Drug-activated cells: From immediate early genes to neuronal ensembles in addiction

Abstract

Beyond their rapid rewarding effects, drugs of abuse can durably alter an individual's response to their environment as illustrated by the compulsive drug seeking and risk of relapse triggered by drug-associated stimuli. The persistence of these associations even long after cessation of drug use demonstrates the enduring mark left by drugs on brain reward circuits. However, within these circuits, neuronal populations are differently affected by drug exposure and growing evidence indicates that relatively small subsets of neurons might be involved in the encoding and expression of drug-mediated associations. The identification of sparse neuronal populations recruited in response to drug exposure has benefited greatly from the study of immediate early genes (IEGs) whose induction is critical in initiating plasticity programs in recently activated neurons. In particular, the development of technologies to manipulate IEG-expressing cells has been fundamental to implicate broadly distributed neuronal ensembles coincidently activated by either drugs or drug-associated stimuli and to then causally establish their involvement in drug responses. In this review, we summarize the literature regarding IEG regulation in different learning paradigms and addiction models to highlight their role as a marker of activity and plasticity. As the exploration of neuronal ensembles in addiction improves our understanding of drug-associated memory encoding, it also raises several questions regarding the cellular and molecular characteristics of these discrete neuronal populations as they become incorporated in drug-associated neuronal ensembles. We review recent efforts towards this goal and discuss how they will offer a more comprehensive understanding of addiction pathophysiology.

Keywords: Activity-dependent transcription; Addiction; Arc; Fos; IEG; Neuronal ensembles; Reward circuit.

Fig. 1

Permanent tagging of cocaine-activated cells in NAc of Arc-CreER^T2 mice. Arc-CreER^T2 mice were crossed with a GFP reporter line to induce stable GFP expression in activated Arc+ neurons. (A) Schematic representation of the Arc-CreERT2 strategy applied to cocaine-activated cell tagging. On the day of tagging (day 0), animals are injected concomitantly with cocaine and tamoxifen. Cocaine triggers the activation of both the endogenous and artificially-expressed Arc promoters controlling, respectively, endogenous Arc and CreER^T2 transcription. This leads to increased expression of both Arc and CreER^T2 proteins in activated Arc+ cells. Upon tamoxifen binding, CreER^T2 enters the nucleus where the Cre-dependent removal of a floxed-STOP cassette triggers GFP expression. The persistent expression of GFP in cells activated initially by cocaine allows for their visualization at any future time (e.g., 7days). (B–C) On day 0, Arc-CreER^T2 mice were treated concomitantly with tamoxifen + cocaine or tamoxifen + saline as a control. Tissue was collected on day 7 post-tagging to visualize cells previously activated by cocaine. TAM = Tamoxifen. (B) Representative confocal images of GFP immunostaining in the NAc at day 7 post-tagging showing a higher number of activated cells in the cocaine-treated group. Scalebar = 50μm. (C) Fluorescence-activated nuclei sorting (FANS) strategy to isolate GFP+ nuclei from NAc punches at day 7 post-tagging. Cocaine-treated group shows a larger population of activated (GFP+) cells in comparison to saline-treated group.