Persistent postsurgical pain associated with neurological injuries is resistant to treatment and occurs in 10 to 50 percent of patients after common surgical procedures. Although estimated to affect less than 5% of the population, chronic neuropathic pain causes significant physical, mental, and social dysfunction, negatively impacting patient well-being. Chronic pain syndromes are associated with long-term drug dependence, drug abuse and addiction, burdening our healthcare system with around $150 billion in costs annually. If the genetic and molecular mechanisms underlying the acute to chronic pain transition (ACPT) in susceptible individuals were understood, better preventative strategies and treatments would be available. We undertook unbiased genome-wide and transcriptome-wide experimental approaches to identify susceptibility genes underlying ACPT, using haplotype association mapping (HAM) of neuropathic pain behaviors to identify clinical quantitative trait loci (cQTL), and transcriptome analysis to identify expression quantitative trait loci (eQTL) within the dorsal root ganglion (DRG). Based on these two independent approaches, carbonic anhydrase 8 (Car8) was identified as a compelling candidate susceptibility gene (see Preliminary Studies). CAR8 lacks enzyme activity and inhibits inositol 1, 4, 5-triphosphate (IP3) binding to the inositol 1, 4, 5-triphosphate receptor, type 1 (ITPR1), which is important for intracellular calcium signaling and synapse formation. We first identified the Car8 gene candidate in cQTL mapping studies of mechanical allodynia after peripheral nerve injury. Independently, eQTL studies pinpointed Car8 as differentially expressed at Baseline in DRG, and as a major cis-regulator of other genes. We hypothesize that DNA polymorphisms at the Car8 locus alter its expression and regulate the expression of other genes, and variable expression of CAR8 serves as a putative mechanism of susceptibility to neuropathic pain after peripheral nerve injury. Our next major step is to show if CAR8 cellular localization and differential expression are directly associated with susceptibility to ACPT after peripheral nerve injury. We will test our hypotheses by 1) studying if differential expression of Car8 after peripheral nerve injury in null mutant and wild type mice influences susceptibility to neuropathic pain behavior associated with ACPT, which will show if Car8 is a major susceptibility gene in ACPT after peripheral nerve injury. We will also 2) define the cellular localization and quantify CAR8 and the ITPR1 protein and gene expression in the peripheral nerve, DRG, and spinal cord during ACPT, to better understand how Car8 acts as a cis-regulator of gene expression. These experiments may pinpoint the sites of action involving Car8 cis- regulation and may explain how CAR8 regulates neuronal activity and synaptic functioning during ACPT. Our studies are the first to give insight into whether altering the biological pathways related to CAR8 could be beneficial in developing new preventative or therapeutic strategies for neuropathic pain. Additionally, these studies may help us to understand treatment resistance and avert long-term drug dependence. PUBLIC HEALTH RELEVANCE: Chronic pain in susceptible individuals arises after nerve injury from trauma or common surgical procedures;it is often resistant to treatment. This negatively impacts the overall well-being of patients, leading to drug abuse and addiction while imposing a tremendous burden on our healthcare systems, with an average of $150 billion dollars spent every year. Our innovative studies will define the role of what may be a major new susceptibility gene, resulting in possible new preventative or therapeutic strategies for chronic pain that may help prevent long-term drug dependence.