Role of Neuromedin S‐expressing Ventral Tegmental Area Neurons in Morphine Behavior

Abstract

ObjectiveOpioid dependence and addiction constitute a major health and economic burden, but our limited understanding of the underlying neurobiology limits better interventions. Alteration in the activity and output of dopamine (DA) neurons in the ventral tegmental area (VTA) is known to contribute to drug effects, but the mechanisms underlying these changes in VTA DA function remain relatively unexplored. We used TRAP to identify gene expression changes in VTA DA neurons following chronic morphine and found that Neuromedin S (NMS) is enriched in VTA DA neurons, and its expression is robustly increased by morphine. Neuromedin U receptor 2 (NMUR2), the receptor with which NMS interacts, is expressed within target regions of VTA DA neurons, like the nucleus accumbens (NAc). However, whether all VTA DA neurons express NMS, and their potential functional impact has yet to be determined. We hypothesize that NMS neurons represent a novel subset of VTA neurons that contribute to morphine‐elicited behavior. Specifically in these studies, we hypothesize that activating and inhibiting VTA‐NMS neurons will promote and inhibit morphine behaviors, respectively.MethodsAdult male and female NMS‐Cre mice and wild‐type littermates were used. Cre‐dependent viral vectors (AAV‐DIO‐mCherry, AAV‐DIO‐hM3Dq‐mCherry, AAV‐DIO‐hM4Di‐mCherry) were stereotaxically injected into the VTA to allow for DREADD‐mediated activation (Dq) or inhibition (Di) of VTA‐NMS neurons. Behavioral analyses were completed two weeks after surgery. Locomotor activity was assessed following saline (d1), saline + Clozapine‐N‐oxide (CNO, 0.3mg/kg, ip, d2‐d3), and morphine (15mg/kg, ip) + CNO (d4‐d8). A morphine + CNO challenge was done 1 week following d8. Immunohistochemistry was completed to validate viral expression and specific targeting of the VTA. Repeated‐measures two‐way ANOVA was used to determine significant differences (p<0.05) in locomotor behavior.ResultsWe found that a subset of VTA DA neurons express NMS (<5%) and that they exhibit diverse projection targets, including the NAc. Using DREADDs, we found that activation (Dq, n=15‐17/group) or inhibition (Di, n=15‐16/group) of VTA‐NMS neurons did not affect general locomotor activity or elicit CNO‐conditioned place preference or aversion. We find that both male (n=19,20) and female Dq mice (n=17,16) exhibit increased morphine‐induced locomotor activity. Additionally, locomotor response to a challenge morphine + CNO injection was significantly increased in Dq mice compared to controls (n=20,16). In contrast, male Di mice show a trend for the opposite effect, with decreased morphine‐elicited locomotor activity (n=20,20) and decreased response to challenge (n=20,20).ConclusionsVTA‐NMS neuron activation promotes morphine‐elicited behaviors including locomotion and sensitization. Future studies will determine if NMS‐VTA neuron inhibition decreases morphine behavior and whether VTA‐NMS neuronal activity modulates morphine conditioned place preference. Our current data suggest that VTA‐NMS neurons represent a subset of VTA neurons that may be functionally relevant for morphine responses.