Sex differences in the structure and function of the vasopressin system in the ventral pallidum are associated with the sex-specific regulation of social play behavior in juvenile rats

Abstract

Vasopressin (AVP) regulates various social behaviors, often in sex-specific ways, including social play behavior, a rewarding behavior displayed primarily by juveniles. Here, we examined whether and how AVP acting in the brain's reward system regulates social play behavior in juvenile rats. Specifically, we focused on AVP signaling in the ventral pallidum (VP), a brain region that is a part of the reward system. First, we examined the organization of the VP-AVP system in juvenile rats and found sex differences, with higher density of both AVP-immunoreactive fibers and AVP V1a receptor (V1aR) binding in males compared to females while females show a greater number of V1aR-expressing cells compared to males. We further found that, in both sexes, V1aR-expressing cells co-express a GABA marker to a much greater extent (approx. 10 times) than a marker for glutamate. Next, we examined the functional involvement of V1aR-expressing VP cells in social play behavior. We found that exposure to social play enhanced the proportion of activated V1aR-expressing VP cells in males only. Finally, we showed that infusion of a specific V1aR antagonist into the VP increased social play behaviors in juvenile male rats while decreasing these behaviors in juvenile female rats. Overall, these findings reveal structural and functional sex differences in the AVP-V1aR system in the VP that are associated with the sex-specific regulation of social play behavior.

Keywords: Juvenile rats; Sex differences; Social play behavior; Vasopressin.

Figure 1.. Sex differences in AVP-ir fiber and V1aR binding densities in the VP of juvenile rats.

(A) Density measures were taken across four regions from rostral to caudal within the VP; Image sampling in each of the four regions are shown on modified rat brain atlas templates (Paxinos and Watson, 2007); VP is outlined in green; red-filled boxes represent sampling locations for fiber analyses; blue circles represent sampling locations for receptor binding analyses; numbers to the left of the atlas templates represent distances from Bregma (Paxinos and Watson, 2007). (B) Representative images from the right hemisphere of the VP showing AVP-ir fibers in a male and a female juvenile rat. Averages of the four sampling locations show that males have denser AVP-ir fibers than females. (C) Representative images from the right hemisphere of the VP showing V1aR binding in a male and female juvenile rat. Averages of the four sampling locations show that males have denser V1aR binding than females. Data are shown as mean ± SEM and collapsed across sampling locations to highlight sex effect; *p < 0.05, mixed-model ANOVA. ac = anterior commissure.

Figure 2.. Sex difference in the number of v1aR-expressing cells in the VP in juvenile rats.

(A) Image sampling locations in the four regions from rostral to caudal within the VP shown on modified rat brain atlas templates (Paxinos and Watson, 2007); At each region, one dorsal and one ventral image were taken; VP is outlined in green; red-filled boxes represent sampling locations for v1aR+, gad1+ (marker for GABAergic cells), and slc17a6+ (marker for glutamatergic cells). (B) Representative images from the right hemisphere showing v1aR mRNA in the VP; white arrowheads indicate v1aR+ cells. Females show a greater number of v1aR+ cells (C) and greater v1aR mRNA fluorescent intensity in the VP compared to males (D). (E) Representative images from the right hemisphere showing v1aR, gad1, and slc17a6 mRNA in the VP. Yellow arrowheads indicate v1aR+ cells that co-express gad1 and pink arrowheads indicate v1aR+ cells that co-express slc17a6. There was no sex difference in the proportion of v1aR+ cells that co-expressed gad1 (F) or slc17a6 (G). Data are shown as mean ± SEM and collapsed across sampling regions to highlight the main effect of sex; *p < 0.05, ****p < 0.0001, mixed-model ANOVA.

Figure 3.. Sex-specific activation of VP in response to social play exposure.

(A) There was no sex difference in the duration of social play between male and female juvenile rats. (B) Image sampling locations in the four regions from rostral to caudal within the VP shown on modified rat brain atlas templates (Paxinos and Watson, 2007). At each region, a dorsal and ventral image were taken. VP is outlined in green; red-filled boxes represent sampling locations for fos+ and v1aR+ cells. Because the number of fos+ cells was similar across sampling regions in all groups, data were collapsed across sampling regions to highlight sex and play condition effects. (C) Exposure to social play enhanced the number of fos+ cells in the VP of males, but not of females. (D) Time spent engaging in social play did not significantly correlate with the number of fos+ cells in males (blue) or females (red). However, there was a trend in the opposite direction between males and females, such that males showed a trending negative correlation while females showed a trending positive correlation between the number of fos+ cells and social play duration. Data are shown as mean ± SEM; *p < 0.05, Bonferroni post hoc comparisons.

Figure 4.. Sex-specific changes in activation of v1aR-expressing cells in the VP in response to social play behavior.

(A) Representative images of fos mRNA (in green) and v1aR mRNA (in red) staining in VP cells of males and females in the “No Social Play” and “Social Play” groups. Yellow arrowheads refer to v1aR+ cells that co-express fos. (B) Females exposed to social play showed a greater number of fos+ cells that co-expressed v1aR compared to males that were not exposed to social play. (C) Time spent engaging in social play did not significantly correlate with the number of fos+ cells that co-expressed v1aR in males (blue) or females (red). (D) Males exposed to social play showed a greater proportion of v1aR+ cells that co-expressed fos compared to males that were not exposed to social play and females that were exposed to social play. Females that were exposed to social play showed a similar proportion of v1aR+ cells that co-expressed fos compared to females that were not exposed to social play. (E) Time spent engaging in social play did not significantly correlate with the proportion of v1aR+ cells that co-expressed fos in males (blue) or females (red). Note that the number of v1aR+ cells co-expressing fos and proportion of v1aR+ cells that co-expressed fos were similar across sampling regions in all groups; thus data were collapsed across sampling regions to highlight sex and play condition effects. Data are shown as mean ± SEM; *p < 0.05, Bonferroni post hoc comparisons.

Figure 5.. Sex-specific regulation of social play behavior by pharmacological blockade of V1aR signaling in the VP.

The V1aR antagonist infused into the VP increased (A) social play duration, (B) the number of nape attacked, and (C) the number of pins in male juvenile rats and reduced these parameters in female juvenile rats compared to vehicle treatment and compared to AVP treatment (A-C for males, A only for females). (D) AVP-treated male and female rats showed a higher duration of social investigation compared to V1aR-antagonist treated rats. (E) The duration of allogrooming was not affected by V1aR antagonist or AVP treatment. (F) V1aR antagonist-treated males showed a decrease in the duration of non-social cage exploration compared to vehicle-treated males and V1aR antagonist-treated females. Data are shown as mean ± SEM; *p < 0.05, ***p < 0.001, ****p < 0.0001, Bonferroni post hoc comparisons; #p < 0.05 Bonferonni post hoc comparison collapsed across sex.