| Drug addiction is a chronic relapsing encephalopathy that is characterized by compulsive drug seeking behavior and taking despite severe adverse consequences. Once an individual becomes addicted to a drug of abuse, there are few effective clinical treatments, and most addicts relapse within a short period of time. The mechanisms of cocaine addiction are very obscure and complicated. Increasing evidence suggests that abnormality of gene expression in the brain reward regions is one of the most important mechanisms involved in a state of addiction caused by drugs of abuse.Chromatin remodeling epigenetically controls gene transcription, indicating that complex mechanisms regulate the accessibility of genes to the transcriptional machinery. It is regarded that increased histone acetylation is associated with DNA relaxation and elevated transcriptional activity, whereas decreased acetylation, brought about by histone deacetylases (HDACs) and methyl-CpG-binding proteins, results in tighter DNA coiling and gene silencing. Nowadays, some researches indicate that histone modification plays a critical in cocaine-induced neuronal and behavioral plasticity. However, research has not completely clarified the epigenetic mechanism in the different phases of drug self-administration including the initial, reinforcement, incubation and reinstatement stages.Intravenous drug self-administration, which better models the human syndrome especially the addicted-state changes, such as increased motivation for the drug, is a useful tool for exploring the neurobiology of drug positive reinforcement in experimental animals. In this study, rats were subjected to cocaine self-administration under fixed-ratio (FR) or progressive-ratio (PR) schedule. The role of histone acetylation and histone-acetylation-mediated gene transcriptional activation in the NAc during the incentive motivation for cocaine following a chronic phase of drug self-administration was examined. Our results showed that:1. HDAC inhibitor infusions into the NAc shell increase the motivation for cocaine self-administration. Daily histone deacetylase (HDAC) inhibitor (TSA or SAHA) infusions in the nucleus accumbens (NAc) shell, but not the core or medial prefrontal cortex (mPFC), caused an upward shift in the dose-response and dose-intake curve under fixed-ratio schedule, and increased the break point under progressive-ratio schedule, indicating enhanced motivation for self-administered drug. These data indicated intra-NAc-shell infusions of HDAC inhibitor enhance motivation for rat to obtain cocaine, which may be attributable to the potentiating of histone acetylation induced by sustained HDAC inhibition.2. HDAC inhibitor augments histone acetylation elevation induced by chronic cocaine experience in the NAc shell. Histone acetylation levels following the acute and chronic administration of cocaine were evaluated by western blotting and immunochemistry analysis. The acetylation levels of H3 and H4 were elevated specifically in the NAc shell, but not in the NAc core or the mPFC following chronic intravenous cocaine self-administration experience. Furthermore, Chronic-cocaine-stimulated increases in histone acetylation in the NAc shell were notably augmented by HDAC inhibitor pretreatment. These data supports the notion that increased histone acetylation in the NAc shell induced by chronic cocaine self-administration is associated with motivation for drug reinforcement.3. Neutralizing the chronic-cocaine-induced increase in histone modification by the bilateral overexpression of HDAC4 in the NAc shell reduced drug motivation. To gain further insight into the relationship between histone acetylation and the motivation for cocaine self-administration, we investigated the effect of neutralizing cocaine-stimulated increase in histone acetylation by overexpressing of HDAC4. Intra-NAc shell overexpression of full-length HDAC4, but not its truncated form, which lacks deacetylase activity, strongly inhibited the cocaine-induced increase in histone acetylation, and induced a downward shift in dose-response and dose-intake curves under FR schedule, and decreased break point under PR schedule, indicating reduced motivation for drug. The effect of HDAC4 expression on the motivation for cocaine self-administration was dependent on its deacetylation activity. However, this effect was not observed when HDAC4 overexpression in the NAc core or the mPFC. These results confirm that the elevation of histone acetylation in the NAc shell is critical for drug motivation.4. Motivation for cocaine is associated with H3 acetylation in the NAc shell. Chromatin immunoprecipitation (ChIP) assays were performed to investigate the association between the motivation for cocaine and the transcriptional activation of addiction-related genes by H3 acetylation in the NAc shell. As is shown in ChIP assay, chronic-cocaine-induced gene transcriptional activation occured primarily in the NAc shell and was predominantly modulated by the acetylation of H3. Futhermore, motivation for cocaine was associated with transcriptional activation of addiction-related genes modulated by H3 acetylation of NAc shell. These data indicated that drug-induced gene transcriptional activation in the NAc shell, modulated by H3 acetylation, is a potential mechanism underlying the motivation for cocaine.5. Localized CaMKⅡαknockdown in the NAc shell decreases the motivation for cocaine. Among the genes activated by chronic cocaine experiences, the expression of CaMKIIa, but not CaMKⅡβ, correlated positively with motivation for the drug. Lentivirus-mediated shRNA knockdown experiments demonstrated that CaMKⅡα, but not CaMKⅡβ, in the NAc shell is essential for the maintenance of motivation to self-administer cocaine. These findings suggest that chronic-drug-use-induced transcriptional activation of genes such as CaMKIIa, modulated by H3 acetylation in the NAc, is a critical regulatory mechanism underlying motivation for drug reinforcement.Taken together, our results establish an important role for the chronic-cocaine-induced increase in histone H3 acetylation and the transcriptional activation of the gene encoding CaMKⅡαin the shell of the NAc in the motivation for cocaine reinforcement firstly, and suggest a new therapeutic mechanism for anti-addiction treatment.
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