Activation of mTORC1 Signaling Cascade in Hippocampus and Medial Prefrontal Cortex Is Required for Antidepressant Actions of Vortioxetine in Mice

Abstract

Background: Although thought of as a multimodal-acting antidepressant targeting the serotonin system, more molecules are being shown to participate in the antidepressant mechanism of vortioxetine. A previous report has shown that vortioxetine administration enhanced the expression of rapamycin complex 1 (mTORC1) in neurons. It has been well demonstrated that mTORC1 participates in not only the pathogenesis of depression but also the pharmacological mechanisms of many antidepressants. Therefore, we speculate that the antidepressant mechanism of vortioxetine may require mTORC1.

Methods: Two mouse models of depression (chronic social defeat stress and chronic unpredictable mild stress) and western blotting were first used together to examine whether vortioxetine administration produced reversal effects against the chronic stress-induced downregulation in the whole mTORC1 signaling cascade in both the hippocampus and medial prefrontal cortex (mPFC). Then, LY294002, U0126, and rapamycin were used together to explore whether the antidepressant effects of vortioxetine in mouse models of depression were attenuated by pharmacological blockade of the mTORC1 system. Furthermore, lentiviral-mTORC1-short hairpin RNA-enhanced green fluorescence protein (LV-mTORC1-shRNA-EGFP) was adopted to examine if genetic blockade of mTORC1 also abolished the antidepressant actions of vortioxetine in mice.

Results: Vortioxetine administration produced significant reversal effects against the chronic stress-induced downregulation in the whole mTORC1 signaling cascade in both the hippocampus and mPFC. Both pharmacological and genetic blockade of the mTORC1 system notably attenuated the antidepressant effects of vortioxetine in mice.

Conclusions: Activation of the mTORC1 system in the hippocampus and mPFC is required for the antidepressant actions of vortioxetine in mice.

Keywords: Chronic social defeat stress; chronic unpredictable mild stress; depression; mammalian target of rapamycin complex 1; vortioxetine.

Figure 1.

The experimental design of the whole study was provided.

Figure 2.

Administration of vortioxetine significantly reversed the chronic social defeat stress (CSDS)-decreased mammalian target of rapamycin complex 1 (mTORC1) signaling cascade in both the hippocampus and mPFC. (A) The antidepressant efficacy of vortioxetine in the CSDS model of depression, which was evaluated by the FST, TST, SPT, and social interaction test (n = 10). (B) The use of vortioxetine notably prevented the CSDS-induced decrease in hippocampal p-mTORC1, p-4E-BP-1, p-S6K1, p-AKT, p-ERK1/2, PSD95, and synapsin-1 expression. (C) The use of vortioxetine also fully prevented the CSDS-induced decrease in the expression of p-mTORC1, p-4E-BP-1, p-S6K1, p-AKT, p-ERK1/2, PSD95, and synapsin-1 in the mPFC (n = 5). The data are expressed as means ± SEM, **P < .01. The comparisons were made by 2-way ANOVA followed by post-hoc Bonferroni test.

Figure 3.

Administration of vortioxetine significantly reversed the chronic unpredictable mild stress (CUMS)-decreased mammalian target of rapamycin complex 1 (mTORC1) signaling cascade in both the hippocampus and mPFC. (A) The antidepressant efficacy of vortioxetine in the CUMS model of depression, which was evaluated by the FST, TST, and SPT (n = 10). (B) The usage of vortioxetine notably prevented the CUMS-induced decrease in the hippocampal p-mTORC1, p-4E-BP-1, p-S6K1, p-AKT, p-ERK1/2, PSD95, and synapsin-1 expression (n = 5). (C) The use of vortioxetine also fully prevented the CUMS-induced decrease in the expression of p-mTORC1, p-4E-BP-1, p-S6K1, p-AKT, p-ERK1/2, PSD95, and synapsin-1 in the mPFC (n = 5). The data are expressed as means ± SEM, **P < .01. The comparisons were made by 2-way ANOVA followed by post-hoc Bonferroni test.

Figure 4.

Co-administration with the pharmacological inhibitors of the mammalian target of rapamycin complex 1 (mTORC1) signaling cascade notably attenuated the antidepressant actions of vortioxetine in the chronic social defeat stress (CSDS) model. (A) Detailed schematic timeline of the experimental design is provided. (B) I.c.v. infusion of LY294002, U0126, and rapamycin all significantly blocked the downregulating effects of vortioxetine on the immobility of mice subjected to CSDS in the FST (n = 10). (C) I.c.v. infusion of LY294002, U0126, and rapamycin also blocked the downregulating effects of vortioxetine on the immobility of mice subjected to CSDS in the TST (n = 10). (D) Co-administration with LY294002, U0126, and rapamycin all attenuated the promoting effects of vortioxetine on the sucrose preference of mice subjected to CSDS (n = 10). (E) Co-administration with LY294002, U0126, and rapamycin also attenuated the promoting effects of vortioxetine on the social interaction of mice subjected to CSDS (n = 10). The data are expressed as means ± SEM, **P < .01. The comparisons were made by 1-way ANOVA followed by post-hoc Tukey test.

Figure 5.

Co-administration with the pharmacological inhibitors of the mammalian target of rapamycin complex 1 (mTORC1) signaling cascade notably attenuated the antidepressant actions of vortioxetine in the chronic unpredictable mild stress (CUMS) model. (A) Detailed schematic timeline of the experimental design is provided. (B) I.c.v. infusion of LY294002, U0126, and rapamycin all significantly blocked the downregulating effects of vortioxetine on the immobility of mice subjected to CUMS in the FST (n = 10). (C) I.c.v. infusion of LY294002, U0126, and rapamycin also blocked the downregulating effects of vortioxetine on the immobility of mice subjected to CUMS in the TST (n = 10). (D) Co-administration with LY294002, U0126, and rapamycin all attenuated the promoting effects of vortioxetine on the sucrose preference of mice subjected to CUMS (n = 10). (E) Co-administration with LY294002, U0126, and rapamycin also attenuated the promoting effects of vortioxetine on the social interaction of mice subjected to CUMS (n = 10). The data are expressed as means ± SEM, **P < .01. The comparisons were made by 1-way ANOVA followed by post-hoc Tukey test.

Figure 6.

Co-administration with the pharmacological inhibitors of the mammalian target of rapamycin complex 1 (mTORC1) signaling cascade notably attenuated the enhancing effects of vortioxetine on this cascade in mouse models of depression. (A) Mice treated with CSDS + vortioxetine + LY294002/U0126/rapamycin showed significantly less protein expression of p-mTORC1, p-4E-BP-1, and p-S6K1 in the hippocampus than mice treated with CSDS + vortioxetine (n = 5). (B) Mice treated with CSDS + vortioxetine + LY294002/U0126/rapamycin also showed less protein expression of p-mTORC1, p-4E-BP-1, and p-S6K1 in the mPFC than mice treated with CSDS + vortioxetine (n = 5). (C) Mice subjected to CUMS + vortioxetine + LY294002/U0126/rapamycin exhibited evidently lower protein levels of p-mTORC1, p-4E-BP-1, and p-S6K1 in the hippocampus than mice subjected to CUMS + vortioxetine (n = 5). (D) Mice subjected to CUMS + vortioxetine + LY294002/U0126/rapamycin also exhibited lower protein levels of p-mTORC1, p-4E-BP-1, and p-S6K1 in the mPFC than mice subjected to CUMS + vortioxetine (n = 5). The data are expressed as means ± SEM, **P < .01. The comparisons were made by 1-way ANOVA followed by post-hoc Tukey test.

Figure 7.

Lentiviral-mammalian target of rapamycin complex 1-short hairpin RNA-enhanced green fluorescence protein (LV-mTORC1-shRNA-EGFP) was stereotactically injected into both the hippocampus and medial prefrontal cortex (mPFC) regions of mice, and its knockdown efficacy was confirmed by western blotting. (A) The fluorescence of a fixed hippocampus slice that expressed mTORC1-shRNA. Corresponding western-blot analysis shows the knockdown effects of mTORC1-shRNA on mTORC1 in the hippocampus (n = 5). (B) The fluorescence of a fixed mPFC slice that expressed mTORC1-shRNA. Corresponding western-blot analysis shows the knockdown effects of mTORC1-shRNA on mTORC1 in the mPFC (n = 5). The scale bar is 200 μm for representative images and 25 μm for enlarged images. The data are expressed as means ± SEM, **P < .01. The comparisons were made by 1-way ANOVA followed by post-hoc Tukey test.

Figure 8.

Co-administration with mammalian target of rapamycin complex 1-short hairpin RNA fully abolished the antidepressant actions of vortioxetine in the chronic social defeat stress (CSDS) model. (A) Detailed schematic timeline of the experimental design is provided. (B) Stereotactic infusion of mTORC1-shRNA notably prevented the protecting effects of vortioxetine against the CSDS-induced behavior of helplessness in mice in the FST (n = 10). (C) Stereotactic infusion of mTORC1-shRNA also prevented the protecting effects of vortioxetine against the CSDS-induced behavior of helplessness in mice in the TST (n = 10). (D) The use of mTORC1-shRNA significantly blocked the protecting effects of vortioxetine against the CSDS-induced behavior of anhedonia in the SPT (n = 10). (E) The use of mTORC1-shRNA also blocked the protecting effects of vortioxetine against the CSDS-induced behavior of social avoidance in the social interaction test (n = 10). The data are expressed as means ± SEM, **P < .01. The comparisons were made by 1-way ANOVA followed by post-hoc Tukey test.

Figure 9.

Co-administration with mammalian target of rapamycin complex 1-short hairpin RNA fully abolished the antidepressant actions of vortioxetine in the chronic unpredictable mild stress (CUMS) model. (A) Detailed schematic timeline of the experimental design is provided. (B) Stereotactic infusion of mTORC1-shRNA notably prevented the reversal effects of vortioxetine against the CUMS-induced behavioral changes in mice in the FST (n = 10). (C) Stereotactic infusion of mTORC1-shRNA also prevented the reversal effects of vortioxetine against the CUMS-induced behavioral changes in mice in the TST (n = 10). (D) The use of mTORC1-shRNA significantly blocked the reversal effects of vortioxetine against the CUMS-induced behavioral changes in mice in the SPT (n = 10). The data are expressed as means ± SEM, **P < .01. The comparisons were made by 1-way ANOVA followed by post-hoc Tukey test.