Elucidating the Role of Gαz in the Behavioral Response to Opioids

Project Summary/Abstract Chronic pain and pain-related diseases affect more than 50 million adults in the United States. Most of the available pain therapies, including opioids, carry severe risks, such as the development of physical dependence and addiction. This lack of broadly effective and safe pain medications necessitates the search for novel pain management therapeutics. The identification of signaling proteins downstream of mu-opioid receptor (MOR) may provide a way to optimize the actions of opioid analgesics by promoting signaling pathways mediating analgesia, but not addiction-like behaviors. Previous work from our lab identified members of the G alpha (Gα) family and Regulator of G protein Signaling (RGS) protein family that are essential for the antinociceptive effects of multiple clinically used opioids. These include the G alpha z (Gαz) protein, an understudied member of the Gαi/o family, which preliminary data suggest plays a central role in the mouse ventrolateral periaqueductal gray (vlPAG) in mediating functional responses to MOR agonists in models of acute pain. Based on preliminary data, we hypothesize that Gαz mediates MOR signaling in a brain region- specific manner, and we will determine if Gαz signaling is involved in multiple behavioral effects of clinically prescribed opioids, or if it selectively modulates antinociceptive responses. To test this, we will utilize conditional knockout models to downregulate Gαz in the vlPAG and in the nucleus accumbens (NAc), a key component of the reward pathway, and investigate its regional role on the various behavioral and biochemical responses to opioids. In Aim 1, we will use qPCR, western blot analysis, RNAscope, and RNA sequencing to characterize the expression of Gαz and downstream pathways mediating response to opioids in the PAG and NAc. In Aim 2, we will use stereotaxic surgery, transgenic mouse lines, behavioral assays, and models of post- operative pain to determine the region-specific role of Gαz in the behavioral responses to opioids. We will use the hot plate assay to measure opioid antinociception, the Hargreaves assay to measure thermal hyperalgesia following opioid treatment, and the Conditioned Place Preference assay to measure opioid reward. To monitor sensory and affective signs of opioid withdrawal, we will use the Hargreaves assay, the marble-burying assay, the light-dark box assay, and the novelty suppressed feeding paradigm. We will also use the paw-incision model to determine the role of Gαz on opioid antinociception under pain states. Data collected from this proposal will provide a comprehensive understanding of the region and cell-specific role of Gαz signaling in response to opioids, contributing to the search for more efficacious and less deleterious pain medications. In addition, our proposed plan provides an opportunity for diverse training in molecular, cellular, and behavioral neuroscience, combined with bioinformatics and translational medicine, which will hone the skills required for success as an independent physician scientist.