Sex and hormonal status influence the anxiolytic-like effect of oxytocin in mice

Abstract

Anxiety and depression are highly prevalent psychiatric disorders, affecting approximately 18% of the United States population. Evidence indicates that central oxytocin mediates social cognition, social bonding, and social anxiety. Although it is well-established that oxytocin ameliorates social deficits, less is known about the therapeutic effects of oxytocin in non-social contexts. We hypothesized that positive effects of oxytocin in social contexts are attributable to intrinsic effects of oxytocin on neural systems that are related to emotion regulation. The present study investigated the effect of intracerebroventricular (ICV) oxytocin administration (i.e., central action) on anxiety- and depression-like behavior in C57Bl/6J mice using non-social tests. Male and female mice received an ICV infusion of vehicle or oxytocin (100, 200, or 500 ng), then were tested in the elevated zero maze (for anxiety-like behavior) and the tail suspension test (for depression-like behavior). Oxytocin dose-dependently increased open zone occupancy and entries in the elevated zero maze and reduced immobility duration in the tail suspension test in both sexes. Oxytocin decreased anxiety and depression-like behavior in male and female mice. The observed effect of oxytocin on anxiolytic-like behavior appeared to be driven by the males. Given the smaller anxiolytic-like effect of oxytocin in the female mice and the established interaction between oxytocin and reproductive hormones (estrogen and progesterone), we also explored whether oxytocin sensitivity in females varies across estrous cycle phases and in ovariectomized females that were or were not supplemented with estrogen or progesterone. Oxytocin reduced anxiety-like behavior in female mice in proestrus/estrus, ovariectomized females (supplemented or not with estrogen or progesterone), but not females in metestrus/diestrus. Additionally, oxytocin reduced depression-like behavior in all groups tested with slight differences across the various hormonal statuses. These results suggest that the effect of oxytocin in depression- and anxiety-like behavior in mice can be influenced by sex and hormonal status.

Keywords: Anxiety; Depression; Estrous cycle; Oxytocin; Reproductive hormones; Sex differences.

Graphical abstract

Fig. 1

Procedure & experiment timeline.

Fig. 2

Oxytocin (OXT) decreased anxiety-like behavior in the elevated zero maze, with a suggestion of a larger effect in males. Oxytocin dose-dependently increased open zone occupancy (A, B) and the number of open zone entries (C, D) in male and female mice. The plots show pooled male and female data for open zone occupancy (A) and entries (C) with females represented as ♀ and males as ♂. Separate plots for male and female open zone occupancy (B) and entries (D) are also shown. Two-way ANOVAs were calculated to analyze the male vs. female data. ^#p < 0.05, main effect of sex. The significant main effect of treatment for open zone occupancy and entries was followed by the Holm-Sidak's multiple comparison test. *p < 0.05, difference from vehicle (VEH) in pooled plots. Because the sex × treatment interaction approached significance for open zone occupancy and entries, the Holm-Sidak's multiple comparison test was used to further assess the male and female data separately; ^trp = 0.09, ^☨p < 0.05, difference from VEH in separate plots. See Section 3.1 (Results) for more details.

Fig. 3

Oxytocin decreased anxiety-like behavior in females in proestrus/estrus, but not in females in metestrus/diestrus. Oxytocin increased open zone entries (C, D), but not open zone occupancy (A, B) in pooled data. Further analysis demonstrates that oxytocin increased open zone entries in females in proestrus/estrus, but not in females in metestrus/diestrus. The plots show pooled data for females in proestrus/estrus and females in metestrus/diestrus for open zone occupancy (A) and entries (C) with female in proestrus/estrus represented as ▲ and females in metestrus/diestrus represented as ▽. Separate plots for females in proestrus/estrus and females in metestrus/diestrus in open zone occupancy (B) and entries (D) are also shown. Two-way ANOVAs were calculated to analyze females in proestrus/estrus vs. females in metestrus/diestrus data. The Holm-Sidak's multiple comparison test followed the main effect of treatment for open zone entries. *p < 0.05, difference from vehicle (VEH) in pooled plots. Because the estrous cycle phase × treatment interaction approached significance for open zone entries, Holm Sidak's multiple comparison tests were used to further assess the females in proestrus/estrus and females in metestrus/diestrus data separately; ^☨p < 0.05, difference from VEH in separate plots. See Section 3.2 (Results) for details. Representative pictures of vaginal cytology across estrous cycle phases are shown in E (proestrus), F (estrus), G (metestrus), and H (diestrus).

Fig. 4

Oxytocin decreased anxiety-like behavior in ovariectomized (OVX) females in the elevated zero maze. The pooled data demonstrate that oxytocin increased open zone occupancy and open zone entries in the elevated zero maze. The plots show pooled data for non-supplemented, estrogen-supplemented and progesterone-supplemented OVX females for open zone occupancy (A) and entries (C) with non-supplemented ovariectomized females represented as ○, estrogen-supplemented ovariectomized females represented as □, and progesterone-supplemented ovariectomized females represented as ◇. Separate plots for non-supplemented, estrogen-supplemented, and progesterone-supplemented ovariectomized females in open zone occupancy (B) and entries (D) are also shown. Two-way ANOVAs were calculated to analyze the non-supplemented vs. estrogen-supplemented vs. progesterone-supplemented ovariectomized female data. Holm-Sidak's multiple comparison tests were used to follow-up the significant main effect of treatment for open zone occupancy and entries. *p < 0.05, ^trp = 0.07, difference from vehicle (VEH) in pooled plots. See Section 3.2 (Results) for details.

Fig. 5

Oxytocin (OXT) decreased depression-like behavior in the tail suspension test in male and female mice. Both males and females exhibited a significant decrease in immobility duration following treatment with oxytocin (A, B). The plots show male and female pooled data (A) using ♂ to represent males and ♀ to represent females. Separate data for males and females are also shown (B). Two-way ANOVAs were calculated to analyze the male vs. female data; ^#p < 0.05, main effect of sex. The Holm-Sidak's multiple comparison test followed the significant main effect of treatment for immobility duration. *p < 0.05, difference from vehicle (VEH) in pooled plots. See Section 3.3 (Results) for details.

Fig. 6

Oxytocin (OXT) decreased depression-like behavior in the tail suspension test in female mice in proestrus/estrus and metestrus/diestrus phases of the estrous cycle. Oxytocin reduced immobility duration of females regardless of estrous cycle. The plots show pooled data (A) for females in proestrus/estrus, represented as ▲, and females in metestrus/diestrus, represented as ▽. Separate plots for females in proestrus/estrus and females in metestrus/diestrus are also shown (B). Two-way ANOVAs were calculated to analyze the females in proestrus/estrus vs. females in metestrus/diestrus data. The Holm-Sidak's multiple comparison test followed the main effect of treatment for immobility duration.*p < 0.05, difference from vehicle (VEH) in pooled plots. Because the estrous cycle phase × treatment interaction approached significance for immobility duration, Holm-Sidak's multiple comparison tests were used to further assess the females in proestrus/estrus and females in metestrus/diestrus data separately; ^☨p < 0.05, difference from VEH in separate plots. See Section 3.4 (Results) for details.

Fig. 7

Oxytocin decreased depression-like behavior in the tail suspension test in ovariectomized female mice. Pooled data demonstrate that oxytocin reduce immobility duration for all ovariectomized female groups tested. The plots show pooled data for ovariectomized female mice, supplemented or not (○) with estrogen (□) or progesterone (◇) for immobility duration (A). Separate plots for non-supplemented, estrogen-supplemented, and progesterone-supplemented ovariectomized female mice are also shown (B). Two-way ANOVAs were calculated to analyze the non-supplemented vs. estrogen-supplemented vs. progesterone-supplemented ovariectomized female data. The Holm-Sidak's multiple comparison test followed the significant main effect of treatment for immobility duration. *p < 0.05, difference from vehicle (VEH) in pooled plots. Because the hormone supplementation × treatment interaction approached significance for open zone entries, Holm-Sidak's multiple comparison tests were used to further assess the non-supplemented, estrogen-supplemented, and progesterone-supplemented ovariectomized female data separately; ^☨p < 0.05, difference from VEH in separate plots. See Section 3.4 (Results) for details.

Fig. 8

Sex and hormonal status influence baseline anxiety-like behavior in the elevated zero maze, but not baseline depression-like behavior in the tail suspension test. There was a significant effect of sex, estrous cycle, and hormone supplementation on open zone entries in vehicle-treated mice. Naturally cycling female mice demonstrated increased open zone entries compared to male mice. Whereas ovariectomized female mice (supplemented or not with estrogen or progesterone) demonstrated decreased open zone entries compared to naturally cycling female mice and not significant difference compared to male mice. The open zone occupancy measure in the elevated zero maze and the immobility duration measure in the tail suspension test demonstrated no significant effect of sex, estrous cycle or hormone supplementation in vehicle-treated mice. The plots represent baseline open zone occupancy, open zone entries and immobility duration of males (♂), females (♀), females in proestrus/estrus (P/E females; ▲), females in metestrus/diestrus (M/D females; ▽), non-supplemented ovariectomized females (OVX; ○), estrogen supplemented ovariectomized females (OVX + E2; □) and progesterone supplemented ovariectomized females (OVX + P4; ◇). One-way ANOVAs were used to analyze the effect of experimental group on baseline open zone occupancy, entries, and immobility duration. The Holm’Sidak's multiple comparison test followed the significant main effect of experimental group for open zone entries. *p < 0.05, compared with males; ^☨p < 0.05, ^trp = 0.05, compared with females. See Section 3.5 (Results) for details.