Fig. 4.

Developmental milestone and juvenile behavior outcomes in FCG mice following early or late MIA A Neonatal body weight trajectory is reduced in late MIA-exposed and XXM mice compared to saline controls. B Juvenile body weight is higher in gonadal males (XXM, XYM) than females (XXF, XYF) across treatments. C Righting reflex latency is delayed in both early and late MIA groups across genotypes. D-F. Ultrasonic vocalization (USV) measures reveal treatment by gonadal sex interactions. Late MIA decreased average USV call frequency and early MIA increases downward/upward call types in gonadal females at PND 4. G-J Three-chamber social interaction tests show gonadal females exhibit greater social (gonadal sex effect, F(1,130) = 68.58, p < 0.001). and novelty preferences (gonadal sex effect, F(1,131) = 23.64, p < 0.001, which is reduced by early MIA (treatment × gonadal sex interaction, F(2,130) = 3.85, p = 0.024), shifting behavior toward gonadal male-like patterns. Early MIA also increases middle chamber activity in XY mice (treatment × sex chromosome interaction, F(2,131) = 5.804, p = 0.004). K-L Marble burying behavior is altered by MIA, eliminating the baseline gonadal female advantage; XYM mice bury more marbles than XYF (treatment × gonadal sex interaction emerged, F(2,131) = 3.258, p = 0.042). M-Q Open field test results show early MIA reduces outer zone time in XYF mice. XX mice spent more time in the center and travel greater distances than XY mice (three-way interaction, F(2,131) = 3.305, p = 0.040). R-V Elevated plus maze parameters indicate sex chromosome and gonadal sex differences in anxiety-like behavior, including latency to enter closed arms, time spent in open and closed arms, and entry counts. Main effects of treatment and gonadal sex indicated “t” and “g”, respectively. Two and three-way ANOVA with Tukey’s post hoc tests, n = 11–12/group. Data are mean ± SEM. * p < 0.05, ** p < 0.01, *** p < 0.001. Heatmaps and graphs created with BioRender or GraphPad Prism