The kappa opioid receptor (KOR) is part of the opioid neuromodulatory system that influences pain and mood perception. KOR agonists have been recognized for their analgesia properties but are also associated with dysphoria, limiting their potential therapeutic use. Deciphering the intracellular signaling events activated by KOR that modulate the therapeutic and aversive effects may help aid in the development of novel compounds. KOR is a G protein coupled receptor (GPCRs) that is endogenously activated by opioid peptides. The balance of heterotrimeric G protein activation and deactivation is central in dictating its cellular and behavioral responses. Regulators of G protein signaling (RGS) proteins serve as an endogenous antagonist of GPCR signaling, determining the extent, and timing of their signaling and may offer a way to fine tune intracellular signaling. Our preliminary data demonstrates that the R7 RGS family modulates KOR signaling and influences KOR-mediated behaviors. In this proposal we will characterize the role of RGS7 on KOR signaling by identifying the neuronal population that drives KOR-mediated aversion and determine its impact of phosphorylation on KOR signaling. In Aim 1, we will identify the neuronal population that RGS7 acts to modulate KOR-mediated aversion. This aim will utilize a well-established conditional place aversion test and novel operant task to assess responses to aversive stimuli. Aim 2 will investigate the impact phosphorylated RGS7 has on KOR signaling and KOR- mediated aversion. We will determine the extent to which phosphorylation affects the interaction with its binding partner R7-binding protein (R7BP), which is required for the stability and catalytic activity of R7 RGS proteins. In Aim 3 we will determine the contribution of G protein signaling on KOR-mediated behaviors. This proposal will utilize a combination of transgenic mouse model as well as viral and chemogenetic approaches to target RGS protein in a cell-specific manner and employ novel aversion behavioral tests. At the successful completion of the proposed research, the expected outcomes are to determine the contribution of the RGS-G protein pathway towards specific KOR-mediated behaviors and delineate the cell-specific contributions. The proposed research is innovative as we use an operant paradigm to employ aversive stimuli that allows us to address the long-term effects within a homecage environment. These results provide a strong basis for furthering our knowledge of KOR signaling and to delineate the impact of RGS-G protein signaling on aversive behaviors which is expected to have significant impact on the future therapeutic development.