Effects of Age and Ovarian Hormone Loss on Neurogenesis and Associated E2/P4 Receptor Expression in the Female Brain

Abstract

Neural stem and progenitor cells (NSPCs) in the adult mammalian brain produce new neurons throughout life. It is known that this neurogenic process is regulated by age and sex-dependent mechanisms, however, these mechanisms are not fully understood. Our previous work in male rats identified a critical period of NSPC regenerative decline, between 13 and 15 months of age, and identified the reduced expression of the redox-sensitive nuclear factor (erythroid-derived 2)- like 2 (Nrf2) transcription factor as a key mediator of this phenomenon. Based on these findings, this work utilized advancing age groups of intact and ovariectomized (OVX) female F344 rats (2, 6, 9, and 14 months) to better understand the roles of age and the ovarian sex hormones, 17β- estradiol (E2) and progesterone (P4), in regulating aging NSPC function. This thesis investigates effects of age and circulating E2/P4 loss on adult neurogenesis and sex hormone receptor expression within the two mammalian NSPC niches – the forebrain subventricular zone (SVZ) and the subgranular zone in the hippocampal dentate gyrus (DG). Unbiased stereological quantification of BrdU and DCX immunolabeling revealed age-based reductions in SVZ neurogenesis by 14 months with OVX-induced declines observed at 9 months compared to intact animals of the same age. In the DG, BrdU and DCX declined earlier, by 6 months of age. Western blotting was used to assess the relative expression of estrogen receptors (ERα, ERβ) and progesterone receptor (PR), revealing a general increase in ER expression with advancing age, as well as region-specific effects of OVX: a significant increase in ER expression at 9 months in the SVZ and a significant decrease at 6 months in the hippocampus, with unique alterations to PR expression. Collectively, these findings identified a particular period of vulnerability, around 9 months of age, in which adult neurogenesis and sex hormone receptor expression is significantly impacted by OVX, particularly in the SVZ. Given the antibody-based challenges in assessing the cell specificity of ER and PR receptor expression we employed RNA fluorescent in situ hybridization (FISH) to probe more abundant ERβ mRNA transcripts, in the SVZ and DG. Our gene of interests, Nfe2l2, which we had previously discovered to play a key role in the aging of male NSPC aging, was also analyzed. This pilot study provided first information on how Erβ and Nfe2l2 RNA expression may change in the SVZ and DG. Our findings broadly show niche-specific ER and PR responses to advancing age and acute ovarian hormone loss, and demonstrate technical challenges in investigating low-abundance, dynamic protein targets in the brain.