Project Summary The goal of our research is to understand the cellular and molecular mechanisms of chronic itch, a disease that interferes with normal daily activity and can have serious clinical consequences. Many pathological conditions can lead to chronic itch such as localized skin diseases or more systemic conditions like cholestasis and kidney failure. Itch can also be a side effect of many therapeutic drugs. Current therapies including antihistamines are ineffective in most chronic itch conditions suggesting the involvement of histamine independent pathways. A major hurdle in understanding histamine-independent itch is the lack of knowledge about the receptors directly activated by non-histaminergic stimuli. Primary sensory neurons in dorsal root ganglia (DRG) play an essential role in detecting itch. Our lab has identified members of a G protein-coupled receptor (GPCR) Mrgpr family that are specifically expressed in DRG and function as itch receptors. Recently we showed that MrgprA1 in mice and its human homologue MRGPRX4 function as novel itch receptors, detecting bilirubin and bile acids (BAs). More importantly, we demonstrated that MrgprA1/X4 play an essential role for mediating cholestatic pruritus (itch), a condition resulting in elevated bilirubin and BAs due to bile duct blockage. Our preliminary data suggest that MrgprA1 are novel receptors for various therapeutic drugs and mediates drug-induced itch side effect. Preliminary data show that MrgprA1 also contributes to allergic contact dermatitis (ACD). Furthermore, previous studies and our preliminary data suggest BAs are also mediators for uremic pruritus. In this proposal, we will take a multidisciplinary approach to test the hypothesis that MrgprA1/X4 are itch receptors and mediate itch caused by different disease conditions including medication caused itch side effect, ACD, and kidney disease which dramatically impact the quality of patient's life and the underlying mechanisms remain unclear. In Aim I, we will test the hypothesis that some of drugs with itch side effect are pruritogens themselves by directly activating pruriceptive (itch sensing) neurons via MrgprA1 using mouse genetics, pharmacology, in vitro Ca2+ imaging, electrophysiology, and in vivo DRG imaging. In Aim II, we will test the hypothesis that MrgprA1 in sensory nerves and its peptide agonist NPFF released from nearby mast cells in the skin contributes to ACD itch using different ACD mouse models combined with mouse genetics and pharmacological approaches. In addition, we will determine whether pruriceptive neurons are activated under ACD conditions and if so whether MrgprA1 and NPFF mediate the activation using in vivo DRG GCaMP imaging. In Aim III, we will employ two uremic itch mouse models, an adenine-induced or a uremic itch patient plasma injection model to test the hypothesis that BAs mediate uremic itch by activating MRGPRX4 expressed in mouse DRG neurons. Moreover, we will identify other itch mediators...