Nucleus accumbens inputs and cell types mediating the incubation of oxycodone craving - Project Summary Individuals with Opioid Use Disorder continue to remain vulnerable to relapse due to craving for the drug during abstinence. I will study oxycodone (Oxy) craving using the incubation of craving model, in which rats show progressive intensification of cue-induced seeking for drugs of abuse including Oxy during the weeks after ending drug self-administration. Craving then remains high for an additional period before declining. Our published behavioral data show that incubated Oxy seeking requires upregulation of Ca2+-permeable AMPA receptors (CP- AMPAR) on medium spiny neurons (MSN) in both Core and Shell subregions of the nucleus accumbens (NAc). Our preliminary data show this occurs on both of the main MSN subtypes: DA D1R-expressing MSN (D1+) and adenosine A2a/DA D2R-expressing MSN (A2a/D2+ MSN; A2a and D2 receptors co-localize). I propose to extend our knowledge of pathways (Aim 1) and MSN subtypes (Aim 2) involved in Oxy incubation. One candidate input originates in the basolateral amygdala (BLA). Our preliminary electrophysiological data show elevated CP- AMPAR levels in both BLA-Core and BLA-Shell pathways after Oxy incubation. Besides our data, there have been no studies of these pathways in Oxy incubation. Therefore, in Aim 1, I will use whole cell patch clamp recordings to measure the contribution of CP-AMPARs to synaptic transmission in the BLA-Core and BLA-Shell pathways onto D1+ or A2a+ MSN. I will use D1-Cre and A2a-Cre rats crossed with TdTomato reporter rats (D1- Td, A2a-Td) to selectively patch D1+ or A2a+ MSN. I hypothesize that CP-AMPAR upregulation will occur on both MSN subtypes. I will then determine if inhibition of these pathways disrupts incubated cue-induced Oxy seeking. The inhibitory DREADD AAV-hM4Di will be infused into BLA, and CNO will be delivered into Core or Shell to inhibit nerve terminals prior to a seeking test. I hypothesize that both BLA-Core and BLA-Shell pathways contribute to Oxy incubation. For Aim 2, it is known that the mu-opioid receptor (MOR) is expressed on a subset of D1+ MSN and A2a+ MSN in NAc. However, the potential role of these MOR+ MSN in incubated Oxy seeking has not been investigated. First, I will determine excitatory synaptic properties of MOR+ MSN compared to MOR- MSN after Oxy incubation. Cre-dependent AAV expressing eYFP under control of the MOR promoter will be infused in Core or Shell of D1-Td or A2a-Td rats, enabling differentiation of MOR+/D1+, MOR-/D1+, MOR+/A2a+ and MOR-/A2a+ MSN. For each, I will determine synaptic properties (paired pulse ratio, AMPA/NMDA, and CP- AMPAR levels). Second, I will determine if MOR+ MSN are necessary for incubated Oxy seeking by infusing AAV-DIO-hM4Di into Core or Shell of MOR-Cre rats. I hypothesize that MOR+ MSN will exhibit distinct plasticity and contribute to Oxy incubation. This project will address a literature gap for understanding incubation of Oxy craving while providing rigorous training in electrophysiology, chemogenetics and experimental design. This proposal will also advance my professional development through writing manuscripts and attending professional conferences, enabling me to become a well-rounded scientist competitive for future PI or staff-scientist positions.
