Eicosanoids, the metabolic products of arachidonic acid, are potent mediators of various components of inflammation, including increased sensitivity to noxious stimuli. Although the mechanisms by which eicosanoids cause inflammation remain unknown, evidence suggests that they activate or facilitate activation of small diameter sensory neurons. This activation and the subsequent release of neuropeptides from these neurons results in neurogenic inflammation and enhanced pain sensation. The studies outlined in this proposal will examine the effects of various eicosanoids on the initiation of neurogenic inflammation by measuring their ability to alter the resting or stimulated release of the neuropeptides, substance P (SP) and calcitonin gene-related peptide (CGRP) from sensory neurons. Studies also will attempt to elucidate the cellular mechanisms of eicosanoid actions by determining cause-effect relationships between changes in peptide release and second messenger systems. Regulation of peptide release by eicosanoids will be studied using two experimental models, in vitro release from spinal cord slices and release from rat sensory neurons grown in culture. The former model allows examination of eicosanoid effects at the level of sensory input to the spinal cord, whereas the latter affords the opportunity to study the cellular mechanisms of eicosanoid actions on isolated sensory neurons. Neuronal tissues will be exposed to eicosanoids and the amount of SP and CGRP released from these preparations measured by radioimmunoassay. To assess the sensitization of sensory neurons, release evoked by various chemical stimuli will be determined in the presence an absence of eicosanoids. Using sensory neurons in culture, second messenger concentrations in cells will be measured in the presence and absence of sensitizing concentrations of eicosanoids. Intracellular content of second messengers will be altered to determine if these manipulations attenuate or potentiate eicosanoid effects on peptide release. In this manner, we can determine which signal transduction pathways mediate eicosanoid effects on peptide release from sensory neurons. Finally, we will determine if the eicosanoid-induced alterations in peptide release correlate with an changes in the phosphorylation of neuronal proteins from isolated sensory neurons. These studies will provide basic information as to the mechanisms of eicosanoid actions on sensor neurons. Furthermore, experiments will establish whether eicosanoid actions at the level of the spina cord are important in altering 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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