Project Summary Millions of people suffer from neuropathic pain and the current treatment protocols and therapies are limited in their effectiveness. There is a clear need for the development of novel, non-addictive treatment strategies for neuropathic pain. Dorsal root ganglion (DRG) neuronal hyperexcitability is central to the pathology of neuropathic pain, so much so that application of local anesthetics is currently considered as a first line treatment strategy for neuropathic pain. However, studies on the efficacy to these local anesthetic patches has been mixed. Notwithstanding, the coordinated movement of ion channels, especially voltage-dependent sodium channels, from intracellular pools to the sites of nerve injury has been suggested to be the underlying cause of electrogenesis and ectopic firing in neuropathic pain. Thus instead of blocking sodium channels, an alternative method to treating neuropathic pain could be to disrupt their trafficking. Recent studies have indicated that WW domain-containing ubiquitin ligases are downregulated during nerve injury causing an increase in membrane targeting of sodium channels. The scaffold proteins responsible for sodium channel targeting and membrane stabilization in DRG neurons have not been fully characterized although our recently published studies identified the Magi1 scaffold protein a potential candidate. Using electrophysiological, biochemical, molecular, pain behavioral assays, and a novel in vivo method to knockdown genes in DRG neurons, we will test the hypotheses: MAGI1, a PDZ and WW domain-containing scaffold protein, is critical for NaV membrane expression in rodent and human DRG neurons and targetable for the treatment of neuropathic pain. The specific aims are: (1) To study the effects of Magi1/MAGI1 knockdown on rodent and human DRG neuronal excitability and to determine if Magi1 deficiency impacts the development of neuropathic pain during nerve injury. (2) To investigate how Magi1 is regulated by neuropathic pain and whether downregulating Magi1 can provide analgesia after established neuropathic pain. (3) To demonstrate that local application of WW domain peptidomimetics can reduce neuropathic pain behavior. This research project will validate MAGI1 as a therapeutic target to treat neuropathic pain.