Differential Effects of Cocaine Self-Administration Regimens on Incubation of Cocaine Craving and Nucleus Accumbens Neuronal Ensembles Activated by Cocaine-Associated Context

Abstract Drug addiction develops in a subset of users following repeated exposure, influenced by biopsychosocial factors. Rodent self-administration protocols, varying in drug access times, are used to study both controlled and compulsive drug-taking behaviors and their neurobiological underpinnings. Drug-associated cues and environmental contexts are well-established triggers for relapse, with susceptibility to these stimuli peaking during early abstinence and remaining elevated, thereby increasing relapse risk. This phenomenon, known as incubation of craving, has been replicated across substances in animal models. The associative learning between drug effects and contextual cues is encoded by neuronal ensembles activated by drug-associated stimuli, driving craving, seeking, and relapse. Neuronal ensembles in the nucleus accumbens (NAcc) are strongly involved in drug seeking, with parvalbumin-expressing fast-spiking interneurons playing a key role in this associative learning process. We investigated activation patterns in the NAcc triggered by cocaine-related cues following restricted- or extended-access self-administration and examined how forced abstinence alters these patterns, contributing to the incubation of cocaine craving. We also analyzed the engagement of parvalbumin-positive interneurons in NAcc neuronal ensembles before and after forced abstinence. Our findings show that the extended access protocol more effectively induced the incubation of cocaine craving. Neuronal activation in the NAcc core increased after thirty days of forced abstinence in both groups, with extended access rats showing consistently higher activation. Forced abstinence also increased NAcc shell activation, with no differences between protocols. NAcc core activation, but not shell, was influenced by cocaine consumption during training. Notably, extended access rats exhibited reduced parvalbumin interneurons activation following thirty days of forced abstinence. Based on these findings, we speculate that the transition from occasional to compulsive drug-taking may be driven by molecular changes in the NAcc core that enhance its responsiveness to drug-related cues. Additionally, the incubation of craving could be linked to impaired inhibitory control of NAcc core medium spiny neurons by PV interneurons. Competing Interest Statement The authors have declared no competing interest. Footnotes Correct name of one author from Casso Moraes Loss to Cássio Morais Loss