Dysregulation of histone modifying enzymes has been associated with numerous psychiatric disorders. Alterations in G9a (Ehmt2), a histone methyltransferase that catalyzes the euchromatic dimethylation of histone H3 at lysine 9 (H3K9me2), has been implicated recently in mediating neural and behavioral plasticity in response to chronic cocaine administration. Here, we show that chronic morphine, like cocaine, decreases G9a expression, and global levels of H3K9me2, in mouse nucleus accumbens (NAc), a key brain reward region. In contrast, levels of other histone methyltransferases or demethylases, or of other methylated histone marks, were not affected in NAcbychronic morphine.Through viral-mediated gene transferand conditional mutagenesis,wefound that overexpressionofG9a in NAc opposes morphine reward and locomotor sensitization and concomitantly promotes analgesic tolerance and naloxone-precipitated withdrawal, whereas downregulationofG9ainNAc enhances locomotor sensitization and delays the developmentofanalgesic tolerance. We identified downstream targetsofG9a by providinga comprehensive chromatin immunoprecipitation followed by massively parallel sequencing analysis ofH3K9me2 distribution in NAc in the absence and presence ofchronic morphine. These data provide novel insight into the epigenomic regulation of H3K9me2 by chronic morphine and suggest novel chromatin-based mechanisms through which morphine-induced addictive-like behaviors arise. © 2012 the authors.