Nucleus accumbens cholinergic interneurons and cue-induced cocaine craving - Project Summary A major problem for persons suffering from addiction is persistent vulnerability to relapse, even after long periods of abstinence. In the `incubation of cocaine craving' model of relapse, rats self-administer cocaine using a Long Access procedure, and then experience a prolonged abstinence period. During abstinence, rats exhibit a progressive intensification (incubation) of cue-induced cocaine craving. Plasticity involving medium spiny neurons in the nucleus accumbens core (NAcc) is required for the expression of `incubated' cue-induced craving. However, to date no incubation studies have focused on another cell type in the NAcc, the Cholinergic Interneuron (CIN). CINs make up only 1-2% of NAcc neurons, but are critically involved in learning, memory, and motivated behaviors. CINs are tonically active, and considerable work now supports the idea that a reduction in their activity augments motivated behavior. Other work has shown that dopamine D2 receptors (D2-R) on CINs reduce CIN activity and this correlates with greater motivated behavior; furthermore, D2-R are upregulated in NAc following cocaine self-administration (SA). Finally, striatal CINs are unique in exhibiting steady-state activation of the Integrated Stress Response (ISR) pathway, linked to their tonic firing. The ISR maintains cellular homeostasis by regulating protein translation. In other cell types, the level of ISR activity regulates motivation for cocaine. This has not been tested for CINs, but the ISR does affect D2-R signaling in these neurons. Integrating these findings, the over-arching hypothesis to be tested here is that cocaine SA and a subsequent abstinence period leads to increased D2-R expression on NAcc CINs. This leads to a potentiated inhibition of CINs in response to DA released during cue presentation, increasing cocaine seeking and leading to reduced ISR activity and translational reprogramming in CINs. Together these changes contribute to increased motivation for cocaine-paired cues. To test this hypothesis, transgenic rats and Cre-driven viruses will be used to selectively monitor and manipulate NAcc CINs. Aim 1 will use fiber photometry to determine the effect of cocaine SA and an abstinence period on the Ca2+ response (a proxy for activity) to a cocaine-paired cue in CINs, and how D2-R regulate this response. Recordings will be performed during cue-induced seeking tests on withdrawal day (WD) 1, prior to incubation, and WD40 (after incubation). Aim 2 will determine the role that CINs play in the expression of `incubated' seeking, through bidirectionally manipulating CINs via chemogenetics prior to WD1 or WD40 seeking tests. Aim 3 will explore how incubation of cocaine craving changes protein translation in CINs, using cell type specific translating ribosome affinity purification to identify active translating mRNAs (including mRNAs for the D2-R and markers of ISR activity) and a new viral tool (SPOTlight) to measure ISR activity. While this work is underway, I will participate in a multi-faceted training plan to develop the non-bench skills needed to reach my goal of becoming an independent PI in an academic setting. I will be supported by my Mentor, co- Mentor, and other Advisory Committee members, who have complementary expertise related to my goals.
