BACKGROUND: Marijuana use by teenagers often predates the use of harder drugs, but the neurobiological underpinnings of such vulnerability are unknown. Animal studies suggest enhanced heroin self-administration (SA) and dysregulation of the endogenous opioid system in the nucleus accumbens shell (NAcsh) of adults following adolescent ?(9)-tetrahydrocannabinol (THC) exposure. However, a causal link between proenkephalin (Penk) expression and vulnerability to heroin has yet to be established. METHODS: To investigate the functional significance of NAcsh Penk tone, selective viral-mediated knockdown and overexpression of Penk was performed, followed by analysis of subsequent heroin SA behavior. To determine whether adolescent THC exposure was associated with chromatin alteration, we analyzed levels of histone H3 methylation in the NAcsh via chromatin immunoprecipitation at five sites flanking the Penk gene transcription start site. RESULTS: Here we show that regulation of the Penk opioid neuropeptide gene in NAcsh directly regulates heroin SA behavior. Selective viral-mediated knockdown of Penk in striatopallidal neurons attenuates heroin SA in adolescent THC-exposed rats, whereas Penk overexpression potentiates heroin SA in THC-naÔve rats. Furthermore, we report that adolescent THC exposure mediates Penk upregulation through reduction of histone H3 lysine 9 (H3K9) methylation in the NAcsh, thereby disrupting the normal developmental pattern of H3K9 methylation. CONCLUSIONS: These data establish a direct association between THC-induced NAcsh Penk upregulation and heroin SA and indicate that epigenetic dysregulation of Penk underlies the long-term effects of THC.