Predator odor exposure of rat pups has opposite effects on play by juvenile males and females

Abstract

Juvenile social play behavior is one of the earliest sexually differentiated behaviors to emerge. In rats, as with most other species that play, males engage in more rough-and-tumble play compared to females. Exposure to early life adversity is a major driver of adult health and can manifest differently in males and females. However, the effects of adverse early life exposure on play behavior in the juvenile period are poorly understood. To address this, male and female neonatal rats were exposed to predator odor (PO), for 5min/day on PN1-PN3. At the time of exposure to PO, both male and female pups suppressed ultrasonic vocalization and displayed more freezing behavior. Circulating corticosterone increased in males immediately following PO exposure but not in females. The enduring effects of PO exposure were opposite in males compared to females in that PO exposed males decreased social play, while PO exposed females increased play behavior compared to same sex controls. PO exposure did not significantly affect cell genesis in the neonatal dentate gyrus of either sex. PO exposure did not affect anxiety-like behavior assessed in the juvenile period or in adulthood, nor did it affect social interactions in adulthood. This work provides new insight into how sex may interact with adverse early life events to contribute to development of the social consequences of such exposures.

Keywords: Anxiety; Corticosterone; Development; Early life adversity; Predator odor; Sex differences; Social play.

Fig. 1

Timeline of the sequence of exposures and assessments for each cohort of animals.

Fig. 2

Rat pups modulate their behavior in response to the fearful stimulus of predator odor. Male and females pups were exposed to PO on PN1-PN3 for analysis of freezing behavior and on PN4 for detection of USV suppression. (A) Exposure to PO significantly reduced the number of USVs emitted in both males and females. (B) In both sexes, the amplitude of USVs produced in the presence of PO were marginally decreased with a large effect size (F[1,7] = 5.545, p = 0.0507; ${\mathrm{\eta }}_{\mathrm{p}}^2=0.442$). (C) When exposed to PO, both male and female pups increased freezing behavior. (Data expressed as mean ± SEM. Control male group, n = 3; PO male group, n = 2; control female group, n = 3; female group, n = 3; two-way ANOVA: *p < 0.05, **p < 0.01.)

Fig. 3

Corticosterone levels increased only in males following PO exposure. (A) There was no sex difference in corticosterone among animals exposed to maternal control bedding. In males, exposure to PO caused an increase in corticosterone, while female corticosterone did not change in response to the exposure. (B) As previously reported (Zhang et al., 2008; Bowers et al., 2010), there is a higher rate of cell proliferation in BrdU + cells in the dentate gyrus of neonatal males compared to females as indicated by BrdU + cell number. PO exposure resulted in a trend towards reduction of cellular proliferation in the dentate gyrus with a medium effect size (F[1,40] = 3.369, p = 0.0739; ${\mathrm{\eta }}_{\mathrm{p}}^2=0.078$). (C) Hippocampal BDNF protein did not significantly differ between any conditions. (Data expressed as mean ± SEM. Corticosterone assessment: control male group, n = 8; PO male group, n = 7; control female group, n = 6; female group, n = 6; BrdU analysis: control male group, n = 10; PO male group, n = 11; control female group, n = 12; PO female group, n = 11; twoway ANOVA and pairwise comparisons with Bonferroni correction: *p < 0.05, **p < 0.01.)

Fig. 4

Neonatal PO exposure has opposite effects on later social play in males and females. Male and female pups that were exposed to PO were assessed for play behavior on PN28–PN32 with a consistent matched sex and exposure partner. (A) The average time spent playing per day was greater in males than females. Males exposed to PO engaged in less play and females more. (B) Males engaged in more play events per day than females. Males exposed to PO had a decreased frequency of play, while females exposed to PO increased play frequency. Analysis of specific play behaviors including pouncing (C), pinning (D), boxing (E) and (F) chasing revealed similar patterns, decreasing in males exposed to PO and increasing in PO exposed females. (Data expressed as mean ± SEM. Control male group, n = 10; PO male group, n = 12; control female group, n = 10; PO female group, n = 12; two-way ANOVA and pairwise comparisons with Bonferroni correction: *p < 0.05, **p < 0.01.)

Fig. 5

Exposure to predator odor during postnatal development does not affect anxiety-like behavior in adolescence. Male and female pups subject to PO exposure from PN1–PN3 and unexposed controls were assessed for anxiety-like behavior in the open field test on PN26, the elevated plus maze on PN33 and the light/dark box on PN34. (A) All animals traveled the same distance in the OFT and there were no differences in time spent in the center (B) or entries into the center (C). In the EPM, neither PO exposure nor sex altered (D) time spent in the open arms or (E) number of entries in the open arms. The L/D box, revealed no effect of predator odor exposure on (F) time spent in the light zone or (G) entries into the light zone, however females spent more time and entered into the light zone more than males. (Data expressed as mean ± SEM. Control male group, n = 10;PO male group, n = 12; control female group, n = 10; PO female group, n = 12; two-way ANOVA and pairwise comparisons with Bonferroni correction: **p < 0.01.)

Fig. 6

Adult social and anxiety-like behaviors are not affected by early PO exposure. Male and female pups exposed to PO were retested for social and anxiety-like behaviors in adulthood. (A) Distance traveled was greater among unexposed females and decreased in both males and females exposed to PO. (B) Unexposed females also displayed more entries in the center zone, whereas PO exposed male and females entered this zone less. (C) Time spent in the center zone did not significantly differ between groups. (D) The interaction ratio of the SIT showed no significant effects of either variable. In the EPM, groups did not differ with respect to (E) time spent in the open arms or (F) number of entries in the open arms of the elevated plus maze. Behavior in the L/D box showed no significant effect of any condition on (G) time spent in the light zone or (H) number of entries into the light zone of the light/dark box. (Data expressed as mean ± SEM. For OFT, EPM and L/D Box control male group, n = 6; PO male group, n = 6; control female group, n = 6; PO female group, n = 6; for SIT control male group, n = 4; PO male group, n = 6; control female group, n = 4; PO female group n = 6 two-way ANOVA.)