Microglial depletion worsens lesion in female but not male C57BL/6J mice after P10 hypoxia-ischemia

Abstract

Background: Rodent models for perinatal hypoxic ischemic (HI) encephalopathy have reported sex differences such as males having larger brain lesions than females after the same injury. Microglia, the resident immune cells of the brain that have distinct developmental trajectories and gene expression patterns by sex, likely play a different role in males and females following HI. However, there is conflicting literature on whether depletion of microglia worsens or improves HI-induced lesions, and whether this differs by sex. We tested the effect of pharmacologic microglial depletion on lesion size and developmental milestones. Methods: C57BL/6J mouse pups received daily intraperitoneal injections from postnatal day 7 (P7) to P12 of either 25 mg/kg PLX3397 (a CSF1R inhibitor) or vehicle (Veh). At P10, pups either underwent Hypoxic-Ischemic (HI) insult using a modified Vannucci procedure, or a Sham insult. This resulted in four groups tested (Veh-Sham, Veh-HI, PLX-Sham, PLX-HI); all groups included both male and female mice. Behavioral testing was performed both pre-HI (forelimb grasping [P8, P9]) and post-HI (open field traversal [P12], gross behavior and appearance in homecage and new environment [P13]). Brains were collected at P13, fixed, and sectioned, then either immunolabeled for Iba1 or stained with cresyl violet for injury scoring. Results: Immunohistochemistry for Iba1 demonstrated >95% depletion of microglia at either P10 or P13 in all PLX3397-treated mice; depletion did not differ by sex. In the hippocampus, Female PLX-HI mice had worse cresyl violet lesion scores than Female Veh-HI mice; this was not true in male mice, where there was a trend in the opposite direction. Female PLX-HI mice also had worse lesion scores than Male PLX-HI mice. There were no differences among groups in forelimb grasp, open field traversal times, or weight gain. Conclusion: Microglial depletion worsens HI-induced injury in female mice but not in male mice.