Temporomandibular Joint Nociception Research Articles

Antinociceptive Efficacy of 15-Deoxy-Δ12,14-Prostaglandin J2 Therapy in Response to Experimentally Induced Temporomandibular Joint Arthritis: A Systematic Review of Studies in Rats

The aim of the present systematic review was to assess the antinociceptive efficacy of 15-deoxy-Δ12,14-prostaglandin J2 (15d-PGJ2) therapy in rats with experimentally induced temporomandibular joint (TMJ) arthritis. The focused question was “Is 15d-PGJ2 therapy effective in the management of TMJ nociception?” Indexed databases were searched without time and language restrictions up to and including September 2023 using different key words. Original studies were included. Risk of Bias (RoB) was assessed using the SYRCLE tool. Six studies performed in male Wistar rats with experimentally induced TMJ arthritis were included. The observation or follow-up period ranged between 45 min and 14 days. Four studies reported that 15d-PGJ2 therapy retards the production of proinflammatory cytokines in TMJ tissues. Four studies reported that 15d-PGJ2 therapy inhibits leukocyte migration and plasma extravasation in TMJ tissues. In one study, the expression of decay-accelerating factor in TMJ tissues increased after 15d-PGJ2 therapy. One study showed that 15d-PGJ2 inhibits nociception in a dose-dependent manner via the activation of peripheral kappa/delta opioid receptors. Prior sample-size-estimation (SSE) was performed in none of the studies and all studies had a high RoB. Due to a high RoB, methodological variations, and the absence of prior SSE within the included studies, it is demanding to derive an absolute verdict regarding the antinociceptive efficacy of 15d-PGJ2 therapy in response to experimentally induced TMJ arthritis.

Effect of propranolol on temporomandibular joint pain in repeatedly stressed rats.

Stress substantially increases the risk of developing painful temporomandibular disorders (TMDs) by influencing the release of endogenous catecholamines. Propranolol, an antagonist of β-adrenergic receptors, has shown potential in alleviating TMD-associated pain, particularly when the level of catecholamines is elevated. The aim of this study was to explore whether intra-articular propranolol administration is effective in diminishing temporomandibular joint (TMJ) pain during repeated stress situations. Additionally, we investigated the effect of repeated stress on the expression of genes encoding β-adrenoceptors in the trigeminal ganglion. In the present study, rats were exposed to a stress protocol induced by sound, then to the administration of formalin in the TMJ (to elicit a nociceptive response), followed immediately afterward by different doses of propranolol, after which the analgesic response to propranolol was evaluated. We also assessed the levels of beta-1 and beta-2 adrenergic receptor mRNAs (Adrb1 and Adrb2, respectively) using reverse transcription-quantitative PCR (RT-qPCR). Our findings revealed that propranolol administration reduces formalin-induced TMJ nociception more effectively in stressed rats than in non-stressed rats. Furthermore, repeated stress decreases the expression of the Adrb2 gene within the trigeminal ganglion. The findings of this study are noteworthy as they suggest that individuals with a chronic stress history might find potential benefits from β-blockers in TMD treatment.

Effect of sound-induced repeated stress on the development of pain and inflammation in the temporomandibular joint of female and male rats.

Temporomandibular disorder (TMD) is a common painful condition of the temporomandibular joint (TMJ) and associated structures. Stress is a significant risk factor for developing this painful condition that predominantly affects women. This study aimed to test the hypothesis that stress increases the risk of developing TMJ pain by facilitating inflammatory mechanisms in female and male rats. To test this hypothesis, we evaluated TMJ carrageenan-induced expression of pro-inflammatory cytokines and migration of inflammatory cells and TMJ formalin-induced nociception in female and male rats submitted to a repeated stress protocol induced by sound. We found that sound-induced repeated stress facilitates TMJ inflammation and contributes to TMJ nociception development equally in females and males. We conclude that stress is a risk factor for developing painful TMJ conditions in males and females, at least in part, by favoring the inflammatory process similarly in both sexes.

Biological and Molecular Docking Evaluation of a Benzylisothiocyanate Semisynthetic Derivative From Moringa oleifera in a Pre-clinical Study of Temporomandibular Joint Pain.

ObjectiveMoringa oleifera possesses multiple biological effects and the 4-[(4′-O-acetyl-α-L- rhamnosyloxy) benzyl] isothiocyanate accounts for them. Based on the original isothiocyanate molecule we obtained a semisynthetic derivative, named 4-[(2′,3′,4′-O-triacetyl-α-L-rhamnosyloxy) N-benzyl] hydrazine carbothioamide (MC-H) which was safe and effective in a temporomandibular joint (TMJ) inflammatory hypernociception in rats. Therefore, considering that there is still a gap in the knowledge concerning the mechanisms of action through which the MC-H effects are mediated, this study aimed to investigate the involvement of the adhesion molecules (ICAM-1, CD55), the pathways heme oxygenase-1 (HO-1) and NO/cGMP/PKG/K+ATP, and the central opioid receptors in the efficacy of the MC-H in a pre-clinical study of TMJ pain.MethodsMolecular docking studies were performed to test the binding performance of MC-H against the ten targets of interest (ICAM-1, CD55, HO-1, iNOS, soluble cGMP, cGMP-dependent protein kinase (PKG), K+ATP channel, mu (μ), kappa (κ), and delta (δ) opioid receptors). In in vivo studies, male Wistar rats were treated with MC-H 1 μg/kg before TMJ formalin injection and nociception was evaluated. Periarticular tissues were removed to assess ICAM-1 and CD55 protein levels by Western blotting. To investigate the role of HO-1 and NO/cGMP/PKG/K+ATP pathways, the inhibitors ZnPP-IX, aminoguanidine, ODQ, KT5823, or glibenclamide were used. To study the involvement of opioid receptors, rats were pre-treated (15 min) with an intrathecal injection of non-selective inhibitor naloxone or with CTOP, naltrindole, or norbinaltorphimine.ResultsAll interactions presented acceptable binding energy values (below −6.0 kcal/mol) which suggest MC-H might strongly bind to its molecular targets. MC-H reduced the protein levels of ICAM-1 and CD55 in periarticular tissues. ZnPP-IX, naloxone, CTOP, and naltrindole reversed the antinociceptive effect of MC-H.ConclusionMC-H demonstrated antinociceptive and anti-inflammatory effects peripherally by the activation of the HO-1 pathway, as well as through inhibition of the protein levels of adhesion molecules, and centrally by μ and δ opioid receptors.

Limonene, a citrus monoterpene, non-complexed and complexed with hydroxypropyl-β-cyclodextrin attenuates acute and chronic orofacial nociception in rodents: Evidence for involvement of the PKA and PKC pathway

BackgroundChronic orofacial pain is a serious public health problem with a prevalence of 7–11% in the population. This disorder has different etiologies and characteristics that make pharmacological treatment difficult. Natural products have been shown to be a promising source of treatments for the management of chronic pain, as an example the terpenes. PurposeThe aim of this study was to evaluate the anti-nociceptive and anti-inflammatory effects of one of these terpenes, d-limonene (LIM - a common monoterpene found in citrus fruits) alone and complexed with hydroxypropyl-β-cyclodextrin (LIM/HPβCD) in preclinical animal models. MethodsOrofacial pain was induced by the administration of hypertonic saline on the corneal surface, the injection of formalin into the temporomandibular joint (TMJ), or chronic constriction injury of the infraorbital nerve (CCI-IoN). The study used male Wistar rats and Swiss mice treated with LIM (50 mg/kg), LIM/HPβCD (50 mg/kg), vehicle (control), gabapentin or morphine, and eyes wiping (induced by hypertonic saline), face rubbing (formalin-induced in TMJ) or mechanical hyperalgesia (provoked by CCI-IoN) were assessed. Additionally, ELISA was used to measure TNF-α, and western blot analysis to assess levels of PKAcα, NFκB, p38MAPK and phosphorylated PKC substrates. Serum levels of aspartate aminotransferase (AST) and alanine transferase (ALT) were also evaluated. ResultsLIM and LIM/HPβCD significantly reduced (p < 0.001) corneal nociception and formalin-induced TMJ nociception. In addition, both substances attenuated (p < 0.001) mechanical hyperalgesia in the CCI-IoN model. The antinociceptive effect induced by LIM and HPβCD/LIM was associated with decreased TNF-α levels, downregulation of the NFκB and p38MAPK signalling pathways and reduced PKC substrate phosphorylation and PKA immunocontent. Moreover, the results demonstrated that complexation with HPβCD was able to decrease the therapeutic dose of LIM. ConclusionLIM was found to be a promising molecule for the treatment of orofacial pain due to its capacity to modulate some important mediators essential to the establishment of pain, and HPβCD can be a key tool to improve the profile of LIM.

P2X7-induced nociception in the temporomandibular joint of rats depends on inflammatory mechanisms and C-fibres sensitization.

P2X7 receptors are responsible for triggering inflammatory responses contributing to processes of pain in articular tissues. This study aimed to investigate whether the activation of the P2X7 receptor located in the temporomandibular joint (TMJ) tissues induces nociception through an inflammatory mechanisms and/or the activation of C-fibres (small-diameter primary afferents) of rats' TMJ. The TMJ hypernociception induced by the activation of P2X7 receptor was assessed by measuring the behavioural nociceptive responses. After behavioural experiments, the animals were terminally anaesthetized and periarticular tissues were removed and homogenate for enzyme-linked immunosorbent assay, leukocyte infiltration and western blotting analysis. The nonselective P2X7 receptor agonist BzATP induced a dose-dependent TMJ nociception, which was blocked by the selective P2X7 receptor antagonist A-438079. The co-administration of the selective β2-adrenoceptor antagonist (ICI-118,551) and the pre-treatment with cyclooxygenase inhibitor indomethacin or with the nonspecific selectin inhibitor Fucoidan significantly reduced BzATP-induced TMJ nociception. BzATP also induced an increase of pro-inflammatory cytokines TNFα, IL-1β and CINC-1 levels, as well as leukocyte recruitment in TMJ tissue, effects that were reduced by A-438079. Moreover BzATP-induced TMJ nociception was inhibited in rats neonatal-treated with Capsaicin (depleting C-fibers). Finally, BzATP-induced an increase in TRPV1 expression in TMJ tissue. These findings suggest that P2X7 receptor activation in TMJ of rats induces nociceptive responses mediated by sympathomimetic amines, prostaglandins, leukocyte migration and increased levels of pro-inflammatory cytokines. Furthermore, the P2X7 receptor activation induces nociceptive responses dependent on the activation of the primary afferent nociceptors of rats' TMJ. The activation of P2X7 receptors has an essential role in TMJ nociception and could be an interesting target to control the inflammatory pain in temporomandibular disorders.

Progesterone Attenuates Allodynia of Inflamed Temporomandibular Joint through Modulating Voltage-Gated Sodium Channel 1.7 in Trigeminal Ganglion.

Background Women with temporomandibular disorders (TMDs) experience some amelioration of pain during pregnancy. Progesterone increases dramatically and steadily during pregnancy. Sodium channel 1.7 (Nav1.7) plays a prominent role in pain perceptions, as evidenced by deletion of Nav1.7 alone leading to a complete loss of pain. In a previous study, we showed that Nav1.7 in trigeminal ganglion (TG) is involved in allodynia of inflamed temporomandibular joint (TMJ). Whether progesterone modulates allodynia of inflamed TMJ through Nav1.7 in TG remains to be investigated. Methods The effects of progesterone on sodium currents of freshly isolated TG neurons were examined using whole-cell recording. Female rats were ovariectomized and treated with increasing doses of progesterone for 10 days. Complete Freund's adjuvant was administered intra-articularly to induce TMJ inflammation. TMJ nociceptive responses were evaluated by head withdrawal thresholds. Real-time PCR and Western blotting were used to examine Nav1.7 mRNA and protein expression in TG. Immunohistofluorescence was used to examine the colocalization of progesterone receptors (PRα/β) and Nav1.7 in TG. Results Whole-cell recording showed that progesterone could attenuate sodium currents. Moreover, progesterone dose-dependently downregulated Nav1.7 mRNA expression and reduced the sensitivity of TMJ nociception in ovariectomized rats. Furthermore, treatment with progesterone attenuated allodynia of inflamed TMJ in a dose-dependent manner and repressed inflammation-induced Nav1.7 mRNA and protein expression in ovariectomized rats. The progesterone receptor antagonist, RU-486, partially reversed the effect of progesterone on allodynia of inflamed TMJ and TMJ inflammation-induced Nav1.7 mRNA and protein expression. Conclusion Progesterone, by modulating trigeminal ganglionic Nav1.7, may represent a promising agent to prevent allodynia of inflamed TMJ.

Improved efficacy of naproxen-loaded NLC for temporomandibular joint administration

Inflammatory conditions of the temporomandibular joint (TMJ) and peripheral tissues affect many people around the world and are commonly treated with non-steroidal anti-inflammatory drugs (NSAIDs). However, in order to get desirable results, treatments with NSAIDs may take weeks, causing undesirable side effects and requiring repeated administration. In this sense, this work describes the development of an optimized nanostructured lipid carrier (NLC) formulation for intra-articular administration of naproxen (NPX). An experimental design (23) selected the best formulation in terms of its physicochemical and structural properties, elucidated by different methods (DLS, NTA, TEM, DSC, and ATR-FTIR). The chosen formulation (NLC-NPX) was tested on acute inflammatory TMJ nociception, in a rat model. The optimized excipients composition provided higher NPX encapsulation efficiency (99.8%) and the nanoparticles were found stable during 1 year of storage at 25 °C. In vivo results demonstrated that the sustained delivery of NPX directly in the TMJ significantly reduced leukocytes migration and levels of pro-inflammatory cytokines (IL-1β and TNF-α), for more than a week. These results point out the NLC-NPX formulation as a promising candidate for the safe treatment of inflammatory pain conditions of TMJ or other joints.

Sex Differences in Estradiol Secretion by Trigeminal Brainstem Neurons.

Estrogen status is a significant risk factor in the development of temporomandibular joint disorders (TMD). Classically, estrogen status is thought to derive mainly from ovarian sources; however, it is well known that estradiol (E2) also is synthesized by neurons in the brain. This study tested the hypothesis that E2 is produced by neurons in trigeminal subnucleus caudalis (Vc), the principal site of termination for sensory afferents that supply the temporomandibular joint (TMJ), to modify evoked responses in a model of TMJ nociception in male and female rats. Intra-TMJ injection of the small fiber excitant, allyl isothiocyanate (AIC), increased the levels of E2 collected from microdialysis probes sites at Vc of ovariectomized (OvX) female rats, ipsilateral to the stimulus, whereas males displayed no change. Dialysate levels of E2 collected from probe sites in the contralateral Vc or cerebellum in OvX rats were not affected by TMJ stimulation. Reverse dialysis of anastrozole, an aromatase (ARO) inhibitor, via the probe reduced perfusate levels of E2 in Vc. Systemic administration of letrozole, a non-steroid ARO inhibitor, for 4 days prevented TMJ-evoked increases in masseter muscle electromyography (MMemg) activity. ARO-positive neurons were distributed mainly in superficial laminae (I-III) at Vc and cell counts revealed no significant difference between OvX and male rats. Intra-TMJ injection of AIC revealed similar numbers of ARO/Fos dual-labeled neurons in OvX and male rats. By contrast, the percentage of ARO neurons co-labeled for glutamic acid decarboxylase (GAD), the biosynthetic enzyme for GABA, was greater in OvX (35%) than male rats (14%). Few ARO-positive neurons were co-labeled for estrogen receptor alpha. These data indicate that E2 is secreted continuously by Vc neurons and that acute stimulation of TMJ nociceptors evokes further secretion in a sex-dependent manner. Reduced TMJ-evoked MMemg activity after ARO inhibition suggests that locally produced E2 by Vc neurons acts via paracrine mechanisms to modify TMJ nociception in female rats.

Distance-based permutation of inter-meal differences as a sensitive test of temporomandibular joint nociception in rats.

Compare non-parametric permutation method using intr-meal rate as endpoint to existing ANOVA method that uses average daily meal duration as an endpoint for detection of chronic pain in Sprague-Dawley rats. Nociception following bilateral temporomandibular joint (TMJ) injection of high-dose of Complete Freunds Adjuvant (CFA, 250 μg/50 μL per side) could be detected in young adult male Sprague-Dawley rats using average daily meal durations as a measure of nociception for up to 19 days (Kramer, Kerins, Schneiderman, & Bellinger, 2010) using ANOVA and multiple comparisons range tests. In this study, we reanalyzed the data using a non-parametric permutation procedure based on absolute differences between intra-meal feeding rate curves. In addition, to that experiment, we injected bilaterally the TMJ of naive rats with either a low-dose CFA (15 μg/50 μL per side, n=6) or saline (50 μL of 0.9%, n=4) and monitored the animals for 7 days. The permutation test of the intra-meal feeding rate detected the presence of nociception in the high-dose CFA treatment group for up to 40 days or twice as long as when using ANOVA on average daily meal durations. The permutation method also detected the low-dose CFA induced nociception with ten-times lower p-values and for several days longer than ANOVA of changes in meal durations. CFA-induced injury resulted in even reduction of intra-meal feeding rate and lengthening of the meals in both high- and low-dose CFA-injected animals. The rate analysis also showed when the rats first started a meal they were experiencing the same level of nociception as at the end of the meal. This demonstrated that intra-meal chewing itself did not alter the level of nociception. These results suggest that permutation tests based on differences in intra-meal feeding rates can be used as a sensitive test to determine and study the temporal patterns of TMJ nociception.

Role of central opioid on the antinociceptive effect of sulfated polysaccharide from the red seaweed Solieria filiformis in induced temporomandibular joint pain

This study aimed to investigate the effect of sulfated polysaccharide from red seaweed Solieria filiformis (Fraction F II) in the inflammatory hypernociception in the temporomandibular joint (TMJ) of rats. Male Wistar rats were pretreated (30min) with a subcutaneous injection (s.c.) of vehicle or FII (0.03, 0.3 or 3.0mg/kg) followed by intra-TMJ injection of 1.5% Formalin or 5-hydroxytryptamine (5-HT, 225μg/TMJ). In other set of experiments rats were pretreated (15min) with an intrathecal injection of the non-selective opioid receptors Naloxone, or μ-opioid receptor antagonist CTOP, or δ-opioid receptor Naltridole hydrochloride, or κ-opioid receptor antagonist Nor-Binaltorphimine (Nor-BNI) followed by injection of FII (s.c.). After 30min, the animals were treated with an intra-TMJ injection of 1.5% formalin. After TMJ treatment, behavioral nociception response was evaluated for a 45-min observation period, animals were terminally anesthetized and periarticular tissue, trigeminal ganglion and subnucleus caudalis (SC) were collected plasma extravasation and ELISA analysis. Pretreatment with F II significantly reduced formalin- and serotonin-induced TMJ nociception, inhibit the plasma extravasation and inflammatory cytokines release induced by 1.5% formalin in the TMJ. Pretreatment with intrathecal injection of Naloxone, CTOP, Naltridole or Nor-BNI blocked the antinociceptive effect of F II in the 1.5% formalin-induced TMJ nociception. In addition, F II was able to significantly increase the β-endorphin release in the subnucleus caudalis. The results suggest that F II has a potential antinociceptive and anti-inflammatory effect in the TMJ mediated by activation of opioid receptors in the subnucleus caudalis and inhibition of the release of inflammatory mediators in the periarticular tissue.

Activational action of testosterone on androgen receptors protects males preventing temporomandibular joint pain

BackgroundTestosterone protects male rats from Temporomandibular Joint (TMJ) pain. This study investigated whether this protective effect is mediated by an organizational action of testosterone during nervous system development, by central estrogen and androgen receptors and by the 5α-reduced metabolite of testosterone, dihydrotestosterone. MethodsA pharmacological approach was used to assess the ability of the androgen receptor antagonist flutamide, the estrogen receptor antagonist ICI 182 780 and the 5-α reductase inhibitor dutasteride to block the protective effect of testosterone, evaluated through the behavioral response induced by a TMJ injection of 0.5% formalin. Flutamide and ICI 182 780 were injected into the medullary subarachnoid space, and dutasteride and testosterone were systemically administered. ResultsThe TMJ injection of 0.5% formalin induced a significant nociceptive behavioral response in gonadectomized male and naïve female, but not in sham gonadectomized male rats, confirming that endogenous testosterone prevents TMJ nociception in males. Testosterone administration prevented formalin-induced TMJ nociception in males gonadectomized either in the neonatal (at the day of birth) or adult period and in naïve female rats, suggesting that the protective effect of testosterone on TMJ nociception does not depend on its organizational actions during critical periods of development. The administration of flutamide and dutasteride but not of ICI 182 780 blocked the protective effect of testosterone. ConclusionsWe conclude that the protective effect of testosterone on TMJ nociception depends on activational actions of dihydrotestosterone on androgen receptors rather than on organizational androgenic actions during central nervous system development or estrogenic actions.

Coactivation of μ- and κ-Opioid Receptors May Mediate the Protective Effect of Testosterone on the Development of Temporomandibular Joint Nociception in Male Rats.

To investigate whether the protective effect of testosterone on the development of temporomandibular joint (TMJ) nociception in male rats is mediated by the activation of central opioid mechanisms. Experiments were performed on 156 male Wistar rats. A pharmacologic approach was used to assess the ability of opioid receptor antagonists infused into the dorsal portion of the brainstem and adjacent to the caudal component (subnucleus caudalis) of the spinal trigeminal nucleus to block the protective effect of testosterone in male rats. The TMJ injection of 0.5% formalin was used as a nociceptive stimulus. One-way or two-way ANOVA was used for data analyses. The injection of 0.5% formalin into the TMJ induced a significant nociceptive behavior in gonadectomized male rats (P < .05), but not in naïve, sham, and testosterone-replaced gonadectomized rats, confirming that testosterone prevents the development of TMJ nociception. The injection of either the nonselective opioid receptor antagonist naloxone (15 μg) or the simultaneous injection of the μ-opioid receptor antagonist Cys2, Tyr3, Orn5, Pen7amide (CTOP, 30 μg) and the κ-opioid receptor antagonist Nor-Binaltorphimine (Nor-BNI, 90 μg) significantly increased the 0.5% formalin-induced behavioral response in sham and testosterone-replaced gonadectomized rats (P < .05) but had no effect in gonadectomized rats. However, the injection of each selective opioid receptor antagonist alone or the simultaneous injection of μ- or κ- and δ-opioid receptor antagonists had no effect. These findings indicate that the protective effect of endogenous testosterone on the development of TMJ nociception in male rats is mediated by the activation of central opioid mechanisms. Furthermore, the coactivation of central μ- and κ-opioid receptors is necessary for testosterone to protect male rats from developing TMJ nociception.

Gonadal hormones modulate the responsiveness to local β-blocker-induced antinociception in the temporomandibular joint of male and female rats.

We have previously demonstrated that blockade of β-adrenoreceptors (β-AR) located in the temporomandibular joint (TMJ) of rats suppresses formalin-induced TMJ nociceptive behaviour in both male and female rats, but female rats are more responsive. In this study, we investigated whether gonadal hormones modulate the responsiveness to local β-blocker-induced antinociception in the TMJ of rats. Co-administration of each of the selective β1 (atenolol), β2 (ICI 118.551) and β3 (SR59230A)-AR antagonists with equi-nociceptive concentrations of formalin in the TMJ of intact, gonadectomized and hormone-treated gonadectomized male and female rats. Atenolol, ICI 118.551 and SR59230A significantly reduced formalin-induced TMJ nociception in a dose response fashion in all groups tested. However, a lower dose of each β-AR antagonist was sufficient to significantly reduce nociceptive responses in gonadectomized but not in intact and testosterone-treated gonadectomized male rats. In the female groups, a lower dose of β1 -AR antagonist was sufficient to significantly reduce nociceptive responses in gonadectomized but not in intact or gonadectomized rats treated with progesterone or a high dose of oestradiol; a lower dose of β2 -AR antagonist was sufficient to significantly reduce nociceptive responses in gonadectomized but not in intact and gonadectomized rats treated with low or high dose of oestradiol. Gonadal hormones may reduce the responsiveness to local β-blocker-induced antinociception in the TMJ of male and female rats. However, their effect depends upon their plasma level, the subtype of β-AR and the dose of β-blockers used.

The combined effects of midazolam and propofol sedation on muscle power

We performed a randomised, crossover study to investigate the effects of intravenous sedation on grip strength and bite force. Twenty male volunteers received a bolus intravenous injection of midazolam (0.02 mg.kg(-1)) together with a 30-min propofol infusion designed to achieve an effect-site concentration of 1.0 μg.ml(-1). Observed variables included bispectral index, observer's assessment of alertness/sedation, correct answer rate of Stroop colour-word test, grip strength and bite force. Grip strength decreased from a median (IQR [range]) of 483 (443-517 [380-586]) N to 358 (280-405 [108-580]) N (p < 0.001) during sedation and recovered following flumazenil administration, while bite force increased from 818 (593-1026 [405-1406]) N to 1377 (1243-1585 [836-2357]) N (p < 0.001) during sedation. Although bite force gradually returned to baseline following flumazenil administration, it remained increased throughout the experimental period. We conclude that bite force increased during intravenous sedation and that this may have clinical implications.

Estrogen status and psychophysical stress modify temporomandibular joint input to medullary dorsal horn neurons in a lamina-specific manner in female rats

Estrogen status and psychological stress contribute to the expression of several chronic pain conditions including temporomandibular muscle and joint disorders (TMJD). Sensory neurons that supply the temporomandibular joint (TMJ) region terminate in laminae I and V of the spinal trigeminal nucleus (Vc/C1–2 region); however, little is known about lamina-specificity and environmental influences on the encoding properties of TMJ brainstem neurons. To test the hypothesis that Vc/C1–2 neurons integrate both interoceptive and exteroceptive signals relevant for TMJ nociception, we recorded TMJ-evoked activity in superficial and deep laminae of ovariectomized rats under high and low estradiol (E2) and stress conditions. Rats received daily injections of low (LE) or high (HE) dose E2 and were subjected to forced swim (FS) or sham swim conditioning for 3days. The results revealed marked lamina-specificity in that HE rats displayed enhanced TMJ-evoked activity in superficial, but not deep, laminae independent of stress conditioning. By contrast, FS conditioned rats displayed increased background firing and TMJ-evoked activity of neurons in deep, but not superficial, laminae independent of E2 status. FS also enhanced TMJ-evoked masseter muscle activity and suggested the importance of deep dorsal horn neurons in mediating evoked jaw muscle activity. In conclusion, E2 status and psychophysical stress play a significant role in modifying the encoding properties of TMJ-responsive medullary dorsal horn neurons with a marked lamina-specificity.

Modulation of temporomandibular joint nociception and inflammation in male rats after administering a physiological concentration of 17β‐oestradiol

Previous studies have shown 17β-estradiol will reduce temporomandibular joint (TMJ) inflammation and hypersensitivity in female rats. Although male rats contain significant amounts of oestradiol, it was unknown whether a physiological concentration of 17β-estradiol would attenuate male TMJ inflammation and nociception. Intact and castrated rats were given a physiological concentration of oestradiol to examine first, if oestradiol will affect male TMJ nociception/inflammation and, second, if administration of oestradiol would act synergistically with endogenous male hormones to attenuate TMJ nociception. The hormonally treated rats were given TMJ injections of complete Freund's adjuvant (CFA) and then nociception was measured using a validated method in which a lengthening in meal duration is directly correlated to the intensity of deep TMJ nociception. Inflammation was assayed by quantitating pro-inflammatory gene expression. Meal duration was significantly lengthened after TMJ CFA injection and this lengthening was significantly attenuated in the castrated but not intact males after administering a physiological concentration of oestradiol. A physiological concentration of 17β-estradiol also significantly increased IL-6 expression in the inflamed TMJ of castrated males while 17β-estradiol did not alter IL-1β, CXCL2 and CCL20 expression. Castration increased pro-inflammatory mediators IL-6, IL-1β and CXCL2 suggesting male sex hormones were anti-inflammatory. Calcitonin gene-related peptide in the trigeminal ganglia was unchanged. Similar to females, male rats with TMJ inflammation showed a reduced nociceptive response after treatment with a physiological concentration of oestradiol suggesting the effects of oestradiol treatment were not constrained by organizational processes in the males.

5-HT induces temporomandibular joint nociception in rats through the local release of inflammatory mediators and activation of local β adrenoceptors

The 5-hydroxytryptamine (serotonin, 5-HT) is an important inflammatory mediator found in high levels in the synovial fluid of the temporomandibular joint (TMJ) of patients with inflammatory pain. In this study, we used the nociceptive behavior responses, measured as flinching the head and rubbing the orofacial region, as a nociceptive assay. We demonstrated that the local blockade of the 5-HT3 receptor and β1 or β2-adrenoceptors, the depletion of norepinephrine in the sympathetic terminals and the local inhibition of cyclooxygenase significantly reduced 5-HT-induced TMJ nociception. These results demonstrated that 5-HT induces nociception in the TMJ region by the activation of β1 and β2 adrenoceptors located in the TMJ region and local release of sympathetic amines and prostaglandins. Therefore, the high levels of 5-HT in the synovial fluid of patients with TMJ inflammatory pain may contribute to TMJ pain by similar mechanisms.

Effect of gonadal steroid hormones on formalin‐induced temporomandibular joint inflammation

We have recently demonstrated that gonadal steroid hormones decrease formalin-induced temporomandibular joint nociception in rats. Given that the attenuation of inflammation is a potential mechanism underlying this antinociceptive effect, we evaluated the effect of gonadal steroid hormones on formalin-induced temporomandibular joint inflammation. Plasma extravasation, a major sign of acute inflammation, and neutrophil migration, an important event related to tissue injury, were evaluated. Formalin induced significantly lower temporomandibular joint plasma extravasation and neutrophil migration in proestrus females than in males and in diestrus females. Since estradiol serum level is high in proestrus females and low in diestrus females and in males, these findings suggest that the high physiological level of estradiol decreases temporomandibular joint inflammation. Estradiol but not progesterone administration in ovariectomized females significantly decreased formalin-induced plasma extravasation and neutrophil migration, an effect that was blocked by the estrogen receptor antagonist ICI 182780. Plasma extravasation and neutrophil migration were not affected by orchiectomy, but testosterone or estradiol administration in orchidectomized males significantly decreased them. The androgen receptor antagonist flutamide blocked the anti-inflammatory effect of testosterone while ICI 182780 blocked that of estradiol in males. Previous intravenous administration of a nonspecific selectin inhibitor significantly decreased formalin-induced temporomandibular joint nociception and neutrophil migration in males, revealing a potent and positive correlation between temporomandibular joint nociception and inflammation. Taken together, these findings demonstrate a pronounced anti-inflammatory effect of estradiol and testosterone in the temporomandibular joint region and suggest that this effect may mediate, at least in part, the antinociceptive effect of these hormones.

Differential ascending projections of temporomandibular joint-responsive brainstem neurons to periaqueductal gray and posterior thalamus of male and female rats

Several craniofacial pain conditions, including temporomandibular joint disorders (TMJDs), are more prevalent in women than men. The basis for sex differences in deep craniofacial pain is not known. The present study compared the magnitude of ascending projections from temporomandibular joint (TMJ)-responsive neurons in trigeminal brainstem with the ventrolateral periaqueductal gray (vlPAG) or posterior nucleus of the thalamus (Po) in males and female rats. Fluorogold (FG) was injected into vlPAG or Po, and TMJ-responsive neurons were identified by Fos-like immunoreactivity (Fos-LI) after mustard oil injection. TMJ-evoked Fos-LI was similar in males and females; however, significant differences in cell counts were seen for FG single-labeled and Fos/FG double-labeled neurons in trigeminal brainstem. After vlPAG injections, the number of FG-labeled neurons in trigeminal subnucleus interpolaris (Vi), ventral interpolaris/caudalis transition (vl-Vi/Vc), and dorsal paratrigeminal region (dPa5) was greater in females than males. The percentage of Fos/FG double-labeled neurons in vl-Vi/Vc and dPa5 after vlPAG injection also was greater in females than males. In contrast, after Po injections, males displayed a greater number of FG-labeled neurons in superficial laminae (Lam I/II) of trigeminal subnucleus caudalis (Vc) and upper cervical spinal cord (C1–2) and deeper laminae (Lam III/V) at C1–2 than females. The percentage of Fos/FG double-labeled neurons in Lam I/II of Vc after Po injection also was greater in males than females. These data revealed significant sex differences in ascending projections from TMJ-responsive neurons in trigeminal brainstem. Such differences may influence the ability of males and females to recruit autonomic reflexes and endogenous pain control circuits relevant for TMJ nociception.