Ovarian status dictates the neuroinflammatory and behavioral consequences of sub-chronic stress exposure in middle-aged female mice

Abstract

Ovarian hormones influence the outcomes of stress exposure and are implicated in stress-related disorders including depression, yet their roles are often complex and seemingly contradictory. Importantly, depression and stress exposure are associated with immune dysregulation, and ovarian hormones have immunomodulatory properties. However, how ovarian hormones can influence the inflammatory outcomes of stress exposure is poorly understood. Here, we examined the effects of long-term ovariectomy on the behavioral and neuroinflammatory outcomes of sub-chronic stress exposure in middle-aged mice. Briefly, sham-operated and ovariectomized mice were assigned to non-stress groups or exposed to 6 days of variable stress. Mice were assessed on a battery of behavioral tests, and cytokine concentrations were quantified in the frontal cortex and hippocampus. In the frontal cortex, postsynaptic density protein-95 expression was examined as an index of excitatory synapse number and/or stability, and phosphorylated mitogen-activated protein kinases (MAPKs) were measured to explore potential cell signaling pathways elicited by stress exposure and/or ovarian hormones. Long-term ovariectomy modified the central cytokine profile by robustly reducing cytokine concentrations in the frontal cortex and modestly increasing concentrations in the hippocampus. Under non-stress conditions, long-term ovariectomy also reduced extracellular signal-regulated kinase (ERK) phosphoprotein expression in the frontal cortex and increased some measures of depressive-like behavior. The effects of sub-chronic stress exposure were however more pronounced in sham-operated mice. Notably, in sham-operated mice only, sub-chronic stress exposure increased IL-1β and IL-6:IL-10 ratio in the frontal cortex and hippocampus and reduced pERK1/2 expression in the frontal cortex. Further, although sub-chronic stress exposure increased anhedonia-like behavior regardless of ovarian status, it increased passive-coping behavior in sham-operated mice only. These data indicate that long-term ovariectomy has potent effects on the central cytokine milieu and dictates the neuroinflammatory and behavioral effects of sub-chronic stress exposure in middle-aged mice. These findings therefore suggest that the immunomodulatory properties of ovarian hormones are of relevance in the context of stress and possibly depression.

Sex Differences and Estradiol Effects in MAPK and Akt Cell Signaling across Subregions of the Hippocampus

“…Rats were subcutaneously administered 1.1 µg/kg 17β-estradiol in sesame oil (low 17β-estradiol; females n = 10, males n = 6, OVX n = 13), 37.3 µg/kg 17β-estradiol in sesame oil (high 17β-estradiol; females n = 10, males n = 7, OVX n = 13), or sesame oil vehicle (females n = 9, males n = 6, OVX n = 10) 30 min prior to tissue extraction. These sample sizes were selected as they have previously produced large effect sizes (Cohen’s d > 0.8) in similar analyses [6, 39]. Estradiol doses were chosen because they result in circulating levels of 17β-estradiol observed on the morning of proestrus (high 17β-estradiol) or during diestrus (low 17β-estradiol) [40], and both doses enhance cell proliferation 30 min after injection in OVX females [6, 30].…”

“…Samples were run in technical replicates, and plates were read with a Sector Imager 2400 (Meso Scale Discovery) using the Discovery Workbench 4.0 (Meso Scale Discovery). Phosphoprotein values were averaged across technical replicates and normalized to total protein concentrations for statistical analyses (as in [39,[43][44][45]). Inter-and intra-assay coefficients of variation were below 10%.…”

“…The present data were normalized to total protein in sample (as is done regularly in the literature: [39,[43][44][45], e.g.). One possible limitation is that phosphorylated proteins were not normalized to the specific protein under investigation (e.g., pERK1/2 normalized to total ERK1/2).…”