Role of mPFC and nucleus accumbens circuitry in modulation of a nicotine plus alcohol compound drug state

Abstract

Combined use of nicotine and alcohol constitute a significant public health risk. An important aspect of drug use and dependence are the various cues, both external (contextual) and internal (interoceptive) that influence drug-seeking and drug-taking behavior. The present experiments employed the use of Designer Receptors Exclusively Activated by Designer Drugs (DREADDs) and complementary Pavlovian drug discrimination procedures (feature-positive and feature-negative training conditions) in order to examine whether medial prefrontal cortex (prelimbic; mPFC-PL) projections to the nucleus accumbens core (AcbC) modulate sensitivity to a nicotine + alcohol (N + A) interoceptive cue. First, we show neuronal activation in mPFC-PL and AcbC following treatment with N + A. Next, we demonstrate that chemogenetic silencing of projections from mPFC-PL to nucleus accumbens core decrease sensitivity to the N + A interoceptive cue, while enhancing sensitivity to the individual components, suggesting an important role for this specific projection. Furthermore, we demonstrate that clozapine-N-oxide (CNO), the ligand used to activate the DREADDs, had no effect in parallel mCherry controls. These findings contribute important information regarding our understanding of the cortical-striatal circuitry that regulates sensitivity to the interoceptive effects of a compound N + A cue.

Keywords: addiction; chemogenetic; drug-discrimination; interoception; poly-drug.

Figure 1

Diagram of Pavlovian discrimination training procedures. (A) Feature positive trained rats received 0.1 ml of 26% sucrose following stimulus light offset on N+A sessions. On water sessions, sucrose was not presented. (B) Feature negative trained rats received 0.1 ml of 26% sucrose following stimulus light offset on water sessions. On N+A session, sucrose was not presented.

Figure 2

c-Fos expression in medial prefrontal cortex – prelimbic region (mPFC; A), nucleus accumbens core (AcbC; B) and nucleus accumbens shell (AcbSh; C) following water or 0.4N+1.0A (0.4 mg/kg nicotine + 1 g/kg alcohol, IG) in rats trained to discriminate N+A from water. Bars are mean (±SEM) c-Fos positive pixels/mm². There was significantly more c-Fos expression in mPFC and AcbC following N+A compared to water. Representative photomicrographs appear above each figure. Scale bars = 250 µm. * - N+A treatment significantly greater than water treatment (p < 0.05).

Figure 3

Substitution curves for the Feature Positive group following mPFC-PL→AcbC silencing. N = 6 for the hM4D group and n = 7 for the mCherry control group. (A) Representative expression maps for hM4D (red) and mCherry (blue), (B) cannulae placements for hM4D (red dots) and mCherry (blue dots) and (C) representative photomicrographs showing representative DREADD and mCherry (top panels, 2X and 20X images, scale bar is 1000 µm) and cannula placements (bottom panels, arrows indicate position of AcbC injector). (D) Mean(+S.E.M.) discrimination score (head entries during the single 15-s light CS minus head entries during 15 seconds before light onset) for the N+A substitution test. CNO significantly decreased discrimination score at the training dose (0.4N/1.0A) compared to vehicle and the mCherry control group suggesting that mPFC→AcbC projections are important for modulating sensitivity to N+A. (E) Mean(+S.E.M.) discrimination score for the nicotine only substitution test. (F) Mean(+S.E.M.) discrimination score for the alcohol only substitution test. CNO significantly increased discrimination score at 0.1, 0.3 and 1.0 g/kg alcohol doses compared to vehicle and mCherry control suggesting that the alcohol component of the compound cue is particularly sensitive to modulation by mPFC→AcbC projections. Solid lines indicate mean discrimination score from 2 water sessions prior to testing. Dashed lines indicate mean discrimination score from 2 N+A sessions prior to testing. +-denotes vehicle condition is significantly different from N+A baseline (p<0.05); *-denotes significant difference from vehicle and mCherry control (p < 0.05).

Figure 4

Substitution curves for the Feature Negative group following mPFC-PL→AcbC silencing. N = 7 for the hM4D group and n = 7 for the mCherry control group. A) Representative expression maps for hM4D (red) and mCherry (blue), B) cannulae placements for hM4D (red dots) and mCherry (blue dots) and (C) representative photomicrographs showing representative DREADD and mCherry (top panels, 2X and 20X images, scale bar is 1000 µm) and cannula placements (bottom panels, arrows indicate position of AcbC injector). (D) Mean (+S.E.M.) discrimination score (head entries during the single 15-s light CS minus head entries during 15 seconds before light onset) for the N+A substitution test. (E) Mean (+S.E.M.) discrimination score for the nicotine only substitution test. CNO significantly decreased discrimination score at the 0.4 and 0.8 mg/kg nicotine doses compared to vehicle and mCherry control suggesting that in FN trained rats, sensitivity to the nicotine component of the compound cue is sensitive to modulation by mPFC→AcbC projections. (F) Mean (+S.E.M.) discrimination score for the alcohol only substitution test. Solid lines indicate mean discrimination score from 2 water sessions prior to testing. Dashed lines indicate mean discrimination score from 2 N+A sessions prior to testing. +-denotes vehicle condition is significantly different from N+A baseline (p<0.05); *-denotes significant difference from vehicle and mCherry control (p<0.05).