Dissecting Paranigral VTA Nociceptin Signaling during Stress and Motivated Behaviors

Abstract ID 93034 Poster Board 149 Despite significant advances in neuroscientific research, the neural circuitry underlying the association between stress and motivation is not clearly defined. Previous research in our lab identified neurons that release the endogenous opioid nociceptin in the paranigral ventral tegmental area (pnVTAPnoc), a region in the brain associated with reward-seeking and motivation. Our lab discovered that activating these neurons will suppress the motivation of mice to obtain a reward (Parker et al., 2019). Following this, our experiments using GCaMP6s, a green calcium indicator, showed that pnVTAPnoc neurons also become activated when mice are exposed to acute stressors such as footshock or predator odor. To further understand how pnVTAPnoc neurons are affected by different forms of stress, we have now evaluated changes to Pnoc and c-Fos expression in pnVTAPnoc neurons at the protein and mRNA-levels following a chronic restraint stress model. While this new data and our lab's prior work collectively suggest that nociceptin release from pnVTAPnoc neurons is engaged during both stress and motivation, the precise mechanisms are still unknown due to a lack of available tools that could allow investigation of nociceptin release dynamics with high spatiotemporal resolution. To address this, we worked with collaborators to develop NOPLight, a genetically encoded biosensor for detection of nociceptin that will allow for investigation of in vivo nociceptin release dynamics. NOPLight is a biochemically modified version of the nociceptin receptor (NOPR) that has been engineered to contain a green fluorescent protein (GFP), and works by coupling the NOPR agonist-induced conformational change to the activation of the GFP, causing it to fluoresce when a NOPR agonist binds to the sensor. In characterizing this sensor, we found that when expressed in the VTA, NOPLight exhibits dose dependent responses to systemic administration of a NOPR agonist. We also found that NOPLight expressed in the VTA dynamically responds to endogenous behaviorally-evoked nociceptin release during reward-seeking in fixed ratio -1, -3, and progressive ratio paradigms. The observed pattern of NOPLight signal closely resembles the pnVTAPnoc neuron calcium activity from our lab's prior study, suggesting that NOPLight detection of endogenously released nociceptin recapitulates the activity of the nociceptin-producing neurons. In ongoing studies, we are piloting simultaneous detection of nociceptin release and nociceptin neuron activity by using dual-color fiber photometry recording of NOPLight and jRCaMP1b respectively during acute stress exposure and in motivated reward-seeking tasks. By using different combinations of reward-seeking and acute stress behavioral assays in freely-moving and head-fixed conditions, we will gain a thorough understanding of how the magnitude and timing of nociceptin neuron activity and nociceptin release are related during behavior. Altogether, these findings provide insight into the dynamics of the less known neuropeptide nociceptin and its role in reward, motivation, and stress as expressed in psychiatric conditions such as depression and substance use disorder. P50 MH119467-02, P30 DA048736, F31 DA059438, Mary Gates Research Scholarship, NAPE Undergraduate Research Training Award.