Opioids target the μ-opioid receptor (MOR) to produce unrivaled pain management, but their addictive properties can lead to severe abuse. We developed a whole-animal behavioral platform for unbiased discovery of genes influencing opioid responsiveness. Using forward genetics in Caenorhabditis elegans, we identified a conserved orphan receptor, GPR139, with anti-opioid activity. GPR139 is coexpressed with MOR in opioid-sensitive brain circuits, binds to MOR, and inhibits signaling to heterotrimeric guanine nucleotide–binding proteins (G proteins). Deletion of GPR139 in mice enhanced opioid-induced inhibition of neuronal firing to modulate morphine-induced analgesia, reward, and withdrawal. Thus, GPR139 could be a useful target for increasing opioid safety. These results also demonstrate the potential of C. elegans as a scalable platform for genetic discovery of G protein–coupled receptor signaling principles.

In the originally published version of this article, Figures 1G and 2B contained errors. In Figure 1G showing representative traces of cAMP responses, the graph of adenosine data was an inadvertent duplication of the dopamine data. In Figure 2B, the representative iMSN responses to dopamine in dendrites were an inadvertent duplication of the data from iMSN responses to dopamine in cell bodies. These figures have now been corrected online and appear below. These graphical errors do not change the data analysis or scientific conclusions of the article.

Modulation of neuronal circuits is key to information processing in the brain. The majority of neuromodulators exert their effects by activating G protein coupled receptors (GPCR) that control the production of second messengers directly impacting cellular physiology. How numerous GPCRs integrate neuromodulatory inputs while accommodating diversity of incoming signals is poorly understood. In this study we develop an in vivo tool and analytical suite for analyzing GPCR responses by monitoring the dynamics of a key second messenger, cAMP with excellent quantitative and spatio-temporal resolution in various neurons. With this imaging approach in combination with CRISPR/Cas9 editing and optogenetics we interrogate neuromodulatory mechanisms of defined populations of neurons in an intact mesolimbic reward circuit and describe how individual inputs generate discrete second messenger signatures in a cell and receptor specific fashion. This offers a resource for studying native neuronal GPCR signaling in real time.

This review discusses the impact of neurotrophins and other trophic factors, including fibroblast growth factor and glial cell line-derived neurotrophic factor, on mood disorders, weight regulation and drug abuse, with an emphasis on stress- and drug-induced changes in the ventral tegmental area (VTA). Neurotrophins, comprising nerve growth factor, brain-derived neurotrophic factor (BDNF), and neurotrophins 3 and 4/5 play important roles in neuronal plasticity and the development of different psychopathologies. In the VTA, most research has focused on the role of BDNF, because other neurotrophins are not found there in significant quantities. BDNF originating in the VTA provides trophic support to dopamine neurons. The diverse intracellular signaling pathways activated by BDNF may underlie precise physiological functions specific to the VTA. In general, VTA BDNF expression increases after psychostimulant exposures, and enhanced BDNF level in the VTA facilitates psychostimulant effects. The impact of VTA BDNF on the behavioral effects of psychostimulants relies primarily on its action within the mesocorticolimbic circuit. In the case of opiates, VTA BDNF expression and effects seem to be dependent on whether an animal is drug-naïve or has a history of drug use, only the latter of which is related to dopamine mechanisms. Social defeat stress that is continuous in mice or intermittent in rats increases VTA BDNF expression, and is associated with depressive and social avoidance behaviors. Intermittent social defeat stress induces persistent VTA BDNF expression that triggers psychostimulant cross-sensitization. Understanding the cellular and molecular substrates of neurotrophin effects may lead to novel therapeutic approaches for the prevention and treatment of substance use and mood disorders.

Social defeat stress causes social avoidance and long-lasting cross-sensitization to psychostimulants, both of which are associated with increased brain-derived neurotrophic factor (BDNF) expression in the ventral tegmental area (VTA). Moreover, social stress upregulates VTA mu-opioid receptor (MOR) mRNA. In the VTA, MOR activation inhibits GABA neurons to disinhibit VTA dopamine neurons, thus providing a role for VTA MORs in the regulation of psychostimulant sensitization. The present study determined the effect of lentivirus-mediated MOR knockdown in the VTA on the consequences of intermittent social defeat stress, a salient and profound stressor in humans and rodents. Social stress exposure induced social avoidance and attenuated weight gain in animals with non-manipulated VTA MORs, but both these effects were prevented by VTA MOR knockdown. Rats with non-manipulated VTA MOR expression exhibited cross-sensitization to amphetamine challenge (1.0 mg/kg, i.p.), evidenced by a significant augmentation of locomotion. By contrast, knockdown of VTA MORs prevented stressinduced cross-sensitization without blunting the locomotor-activating effects of amphetamine. At the time point corresponding to amphetamine challenge, immunohistochemical analysis was performed to examine the effect of stress on VTA BDNF expression. Prior stress exposure Publisher's Disclaimer: This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final citable form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain. NIH Public Access

Opioids exert their analgesic effects by activating the μ‐opioid receptor (MOR), a G protein‐coupled receptor expressed in the nervous system. In addition to clinical pain relief, activation of MOR produces euphoria and leads to dependence. Despite significant process in understanding molecular mechanisms of MOR signaling, the regulatory mechanisms of MOR function are not fully understood. To better understand MOR regulation and to uncover genes influencing MOR signaling, we developed a genetic behavioral platform utilizing nematode C. elegans. Mammalian MOR was transgenically expressed in the nervous system of C. elegans endowing the resulting model (tgMOR) with opioid sensitivity. We validated tgMOR platform showing conservation of pharmacological properties and regulatory mechanisms of MOR mediated behavioral responsiveness. We then performed a large scale forward genetic screen isolating a population of mutants with altered opioid sensitivity. Integrating whole genome sequencing, CRISPR/Cas9 gene editing and transgenic rescue, we identified a set of candidate negative modulators of MOR signaling. One of these genes encoded an orphan GPCR, FRPR‐13. Our phylogenic and functional analysis revealed that mammalian ortholog of FRPR‐13 is GPR139. We found GPR139 to be extensively co‐expressed with MOR in the mammalian nervous system. Knockout of GPR139 in mice enhanced opioid analgesia and reward, but diminished withdrawal. These observations suggest the existence of an “anti‐opioid” system that regulates the extent of MOR signaling in vivo. Our findings further showcase the utility of C. elegans as a scalable platform for genetic discovery of novel GPCR signaling principles. Support or Funding Information This work was supported by an NIH Cutting Edge Basic Research Award (R21DA040406) to B.G. and K.A.M., DA036596 to K.A.M., and an NIH Center of Biomedical Research Excellence Grant (P20GM103638) to University of Kansas Genome Sequencing Core.

The majority of neuromodulators exert their effects by activating G protein coupled receptors (GPCRs) which shape neuronal properties through the regulation of second messengers. The mechanisms of signal decoding at GPCRs amidst diversity of the effects associated with their ctivation remains elusive. Furthermore, there is a paucity of understanding of how GPCRs signal to produce unique responses impacting cellular physiology while converging on a limited set of second messenger signaling cascades. Although significant progress has been made studying GPCRs in reconstituted systems, information decoding processes by GPCRs in an endogenous setting is understood less due to a lack of appropriate tools. Here, we developed a novel in vivo reagent that allows monitoring GPCR signaling in the endogenous setting from genetically defined populations of cells. The approach takes advantage of knock‐in mice engineered to express a genetically encoded FRET‐based cAMP biosensor in a Cre‐dependent fashion. We applied this reagent to study integration of GPCR signals in striatal medium spiny neurons both in culture and in brain slices where intact circuits were activated by optogenetics. Combined with CRISPR/Cas9 genomic editing, this approach enabled us to identify individual receptors and analyze their contributions to downstream signaling in the native environment. We identified key spatial differences in pharmacological responsiveness by comparing dendritic regions with the neuronal cell body as well as plasma membrane and internal cellular signaling compartments. We then conducted comparative analysis of GPCR signaling mediated by two opposing circuits modulated by the neurotransmitter dopamine, revealing the mechanisms of dopaminergic processing as well as major differences in the generation of discrete second messenger signatures. We further demonstrate how cAMP signal processing mechanisms contribute to allostatic adaptations that imbalance tuning between striatal output neurons that may underlie states of reward and aversion.Support or Funding InformationNIH 1R01DK110621, APS STRIDE Undergraduate Summer Research Fellowship 1R25HL115473‐01.This work was supported by NIH grants: DA041207 (to B.S.M.), DA036596 (to K.A.M.), and DA026405 (to K.A.M.).