Perspectives on next steps in classification of oro-facial pain - Part 3: biomarkers of chronic oro-facial pain - from research to clinic.

The purpose of this study was to review the current status of biomarkers used in oro-facial pain conditions. Specifically, we critically appraise their relative strengths and weaknesses for assessing mechanisms associated with the oro-facial pain conditions and interpret that information in the light of their current value for use in diagnosis. In the third section, we explore biomarkers through the perspective of ontological realism. We discuss ontological problems of biomarkers as currently widely conceptualised and implemented. This leads to recommendations for research practice aimed to a better understanding of the potential contribution that biomarkers might make to oro-facial pain diagnosis and thereby fulfil our goal for an expanded multidimensional framework for oro-facial pain conditions that would include a third axis.

Pathophysiology of TMD pain--basic mechanisms and their implications for pharmacotherapy.

This article discusses the pathophysiology of temporomandibular disorders (TMD)-related pain and its treatment with analgesic drugs. Temporomandibular disorders are comprised of a group of conditions that result in temporomandibular joint pain (arthralgia, arthritis) and/or masticatory muscle pain (myofascial TMD). In at least some patients with TMD, a peripheral mechanism contributes to this pain. However, there is often a poor correlation between the severity of TMD-related pain complaints and evidence of definitive tissue pathology. This has led to the concept that pain in some patients with TMD may result from altered central nervous system pain processing and further that this altered pain processing may be attributable to specific genes that are heritable. Psychosocial stressors are also thought to contribute to the development of TMD-related pain, particularly masticatory muscle pain. Finally, substantially more women suffer from TMD than men. Although there are arguably multiple reasons for sex-related differences in the prevalence of TMD, one candidate for the increased occurrence of this disorder in women has been suggested to be the female sex hormone oestrogen. Analgesic drugs are an integral part of the primary treatment for TMD-related pain and dysfunction with more that 90% of treatment recommendations involving use of medications. The most commonly used agents include non-steroidal anti-inflammatory drugs, corticosteroids, muscle relaxants, anxiolytics, opiates and tricyclic antidepressants, however, evidence in support of the effectiveness of these drugs is lacking. Continued research into the pathophysiology of TMD-related pain and the effectiveness of analgesic treatments for this pain is required.

γ-Aminobutyric acid (GABA) oral rinse reduces capsaicin-induced burning mouth pain sensation: An experimental quantitative sensory testing study in healthy subjects.

BACKGROUND: In burning mouth patients, analgesia after oral administration of clonazepam may result from modulation of peripheral γ-aminobutyric acid (GABA) receptors. METHODS: The effect of oral administration of test solutions (water, 0.5 mol/L or 0.05 mol/L GABA, 1% lidocaine) was investigated for the amelioration of pain and sensitivity induced by application of capsaicin (1%, 2 min) to the tongue of thirty healthy male and female subjects in this four-session, randomized, placebo-controlled, double-blinded, cross-over study. Intra-oral quantitative sensory testing was used to assess cold (CDT), warm (WDT) and mechanical (MDT) detection thresholds as well as mechanical (MPT) and heat (HPT) pain thresholds. Capsaicin-induced pain intensity was continuously rated on a 0-10 electronic visual analogue scale (VAS). The area under the VAS curve (VASAUC) after rinsing was calculated for each solution. RESULTS: Capsaicin application on the tongue evoked burning pain with a peak of 4.8/10, and significantly increased CDT and MDT while significantly decreasing WDT, HPT, and MPT. The VASAUC was significantly smaller after oral rinse with 0.05 mol/L GABA, 0.5 mol/L GABA, and 1% lidocaine than after oral rinse with water. Rinse with 0.5 mol/L or 0.05 mol/L GABA were similarly effective in decreasing VASAUC. Rinsing with either 1% lidocaine, 0.5 mol/L or 0.05 mol/L GABA also significantly attenuated the effects of capsaicin on WDT and HPT in a treatment independent manner. There were no sex-related differences in these effects of GABA. CONCLUSIONS: Capsaicin-induced burning tongue pain and decreases in WDT and HPT can be ameliorated by rinsing the mouth with lidocaine and GABA solutions. SIGNIFICANCE: Rinsing the mouth with an oral GABA containing solution ameliorated burning pain and increased heat sensitivity produced by application of capsaicin to the tongue. This finding suggests that GABA can act as a local analgesic agent in the oral cavity.

Sex-related differences in NMDA-evoked rat masseter muscle afferent discharge result from estrogen-mediated modulation of peripheral NMDA receptor activity.

In the present study, the hypothesis that sex-related differences in glutamate-evoked rat masseter muscle afferent discharge may result from estrogen-related modulation of peripheral N-methyl-d-aspartate (NMDA) receptor activity and/or expression was tested by examining afferent fiber discharge in response to masseter injection of NMDA and the expression of NR2A/B subunits by masseter ganglion neurons in male and female rats. The results showed that injection of NMDA into the masseter muscle evoked discharges in putative mechanonociceptive afferent fibers and increased blood pressure that was concentration-dependent, however, a systemic action of NMDA appeared responsible for increased blood pressure. NMDA-evoked afferent discharge was significantly greater in female than in male rats, was positively correlated with plasma estrogen levels in females and was significantly greater in ovariectomized female rats treated with a high dose (5 mug/day) compared with a low dose (0.5 mug/day) of estrogen. Pre-treatment of high dose estrogen-treated-ovariectomized female rats with the Src tyrosine kinase inhibitor PP2 did not affect NMDA-evoked afferent discharge. NMDA-evoked afferent discharge was attenuated by the antagonists ketamine and ifenprodil, which is selective for NR2B containing NMDA receptors. Fewer masseter ganglion neurons expressed the NR2A (16%) subunit as compared with the NR2B subunit (38%), which was expressed at higher frequencies in intact female (46%) and high dose estrogen-treated ovariectomized female (60%) rats than in male (31%) rats. Taken together, these results suggest that sex-related differences in NMDA-evoked masseter afferent discharge are due, at least in part, to an estrogen-mediated increase in expression of peripheral NMDA receptors by masseter ganglion neurons in female rats.

Delta-9-tetrahydrocannabinol decreases masticatory muscle sensitization in female rats through peripheral cannabinoid receptor activation.

BACKGROUND: This study investigated whether intramuscular injection of delta-9-tetrahydrocannabinol (THC), by acting on peripheral cannabinoid (CB) receptors, could decrease nerve growth factor (NGF)-induced sensitization in female rat masseter muscle; a model which mimics the symptoms of myofascial temporomandibular disorders. METHODS: Immunohistochemistry was used to explore the peripheral expression of cannabinoid receptors in the masseter muscle while behavioural and electrophysiology experiments were employed to assess the functional effects of intramuscular injection of THC. RESULTS: It was found that CB1 and CB2 receptors are expressed by trigeminal ganglion neurons that innervate the masseter muscle and also on their peripheral endings. Their expression was greater in TRPV1-positive ganglion neurons. Three days after intramuscular injection of NGF, ganglion neuron expression of CB1 and CB2, but not TPRV1, was decreased. In behavioural experiments, intramuscular injection (10 µL) of THC (1 mg/mL) attenuated NGF-induced mechanical sensitization. No change in mechanical threshold was observed in the contralateral masseter muscles and no impairment of motor function was found after intramuscular injections of THC. In anaesthetized rats, the same concentration of THC increased the mechanical thresholds of masseter muscle mechanoreceptors. Co-administration of the CB1 antagonist AM251 blocked the effect of THC on masseter muscle mechanoreceptors while the CB2 antagonist AM630 had no effect. CONCLUSIONS: These results suggest that reduced inhibitory input from the peripheral cannabinoid system may contribute to NGF-induced local myofascial sensitization of mechanoreceptors. Peripheral application of THC may counter this effect by activating the CB1 receptors on masseter muscle mechanoreceptors to provide analgesic relief without central side effects. SIGNIFICANCE: Our results suggest THC could reduce masticatory muscle pain through activating peripheral CB1 receptors. Peripheral application of cannabinoids could be a novel approach to provide analgesic relief without central side effects.

Sensitivity of rat temporalis muscle afferent fibers to peripheral N-methyl-D-aspartate receptor activation.

The temporalis muscle is a common source of pain in headache and chronic craniofacial pain conditions such as temporomandibular disorders, which have an increased prevalence in women. The characteristics of slowly conducting temporalis afferent fibers have not been investigated. Therefore, the aim of the present study was to examine the characteristics of slowly conducting temporalis muscle afferent fibers and to determine whether these fibers are excited by activation of peripheral N-methyl-D-aspartate receptors. The response properties of a total of 117 temporalis afferent fibers were assessed in male and female rats. A majority of these fibers had high mechanical thresholds and slow conduction velocities (<10 m/s). The mechanical threshold of the temporalis afferent fibers was inversely correlated with afferent conduction velocity, however, no sex-related differences in mechanical threshold were identified. There were also no sex-related differences in N-methyl-D-aspartate-evoked afferent discharge. Indeed, injection of a high concentration (1600 mM) of N-methyl-D-aspartate into the temporalis muscle was necessary to evoke significant afferent discharge. Thirty minutes after the initial injection of N-methyl-D-aspartate into the temporalis muscle, a second injection of N-methyl-D-aspartate produced a response only about 50% as large as the initial injection. Co-injection of ketamine (20 mM) with the second injection of N-methyl-D-aspartate significantly decreased N-methyl-D-aspartate-evoked afferent discharge in both sexes. This concentration of ketamine is greater than that needed to attenuate afferent discharge evoked by injection of glutamate into the masseter muscle. These results suggest that unlike masseter afferent fibers, temporalis afferent fibers are relatively insensitive to peripheral N-methyl-D-aspartate receptor activation.

Increased pain and muscle glutamate concentration after single ingestion of monosodium glutamate by myofascial temporomandibular disorders patients.

BACKGROUND: A randomized, double-blinded, placebo-controlled study was conducted to investigate if single monosodium glutamate (MSG) administration would elevate muscle/serum glutamate concentrations and affect muscle pain sensitivity in myofascial temporomandibular disorders (TMD) patients more than in healthy individuals. METHODS: Twelve myofascial TMD patients and 12 sex- and age-matched healthy controls participated in two sessions. Participants drank MSG (150 mg/kg) or NaCl (24 mg/kg; control) diluted in 400 mL of soda. The concentration of glutamate in the masseter muscle, blood plasma and saliva was determined before and after the ingestion of MSG or control. At baseline and every 15 min after the ingestion, pain intensity was scored on a 0-10 numeric rating scale. Pressure pain threshold, pressure pain tolerance (PPTol) and autonomic parameters were measured. All participants were asked to report adverse effects after the ingestion. RESULTS: In TMD, interstitial glutamate concentration was significantly greater after the MSG ingestion when compared with healthy controls. TMD reported a mean pain intensity of 2.8/10 at baseline, which significantly increased by 40% 30 min post MSG ingestion. At baseline, TMD showed lower PPTols in the masseter and trapezius, and higher diastolic blood pressure and heart rate than healthy controls. The MSG ingestion resulted in reports of headache by half of the TMD and healthy controls, respectively. CONCLUSION: These findings suggest that myofascial TMD patients may be particularly sensitive to the effects of ingested MSG. WHAT DOES THIS STUDY ADD?': Elevation of interstitial glutamate concentration in the masseter muscle caused by monosodium glutamate (MSG) ingestion was significantly greater in myofascial myofascial temporomandibular disorders (TMD) patients than healthy individuals. This elevation of interstitial glutamate concentration in the masseter muscle significantly increased the intensity of spontaneous pain in myofascial TMD patients.

Intramuscular pH modulates glutamate-evoked masseter muscle pain magnitude in humans.

BACKGROUND: This study was conducted to determine whether glutamate-evoked jaw muscle pain is modulated by the acidity and temperature of the solution injected. METHODS: Thirty two participants participated and received injections of high-temperature acidic (HT-A) glutamate (pH 4.8, 48 °C), high-temperature neutral (HT-N) glutamate (pH 7.0, 48 °C) and neutral temperature neutral (NT-N) glutamate (pH 7.0, 38 °C) solutions (0.5 mL) into the masseter muscle. Pain intensity was assessed with an electronic visual analogue scale (eVAS). Numerical rating scale (NRS) scores of unpleasantness and temperature perception, pain-drawing areas, mechanical sensitivity and pressure pain thresholds (PPT) were also measured. Participants filled out the McGill Pain Questionnaire (MPQ). One or two way ANOVAs were used for data analyses. RESULTS: Injection of HT-A glutamate solutions significantly increased the area under the VAS-time curve compared with injection of HT-N glutamate and NT-N glutamate solution (p < 0.040). The duration of glutamate-evoked pain was significantly longer when HT-A glutamate was injected than when NT-N glutamate was injected (p < 0.017). No significant effects of acidity were detected on pain drawings, NRS unpleasantness and heat perception, but there was a significant effect of acidity on MPQ scores and mechanical sensitivity. CONCLUSION: Acidity and temperature modulate glutamate-evoked jaw muscle pain suggesting an interaction between acid sensing and glutamate receptors which could be of importance for understanding clinical muscle pain conditions.

Glutamate-induced sensitization of rat masseter muscle fibers.

In rats, intradermal or intraarticular injection of glutamate or selective excitatory amino acid receptor agonists acting at peripheral excitatory amino acid receptors can decrease the intensity of mechanical stimulation required to evoke nocifensive behaviors, an indication of hyperalgesia. Since excitatory amino acid receptors have been found on the terminal ends of cutaneous primary afferent fibers, it has been suggested that increased tissue glutamate levels may have a direct sensitizing effect on primary afferent fibers, in particular skin nociceptors. However, less is known about the effects of glutamate on deep tissue afferent fibers. In the present study, a series of experiments were undertaken to investigate the effect of intramuscular injection of glutamate on the excitability and mechanical threshold of masseter muscle afferent fibers in anesthetized rats of both sexes. Injection of 1.0 M, but not 0.1 M glutamate evoked masseter muscle afferent activity that was significantly greater than that evoked by isotonic saline. The mechanical threshold of masseter muscle afferent fibers, which was assessed with a Von Frey hair, was reduced by approximately 50% for a period of 30 min after injection of 1.0 M glutamate, but was unaffected by injections of 0.1 M glutamate or isotonic saline. Injection of 25% dextrose, which has the same osmotic strength as 1.0 M glutamate, did not evoke significant activity in or decrease the mechanical threshold of masseter muscle afferent fibers. Magnetic resonance imaging experiments confirmed that injection of 25% dextrose and 1.0 M glutamate produced similar edema volumes in the masseter muscle tissue. Co-injection of 0.1 M kynurenate, an excitatory amino acid receptor antagonist, and 1.0 M glutamate attenuated glutamate-evoked afferent activity and prevented glutamate-induced mechanical sensitization. When male and female rats were compared, no difference in the baseline mechanical threshold or in the magnitude of glutamate-induced mechanical sensitization of masseter muscle afferent fibers was observed; however, the afferent fiber activity evoked by injection of 1.0 M glutamate into the masseter muscle was greater in female rats. The results of the present experiments show that intramuscular injection of 1.0 M glutamate excites and sensitizes rat masseter muscle afferent fibers through activation of peripheral excitatory amino acid receptors and that glutamate-evoked afferent fiber activity, but not sensitization, is greater in female than male rats.

On the reduction of spontaneous and glutamate-driven spinocerebellar and spinoreticular tract neuronal activity during active sleep.

The present study was performed to provide evidence that dynamic neural processes underlie the reduction in dorsal spinocerebellar tract and spinoreticular tract neuron activity that occurs during active sleep. To ascertain the effect of local inhibition on the spontaneous and glutamate-evoked spike discharge of sensory tract neurons, preliminary control tests were performed during the state of quiet wakefulness, where GABA or glycine was co-administered in a sustained fashion during pulsatile release of glutamate to dorsal spinocerebellar tract (n=3) or spinoreticular tract (n=2) neurons. Co-administration of GABA or glycine also resulted in a significant marked suppression of spontaneous spike activity and glutamate-evoked responses of these cells. Extracellular recording experiments combined with juxtacellular application of glutamate were then performed on 20 antidromically identified dorsal spinocerebellar tract and spinoreticular tract neurons in the chronic intact cat as a function of sleep and wakefulness. The glutamate-evoked activity of a group of 10 sensory tract neurons (seven dorsal spinocerebellar tract, three spinoreticular tract), which exhibited a significant decrease in their spontaneous spike activity during active sleep, was examined. Glutamate-evoked activity in these cells was significantly attenuated during active sleep compared with wakefulness. In contrast, the glutamate-evoked activity of a second group of eight sensory tract neurons (four dorsal spinocerebellar tract, four spinoreticular tract), which exhibited a significant increase in their spontaneous spike activity during active sleep, was not significantly altered in a state-dependent manner. These data indicate that, during natural active sleep, a dynamic neural process is engaged onto certain dorsal spinocerebellar tract and spinoreticular tract neurons, which in turn dampens sensory throughput to higher brain centers.

Active sleep-related depolarization of feline trigemino-thalamic afferent terminals.

Presynaptic depolarization of trigemino-thalamic (TGT) terminals may contribute to modulation of ascending oro-facial somatosensory information during active (or rapid eye movement) sleep. The relative excitability of TGT terminals was inferred from changes in the current required to maintain an antidromic firing probability of 50% (EC50) during quiet wakefulness as compared to active sleep. Depolarization or hyperpolarization of TGT terminals was defined as a decrease or increase, respectively, in the EC50. Overall, the EC50 of 8 TGT terminals was reduced by a mean 8.8+/-3.6 microA during active sleep relative to quiet wakefulness. This result suggests that depolarization of TGT terminals, which may act to suppress the transfer of sensory information from the trigeminal nucleus to the thalamus, occurs during active sleep.

Temporomandibular-evoked jaw muscle reflex: role of brain stem NMDA and non-NMDA receptors.

This study investigated the possible involvement of brain stem excitatory amino acid receptor mechanisms and the trigeminal subnucleus caudalis (Vc) in temporomandibular joint (TMJ)-evoked reflex jaw muscle activity. Glutamate injected into the TMJ of anesthetized rats reflexly evoked activity in the jaw muscles. Application of lidocaine, but not saline, to the surface of the caudal brainstem overlying Vc significantly suppressed TMJ-evoked jaw muscle activity, while application of NMDA or non-NMDA receptor antagonists also significantly attenuated jaw muscle activity. These results provide evidence that Vc is a critical relay in the TMJ-evoked reflex activation of the jaw muscles, and that both NMDA and non-NMDA receptor mechanisms may contribute to these effects.

Sex-related suppression of reflex jaw muscle activity by peripheral morphine but not GABA.

The present study examined the effect of peripherally applied morphine and GABA on jaw muscle electromyographic activity reflexly evoked by co-injection of glutamate into the temporomandibular joint (TMJ) of lightly anesthetized Sprague-Dawley rats of both sexes. In male but not female rats, morphine significantly suppressed glutamate-evoked jaw muscle activity in a dose-dependent and naloxone-reversible manner. The median suppressive dose (+/- s.e.) for male rats was 12.7 +/- 3.1 microg (digastric muscle) and 12.6 +/- 1.3 microg (masseter muscle). GABA (5 micromol) significantly reduced glutamate-evoked muscle activity in both sexes. These data suggest that female rats are considerably less sensitive than male rats to the suppressive effects of peripherally applied morphine, but both sexes are equally affected by peripherally applied GABA.

Dorsal spinocerebellar tract neuronal activity in the intact chronic cat.

The ability to electrophysiologically identify the axonal projections of lumbar neurons recorded in chronic unanesthetized intact awake animals is a formidable but essential requirement toward understanding ascending sensory transmission under naturally occurring conditions. Chronic immobilization procedures previously introduced by Morales et al. (1981) for intracellular studies of motoneurons are modified and then integrated with procedures for antidromic cellular identification and extracellular recording of upper (or lower) dorsal lumbar spinocerebellar tract (DSCT) neuronal activity, in conjunction with behavioral state recording and drug microiontophoresis. These implant procedures provide up to 6 months of stable recording conditions and, when combined with other techniques, allow individual DSCT neurons to be monitored over multiple cycles of sleep and wakefulness, following the induction into and recovery from barbiturate anesthesia and/or during the juxtacellular microiontophoretic ejection of inhibitory or excitatory amino acid neurotransmitters. The combination of such techniques allows a comprehensive examination of synaptic transmission through the DSCT and other lumbar sensory pathways in the intact normally respiring cat and its modulation during the general anesthetic state. These techniques permit investigations of the supraspinal controls impinging on lumbar sensory tract neurons during wakefulness and other behavioral states such as active sleep.

Involvement of GABA(A) receptor in modulation of jaw muscle activity evoked by mustard oil application to the rat temporomandibular joint.

The effect of intrathecal administration of the GABA(A) receptor antagonist bicuculline methylbromide on jaw muscle electromyographic (EMG) activity evoked by mustard oil injection into the rat temporomandibular joint was studied. Bicuculline given prior to mustard oil augmented the EMG activity evoked by mustard oil, and "rekindling" of EMG activity was induced by bicuculline given 30 min after mustard oil. These results suggest that central GABA(A) receptors modulate reflex responses to noxious craniofacial stimuli.

Development of inflammation after application of mustard oil or glutamate to the rat temporomandibular joint.

Application of the small-fibre excitant and inflammatory irritant mustard oil or the excitatory amino-acid receptor agonist glutamate to the rat temporomandibular joint (TMJ) region evokes similar changes in jaw-muscle activity, suggesting that peripheral application of glutamate may be nociceptive. Application of mustard oil to the TMJ region is also inflammatory, but, it is not clear if application of glutamate is equally inflammatory. In this study the extent of plasma-protein extravasation and oedema induced by mustard oil application to the TMJ region was compared with that induced by glutamate. Application of mustard oil resulted in plasma-protein extravasation into the TMJ tissues and oedema of the TMJ region. In contrast, glutamate did not cause plasma-protein extravasation or oedema.

Effects of adrenergic agonists and antagonists on tetrodotoxin-induced nerve block.

BACKGROUND AND OBJECTIVES: The relative contributions of alpha(1)-, alpha(2)-, and beta-adrenergic receptors to adrenergic agonists' prolongation of nerve block by tetrodotoxin (TTX) are unknown. We investigated which receptor agonists prolong TTX block, and whether delayed injection of antagonists can interrupt prolonged blocks after coinjection of TTX and agonists. METHODS: Rats received percutaneous sciatic nerve block with 120 micromol/L TTX with and without adrenergic agonists and antagonists. Block duration was assessed by a modified hot-plate test. Functional deficits in the uninjected leg were used to assess systemic distribution of TTX. Data were expressed as medians with 25th and 75th percentiles. RESULTS: Coinjection of 5.5 micromol/L phenylephrine (alpha(1)-specific), 10 micromol/L clonidine (alpha(2)-specific), and 1.1 micromol/L epinephrine (mixed alpha- and beta-agonist) prolonged TTX nerve block, but 5.5 micromol/L isoproterenol (mixed beta-agonist) did not. Yohimbine inhibited TTX block prolongation by clonidine (median inhibitory concentrations, IC(50) = 130 nmol/L); phentolamine similarly inhibited epinephrine (IC(50) = 45 nmol/L). Adrenergic antagonists did not inhibit the prolongation of TTX block by agonists when injected 3 or 6 hours after the initial block. Subcutaneous injection of adrenergic agonists at a remote site did not prolong TTX block, except for a modest prolongation by clonidine. CONCLUSION: TTX block can be prolonged by alpha(1)- and alpha(2)-, but not beta-adrenergic agonists via locally mediated events of relatively brief duration. Delayed injection of adrenergic antagonists does not interrupt the prolonged blocks produced by coinjection of TTX and adrenergic agonists unless administered soon after block is established. Reg Anesth Pain Med 2001;26:239-245.

NGF-induced mechanical sensitization of the masseter muscle is mediated through peripheral NMDA receptors.

Intramuscular injection of nerve growth factor (NGF) in healthy humans mimics some of the symptoms of myofascial temporomandibular disorders (M-TMD). We hypothesized that NGF induces a prolonged myofascial mechanical sensitization by increasing peripheral N-methyl-d-aspartate (NMDA) receptor expression, leading to an enhanced response of muscle nociceptors to endogenous glutamate. Behavioral experiments with an injection of NGF (25 µg/ml, 10 µl) into the masseter muscle reduced the mechanical withdrawal threshold for 1 day in male rats and 5 days in female rats. These results mirror the sex-related differences found in NGF-induced mechanical sensitization in humans. Intramuscular injection with the competitive NMDA receptor antagonist dl-2-amino-5-phosphonovaleric acid (APV, 0.020 g/ml, 10 µl) reversed the mechanical sensitization in male but not in female rats. NGF increased the number of NMDA receptor subtype 2B (NR2B)-expressing rat trigeminal masseter ganglion neurons in both sexes, which peaked at 3 days post injection. There was an association between the levels of NR2B expression and NGF-induced mechanical sensitization. The average soma size of NR2B-expressing neurons increased significantly. Increased expression of neuropeptides (CGRP and SP) was observed in NR2B-expressing masseter ganglion neurons in female but not in male rats. In healthy men and women, comparable basal expression levels of NR2B and SP were found in peripheral fibers from masseter muscle microbiopsies. This study suggests that NGF-induced sensitization of masseter nociceptors is mediated, in part, by enhanced peripheral NMDA receptor expression. Measurement of peripheral NMDA receptor expression may be useful as a biomarker for M-TMD pain.

Glutamate dysregulation in the trigeminal ganglion: a novel mechanism for peripheral sensitization of the craniofacial region.

In the trigeminal ganglion (TG), satellite glial cells (SGCs) form a functional unit with neurons. It has been proposed that SGCs participate in regulating extracellular glutamate levels and that dysfunction of this SGC capacity can impact nociceptive transmission in craniofacial pain conditions. This study investigated whether SGCs release glutamate and whether elevation of TG glutamate concentration alters response properties of trigeminal afferent fibers. Immunohistochemistry was used to assess glutamate content and the expression of excitatory amino acid transporter (EAAT)1 and EAAT2 in TG sections. SGCs contained glutamate and expressed EAAT1 and EAAT2. Potassium chloride (10 mM) was used to evoke glutamate release from cultured rat SGCs treated with the EAAT1/2 inhibitor (3S)-3-[[3-[[4-(trifluoromethyl)ben zoyl]amino]phenyl]methoxy]-L-aspartic acid (TFB-TBOA) or control. Treatment with TFB-TBOA (1 and 10 µM) significantly reduced the glutamate concentration from 10.6 ± 1.1 to 5.8 ± 1.4 µM and 3.0 ± 0.8 µM, respectively (p<0.05). Electrophysiology experiments were conducted in anaesthetized rats to determine the effect of intraganglionic injections of glutamate on the response properties of ganglion neurons that innervated either the temporalis or masseter muscle. Intraganglionic injection of glutamate (500 mM, 3 µl) evoked afferent discharge and significantly reduced muscle afferent mechanical threshold. Glutamate-evoked discharge was attenuated bythe N-methyl-D-aspartate receptor antagonist 2-amino-5-phosphonovalerate (APV) and increased by TFB-TBOA, whereas mechanical sensitization was only sensitive to APV. Antidromic invasion of muscle afferent fibers by electrical stimulation of the caudal brainstem (10 Hz) or local anesthesia of the brainstem with lidocaine did not alter glutamate-induced mechanical sensitization. These findings provide a novel mechanism whereby dysfunctional trigeminal SGCs could contribute to cranial muscle tenderness in craniofacial pain conditions such as migraine headache.

Interactions of glutamate and capsaicin-evoked muscle pain on jaw motor functions of men.

OBJECTIVE: The aim of the study was to investigate the interaction between glutamate and capsaicin-evoked muscle pain on human jaw motor functions. METHODS: Fifteen male volunteers participated. Glutamate or capsaicin or isotonic saline, in a paired-sequence order, was injected randomly into the right or left masseter muscle. Two injections were given in a double-blinded design 25 min apart in one session/week over four weeks: isotonic saline (A1) followed by glutamate (A2), capsaicin (B1) followed by glutamate (B2), isotonic saline (C1) followed by capsaicin (C2), and glutamate (D1) followed by capsaicin (D2). The resting electromyographic (EMG) activity of the right and left masseter muscles, maximum voluntary bite force (MVBF), and maximum voluntary jaw opening (MVJO) were recorded before and after injection and subsequently at 5-min intervals for 50 min. The pain intensity was recorded on a 0-10 numerical rating scale during each MVBF or MVJO jaw function. RESULTS: Resting EMG activity was significantly increased after 5 min of D2 (ANOVA: P=0.028) injection. The percentage change (compared with baseline) in EMG activity was significantly different between D2 (116.1+/-6.1%) and C2 (102.1+/-3.4%) injections (paired t-test: P=0.039). The MVBF and MVJO were significantly decreased after injection of glutamate or capsaicin, however, there was no significant difference in the relative decrease between A2 and B2, or between C2 and D2 at any time point (P>0.152). There was a significantly higher peak pain rating after D2 compared to C2 during MVBF or MVJO (P<0.022), whereas no significant difference in peak pain ratings was found between A2 and B2 (P>0.084). There were significant negative correlations between pain ratings and MVBF or MVJO (Pearson correlation: P<0.001). CONCLUSIONS: The results indicate that intramuscular administration of glutamate and capsaicin induces muscle pain which has the potential to perturb some normal jaw motor functions. SIGNIFICANCE: The present findings suggest that peripheral glutamate and capsaicin receptor mechanisms interact to affect some jaw motor as well as sensory (i.e. pain) functions and provide new insights into the complexity of orofacial pain. Management approaches that target the peripheral nervous system and receptor mechanisms may prevent such changes in jaw motor function.