miR-218 in Adolescence Predicts and Mediates Vulnerability to Stress

Abstract

Background: Adolescence is a period of increased vulnerability to psychiatric disorders, including depression. Discovering novel biomarkers to identify individuals who are at high risk is very much needed. Our previous work shows that the microRNA miR-218 mediates susceptibility to stress and depression in adulthood by targeting the netrin-1 guidance cue receptor gene Dcc in the medial prefrontal cortex (mPFC).

Methods: Here, we investigated whether miR-218 regulates Dcc expression in adolescence and could serve as an early predictor of lifetime stress vulnerability in male mice.

Results: miR-218 expression in the mPFC increases from early adolescence to adulthood and correlates negatively with Dcc levels. In blood, postnatal miR-218 expression parallels changes occurring in the mPFC. Notably, circulating miR-218 levels in adolescence associate with vulnerability to social defeat stress in adulthood, with high levels associated with social avoidance severity. Indeed, downregulation of miR-218 in the mPFC in adolescence promotes resilience to stress in adulthood.

Conclusions: miR-218 expression in adolescence may serve both as a marker of risk and as a target for early interventions.

Keywords: Biomarkers; Chronic social defeat stress; Guidance cues; Netrin-1/DCC pathway; Prefrontal cortex; Resilience.

FIGURE 1.. Developmental expression of miR-218 and Dcc and mRNA in the mPFC.

(A) Timeline of experiment. (B) miR-218 increases from early adolescence (PND21) to adulthood (PND75±15). One-way ANOVA: F_(2,12) = 3.9; p<0.05; Tukey test: *p<0.05; different from PND 21. (C) Dcc mRNA decreases in postnatal life. One-way ANOVA: F_(2,12) = 7.55; p<0.01; Tukey test: *p<0.05; different from PND 21; †p<0.01; different from PND 35. (D) Negative correlation between mPFC levels of Dcc mRNA and miR-218 across the lifespan. (E) The expression of Robo-1 mRNA does not change from adolescence to adulthood. One-way ANOVA: F_(2,12) = 0.35; p=0.71. (F) miR-218 expression in blood increases across the lifespan. One-way ANOVA: F_(2,16)= 4.341; p<0.05; Tukey test: *p<0.05; different from PND 21.

FIGURE 2.. Levels of miR-218 in blood during adolescence correlate with susceptibility to CSDS in adulthood.

(A) Timeline of blood collection and CSDS protocol. (B) Time in the interaction zone in control (CON=9), susceptible (SUS=10), resilient (RES=8) mice. Two-way ANOVA: significant stress by target interaction: F_(2,48)= 9.44; p<0.001; stress: F(2,48)= 1.43; p=0.2; session: F(1,48)= 0.01; p=0.9. Post hoc Tukey test: **p<0.01; different from CON and RES, in session with target present. Inset: Social interaction ratio: One-way ANOVA: F_(2,24)= 14.02; p<0.0001; Tukey test: **p<0.01; different from CON and RES. (C) Blood levels of miR-218 in adolescence and following CSDS in adulthood. All samples were segregated retrospectively into CON, SUS, and RES groups according to the social interaction ratio as in(31,32). Expression was normalized to CON in Adolescent. Two-way ANOVA: Age by stress interaction: F_(2,48)= 7.18; p<0.01; age: F_(1,48)= 1.06; p= 0.31, stress: F_(2,48)= 0.040; p= 0.96. Paired t-test: *p<0.05; SUS different from CON and RES in Adolescent. †p<0.05; SUS different from CON and RES in Adult after CSDS. (D) Negative correlation between adolescent levels of miR-218 in blood and the time in the interaction zone with target present.

FIGURE 3.. Downregulation of miR-218 in adolescence potentially promotes resilience to stress in adulthood.

(A) Schematic illustration of the miR-218 antagomir (Ant-miR-218). (B) Timeline of experiment. (C) Significant downregulation of miR-218 and upregulation of Dcc mRNA in the mPFC following Ant-miR-218 infusion. miR-218: t₍₁₂₎=6.49; *p=0.0001, different from Ant-scrambled; Dcc mRNA: t₍₁₂₎=2.28; *p=0.041, different from Ant-scrambled. (D) Negative correlation of miR-218 and Dcc mRNA. (E) Time in the social interaction zone with target present: Two-way ANOVA: main effect of stress: F_(1,31)= 10.10; p=0.0033; main effect of infusion: F_(1,31)= 8.26; p= 0.0077, and stress by infusion interaction: F_(1,31)= 2.79; p=0.1). Post hoc Tukey test shows reduced time in the social interaction zone in Ant-scrambled-Defeat in comparison to Ant-scrambled-Control, *p<0.05; Ant-miR-218-Control, †p<0.01 and Ant-miR-218-Defeat, †p<0.01. (F) Percentage of susceptible (SUS), and resilient (RES) mice in each group. (G) Trend towards reduced percentage of immobility time in the FST in adult mice that were microinfused with Ant-miR-218 in adolescence (t₍₈₎=2.14; p=0.064).