Deep brain stimulation of the anterior cingulate cortex reduces opioid addiction in preclinical studies

Abstract

Substance Use Disorder (SUD) is a medical condition where an individual compulsively misuses drugs or alcohol despite knowing the negative consequences. The anterior cingulate cortex (ACC) has been implicated in various types of SUDs, including nicotine, heroin, and alcohol use disorders. Our research aimed to investigate the effects of deep brain stimulation (DBS) in the ACC as a potential therapeutic approach for morphine use disorder. Additionally, we measured c-Fos protein expression as an indicator of neural activity in the nucleus accumbens (NAc) and prefrontal cortex (PFC). Our findings indicate that high-frequency (130 Hz) DBS at different amperages, 150 µA and 200 µA in the ACC during the acquisition phase of morphine conditioned place preference (CPP) inhibited the rewarding properties of morphine. Furthermore, DBS at these intensities during the extinction phase facilitated the extinction and mitigated the reinstatement of morphine CPP triggered by drug priming. Morphine conditioning was associated with impaired novel object conditioning (NOR) and locomotor activity. While DBS in the acquisition and extinction phases at both intensities restored NOR memory, only DBS at 200 µA recovered locomotor activity in the open field test. Treatment with DBS at 200 µA decreased c-Fos protein expression in the NAc and PFC (compared to morphine-only group). In conclusion, our data indicate an intensity-dependent effect of ACC DBS on the acquisition, extinction, and reinstatement of morphine-induced CPP in rats. These findings suggest that ACC DBS could be a potential intervention for the treatment of morphine use disorder.

Keywords: Addiction; Anterior cingulate cortex; Deep brain stimulation; Morphine.

Fig. 1

Histological verification of electrode placement in the anterior cingulate cortex (ACC). This coronal brain section illustrates the precise placement of the electrode in the anterior cingulate area (ACA) of a rat. The electrode track is clearly observed within the ACA, a region critical for the study. Key anatomical landmarks, including the Corpus Callosum (CC), caudoputamen (CP), somatomotor area (MO), lateral septal nucleus (LS), and lateral ventricle (VL), are labeled to provide context. This histological evidence ensures the accuracy of electrode targeting, validating the subsequent data obtained from the deep brain stimulation (DBS) experiments.

Fig. 2

The acquisition phase of CPP. (A) Administration of morphine (in sham-DBS group), but not saline, during the conditioning (COND) period resulted in a greater preference for the drug-paired chamber. (B) The alterations in the side preference observed in morphine-conditioned groups over the whole behavioral session. Administering morphine to rats that did not receive DBS (DBS-0) resulted in a noticeable and long-lasting preference for the drug-paired side. Nevertheless, DBS at both 150 µA and 200 µA effectively inhibited the formation of morphine CPP. Reinstatement (REINS) was tested on day 12 in DBS-150 C and DBS-200 C groups and on day 21 in DBS-0 group (see Fig. 3B; data in DBS-0 group are from the homonymous group illustrated in Fig. 3B) after three days of performing open field (OF) and novel object recognition (NOR) tests. (C) The average side preference in each phase across the groups that present statistical proof (2-way repeated measures ANOVA and Bonferroni’s test) for the data shown in (B). Results are shown as mean ± SEM (n = 6 rats in Saline group, 14 in Morphine/DBS-0, 5 in DBS-150 C, and 10 in DBS-200 C). *p < 0.05, ****p < 0.0001.

Fig. 3

The extinction phase of CPP. (A) Changes in side preference along the whole behavioral course. Bars represent reinstatement (REINS) test performed four days after the complete extinction (EXT) of morphine CPP. (B) The average side preference in each phase of CPP across the groups that present statistical proof (2-way repeated measures ANOVA and Bonferroni’s test) for the data shown in (A). The bars for the extinction phase represent the last day of the phase when complete extinction occurred. The findings demonstrated significant differences in side preference between the post-conditioning (PostC) and the extinction phases in all groups, indicating complete extinction of morphine CPP. There was a significant difference in side preference between the extinction and reinstatement phases in the sham-DBS (DBS-0) group, but not in DBS-150E and DBS-200E groups, suggesting that ACC DBS successfully impeded the reinstatement of morphine CPP. **p < 0.01, ***p < 0.001, ****p < 0.0001. (C) The Kaplan-Meier test demonstrated that the DBS-150E and DBS-200E groups had markedly reduced survival rates for morphine CPP (time to complete extinction) compared to the sham-DBS group (median survival: 2, 2.5, and 11 days, respectively, log-rank). Results are shown as mean ± SEM (n = 14 rats in DBS-0 group, 5 in DBS-150E, and 10 in DBS-200E).

Fig. 4

Novel object recognition (NOR) and locomotor activity tests. (A) Exploration time of two identical objects (familiar objects 1 and 2) in the familiarization stage of the NOR test. (B) Exploration time of a familiar (Object 1) and a novel object (Object 3) in the choice stage of the NOR test. (C) Ordinary 1-way ANOVA and Tukey’s tests showed that morphine conditioning in the sham-DBS group resulted in a significant reduction in the recognition index as compared to the saline group. Nevertheless, all DBS groups showed a significant improvement in the recognition index when compared to the sham-DBS group. This suggests that the DBS treatment led to a complete restoration of the NOR memory. (D) 1-way ANOVA and Fisher’s LSD tests demonstrated significant decreases in spontaneous movement in the DBS-0, DBS-150 C, and DBS-150E groups when compared to the saline group. Nevertheless, DBS-200 C and DBS-200E groups had locomotor activity similar to that of saline group. Results are shown as mean ± SEM (n = 6 in Saline and DBS-0 groups, 5 in DBS-150c and DBS-150E groups, and 10 in DBS-200 C and DBS-200E groups). **p < 0.01, ****p < 0.0001 within-group comparison; ^#p < 0.05, ^##p < 0.01, ^###p < 0.001, ^####p < 0.0001 compared to the saline group; ^†p < 0.05, ^†††p < 0.001, ^††††p < 0.0001 compared to DBS-0 group.

Fig. 5

ELISA analysis of c-Fos in the NAc and PFC. The findings indicated that there was an alteration in the expression of c-Fos protein in the NAc (A) and PFC (B). The results of Dunn’s test indicated that there was no statistically significant alteration in c-Fos expression between the sham-DBS group and the saline group. However, the DBS-200 group exhibited a substantial reduction in c-Fos levels compared to the sham-DBS group. Results are shown as mean ± SEM (n = 3–5/group).