Arginine vasopressin (AVP) is a neuropeptide signaling system that is involved in regulating a wide variety of social behaviors in rodents such as aggression (Ferris and Potegal, 1988; Potegal and Ferris, 1989; Caldwell and Albers, 2004; Bosch et al., 2010), sexual behaviors (Södersten et al., 1985; Pedersen and Boccia, 2006), social investigation (Le Moal et al., 1987; Rigney et al., 2019, Rigney et al., 2021), and pair-bonding (Pitkow et al., 2001; Lim et al., 2004b; Lim and Young, 2004). Although most of these studies have focused on the role of AVP signaling in regulating social behaviors in adult rodents, recent studies provide evidence that AVP signaling is involved in regulating social behaviors in juvenile rats, specifically juvenile social play behavior (Veenema et al., 2013; Bredewold et al., 2014; Paul et al., 2014; Reppucci et al., 2018). Social play behavior, also known as play-fighting or rough-and-tumble play (Panksepp and Beatty, 1980; Pellis et al., 2018; VanRyzin et al., 2020), is predominantly displayed by juveniles of various mammalian species, including rats. Social play is the earliest form of peer-to-peer social behavior in rats (Eckerman et al., 1975; Panksepp, 1981) and is essential for the development of social competence later in adulthood (Bekoff, 1974; Taylor, 1980; van den Berg et al., 1999). AVP signaling in different regions in the brain have been shown to regulate the expression of social play behavior in juvenile male and female rats, sometimes in sex-specific ways (Veenema et al., 2013; Paul et al., 2014). For example, administration of an AVP V1a receptor (V1aR) antagonist into the lateral septum (LS) increased social play duration in juvenile males but decreased social play duration in juvenile females (Veenema et al., 2013). In addition, AVP mRNA expression in the bed nucleus of the stria terminalis (BNST) correlated negatively with social play behavior in juvenile males, but not juvenile females (Paul et al., 2014).

One target brain region of AVP inputs is the ventral pallidum (VP; DiBenedictis et al., 2020). AVP-synthesizing neurons in the posterior bed nucleus of the stria terminalis (pBNST) and the posterodorsal region of the medial amygdala (MePD) project to the VP in adult rats (DiBenedictis et al., 2020), with a greater proportion of AVP cells in both the pBNST and MePD that project to the VP in males compared to females (DiBenedictis et al., 2020). In addition, adult males show a higher density of AVP-immunoreactive fibers in the VP compared to adult females (DiBenedictis et al., 2020), providing evidence of sex differences in the structure of the AVP system in the VP in adult rats. Furthermore, AVP signaling in the VP regulates sociosexual motivation in a sex-specific manner in adult rats. In detail, blockade of AVP signaling in the VP decreased investigation of the opposite sex in adult male rats, while increasing investigation of the opposite sex in adult female rats (DiBenedictis et al., 2020). This finding suggests that in adult rats, AVP signaling in the VP facilitates specific social behaviors in males while attenuating these behaviors in females. Together, these findings provide evidence of sex differences in structure and function of the AVP-VP system in adult rats. Here, we aimed to determine whether there are sex differences in the structure and function of the AVP-VP system in juvenile rats.

First, we examined whether there are sex differences in various structural elements of the AVP system in the VP of juvenile rats through analysis of AVP-immunoreactive (AVP-ir) fibers, V1aR binding density, and v1aR-expressing cells. Second, we determined whether there are sex differences in the proportion of V1aR-expressing VP cells co-expressing GABAergic or glutamatergic markers. Third, we determined whether and how exposure to social play alters the neuronal activation of the VP and of V1aR-expressing VP cells using fos as a marker of neuronal activity (Morgan and Curran, 1991). Lastly, we determined the effects of a specific V1aR antagonist and of synthetic AVP infused into the VP on social play expression. Based on the studies discussed above, we hypothesized that there are robust sex differences in the structural anatomy of the AVP system in the VP in juvenile rats, that social play exposure activates V1aR-expressing VP cells differently in males and females, and that the AVP-VP system regulates social play behavior differently in males and females.