Women report greater temporomandibular joint (TMJ) pain when the concentration of estrogen is diminishing. Consistent with these results, estrogen reduces TMJ hypersensitivity in proestrus rats, when estrogen concentrations are highest. In our lab, a large gene array study (>10,000 genes) indicated that gene expression in the rat trigeminal ganglia (TG) was significantly affected by increased estrogen. High proestrus versus low diestrus levels of 17?-estradiol resulted in a 16 to 34 fold increase in the GABAA receptor subunit ?6 (Gabr?6), the glycine receptor subunit ?2 (Glr?2) and the beta-adrenergic receptor subunit ?1 (Adr?1). This significant increase occurred in the TG of na?ve rats and in rats with ligature of the masseter tendon, a model of chronic (>6 months) myogenic TMJ hypersensitivity. Since GABA, glycine and sympathomimetic amines can inhibit hypersensitivity the knock-down of Gabr?6, Glr?2 and Adr?1 expression would be expected to increase hypersensitivity. In preliminary studies siRNA knock-down of Gabr?6 expression in the TG increased hypersensitivity, increased the level of phosphorylated-ERK (p-ERK) in TG neurons and increased neuronal electrical activity. A homology search for estrogen receptor ? and ? (ER ? and ER?) binding sites 10kb of the transcriptional start site for these three genes showed that at least one potential binding site was present but the mechanism regulating the estrogen response of these genes is unknown. Based on this information we hypothesize that 17 ?-estradiol decreased TMJ hypersensitivity at proestrus by increasing Gabr?6, Glr?2 and Adr?1 expression in the TG and that this estrogen effect was due to interaction with the estrogen receptor and sequence proximal of the transcriptional start site. To address this hypothesis we propose two specific aims, in Aim #1 we will determine in the TG the role of Gabr?6, Glr?2 and Adr?1 in modulating TMJ hypersensitivity/cellular activity in both males and females. To complete aim #1 a ligature will be placed on the masseter tendon and TG expression of Gabr?6, Glr?2, and Adr?1 will be reduced through antagonists or siRNA. Hypersensitivity/cellular activity will be assessed at an acute stage (7 days post-ligature) and at a chronic stage (6 months post-ligature) using von Frey filaments, meal duration, electrophysiological recordings and by quantitating p-ERK levels. The goal of Aim #2 will be to characterize the role of ER? and ER? in controlling Gabr?6, Glr?2, and Adr?1 expression using chromatin immunoprecipitation, electrophoretic mobility shift assays and luciferase reporter constructs. We expect to show that an altered expression of these three genes can affect the TMJ nociceptive response and that these genes are responsible, in part, for the decrease in hypersensitivity observed in proestrus rats. We also expect to determine the mechanism by which estrogen modulates expression of these genes in TG cells.
Novel genes by which 17?-estradiol effects TMJ hypersensitivity will be identified, providing a greater understanding of why females suffer greater TMJ pain and allowing for targeted control of expression to reduce orofacial pain.
These studies will evaluate the role of three genes, the GABAA receptor subunit ?6 (Gabr?6), the glycine receptor subunit ?2 (Glr?2) and the beta-adrenergic receptor subunit ?1 (Adr?1). The expression of these three genes was regulated by estrogen and the role of these genes in the underlying molecular pathogenesis of TMJ pain will be evaluated in this application. These experiments will shed valuable insights into an area of TMJ biology and pathology that are poorly understood and that is highly prioritized by the NIDCR.
Novel genes by which 17?- estradiol effects TMJ hypersensitivity will be identified, providing a greater understanding of why females suffer greater TMJ pain and allow for targeted control of expression to reduce orofacial pain.
|Umorin, Mikhail; Stinson, Crystal; Bellinger, Larry L et al. (2016) Genes in the GABA Pathway Increase in the Lateral Thalamus of Sprague-Dawley Rats During the Proestrus/Estrus Phase. J Cell Physiol 231:1057-64|
|Kramer, Phillip R; Umorin, Mikhail; Bellinger, Larry L (2015) Attenuation of myogenic orofacial nociception and mechanical hypersensitivity by viral mediated enkephalin overproduction in male and female rats. BMC Neurol 15:34|
|Kramer, P R; Bellinger, L L (2014) Infusion of GabrÎ±6 siRNA into the trigeminal ganglia increased the myogenic orofacial nociceptive response of ovariectomized rats treated with 17Î²-estradiol. Neuroscience 278:144-53|
|Kramer, P R; Bellinger, L L (2013) Modulation of temporomandibular joint nociception and inflammation in male rats after administering a physiological concentration of 17Î²-oestradiol. Eur J Pain 17:174-84|
|Kramer, P R; Bellinger, L L (2013) Reduced GABAA receptor Î±6 expression in the trigeminal ganglion enhanced myofascial nociceptive response. Neuroscience 245:1-11|