Sexual Dimorphism of Microglia in High Trait Anxiety with a Particular Focus on Synaptic Pruning

Abstract

Microglia are the first responders to disruptions in homeostasis as well as crucial players contributing to synaptic refinement in the central nervous system (CNS). We and others have previously reported alterations in microglia phenotype as well as neuroinflammation in elevated anxiety. Futhermore, anxiety and other forms of stress were shown to alter strength of hippocampal excitatory synapses. Yet, little is known about the role of microglia in the engulfment of excitatory synapses in subjects with innate high anxiety-related behavior (HAB). Given that sexual dimorphism is widely reported in anxiety and depression-like behavior, we aimed to investigate a potential sex bias in the engulfment of vGlut1+ excitatory synapses by microglia in the hippocampus of male and female HABs. We observed an elevated level of engulfment torwards vGlut1+ synapses by HAB microglia specifically in female as compared with male. We also examined various microglia markers driving the process of synaptic pruning such as CX3CR1, IL33R and TREM-2, and showed an elevated expression of these markers by HAB female microglia in the hippocampus. We further investigated composition of different microglia subsets in HAB brains with single-cell sequencing and revealed 11 different microglia subpopulations diversified based on the expression of inflammatory and homeostatic markers in both sexes. We add up to the prominent sex differences by reporting a higher percentage of inflammation and senescence-associated subsets of migroclia as well as by revealing a higher expression of pruning-associated genes – C1qa, C1qb, C1qc, CD68, TREM2- by the resting microglia of HAB females in comparison to males. Due to previously reported effects of minocycline on microglia and behavior, we treated HABs with oral minocycline and observed a significant reduction in the engulfment of vGlut1+ synapses by micorglia in females, but not males suggesting a recovery in the state of over-pruning in females. Our study reveals an altered state of pruning by HAB microglia towards vGlut1+ synapses in a sexually dimorphic manner and resolves the main microglia subtypes associated with high trait anxiety. We provide a basis for future studies focusing more on the sexual dimorphism regarding interplay of microglia and synapses in elevated anxiety along with the modulatory effects of minocycline.