Epigenetic Priming in Drug Addiction

Abstract

Drug addiction is a chronic relapsing brain disorder that is characterized by compulsive drug seeking and continued use despite negative outcomes. Current pharmacological therapies target neuronal receptors or transporters upon which drugs of abuse act initially, yet these treatments remain ineffective for most individuals and do not prevent disease relapse after abstinence. Drugs of abuse, in addition to their acute effects, cause persistent plasticity after repeated use, involving dysregulated gene expression in the brain's reward regions, which are thought to mediate the persistent behavioral abnormalities that characterize addiction. Emerging evidence implicates epigenetic priming as a key mechanism that underlies the long-lasting alterations in neuronal gene regulation, which can remain latent until triggered by re-exposure to drug-associated stimuli or the drug itself. Thus, to effectively treat drug addiction, we must identify the precise epigenetic mechanisms that establish and preserve the drug-induced pathology of the brain reward circuitry.

Figure 1.

Gene expression and its spatiotemporal regulation are central to the development of drug addiction. Activity-dependent changes at the circuit level remodel chromatin within individual neurons to regulate gene expression involved in synaptic and neural plasticity. Circuit activity and neurotransmitter release trigger intracellular signaling cascades such as the PKA or MAP-kinase (MAPK)/extracellular signal-regulated kinase (ERK) pathways that are activated by G protein–coupled receptors, and activated calcium pathways that occur upon receptor stimulation or neural activity, and transmit circuit activity information to the cell nucleus. In the nucleus, DNA is organized by wrapping around histone octamers to form nucleosomes. Only by temporarily unraveling compacted chromatin can the DNA of a specific gene be made accessible to the transcriptional machinery. This process involves the recruitment of chromatin remodeling and modifying enzymes that mediate acute and transient gene expression in response to upstream neural activity. Induction of immediate early genes (e.g., cFos, Egr1, NPas4, Arc) leads to the activation or inhibition of many other transcription factors and nuclear targets, including chromatin-regulatory proteins, that ultimately alter the composition or levels of membrane receptors and many other classes of proteins in neuronal signaling pathways, leading to changes in the excitability and structural connectivity of neurons in the reward circuitry.

Figure 2.

Patterns of time-dependent gene regulation in NAc after cocaine self-administration. The heatmaps show results from RNA-seq analyses of NAc after short- (24 h) or long-term (30 d) withdrawal from chronic self-administration of cocaine. Animals subjected to long-term withdrawal were given an intraperitoneal injection of saline or cocaine and placed back in their original chambers to determine if withdrawal plus context or context/drug re-exposure resulted in transcriptional priming or desensitization of genes. For both re-exposure paradigms, gene expression changes were mostly observed in magnitude and not direction. By applying the analytical technique of pattern analysis, unique gene lists were generated of transcripts that are regulated significantly and uniquely under specific conditions. For example, in the figure, only genes altered by cocaine/context re-exposure to a significantly greater extent than all other conditions are presented. Given that this is a unique list of genes that are up- and down-regulated by cocaine/context re-exposure but not context re-exposure alone, we hypothesize that these genes are primed or desensitized during withdrawal and activated or suppressed by re-exposure to the drug. Upstream regulator analysis predicted CREB and several nuclear receptors as the most prominent transcriptional regulators of these genes, suggesting an important role for these transcription factor families in priming/desensitizing the transcriptional response to cocaine in the NAc. (24 h – Coc) 24 hours after chronic cocaine self-administration, (30 d – Sal-Coc) 30 days after saline self-administration with an acute cocaine challenge (reflects the effects of acute cocaine exposure), (30 d – Coc-Sal) 30 days after cocaine self-administration with an acute saline challenge (reflects the effects of prolonged withdrawal plus context re-exposure), (30 d – Coc-Coc) 30 days after cocaine self-administration with an acute cocaine challenge (reflects the effects of prolonged withdrawal plus context and drug re-exposure). All conditions were compared to the same control group: 24 h after chronic saline self-administration. (Adapted from Walker et al. 2018, with permission from Elsevier, © 2018 Society of Biological Psychiatry.)

Figure 3.

Time-dependent control of gene regulation by drug exposure via epigenetic mechanisms. Chromatin mechanisms not only regulate the acute transcriptional response to drug exposure but further mediate latent effects at many genes that alter their inducibility to future stimuli. Transcription initiates within core promoters at the transcriptional start sites (TSSs) at 5′ ends of genes, which recruit RNA polymerase II (Pol2) and determine the accurate initiation position and direction of transcription. Efficient transcription is supported by enhancer elements (located at some distance from their target genes) that contain transcription factor (TF; “salmon”) binding sites and recruit a combination of TFs with a variety of cofactors to exert their overall regulatory function to control transcription from the targeted core promoter. Neuronal activation upon the initial drug exposure triggers intracellular signaling cascades that activate TFs and many other nuclear targets, including chromatin-regulatory proteins that modify histones and other proteins to regulate DNA accessibility, as well as transcription initiation and elongation. Early evidence suggests that chronic drug exposure causes extremely stable changes at the chromatin level that underlie transcriptional priming (shown) and desensitization (not shown) linked to drug addiction. Such gene priming/desensitization may remain latent during periods of withdrawal, when certain TFs and cofactors are bound to accessible chromatin without changing the steady state mRNA levels. However, future context and/or drug re-exposure can up-regulate a primed gene much faster and to a greater extent, based on the epigenetic changes induced by previous chronic drug exposure at regulatory promoter and enhancer regions.

Figure 4.

Cell type–specific control of chromatin accessibility in NAc by acute and chronic cocaine exposure. ATAC-seq was performed on D1 and D2 MSNs isolated from NAc using FACS in two transgenic mouse lines that express EGFP-RPL10a in either subtype (Drd1a/Drd2a::EGFP-L10a). Each horizontal row reflects a single gene locus centered around its TSS and 1 kb up- and downstream. This ongoing study investigates immediate and long-term changes in chromatin architecture, following acute cocaine (Sal-Coc, 1 h after 20 mg/kg cocaine by intraperitoneal injection) and prolonged withdrawal after chronic cocaine exposure (Coc-Sal, 30 d following 10 d of cocaine injections) as well as after drug challenge in withdrawal animals (Coc-Coc, 1 h after cocaine challenge). Drug-induced changes in chromatin accessibility discriminate D1 from D2 MSNs, and D1-specific chromatin “opening” following chronic cocaine exposure is sustained even during prolonged periods of withdrawal.