Maternal immune activation during the lactational period alters offspring behavior, reproductive development, and immune function in mice

Abstract

Exposure to infection during early life can lead to lasting neurodevelopmental changes. Animal models of maternal immune activation (MIA) typically assess neurobehavioral alterations in offspring following a prenatal inflammatory challenge. However, MIA effects on offspring can extend to challenges that occur during the lactational period. In the present study, we adapted previous methods focused on rats and challenged nursing C57BL/6J mouse mothers on postnatal day (P)8 with either the bacterial mimetic lipopolysaccharide (LPS; 250 μg/kg, i.p.) or saline (control, i.p.). By exposing only the mother to LPS, this modeled a postpartum infection in the dam. Similar to the rat model, lactational MIA did not detrimentally alter maternal care but induced displays of maternal sickness, as expected. While neonatal offspring behaviors (e.g., huddling, ultrasonic vocalizations, negative geotaxis) were unaffected, significant effects of lactational MIA emerged in juvenile (e.g., social preference, accelerated reproductive milestones) and adult (e.g., mechanical allodynia, prepulse inhibition) offspring. In a separate set of animals, the developmental programming potential of lactational MIA on immune function was evident following a "second hit" LPS challenge in adulthood (e.g., altered plasma concentrations of interleukin-6 and leukocytes, including neutrophils, and lymphocytes). These findings confirm the generalizability of the lactational MIA model across species and highlight the importance of supporting caregiver health and wellness across the critical nursing period.

Keywords: lactation; maternal immune activation; nursing; programming; second hit.

Figure 1.. Experimental Timeline.

Created in BioRender. Kentner, A. (2025) https://BioRender.com/86qqgzg

Figure 2.. Maternal and neonatal outcomes following lactational maternal immune activation (MIA) in mice.

A) Mouse dams treated with LPS on postnatal day (P) 8 had higher sickness scores than dams treated with saline, validating MIA. B) LPS dams had a modest and transient decrease in nest quality on the day of MIA challenge, suggesting that maternal care was not strongly compromised. C) Neonatal huddling behavior was not affected by MIA across P9 and P10. Neonatal behaviors including D) the total number of calls emitted during the isolation induced ultrasonic vocalization test and E) negative geotaxis were not affected by MIA (males left-side, females right-side). Saline: grey circles; lipopolysaccharide (LPS): blue squares. Data expressed as mean ± SEM, n = 9 litters/group. *p <0.05; **p< 0.01, Saline versus LPS.

Figure 3.. Juvenile offspring behaviors and reproductive milestones were affected by lactational maternal immune activation (MIA).

A) The day of vaginal opening was accelerated in female LPS compared to saline female mice (top) while B) male preputial separation was not affected (independent t-tests left graphs; Log-rank (Mantel Cox) tests right graphs). Lactational MIA C) increased social preference in males (left-side) while decreasing it in females (right-side). Since D) percent time in the center of the open field was not affected in either male or female offspring, this suggest the social changes were not mediated by anxiety-like behavior. E) Saline: grey circles; lipopolysaccharide (LPS): blue squares. Data expressed as mean ± SEM, n = 9 litters/group. *p<0.05; **p<0.01; ***p<0.001, Saline versus LPS.

Figure 4.. Adult offspring behaviors were affected by lactational maternal immune activation (MIA).

Male and female offspring A) percent prepulse inhibition (%PPI) and B) mean %PPI of the acoustic startle response. C) Lactational MIA decreased mechanical allodynia thresholds (grams) in males (left-side) while increasing it in females (right-side) in the von Frey test. Saline: grey circles; lipopolysaccharide (LPS): blue squares. Data expressed as mean ± SEM, n = 9 litters/group. ###p<0.001, main effect of dB level; *p<0.05; **p<0.01; ***p<0.001, Saline versus LPS.

Figure 5.. Adult offspring have altered immune functioning following lactational maternal immune activation (MIA).

Plasma markers (n = 5-6 litters/group) for A) the number of white blood cells (leukocytes) measured, the absolute number of leukocytes that were neutrophils or lymphocytes (10⁹ cells/liter) directly measured, and B) interleukin (IL)-6 (pg/mL) demonstrating programming effects following the early life inflammatory challenge; one male Saline-LPS animal had undetectable levels of IL-6. C) Plasma corticosterone concentrations (ng/mL; n = 5 litters/group; one female Saline-Saline animal had undetectable levels of this hormone). Saline-Saline: solid circles and light grey shaded bars; Saline-LPS: open circles and solid dark grey bars; LPS-Saline: solid blue squares and light blue shaded bars; LPS-LPS: open squares and dark blue solid bars; lipopolysaccharide (LPS). Data expressed as mean ± SEM, Males on left side and females on right. *p<0.05, **p<0.01, ***p<0.001, Saline-Saline versus Saline-LPS; #p<0.05, ##p<0.01, ###p<0.001, LPS-Saline versus LPS-LPS; ^ap<0.05, Saline-LPS versus LPS-LPS; ^bp<0.05, Saline-Saline versus LPS-Saline; ^cp<0.05, ^ccp<0.01, ^cccp<0.001, main effect of adult LPS challenge.