1. Background and introduction

Diagnostic criteria that are necessary to meet a diagnosis of substance use disorder (SUD) include compulsion to use licit or illicit substances and continuous use in the presence of adverse medical, neurological and/or psychiatric complications (DSM5, 2013). SUDs that are associated with increased morbidity and mortality in susceptible individuals (Degenhardt et al., 2014) are thought to be secondary to maladaptive changes in several brain regions [(Volkow and Boyle, 2018) see also Fig. 1]. Importantly, negative cognitive impact of substance use suggests that therapeutic approaches against SUDs will only be successful if and only if they address the various clinical courses associated with various patient populations because the clinical outcomes of patients are not uniform (Soares and Pereira, 2019). It is to be noted, at the onset, that neuroadaptations and their clinical sequelae are influenced by genetic and environmental factors that also account for epigenetic and transcriptional responses to these drugs in the central nervous system (Fig. 1). Using similar reasoning, several groups of investigators have tried to mimic these disorders in animal models in attempts to develop better pharmacological approaches to treat patients with SUDs (Cadet, 2019). Some of these models include conditioned place preference (CPP) and drug self-administration (SA) (Cadet et al., 2019; Muller, 2018). Molecular studies using various animal models have now provided evidence for the participation multiple gene networks in the development and maintenance of drug taking behaviors (Cadet et al., 2015; Korostynski et al., 2013; Nestler, 2012).

_{Fig. 1. This figure provides a working model of brain regions that are presumed to be involved in the development and maintenance of addictive states. Although the figure shows mainly dopaminergic projections that participate in reward circuits, there are non-reward projections that definitely participate in the maintenance of addiction because these diseased states involved more than rewarding effects of drugs. Future studies will need to emphasize more complex interactions of reward and non-reward pathways in the clinical course of addiction in individual patients and their responses to therapeutic interventions.}

2. Epigenetic regulation of gene expression

Changes in gene expression are regulated by epigenetic changes that include post-translational histone modifications, DNA methylation, and microRNAs (Jackson and Standart, 2007; Mazzio and Soliman, 2012; Murr, 2010). Modifications of N-tails of histones that possess lysine residues include acetylation/deacetylation processes catalyzed reversibly by histone acetyltransferases (HATs) and histone deacetylases (HDACs) (Falkenberg and Johnstone, 2014; Voss and Thomas, 2018). There are three classes of nuclear HATs that have histone acetyltransferase domains and bind acetyl-coenzyme A (Voss and Thomas, 2018). They comprise the GNAT (GCN5-related acetyltransferase), CBP/p300, and the MYST (MOZ, Ybf2/Sas 3, Sas 2, and Tip 60) families of enzymes. HDACs are divided into four classes based on sequence similarities (Mottet and Castronovo, 2008; Yoshida et al., 2017). Those class I (HDAC1, HDAC2, HDAC3, and HDAC8), class IIA (HDAC4, HDAC5, HDAC7, HDAC9), class IIB (HDAC6, HDAC10), class III (Sirtuins 1-7) and Class IV (HDAC11) HDACs. Class I, II and IV HDACs are the classical HDACs that are Zn²⁺-dependent enzymes whereas the sirtuins require NAD⁺-dependent.

Histone methylation, catalyzed by lysine methyltransferases (KMTs), is also an important player in the regulation of gene expression (Black et al., 2012; Hyun et al., 2017). These enzymes include KMT1A (SUV39H1), KMT1B (SUV39H2), KMT1C (G9a), KMT2A (MLL1), KMT2B (MLL2), KMT3A (SET2), KMT3B (NSD1), among others. In addition to the KMTs, protein arginine N-methyltransferases (PRMTs) can catalyze the addition of methyl groups to arginine residues in histones (Cha and Jho, 2012; Wolf, 2009).

Gene expression in the brain is also regulated by DNA methylation (Bayraktar and Kreutz, 2018). DNA contains four bases that are adenine (A), guanine (G), cytosine (C) and thymine (T). Cytosine can be altered at the carbon-5 position by adding a methyl group to form 5-methylcytosine (Jones, 2012; Moore et al., 2013). Mammalian species have enzymes named DNA methyltransferases (DNMTs) (DNMT1, DNMT3A, DNMT3B) that catalyze these reactions (Lyko, 2018). Methylated CpG pairs are found throughout the genome and DNA methylation can participate in repression of transcription (Moore et al., 2013). Methylated DNA can also be oxidized to form hydroxymethylated DNA by ten-eleven translocation (TET) enzymes (Lio et al., 2020). In addition to DNA hydroxymethylation, these enzymes are also involved in the formation of 5-formylcytosine, and 5-carboxylcytosine. TET enzymes and DNA hydroxymethylation have been reported to participate in the regulation of plasticity-related genes, learning, and memory (Kaas et al., 2013; Kumar et al., 2015) that are important to the addiction process.

3. Models of drug administration and post-translational histone acetylation

Because cocaine can alter gene expression (Bannon et al., 2005; Nestler et al., 1993; Yano and Steiner, 2007), several groups of investigators have sought to determine how the drug might impact histone modifications and HAT expression in various brain regions (Levine et al., 2005). were the first to show that cocaine increased the binding of CBP, a histone acetyl-transferase (Bannister and Kouzarides, 1996) that is involved in the regulation of many cellular events (Goodman and Smolik, 2000, Lipinski et al., 2019), at the fosB promoter accompanied by increased fosB mRNA expression. Acute and chronic non-contingent cocaine administration by experimenters caused increased acetylation of histone H3 at lysine 14 (H3K14Ac) and of histone H2B at lysine 12 (H2BK12Ac) in the nucleus accumbens (NAc) of mice (Malvaez et al., 2011), changes that were attenuated in mice with CBP deletion in the NAc. Bilateral deletion of CBP in the NAc also attenuated cocaine conditioned place preference (CPP). Interestingly, however, Madsen et al. (2012) showed that deletion of CBP in the mouse striatum produced increased sensitivity to cocaine and amphetamine.

In addition to experiments using cocaine administered by experimenters, some papers have also used studies in which rodents self-administered cocaine (Freeman et al., 2008; Monsey et al., 2019; Sadakierska-Chudy et al., 2017a; Sadri-Vakili et al., 2010). Freeman et al. (2008) used discrete-trials cocaine self-administration (SA) after rats had established a stable daily pattern of cocaine intake for 5 days (40 infusions within 6 h). During those session, rats could access cocaine during 10-min discrete trials. A trial ended when a cocaine injection (1.5 mg/kg/injection) was collected or 10 min had elapsed. Rats received four discrete trials per hour (i.e., DT4), 24 h per day for 10 days. The authors found that forced abstinence from cocaine self-administration (SA) for one and 10 days was associated with decreased abundance of H3 acetylated at K9 and K14 bound at the EGR1 promoter and decreased EGR1 mRNA levels but increased H3K9/14ac at the promoter gene of Npy and increased NPY expression in the rat mPFC after one day of abstinence. Interestingly, Sadri-Vakili et al. (2010) reported that cocaine SA followed by 7 days of forced abstinence also increased abundance of acetylated H3K9/14Ac at the Bdnf IV promoter and increased mRNA expression in rat medial prefrontal cortex (mPFC). Another group of investigators trained rats to self-administer cocaine using 2-h daily sessions 6 days/week for a minimum of 12 days (Sadakierska-Chudy et al., 2017a). At the end of the experiments, microarray analyses were used to measure gene expression in the rat prefrontal cortex (PFC). They found that mRNA levels of several epigenetic enzymes were significantly impacted in the PFC of rats that had self-administered cocaine. They also found significant increases in acetylated histone H3K9 and H4K8 but not H3K27 or H4K5 (Sadakierska-Chudy et al., 2017a). The role of HATs in cocaine-associated behaviors was also tested in cocaine SA experiments by Monsey et al. (2019). In those experiments, rats underwent 12 days of cocaine self-administration (SA) training using 1-h sessions. Monsey et al. (2019) reported that injections of garcinol, an inhibitor of KATs of p300/CBP and PCAF families (Arif et al., 2009; Balasubramanyam et al., 2004), into the lateral nucleus of the amygdala (LA) was able to block the reconsolidation of a cocaine-cue memory. They also reported that systemic injections of garcinol caused decreased abundance of acetylated histone H3K27 in the LA. Acetylation of histone H3K27 is known to be mediated, in part, by CBP/p300 (Jin et al., 2011). Together, these data suggest that the involvement of identifiable epigenetic events in cocaine-induced behaviors may be specific to certain brain regions.

Injection of another psychostimulant, methamphetamine, also causes changes in histone acetylation (Martin et al., 2012). Methamphetamine injection caused time-dependent increases in acetylated H4K5 (H4K5Ac) and H4K8 (H4K8Ac) levels that were associated with increased expression of the histone acetyltransferase, ATF2. A follow-up study found that acute and chronic METH injections could cause differential changes in striatal gene expression that was associated with increased H4K5Ac binding near the transcriptional start sites (TSSs) of several genes using chromatin immunoprecipitation followed by genome wide sequencing (Cadet et al., 2013). There were also significant positive correlations between gene expression obtained by microarray analysis and H4K5Ac binding by ChIP-Seq in the striatum of control and METH-treated rats. Genes of interest that show increased mRNA levels and H4K5Ac binding include immediate early genes (IEGs) such as Arc, c-fos, Egr 2, Egr 4, JunB, Nr4a3, and Npas4 (Cadet et al., 2013). Other genes of interest are DUSP5, DUSP 14, Neurotensin, and Per 1 (Cadet et al., 2013). These results implicate this epigenetic marker in the molecular effects of methamphetamine in the brain. Those results also suggest that HAT1/KAT1 which is responsible for histone H4 acetylation (Nagarajan et al., 2013, Parthun, 2007) in the long-term effects of this drug since repeated injections of METH are also associated with increased H4K5 acetylation (Cadet et al., 2013). Moreover, Gonzalez et al. (2019) reported that a single injection of METH increased the abundance of acetylated histone H4 on Hdac1 and Hdac10 gene promoters. They also found decreased acetylated H4 on Hdac4 and Hdac5 genes. In contrast, they found that repeated injections of METH increased H4 acetylation on Hdac4 and Hdac5 genes but decreased abundance on Hdac1, Hdac2, and Hdac8 gene promoters. More studies are necessary to further test the role of HATs and histone acetylation in models that use methamphetamine self-administration.

There is evidence that class I HDACs are also differentially regulated in animal models of drug administration. Schroeder et al. (2008) showed that pretreatment of mice with the class I inhibitor, sodium butyrate, in conjunction with a dopamine D1 agonist caused increased TH and BDNF mRNA expression in the ventral midbrain, changes that were associated, unexpectedly, with H3 hypoacetylation at the promoters of these genes. Kennedy et al. (2013) showed that local knockdown of HDAC1, but not of HDAC2 and HDAC3, caused reduction of cocaine-induced locomotor sensitization. In addition, Host et al. (2011) reported that cocaine SA increased HDAC2 mRNA expression in the NAc, dorsal striatum and cingulate cortex. Importantly, reduction of HDAC2 expression by induction of cyclic GMP reduced cocaine SA (Deschatrettes et al., 2013). Another class I HDAC, HDAC3, appears to also participate in cocaine-induced behaviors. Specifically, systemic administration of RGFP966, a selective HDAC3 inhibitor, was able to facilitate extinction of cocaine CPP while increasing acetylation of histone H4 at lysine 8 in the infra-limbic cortex (Malvaez et al., 2013). RGFP increased H3K14 acetylation in the infra-limbic cortex, NAc and hippocampus (Malvaez et al., 2013). RGFP966 treatment decreased HDAC3 binding at the c-fos promoter (Malvaez et al., 2013). Methamphetamine, another illicit drug, also impacts the expression of class I HDACs. Martin et al. (2012) reported that a single methamphetamine injection decreased HDAC1 but increased HDAC2 protein levels in the rodent NAc. In addition, Jayanthi et al. (2014) showed that repeated injections of methamphetamine increased HDAC1 and HDAC2 expression in the dorsal striatum. These increases were associated with hypoacetylation of histone H4 at lysines 4, 8, 12 and 16 (Jayanthi et al., 2014).

Class II HDACs are also involved in the behavioral effects of drugs of abuse. For example, chronic bilateral injection of the HDAC inhibitor, Trichostatin (TSA), in the rat NAc shell caused higher SA rates in rats by influencing, in part, HDAC4 expression because HDAC4 overexpression attenuated the motivation to self-administer cocaine (Wang et al., 2010) Chronic non-contingent cocaine injections decreased HDAC5 expression in the NAc (Renthal et al., 2007) whereas cocaine SA decreased nuclear HDAC5 localization (Host et al., 2011). HDAC5 knockout in rodents increased cocaine CPP and that viral HDAC5 overexpression in the NAc decreased cocaine CPP (Renthal et al., 2007). Dietrich et al. (2012) also showed that repeated cocaine injections induced salt-inducible kinase (SIK1)-dependent HDAC5 phosphorylation in the nucleus and its subsequent shuttling to the cytoplasm. In addition to cocaine, methamphetamine can also impact the expression of these HDACs. Specifically, an acute methamphetamine injection decreased HDAC4 and HDAC7 mRNA levels in the NAc of WT mice (Torres et al., 2016). In contrast, Li et al. (2015) observed significant increases in the mRNA expression of HDAC4 and HDAC5 in the dorsal striatum of rats that had undergone 35 days of withdrawal from methamphetamine SA. Furthermore, striatal HDAC5 overexpression or knockdown respectively increased or decreased methamphetamine seeking after a long interval of withdrawal (Li et al., 2017).

The use of psychostimulants also impacts the expression of class III HDACs that are the SIRTs (Satoh et al., 2017). Renthal et al. (2009) performed chromatin immunoprecipitation (ChIP) to investigate the effects of repeated injections of cocaine in the NAc of mice and found that cocaine increased histone acetylation at Sirt1, and Sirt2 DNA sequences (Renthal et al., 2009). Ferguson et al. (2013) also reported that overexpression of Sirt1 in the NAc increased cocaine CPP whereas its depletion caused decreased CPP (Ferguson et al., 2013). Interestingly, although cocaine administration increased SIRT1 induction in the rodent nucleus accumbens, Ferguson et al. (2015) found that SIRT1 abundance was decreased at gene promoters. These cocaine-induced changes were also accompanied by increased expression of SIRT1 target genes. The authors found that increased SIRT1 expression was associated with deacetylation of FOXO3a and increased expression of known FOXO3a gene targets. Consistent with the results with cocaine, repeated methamphetamine injections also increased Sirt1 and Sirt2 mRNA and protein levels in the rat dorsal striatum (Jayanthi et al., 2014). Consistent with these observations, chronic METH was recently found to increase H4 acetylation binding at the Sirt2 promoter (Gonzalez et al., 2020). These findings thus implicate class III HDACs in the regulation of some genes that might be involved in mediating long-term neuroadaptations to exposure to psychostimulants.

The class IV HDAC, HDAC11, which was cloned and characterized by Gao et al. (2002), is highly expressed in the brain (Takase et al., 2013). Similar to the other HDACs, Host et al. (2011) have reported that cocaine self-administration was accompanied by increased HDAC11 expression. In contrast, acute METH injections caused decreases in HDAC11 mRNA levels in the NAc (Omonijo et al., 2014; Torres et al., 2016).

In addition to the effects of psychostimulants on histone acetylation/deacetylation, opioids have also been reported to cause histone post-translational modifications (Browne et al., 2020). Sheng et al. (2011) had demonstrated that heroin administration was associated with increased histone H3 phosphoacetylation that was essential to heroin CPP. These changes in histone acetylation observed in rodents are consistent with a report of increased acetylation of H3K27 in the post-mortem striatum of heroin addicts (Egervari et al., 2017). Importantly, HDAC inhibition was able to enhance morphine-associated memory formation (Wang et al., 2015), further suggesting a role of histone acetylation in opioid-mediated behaviors.

4. Models of drug administration and post-translational histone methylation

Cocaine-induced changes in gene expression appear to also depend on alterations in histone methylation although this histone mark has been less well investigated in models of addiction. Maze et al. (2010) reported that repeated injections of cocaine reduced global levels of H3K9 dimethylation in the NAc. These decreases were due to G9a repression in a deltaFoB-dependent fashion. They further showed that viral-mediated G9a down-regulation enhanced preference for cocaine. In addition, Jordi et al. (2013) reported that a single cocaine injection induced significant changes in histone methylation at 15 min after the drug administration. Histone H3K9 trimethylation was increased in D1-containing neurons whereas all three methylated forms of H3K9 were increased in D2-containing cells. In addition to methylation at lysine residues, cocaine can also alter arginine methylation (Damez-Werno et al., 2016). Specifically, cocaine SA led to significant decreases in protein-R-methyltransferase-6 (PRMT6) and its associated histone mark, asymmetric dimethylation of R2 on histone H3 (H3R2me2a) in rodents (Damez-Werno et al., 2016). Humans who suffered from cocaine use disorder also showed similar changes in their NAc.

5. Involvement of DNA methylation and hydroxymethylation in models of addiction

Because DNA methylation plays important roles in cognitive functions such as learning and memory (Jobe and Zhao, 2017) that are impacted by drugs of abuse (Bisagno et al., 2016), it was highly likely that these drugs might influence the expression of DNMTs. An acute cocaine injection increased DNMT3A and DNMT3B mRNA levels in the NAc (Anier et al., 2010). LaPlant et al. (2010) reported that DNMT3A mRNA was up-regulated at 4 h but down-regulated at 24 h after a single cocaine injection. Cocaine also increased DNA hypermethylation and increased binding of methyl CpG binding protein 2 (MeCP2) at the protein phosphatase-1 catalytic subunit (PP1c) promoter associated with decreased PP1C mRNA levels (Anier et al., 2010). Repeated cocaine injections also increased DNMT1, DNMT3A, and DNMT3B mRNA levels in the NAc (Anier et al., 2018). Interestingly, mice that self-administered cocaine showed significant decreases in DNMT1 and DNMT3A mRNAs measure 24 h after the last session (LaPlant et al., 2010). Using rats that self-administered cocaine, Wright et al. (2015) found significant increases of DNMT3A in the NAc but not in the prefrontal cortex. Of related interest, rats that underwent cocaine SA also exhibited increased MeCP2 expression in the dorsal striatum (Im et al., 2010). Using genome-wide sequencing, Baker-Andresen et al. (2015) found that rats that self-administered cocaine exhibited 29 differentially methylated regions in a persistent manner in comparison to saline- or passive cocaine-treated animals, with five regions being demethylated and the rest hypermethylated. Methamphetamine also influences the expression of DNMT gene expression. For example, Numachi et al. (2007) repeated injections of the drug increased DNMT1 expression in the NAc and dorsal striatum of Fisher rats. In addition, Jayanthi et al. (2019) have found that methamphetamine SA is accompanied by increased DNA methylation at the DNA sequences of several genes that code for potassium channels.

Interestingly, DNA hydroxymethylation marks have also been investigated after administration of cocaine and methamphetamine. Global levels of 5-hmC were increased after repeated cocaine administration (Anier et al., 2018). Ploense et al. (2018) found that limited access to cocaine decreased DNA hydroxymethylation within the Homer2 promoter in rats (Sadakierska-Chudy et al., 2017b). also reported that abstinence from cocaine was associated with increased global 5-hydroxymethylcytosine (5-hmC) levels and increased Tet 3 transcript levels in the hippocampus. Cadet et al. (2017) used an animal model of drug SA wherein footshocks were used to suppress methamphetamine SA in some but not in other rats. They found that the animals that suppress their intake of the drug had much higher levels of DNA hydroxymethylation at the sequences of several reward-related genes including potassium channels in the NAc. Injections of methamphetamine also produced DNA hypomethylation at sites near the corticotrophin (CRH) transcription start site (TSS) and at intragenic vasopressin (AVP) sequences. methamphetamine also increased DNA hydroxymethylation at these sites. Interestingly, there was increased protein expression of ten-eleven-translocation (TET) enzymes that catalyze DNA hydroxymethylation. Importantly, the TET inhibitor, 1,5-isoquinolinediol (IQD), blocked methamphetamine-induced increases in Crh and Avp mRNA expression.

Changes in DNA methylation have also been reported after opioid exposure. For example, Kozlenkov et al. (2017) reported 1298 differentially methylated CpG sites (DMSs) between heroin users and controls. Hyper-DMSs were enriched in gene bodies and exons but depleted in promoters while hypo-DMSs were enriched in promoters and enhancers. In addition, hyper-DMGs showed preference for genes involved in axonogenesis- and synaptic functions. Similarly, post-mortem brain samples from long-term opioid abusers have been reported to exhibit reduced mu-opioid receptor (OPRM1) expression that was related to altered methylation at a CpG site in the functional SNP variant of the OPRM1 (118A > G) gene. These changes in methylation were also accompanied with abnormal compensatory mechanisms that would have occurred with chronic opioid exposure (Oertel et al., 2012). Besides DNA methylation, epigenetic regulation via repressive histone methylation has also been reported after repeated morphine exposure (Koo et al., 2015). Moreover, the authors reported that chronic morphine administration caused behavioral sensitization that was associated with increased binding of H3K27me3 at the Bdnf-p2 promoter and secondary decreased Bdnf expression in the ventral tegmentum (VTA) (Koo et al., 2015). Taken together, these results implicate drug-induced methylation as a regulator of gene expression and behavioral changes associated with drug taking behaviors.

6. Concluding remarks and future directions

The present literature on the effects of drugs of abuse on epigenetic markers supports the idea that these drugs can have both acute and persistent effects on the expression of several enzymes involved in causing post-translational histone modifications and DNA methylation/hydroxymethylation in the brains of rodents. These changes are accompanied by differential changes in the abundance of modified histones on the promoters or bodies of genes that have been implicated in animal models of addiction. These observations are interesting and are remarkable in terms of supporting the notion that long-term neuroadaptations are responsible for the long-lasting behavioral effects of drugs of abuse. Although these findings are very interesting, it remains to be determined to what extent, they will be translatable to human conditions. Studies being conducted in models that use footshocks to separate groups of rodents that use cocaine, methamphetamine, and opioids into resilient and susceptible rodents may enhance the ability of scientists to translate these results into the development of pharmacotherapeutics that might revert some of the epigenetic changes observed in the brain of drug exposed animals. These novel studies promise to provide a better window to the long-term impact of drug taking behaviors and of withdrawal/abstinence on epigenetic markers (see scheme in Fig. 2). There is, however, a very important need to expand these types of investigations to post-mortem tissues in a fashion similar to studies of post-mortem tissues of patients who suffer from Schizophrenia, major affective disorder, Alzheimer's and Parkinson's diseases, and normal and abnormal aging. Epigenetic studies of post-mortem tissues may help to further clarify what animal models of addiction are more relevant to human situations.

_{Fig. 2. The scheme described here indicates that individual licit and illicit drugs can impact individuals in very specific ways as reported in the clinical literature. Not all individuals who are exposed to drugs become addicted to these drugs and their responses to drugs depend on genetic and environment factors that either protect them or help to escalate their uses of specific initially rewarding agents. This figure also suggests the need to identify the differential and progressive epigenetic and transcriptional consequences of drug exposure that determine paths to compulsive drug use despite adverse consequences or sustained drug use without developing a diagnosis of substance use disorder. The schema also suggests that treatment approaches will depend on pharmacological agents that can block the further development of epigenetic changes or reverse already established epigenetic modifications.}

It is also important to conclude with some suggestion about the potential use of drugs that might impact epigenetic mechanisms in the brain. This is important because these drugs will not impact epigenetic mechanisms and gene transcription in the central nervous system but also in various peripheral organs. At present, there are several drugs that being used as anticancer therapeutics (Nepali and Liou, 2021). These drugs can impact many of the targets that we have discussed in this review including histone acetylation and methylation as well as DNA methylation (Nepali and Liou, 2021; Sarno et al., 2021). Given the levels of plasticity associated with the epigenome, the use of similar compounds in therapeutic interventions against SUDs seems quite warranted. However, the translation of these ideas to clinical practice will require great investments of resources to investigate the precise doses of specific epigenetic drugs and their combinations to treat specific SUD. Moreover, it will be important to carefully calculate the benefits of using these drugs in contrast to adverse events encountered during their clinical use. Some of these negative consequences may be avoided by the use of pro-drugs that are transformed to their active forms in the brain. Because drugs of abuse appear to influence multiple epigenetic targets, it will be important to generate therapeutic mixtures that might influence different targets at the same time. This poly-pharmacological approach has been used successfully in medicine and psychiatry for many years. Finally, the future use of epigenetic agents promises to add in the clinicians’ armamentarium against SUDs because the repeated oral intake or intravenous injections of licit and illicit drugs are associated with abnormalities in multiple epigenetic pathways. This approach may be more productive beyond approaches that use single drugs that block or stimulate single neurotransmitter receptors.

Summary

Substance use disorder (SUD) diagnosis necessitates criteria encompassing compulsive substance use despite adverse medical, neurological, or psychiatric consequences. SUDs, linked to heightened morbidity and mortality, stem from maladaptive alterations across diverse brain regions. The negative cognitive impact of substance use underscores the necessity of therapeutic interventions tailored to individual clinical courses, as patient outcomes vary considerably. Neuroadaptations and their clinical manifestations are shaped by genetic and environmental influences, including epigenetic and transcriptional responses within the central nervous system. Animal models, such as conditioned place preference (CPP) and drug self-administration (SA), have been employed to investigate SUDs and develop improved pharmacological treatments. Molecular studies using these models reveal the involvement of multiple gene networks in the development and maintenance of drug-seeking behaviors.

Epigenetic Regulation of Gene Expression

Gene expression is regulated epigenetically through post-translational histone modifications, DNA methylation, and microRNAs. Histone modifications, particularly acetylation/deacetylation, are catalyzed by histone acetyltransferases (HATs) and histone deacetylases (HDACs), respectively. HATs are categorized into GNAT, CBP/p300, and MYST families, while HDACs are classified into four classes (I-IV) based on sequence homology. Histone methylation, involving lysine methyltransferases (KMTs) and protein arginine N-methyltransferases (PRMTs), also plays a significant role. DNA methylation, catalyzed by DNA methyltransferases (DNMTs), modifies cytosine bases, influencing transcriptional repression. Ten-eleven translocation (TET) enzymes mediate DNA hydroxymethylation, impacting plasticity-related genes crucial to addiction.

Models of Drug Administration and Post-Translational Histone Acetylation

Cocaine's impact on gene expression has prompted investigations into its effects on histone modifications and HAT expression. Studies have demonstrated cocaine-induced increases in histone acetylation (H3K14Ac, H2BK12Ac) in the nucleus accumbens (NAc), effects attenuated by CBP deletion. Cocaine self-administration studies reveal alterations in histone acetylation levels (H3K9Ac, H3K14Ac, H4K8Ac, H3K27Ac) across various brain regions following abstinence, implicating specific epigenetic events in cocaine-associated behaviors. Methamphetamine administration also induces changes in histone acetylation, affecting the expression of various HDAC classes (I, II, III, IV), with alterations in histone acetylation patterns correlating with changes in gene expression.

Models of Drug Administration and Post-Translational Histone Methylation

Cocaine's influence on histone methylation has been studied less extensively, but studies indicate reductions in global H3K9 dimethylation and alterations in methylation patterns across different neuronal populations. Cocaine self-administration also affects arginine methylation through changes in PRMT6 levels and H3R2me2a.

Involvement of DNA Methylation and Hydroxymethylation in Models of Addiction

Cocaine and methamphetamine influence the expression of DNMTs, impacting DNA methylation patterns. Studies reveal drug-induced changes in DNA methylation levels at various gene loci associated with addiction-related behaviors. DNA hydroxymethylation is also affected by these drugs, with alterations in 5-hmC levels observed following drug administration and abstinence. Opioid exposure also results in changes in DNA methylation, including altered mu-opioid receptor expression.

Concluding Remarks and Future Directions

The literature suggests both acute and persistent effects of drugs of abuse on epigenetic markers, potentially leading to long-lasting behavioral changes. Translational studies in animal models that distinguish between resilient and susceptible individuals are crucial for developing targeted pharmacotherapeutics. Future research should also expand to human post-mortem studies to validate animal model findings and enhance understanding of SUDs. The use of epigenetic drugs, currently employed in cancer treatment, holds promise for SUD therapeutics, but careful dose optimization, safety profiling, and potential use of prodrugs are essential considerations. A poly-pharmacological approach, targeting multiple epigenetic pathways concurrently, may prove particularly effective.

Summary

Substance use disorder (SUD) diagnosis necessitates identifying compulsive substance use despite adverse consequences. These disorders, linked to increased morbidity and mortality, stem from maladaptive brain region changes. Effective SUD therapies must address diverse clinical presentations, as patient outcomes are heterogeneous. Neuroadaptations and their clinical consequences are influenced by genetic and environmental factors, including epigenetic and transcriptional responses to drugs within the central nervous system. Animal models, such as conditioned place preference (CPP) and drug self-administration (SA), aid in developing better pharmacological treatments, highlighting the involvement of multiple gene networks in drug-seeking behavior.

Epigenetic Regulation of Gene Expression

Gene expression is regulated epigenetically through histone modifications, DNA methylation, and microRNAs. Histone modifications, including acetylation/deacetylation, are catalyzed by histone acetyltransferases (HATs) and histone deacetylases (HDACs). HATs are classified into GNAT, CBP/p300, and MYST families, while HDACs are categorized into four classes based on sequence similarities (I-IV) and their dependence on Zn2+ or NAD+. Histone methylation, crucial for gene expression regulation, is performed by lysine methyltransferases (KMTs) and protein arginine N-methyltransferases (PRMTs). DNA methylation, the addition of a methyl group to cytosine, is catalyzed by DNA methyltransferases (DNMTs) and impacts transcription repression. Ten-eleven translocation (TET) enzymes oxidize methylated DNA, influencing plasticity-related genes crucial for addiction.

Models of Drug Administration and Post-Translational Histone Acetylation

Cocaine's effects on gene expression and histone modifications have been extensively studied. Cocaine increases CBP binding at the fosB promoter, leading to increased fosB mRNA expression. Acute and chronic cocaine administration increases histone H3 and H2B acetylation in the nucleus accumbens (NAc), effects attenuated by CBP deletion. Cocaine self-administration studies reveal altered histone acetylation patterns upon abstinence, varying across brain regions. Methamphetamine also modifies histone acetylation, impacting immediate early genes (IEGs) and epigenetic enzyme expression. Class I HDACs are differentially regulated in response to psychostimulant exposure. Class II HDACs, like HDAC4 and HDAC5, are involved in cocaine-induced behaviors, with their expression levels influencing cocaine self-administration. Class III HDACs (SIRTs) also participate in the response to psychostimulants, affecting gene expression and neuroadaptations. Finally, HDAC11, a class IV HDAC, shows altered expression following cocaine and methamphetamine exposure. Opioids also induce histone post-translational modifications, influencing drug-associated memory.

Models of Drug Administration and Post-Translational Histone Methylation

Cocaine alters histone methylation, with repeated injections reducing global H3K9 dimethylation. A single cocaine injection induces changes in H3K9 methylation, differing across neuronal subtypes. Cocaine self-administration also decreases PRMT6 and H3R2me2a.

Involvement of DNA Methylation and Hydroxymethylation in Models of Addiction

Cocaine impacts DNMT expression, altering DNA methylation patterns and MeCP2 binding. Cocaine self-administration studies reveal varied effects on DNMT expression. Methamphetamine also modifies DNMT expression and DNA methylation at potassium channel genes. Cocaine and methamphetamine administration influence DNA hydroxymethylation, with varied effects depending on drug exposure and brain region. Opioid exposure also alters DNA methylation and histone methylation, affecting gene expression relevant to addiction.

Concluding Remarks and Future Directions

Studies demonstrate that drugs of abuse acutely and persistently affect epigenetic enzymes, leading to modified histone and DNA methylation patterns. These epigenetic alterations underlie long-term behavioral changes, but their translatability to human conditions requires further investigation. Future research should focus on identifying epigenetic biomarkers of addiction resilience and susceptibility, informing the development of targeted pharmacotherapies to reverse or prevent epigenetic changes. The use of epigenetic drugs, currently employed in cancer treatments, holds promise for SUD treatment, necessitating careful dose optimization and consideration of adverse effects. A poly-pharmacological approach, targeting multiple epigenetic pathways, may prove superior to single-target strategies.

Summary

Substance use disorder (SUD) involves compulsive substance use despite negative consequences. This is linked to brain changes, influenced by genetics and environment, affecting multiple gene networks. Animal models, such as conditioned place preference (CPP) and drug self-administration (SA), help researchers study these changes and develop treatments.

Epigenetic Regulation of Gene Expression

Gene expression is controlled by epigenetics, including histone modifications (acetylation/deacetylation, methylation), and DNA methylation/hydroxymethylation. Histone acetyltransferases (HATs) and deacetylases (HDACs) regulate acetylation. Methylation is controlled by lysine methyltransferases (KMTs) and protein arginine N-methyltransferases (PRMTs). DNA methylation involves adding methyl groups to cytosine, catalyzed by DNA methyltransferases (DNMTs); ten-eleven translocation (TET) enzymes create hydroxymethylated DNA.

Models of Drug Administration and Post-Translational Histone Acetylation

Studies using cocaine and methamphetamine show these drugs alter histone acetylation. Cocaine affects histone acetylation in the nucleus accumbens (NAc), impacting conditioned place preference (CPP). Methamphetamine causes changes in histone acetylation and the expression of HATs and HDACs in the striatum, affecting gene expression. Different classes of HDACs (I, II, III, IV) are differentially impacted by cocaine and methamphetamine.

Models of Drug Administration and Post-Translational Histone Methylation

Cocaine alters histone methylation, reducing H3K9 dimethylation and affecting cocaine preference. Methamphetamine also affects histone methylation, with changes occurring in both lysine and arginine residues.

Involvement of DNA Methylation and Hydroxymethylation in Models of Addiction

Cocaine and methamphetamine affect DNA methylation and hydroxymethylation. Cocaine alters DNMT expression and DNA methylation. Methamphetamine impacts DNA methylation of genes related to potassium channels. Both drugs impact DNA hydroxymethylation levels and TET enzyme expression, potentially affecting reward-related genes. Opioids also show similar epigenetic changes.

Concluding Remarks and Future Directions

Studies show drugs of abuse impact histone modifications and DNA methylation/hydroxymethylation, leading to long-lasting behavioral effects. Future research should focus on translating these findings to humans and developing therapies targeting epigenetic mechanisms. The use of epigenetic drugs, potentially in combination, shows promise, but careful consideration of dosage and side effects is necessary.

Summary

Drug addiction is a serious problem. Scientists are studying the brain to understand how drugs affect it and cause addiction. They are looking at tiny changes in the brain's instructions (genes) that can be caused by drugs. These changes, called epigenetic changes, affect how genes work. Scientists use animal studies to learn more about these changes. They give drugs to animals, such as cocaine or methamphetamine, and see how it affects their brains and behavior. They look at things like how the genes turn on and off and other small changes in the brain's chemicals. This research might help develop better ways to treat drug addiction.

Epigenetic Regulation of Gene Expression

Our bodies have tiny instructions called genes that tell our cells what to do. Genes can be turned on or off like a light switch. Epigenetics are like little switches that control these genes. One way this happens is by changing the proteins around the genes. There are also things called DNA methyltransferases (DNMTs) that can add a special tag to the genes, changing how they work. Scientists are studying these switches to see how they affect drug addiction.

Models of Drug Administration and Post-Translational Histone Acetylation

Scientists give drugs to animals (like mice and rats) to study how drugs change the brain. They look at how the drugs change the small switches and tags on the genes. For example, cocaine can change how certain proteins work, affecting genes related to addiction. Some experiments use animals that give themselves drugs, while others give the drugs to the animals directly. These studies help scientists understand how drugs affect the brain's chemistry and lead to addiction.

Models of Drug Administration and Post-Translational Histone Methylation

Besides changes in proteins, drugs also affect little tags called methyl groups that stick to the genes. Cocaine, for example, changes these methyl tags, and this affects how genes work. This is another way drugs alter brain function and contribute to addiction. These small changes in the genes' tags have a big effect on the brain.

Involvement of DNA Methylation and Hydroxymethylation in Models of Addiction

Drugs can also affect the DNA itself by adding or removing special tags, affecting how our genes work. Cocaine and methamphetamine can change these tags, and this affects genes involved in addiction. These changes can be long-lasting. Studying these changes helps scientists better understand drug addiction.

Concluding Remarks and Future Directions

Scientists have learned a lot about how drugs change the brain's tiny switches and tags on genes. These changes are important because they help explain why addiction lasts. Further studies are needed to understand how these findings apply to people and to develop new treatments for drug addiction. Scientists hope to use this knowledge to find better ways to help people overcome drug addiction.