Ventral medial prefrontal cortex neuronal ensembles mediate context-induced relapse to heroin

Abstract

In a rat model of context-induced relapse to heroin, we identified sparsely distributed ventral medial prefrontal cortex (mPFC) neurons that were activated by the heroin-associated context. Selective pharmacogenetic inactivation of these neurons inhibited context-induced drug relapse. A small subset of ventral mPFC neurons formed neuronal ensembles that encode the learned associations between heroin reward and heroin-associated contexts; re-activation of these neuronal ensembles by drug-associated contexts during abstinence provoked drug relapse.

Figure 1. Context-induced reinstatement of heroin seeking is associated with Fos induction in dorsal and ventral mPFC

(a) Reinstatement test: Left column: Total number of active lever and inactive lever presses in rats in which heroin seeking was extinguished in the same (Control A-A-A) or different (Renewal A-B-A) context as their heroin self-administration context. ANOVA showed a significant Experimental Group (Control [AAA], Renewal [ABA]) × Lever (Active, Inactive) interaction effect (F_1,18=39.9, p=0.0001). Right column: Time course of active lever-presses. (b) Fos: Number of Fos-immunoreactive nuclei per mm² in dorsal and ventral mPFC (right panel: area of quantification). ANOVA shows a significant effect of Experimental Group (F_1,18=9.4, p=0.007), but not mPFC region (dorsal, ventral) or Experimental Group × mPFC region. (c) Fos+NeuN double labeling: Representative photomicrographs of Fos+NeuN for dorsal and ventral mPFC (see small squares in the picture in 1b for approximate areas). Data are depicted as mean±SEM. * Different from Control, p<0.05, n=9–11 per group in 1a and 1b, and n=4 per group in 1c.

Figure 2. Non-selective global inhibition of the majority of ventral but not dorsal mPFC neurons by muscimol+baclofen decreased context-induced reinstatement of heroin seeking

Total lever-presses (left panel): Number of active lever-presses after bilateral injections of vehicle or muscimol+baclofen (0.03 nmol+0.3 nmol/side) into dorsal (a) or ventral (b) mPFC 5–10 min before exposure to the heroin or extinction context. ANOVA showed a significant Drug Dose (vehicle or muscimol+baclofen) × mPFC Area (dorsal or ventral) × Test Context (Heroin [A] or Extinction [B]) interaction effect (F_1,29=4.0, p=0.05). Time course (right panel): Number of active lever-presses (c) Cannulae placement: Approximate placement and representative pictures of injector tips. Data are depicted as mean±SEM. * Different from Vehicle, p<0.05, n=7–10 per group. Cg1, cingulate area 1; PrL, prelimbic cortex; IL, infralimbic cortex.

Figure 3. Ventral mPFC Daun02 injections after exposure to the heroin-associated context during induction day decreased subsequent context-induced reinstatement of heroin seeking

(a) Timeline of the experimental procedure: Heroin self-administration training (A context), extinction of heroin seeking (B context), induction (A or B context), and context-induced reinstatement test (A context). During the “induction” day, Daun02 (2 µg/side) or vehicle was injected into the ventral mPFC 90 min after 30 min exposure to the Heroin (A) context or the Extinction (B) context. (b–c) Daun02 injections in the heroin-associated context or the extinction-associated context: Total active lever-presses and ventral mPFC β-gal expression. ANOVAs of active lever-presses and β-gal counts revealed significant Daun02 Condition (vehicle, Daun02) × Induction Day Context (Heroin [A] context, Extinction [B] context) interaction effects (F_1,53=12.6, p=0.001 and F_1,53=4.3, p=0.042, respectively). (d) Representative images of X-gal staining. Visualization of β-gal -labeled nuclei in ventral mPFC. Dotted red lines indicate approximate area of injector tip. Data are depicted as mean±SEM. * Different from Vehicle, p<0.05, n=10-18 per group.