-specific manner to sex steroid treatment. These data support the role of PORF-1 and PORF-2 in gender-dependent brain function in the adult female rat.