The long term goal of this study is to determine the impact of gender differences and sex hormones on gene(s) and molecular pathways leading to and effecting acute and persistent joint pain as observed in the temporomandibular joint (TMJ) and associated peripheral tissues. The objective of this application is to determine the peripheral effect that steroid sex hormones have on acute pain and identify nociception gene(s) that function in pain pathways as modeled in the TMJ. Our central hypothesis is that changes in physiological estrogen, progesterone and testosterone concentrations modulates acute pain as modeled in the TMJ through action at peripheral tissue(s) within and surrounding the joint and/or within the trigeminal ganglia. To test our central hypothesis and attain the objective of this application we propose the following two Specific Aims first, we will characterize pain in the TMJ and surrounding region related to endogenous and administered estrogen and progesterone in females. Second, we will characterize gender differences in pain in the TMJ and surrounding region by determining the modulatory effect of sex steroids in males. The working hypothesis for Specific Aim one is that cyclical plasma levels of estrogen and/or progesterone in females modulates TMJ pain, in part, through changes in gene expression in the joint and/or trigeminal ganglia. To test the working hypothesis the experimental approach will consist of injecting complete Freund's adjuvant (CFA) into the superior TMJ space. The ovariectomized Sprague-Dawley rats will have been treated with physiological doses of 17-beta estradiol and/or progesterone with and without hormone receptor antagonists. The rats will be first tested for TMJ pain using an established non-invasive behavioral procedure. This test uses the time required to eat a meal (i.e., meal duration), as a behavioral measure of pain. Second, changes in the transcript levels of nearly four thousand genes will be measured in the TMJ and trigeminal ganglia of these treated animals with gene microarrays. Third, candidate gene(s) from the gene array studies that were 1) modulated by hormonal treatment and 2) function in pain related pathways will be further quantitated by real-time PCR, ELISA and/or westerns to measure transcript and protein levels. The working hypothesis for Specific Aim two is that low non-cyclical plasma levels of estrogen and/or high testosterone in the males modulates pain through changes in gene expression in the joint and/or trigeminal ganglia. To test the working hypothesis the experimental approach will consist of measuring pain (i.e., meal duration) on castrated and intact male rats treated with and without estrogen before and after injecting CFA in the TMJ. Upon sacrifice transcript levels will be measured. We expect these data will be used for the evaluation and treatment of gender-related TMJ disorders and for mechanistic studies into hormonal action on joint pain. The rationale for completing these proposed studies is that once the hormones influencing pain sensitivity have been characterized and the gene(s) modulated by these hormonal effects in the joint and trigeminal ganglion identified, targets for pharmaceutical intervention of pain in the TMJ can be pursued.
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