Characterization of GalR1 and MOR mRNA Co-Expression in the Nucleus Accumbens Open Access

Abstract

The neuropeptide galanin is a potential therapeutic target for opioid use disorder and has been shown to be protective against opioid reward and withdrawal symptoms via a potential μ-opioid-galanin receptor (MOR-GalR1) heteromer mechanism. However, the distribution of MOR-GalR1 heteromers within addiction-related circuitry is unknown. Furthermore, it is unclear whether heteromer expression changes in response to chronic opioid exposure, and how these potential changes may affect behavior. The current study utilizes an oral fentanyl self-administration paradigm and RNAscope in situ hybridization to characterize the baseline expression of μ-opioid receptors (MOR) and galanin receptor 1 (GalR1) mRNA in the nucleus accumbens (NAc) and to investigate changes in their abundance following chronic opioid exposure. RNAscope demonstrated successful in situ hybridization of individual galanin receptor 1 and μ-opioid receptor mRNA in the nucleus accumbens and demonstrated the capacity for nucleus accumbens GABAergic neurons to express the MOR-GalR1 heteromer. In addition, RNAscope also demonstrated dynamic expression of galanin receptor 1 and μ-opioid receptor mRNA in GABAergic and non-GABAergic neurons following chronic fentanyl exposure. Together, these results provide evidence for a subpopulation of neurons in the brain to have the capacity to express functional GalR1-MOR heteromers, which may have important implications for the treatment of opioid use disorder.

Table of Contents

Table of Contents 

Introduction ………………………………….………………………..…………………..………………..………….….1 

The Opioid Epidemic ………………………………………………..……………………………………………….……1 

Opioid Signaling ………………………………..…………………………..……………………………….……….……3 

The Mesolimbic Dopamine System ………………………..………………………..…………………….....…….…3 

Galanin as a Potential Therapeutic Target ..…………...…………...…………...……………............….…...…5 

Materials and Methods ..…………...…………..…………...…………....………………..………………….……….8 

Animals ……..…………..…………..…………..…………..…………..…………..…………..…………….………....8 

Drugs ..…………...…………...…………...…………..…………...…………..………………………....…..………….8 

Oral Self-Administration ..………………..…………………………………………….…………………..…..………9 

Tissue Collection ………..…………………………………………….………………………….………..….….………9 

Cryosectioning ………..…………………………………………….………………………….……………….………..10 

RNAscope Multiplex Fluorescent Assay ………..…………………………………………….………….......…….10 

Confocal Microscopy ………..…………………………………………….………………………….……...………….11 

Image Analysis ………..…………………………………………….………………………….……………….………..11 

Statistical Analysis ………..…………………………………………….………………………….……………….…..12 

Results …………………………….………..…………………………………………….………………………….....…13 

Oral Self-Administration of Water, Saccharin, and Fentanyl Reveal 

No Differences in Liquid Intake ………………………………………..…….……………….……………….....… 13 

Characterization of GalR1 and MOR mRNA in Gad1+ and Gad1- Nuclei 

in Saccharin Mice …….………….…………………….……………….……………………………….……….………13 

Characterization of GalR1 and MOR mRNA in Gad1+ and Gad1- Nuclei 

in Fentanyl Mice ………..……………….……………………………………………………………………..………..14 

Chronic Exposure to Fentanyl Decreases GalR1 Expression in Gad1+ and Gad1- Nuclei 

Expressing GalR1 mRNA, but not MOR or GalR1+MOR, in the NAc ………………….……..…................15 

Figures and Graphs ………..…………………………………………….……………………….…….……………….17 

Figure 1 ………..………..……………………….………………………….…………………………….……………….17 

Figure 2 ………..………..……………………….………………………….………………………………….………….18 

Figure 3 ………..………..……………………….………………………….………………………………….………….21 

Figure 4 ………..………..……………………….………………………….…………………………………….……….23 

Discussion ………..……………………………………………….…………………….……………….…………….….25 

Future Directions ………..………………………………………………………….……………….……………….….28 

Conclusion…………………………………………….……………………….……………….…………………….……29 

References ………..…………………………………………….……………………….……………….…..……………30 

About this Honors Thesis

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School	Emory College
Department	Neuroscience and Behavioral Biology
Degree	B.S.
Submission	Honors Thesis
Language	English
Research Field	Biology, Neuroscience
Keyword	opioid addiction behavior molecular biology genetics
Committee Chair / Thesis Advisor	David Weinshenker, Emory University
Committee Members	Melvin Konner, Emory University Shannon Gourley, Emory University

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