Hypocretin in the nucleus accumbens shell modulates social approach in female but not male California mice

Abstract

The hypocretin (Hcrt) system modulates arousal and anxiety-related behaviors and has been considered as a novel treatment target for stress-related affective disorders. We examined the effects of Hcrt acting in the nucleus accumbens shell (NAcSh) and anterodorsal bed nucleus of the stria terminalis (adBNST) on social behavior in male and female California mice (Peromyscus californicus). In female but not male California mice, infusion of Hcrt1 into NAcSh decreased social approach. Weak effects of Hcrt1 on social vigilance were observed in both females and males. No behavioral effects of Hcrt1 infused into the adBNST were observed. Analyses of sequencing data from California mice and Mus musculus NAc showed that Hcrtr2 was more abundant than Hcrtr1, so we infused the selective Hcrt receptor 2 antagonist into the NAcSh, which increased social approach in females previously exposed to social defeat. A calcium imaging study in the NAcSh of females before and after stress exposure showed that neural activity increased immediately following the expression of social avoidance but not during freezing behavior. This observation is consistent with previous studies that identified populations of neurons in the NAc that drive avoidance. Intriguingly, calcium transients were not affected by stress. These data suggest that hypocretin acting in the NAcSh plays a key role in modulating stress-induced social avoidance.

Fig. 1. Hcrt1 infusion in the NAcSh reduced social approach and vigilance in stress-naïve female California mice.

Timeline of experiment and mechanism of action for Hcrt1 (A). Infusion of 30 ng or 300 ng Hcrt1 in the NAcSh reduced social approach in females, but not males (B). Infusion of 30 ng but not 300 ng Hcrt1 increased social vigilance in females and males (C). Intra-NAcSh infusion of 30 ng and 300 ng Hcrt1 decreased approach of a novel empty cage during the acclimation phase in both sexes (D) but had no effects on vigilance (E). Infusion of Hcrt1 in the NAcSh had no effect on distance traveled (F) or time spent in the center (G) during the open field phase. Schematics representing injection sites (red shading) of successful cannula placement in the NAcSh (H). *p < 0.05, **p < 0.01, ***p < 0.001 vs saline. Group n’s Male/saline n = 8, male/Hcrt1 30 ng n = 7, male/Hcrt1 300 ng n = 8, male/miss n = 7, female/saline n = 8, female/Hcrt1 30 ng n = 7, female/Hcrt1 300 ng n = 8, female/miss, n = 7.

Fig. 2. Selective HcrtR2 antagonist infusion in the NAcSh increased social approach but not social vigilance behaviors in stressed female California mice.

In male Mus, NAc cells were organized into 8 cell types: astrocytes (Astro), oligodendrocytes (Oligo), endothelial cells (Endo), interneurons (IN), dopamine receptor 1-expressing cells (D1), and dopamine receptor 2-expressing cells (D2). Hcrtr2 is primarily expressed in the dopamine D1 receptor medium spiny neurons and interneurons, whereas Hcrtr1 is primarily expressed in the D1 and D2 medium spiny neurons (A). A Dotplot shows that Hcrtr1 and Hcrtr2 have distinct expression patterns across all cell types in male Mus. Hcrtr1 has the highest average and percent expression in the interneurons, whereas Hcrtr2 has the highest average and percent expression in D1 neurons (B). In the nucleus accumbens but not medial prefrontal cortex or ventral tegmental area Hcrtr2 was more abundant than Hcrtr1 (C). Timeline of experiment in stressed female California mice and schematic of the mechanism of action for selective HcrtR2 antagonist TSC OX2 29 (D). Infusion of TSC OX2 29 in the NAcSh of female California mice previously exposed to social defeat stress increased social approach (E) but had no effects on social vigilance behavior (F), behaviors during the acclimation phase (G). *p < 0.05, +p = 0.06 vs. saline. ***p < 0.001 vs. Hcrtr1. Group n’s: gene expression; male/control n = 7, male/stress n = 6, female/control n = 8, female/stress n = 7, pharmacology; female/saline n = 9, female/OX2 29 n = 9.

Fig. 3. Hcrt1 infusion in the adBNST had no behavioral effects during the social interaction test in male or female California mice.

Timeline of experiment and schematic of mechanism of action for Hcrt1 (A). Infusion of 300 ng Hcrt1 in the adBNST in male and female California mice had no effects on social approach (B), acclimation approach (C), locomotion (D), social vigilance (E), acclimation vigilance (F) or center time during the open field (G). Schematic representing injection sites (red shading) of successful cannula placement in the NAcSh (H). Group n’s Male/saline n = 10, male/Hcrt1 n = 10, female/saline n = 8, female Hcrt1 n = 11.

Fig. 4. Increased nucleus accumbens shell activity coincides with behavioral avoidance.

Experimental timeline for fiber photometry observations of GCaMP6 in the NAcSh of female California mice (A). GCaMP6f signals were significantly stronger in the 2 s period following the initiation of avoidance of a target mouse (B–D). No changes in GCaMP6f signals were observed following bouts of freezing (E–G) in the presence of non-aggressive or aggressive target mice. *p < 0.05 versus baseline. Seven females were used for these analyses.