Medial anterior prefrontal cortex stimulation downregulates implicit reactions to threats and prevents the return of fear

Abstract

Downregulating emotional overreactions toward threats is fundamental for developing treatments for anxiety and post-traumatic disorders. The prefrontal cortex (PFC) is critical for top-down modulatory processes, and despite previous studies adopting repetitive transcranial magnetic stimulation (rTMS) over this region provided encouraging results in enhancing extinction, no studies have hitherto explored the effects of stimulating the medial anterior PFC (aPFC, encompassing the Brodmann area 10) on threat memory and generalization. Here we showed that rTMS over the aPFC applied before threat memory retrieval immediately decreases implicit reactions to learned and novel stimuli in humans. These effects enduringly persisted 1 week later in the absence of rTMS. No effects were detected on explicit recognition. Critically, rTMS over the aPFC resulted in a more pronounced reduction of defensive responses compared to rTMS targeting the dorsolateral PFC. These findings reveal a previously unexplored prefrontal region, the modulation of which can efficiently and durably inhibit implicit reactions to learned threats. This represents a significant advancement toward the long-term deactivation of exaggerated responses to threats.

Keywords: TMS; anterior prefrontal cortex; dorsolateral prefrontal cortex; human; neuroscience; threat generalization; threat memory; transcranial magnetic stimulation.

Figure 1.. Schematic diagram depicting the experimental outline and repetitive transcranial magnetic stimulation (rTMS) conditions.

In the first session (day 1, context A), participants underwent a single-cue threat conditioning in which a tone (CS) was paired with a mild electrical shock (US). In the second session (day 8, context B), a 1 Hz-rTMS procedure was actively applied over the medial anterior prefrontal cortex (aPFC, n = 30; aPFC-E, n = 21), sham-applied over the same site (sham, n = 30; sham-E, n = 21), actively applied over the left occipital cortex (OC, n = 30) and over the left dorsolateral prefrontal cortex (dlPFC, n = 30). In the implicit conditions (aPFC, sham, OC, dlPFC), subjects underwent an implicit test during which they were presented with the CS and two new stimuli (NS₁ and NS₂) and then an unconditioned threat test while being recorded in their skin conductance responses (SCRs). In the explicit conditions (aPFC-E, sham-E), participants underwent an explicit two-alternative forced-choice (2AFC) recognition task during which they were presented with tone pairs each composed of the CS and one of the two NSs, and they were asked to recognize the CS providing a confidence level for each choice. Last, participants underwent a 2AFC perceptual discrimination test, in which they had to judge whether the two tones in each pair (CS and/or NSs) were ‘the same tone’ or ‘different tones’. The third session (day 15, context A) was identical to the second one except for the absence of the rTMS.

Figure 2.. Effects of repetitive transcranial magnetic stimulation (rTMS) over the anterior prefrontal cortex (aPFC) on immediate and remote implicit threat memory, threat generalization to new stimuli, and overall electrodermal responsivity.

(A) Simulation of rTMS effects on the neural tissue of the medial aPFC (medial Brodmann area 10 [BA 10]), performed with SimNIBS 4.0 software. The magnitude of the electric field is expressed in V/m. (B, C) Dot plot and line chart representing the mean skin conductance responses (SCRs) elicited by the CS during the conditioning session and test session in the two different conditions. Groups’ reactions were not different during the conditioning phase, whereas during the test phase the group stimulated over the aPFC (n = 30) showed attenuated implicit reactions relative to the sham condition (n = 30). The aPFC group displayed reduced autonomic reactions to the CS from conditioning to test, while the sham group showed an increase in defensive responses. (D, E) Implicit reactions to all the tones (NS₁, CS, and NS₂) during the test session were decreased in the aPFC group relative to the sham group. Although we found a significant main effect of group and no group × tone interaction effect, we reported the statistical significance marks of simple main effects. (F) Implicit reactions to the US₂ during the test session were not different between conditions, showing no rTMS effects on the overall electrodermal responsivity. (G, H) In the follow-up session, the aPFC group enduringly demonstrated reduced implicit reactions to the CS relative to the sham group and to the conditioning phase. (I) Implicit reactions to the US₂ during the follow-up session were not different between groups. *p<0.05, **p<0.01, ***p<0.001. All data are mean and SEM. 2 × 2 mixed ANOVA followed by Bonferroni-adjusted post hoc comparisons (B, C, G, H); 2 × 3 mixed ANOVA followed by Bonferroni-adjusted post hoc comparisons (D, E); Student’s unpaired t-test (F, I).

Figure 2—figure supplement 1.. Implicit reactions during preconditioning (CS) and conditioning (CS, US) in the anterior prefrontal cortex (aPFC) and sham groups.

(A) Dot plot representing the mean skin conductance responses (SCRs) elicited by the CS during the preconditioning phase in the aPFC group (n = 30) compared with the sham group (n = 30). Implicit reactions were not significantly different. (B) Mean SCRs elicited by the US during the conditioning phase in the aPFC group compared with the sham group. Responses were not significantly different. (C) Mean SCRs evoked by the CS over the 15 trials of the conditioning phase, and averaged SCRs (trials 1–15) in the aPFC and sham groups. Autonomic reactions were not significantly different. All data are mean and SEM. Student’s unpaired t-test (A–C); 2 × 15 mixed ANOVA (C).

Figure 2—figure supplement 2.. Effects of a discomfort-inducing procedure on immediate and remote implicit threat memory.

(A) Mean discomfort ratings provided by the subjects of the ctrl discomfort group (n = 10) during the 10 min discomfort-inducing procedure. (B) Dot plot representing the mean skin conductance responses (SCRs) elicited by the CS during the conditioning phase and the test phase in the ctrl discomfort group and the sham group (n = 30). The groups’ reactions were not significantly different during the conditioning phase as well as during the test phase (2 × 2 mixed ANOVA; main effect of group: F_(1,38) = 0.712, p=0.404; main effect of phase: F_(1,38) = 1.713, p=0.198; group × phase interaction: F_(1,38) = 0.956, p=0.335). (C) In the follow-up session, SCRs to the CS did not differ between groups (2 × 2 mixed ANOVA; main effect of group: F_(1,38) = 1.335, p=0.255; main effect of phase: F_(1,38) = 0.042, p=0.838; group × phase interaction: F_(1,38) = 0.175, p=0.678). All data are mean and SEM.

Figure 3.. Effects of repetitive transcranial magnetic stimulation (rTMS) over the anterior prefrontal cortex (aPFC) on immediate and remote explicit threat memory.

(A) During the test session, explicit recognition patterns were not different between the group stimulated over the aPFC (n = 21) and the sham group (n = 21). (B) During the test session, confidence ratings did not differ between the two conditions. (C) During the follow-up session, aPFC-E and sham-E groups identified the CS between the NSs in a not different manner. (D) During the follow-up session, aPFC-E and sham-E groups were not differently confident about their explicit choices. All data are mean and SEM. Student’s unpaired t-test (A–D).

Figure 4.. Selective effects of repetitive transcranial magnetic stimulation (rTMS) over the anterior prefrontal cortex (aPFC) and the left occipital cortex (OC) on the defensive responses to threat-predictive cues.

(A) Simulation of rTMS effects on the neural tissue of the left OC (BA 18/19), performed with SimNIBS 4.0 software. The magnitude of the electric field is expressed in V/m. (B, C) Dot plot and line chart representing the mean skin conductance responses (SCRs) elicited by the CS during the conditioning session and test session in the OC group (n = 30) compared with the same aPFC group of Figure 2 (n = 30). The two groups did not differently respond during the conditioning phase, but during the test phase the group stimulated over the aPFC showed weaker reactions than the OC group. While the defensive reactions of the aPFC group decreased from conditioning to test, those of the OC group remained not differently high. (D, E) Implicit reactions to NSs during the test session did not differ between groups. In the OC group, the responses elicited by the NS₂ were lower than those evoked by the CS and the NS₁. (F) Implicit reactions to the US₂ during the test session were not different between groups. (G, H) In the follow-up session, the aPFC group persisted in showing reduced implicit reactions to the CS relative to the OC group. Defensive reactions of both groups decreased from the conditioning phase. Although we found a significant main effect of phase and no group × phase interaction effect, we reported the statistical significance marks of simple main effects. (I) Implicit reactions to the US₂ during the follow-up session were not different between groups. *p<0.05, ***p<0.001. All data are mean and SEM. 2 × 2 mixed ANOVA followed by Bonferroni-adjusted post hoc comparisons (B, C, G, H); 2 × 3 mixed ANOVA followed by Bonferroni-adjusted post hoc comparisons (D, E); Student’s unpaired t-test (F, I).

Figure 4—figure supplement 1.. Implicit reactions during preconditioning (CS) and conditioning (CS, US) in the anterior prefrontal cortex (aPFC) and occipital cortex (OC) groups.

(A) Dot plot representing the mean skin conductance responses (SCRs) elicited by the CS during the preconditioning phase in the aPFC group (n = 30) compared with the OC group (n = 30). Implicit reactions were not significantly different. (B) Mean SCRs elicited by the US during the conditioning phase in the aPFC group compared with the OC group. Responses were not significantly different. (C) Mean SCRs evoked by the CS over the 15 trials of the conditioning phase, and averaged SCRs (trials 1–15) in the aPFC and OC groups. Autonomic reactions were not significantly different. All data are mean and SEM. Student’s unpaired t-test (A–C); 2 × 15 mixed ANOVA (C).

Figure 5.. Different effects of repetitive transcranial magnetic stimulation (rTMS) over the anterior prefrontal cortex (aPFC) and the left dorsolateral prefrontal cortex (dlPFC) on immediate and remote implicit threat memory.

(A) Simulation of rTMS effects on the neural tissue of the left dlPFC (BA 8/9), performed with SimNIBS 4.0 software. The magnitude of the electric field is expressed in V/m. (B, C) Dot plot and line chart representing the mean skin conductance responses (SCRs) elicited by the CS during the conditioning session and test session in the dlPFC group (n = 30) compared with the same aPFC group of Figure 2 (n = 30). The two conditions did not differently react during the conditioning phase, whereas during the test phase the group stimulated over the aPFC displayed lower reactions than the dlPFC group. Implicit reactions of the aPFC group decreased from conditioning to test, while those of the dlPFC group increased. (D, E) Implicit reactions to NSs during the test session did not differ between groups. In the dlPFC group, the responses elicited by the NS₂ were lower than those evoked by the CS. (F) The two groups did not differently react to the US₂ during the test session. (G, H) In the follow-up session, the aPFC group persisted in more dimly reacting to the CS relative to the dlPFC group and to the conditioning phase. (I) Implicit reactions to the US₂ during the follow-up session were not different between groups. *p<0.05, **p<0.01, ***p<0.001. All data are mean and SEM. 2 × 2 mixed ANOVA followed by Bonferroni-adjusted post hoc comparisons (B, C, G, H); 2 × 3 mixed ANOVA followed by Bonferroni-adjusted post hoc comparisons (D, E); Student’s unpaired t-test (F, I).

Figure 5—figure supplement 1.. Implicit reactions during preconditioning (CS) and conditioning (CS, US) in the anterior prefrontal cortex (aPFC) and dorsolateral prefrontal cortex (dlPFC) groups.

(A) Dot plot representing the mean skin conductance responses (SCRs) elicited by the CS during the preconditioning phase in the aPFC group (n = 30) compared with the dlPFC group (n = 30). Implicit reactions were not significantly different. (B) Mean SCRs elicited by the US during the conditioning phase in the aPFC group compared with the dlPFC group. Responses were not significantly different. (C) Mean SCRs evoked by the CS over the 15 trials of the conditioning phase, and averaged SCRs (trials 1–15) in the aPFC and dlPFC groups. Autonomic reactions were not significantly different. All data are mean and SEM. Student’s unpaired t-test (A–C); 2 × 15 mixed ANOVA (C).