Lipases are critically important in signal transduction in that they produce a number of first and second messengers that mediate numerous physiological processes. These lipases include phospholipase A2s which liberate arachidonic acid (the rate limiting step in eicosanoid biosynthesis), phospholipase Cs which liberate inositol trisphosphate and diacylglycerols (DAGs), and phospholipase Ds which synthesize phosphatidic acids (PAs) and lysophosphatidic acids (LPAs). Although much work has focused on the involvement of products of PLA2s in pain and inflammation, few studies have examined whether other lipase products modulate neurogenic inflammation and nociception. A major mechanism to account for both neurogenic inflammation and enhanced pain sensitization involves augmenting the activity of small diameter sensory neurons with the subsequent release of neuropeptides from these neurons. Consequently, the studies outlined in this proposal will examine the effects of various lipase products on sensitization of sensory neurons. We also will determine whether select inflammatory mediators augment peptide release from sensory neurons and whether these actions are mediated by lipase products. Studies also will determine whether peripheral inflammation increases the activity of the PLC and PLD signaling pathways and thus contributes to long-term sensitization. The role of lipase products will be studied using two experimental models: (1) rat sensory neurons grown in culture, and (2) spinal cord and dorsal root ganglia (DRGs) from rats with unilateral inflammation. The former model provides a unique opportunity to examine the mechanisms of action of lipase products on sensory neurons and the transduction cascades activated by inflammatory mediators without significant interference from other types of cells. The latter model affords the opportunity to study actions of lipases at the level of sensory input to the central nervous system in naive animals or during chronic inflammation. We will manipulate the lipase transduction cascades by using drugs that inhibit specific enzymes in the pathways, by reducing expression of specific proteins with small linterfering RNA, and by over expressing proteins by infection with adenoviral constructs. The ability of lipase products to augment stimulated release of the neuropeptides, substance P (SP) and calcitonin gene-related peptide (CGRP) from sensory neurons and to increase excitability will be used as indices of sensitization. These studies will provide basic information as to the transduction cascades that mediate neuronal sensitivity during inflammation. This knowledge is critically important for understanding the process of neurogenic inflammation and in ultimately designing new drug therapies for the management of inflammatory diseases.
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