The main objective of this research grant application is to direct our research efforts towards a more accurate and basic understanding of the molecular events that occur in several biologically important endogenous peptidergic pathways - enkephalinergic, endorphinergic, dynorphinergic, and substance P-ergic. Each one of these four peptidergic pathways consists of a large precursor, intermediate precursors, the working peptide itself, and several inactive metabolites. Each step in each peptide pathway is, in turn, mediated by appropriate proteolytic enzymes. Metabolism is directed along tissue-specific pathways. We will provide a metabolic profile of all of the constituents of these four peptidergic pathways as a background against which we will monitor the effects of stressor pain on these peptidergic systems. The hypothesis is that these three opinoid peptidergic pathways are mobilized following an appropriate stimulus, and that an inverse relationship exists between the three opioid pathways versus the substance P-ergic pathway. The biological tissues that will be studied in thisinvestigation include: human and canine tooth pulp, cerebrospinal fluid, and selected brain regions. The various analytical assay methods that will be utilized include RP-HPLC, radioimmunoassay, radiorecptor assay, and our novel mass spectrometric procedure. The MS procedure that was developed in our labe has the highest level of structural specificity that is available for endogenous peotide measurement. The purpose of this study is to provide a firmer molecular basis for our understanding of these cellular processes that operate in pain, stress, addiction, and other brain-related phenomena.
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