Splenius muscle activities induced by temporomandibular joint stimulation in rats

Recent studies show that temporomandibular joint disorders cause hyperalgesia and deficits in the postural control of cervical region. However, the effects of specific modalities of receptors in the temporomandibular joint area on these phenomena are still unclear. In this study, we investigated the neck muscle activities while natural mechanical stimulation was applied to the temporomandibular joint. Single motor unit activities were recorded bilaterally from the splenius muscles in 22 Wistar rats. Mechanical stimulation applied to the left temporomandibular joint elicited tonic discharges in the left or right splenius muscle. The mean threshold values for mechanical stimulation were 48.1 (+/-16.2 S.E.M.) and 54.1 mN (+/-16.3 S.E.M.) for left and right sides, respectively. It is suggested that the temporomandibular joint mechanoreceptors not only affect the motor unit activities of neck muscles, but also are concerned in the regulation of postural control of the head.     

Unilateral application of an inflammatory irritant to the rat temporomandibular joint region produces bilateral modulation of the jaw-opening reflex

The aim of this study was to determine the effect of unilateral acute inflammation of craniofacial deep tissues on the ipsilateral and contralateral jaw-opening reflex (JOR). The effects of mustard oil (MO), injected into the temporomandibular joint region, were tested on the JOR recorded in the digastric muscle and evoked by low-intensity electrical stimulation of the ipsilateral and contralateral inferior alveolar nerve in anesthetized rats. The MO injection induced a long-lasting suppression of the amplitude of both ipsilaterally and contralaterally evoked JOR, although the latency and duration of the JOR were unaffected. The suppressive effect was more prominent for the contralaterally evoked JOR, and observed even when background activity in the digastric muscle was increased by the MO injection. The results indicate that changes in the JOR amplitude following MO injection do not simply reflect alterations in motoneuronal excitability, and suggest that inflammation of deep craniofacial tissues modulates low-threshold sensory transmission to the motoneurons.     

Influence of sex on reflex jaw muscle activity evoked from the rat temporomandibular joint

Injection of glutamate into the rat temporomandibular joint (TMJ) evoked a concentration-dependent increase in jaw muscle activity. We investigated whether there are sex-related differences in glutamate-evoked jaw muscle activity that are mediated by sex hormones and whether prior injection of glutamate into the TMJ alters the magnitude of jaw muscle activity evoked by a subsequent injection of the algesic and inflammatory compound mustard oil (MO) into the TMJ. The magnitude of glutamate-evoked digastric and masseter muscle activity was significantly greater in female than male rats when 1000 mM glutamate was injected into the TMJ. Gonadectomy significantly reduced the magnitude of glutamate-evoked digastric muscle activity in female rats. Treatment of gonadectomized female rats with estrogen (20 microg/day) increased the magnitude of glutamate-evoked digastric muscle activity. Glutamate-evoked jaw muscle activity in gonadectomized and estrogen-treated gonadectomized males was not significantly different from intact males. Prior injection of glutamate over a concentration range of 10-1000 mM significantly increased digastric muscle activity evoked by MO injection into the TMJ 30 min later. In contrast, MO-evoked masseter muscle activity was significantly increased by prior injection of 250 mM glutamate only. There were, however, no sex-related differences in the enhancement of MO-evoked jaw muscle activity by prior injection of glutamate. These findings indicate that there are sex-related differences in glutamate-evoked jaw muscle activity that are dependent on female sex hormones, and increased glutamate concentrations sensitize the TMJ to noxious chemical stimuli.     

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.     

Peptidergic nociceptive axon visualization in whole-mount preparations of cornea and tympanic membrane in rat

A whole-mount preparation is described for visualizing unmyelinated peptide immunoreactive axons in two specialized tissues, thin enough for tracing axons to their terminal knobs. These tissues, the cornea and the mucosal surface of the tympanic membrane, are known to possess specialized nociceptor innervation and can be used for selecting sensory endings labeled by peroxidase reaction product and embedded in plastic for more detailed analysis. Rat calcitonin gene-related peptide antibody proved a particularly robust primary antibody found in a substantial proportion of sensory unmyelinated axons in a variety of fixation and preparative conditions.     

An intrinsic neuronal-like network in the rat pineal gland

Recent studies have shown that in rat pineal glands kept in vitro action potential-producing cell clusters are demonstrable. To test whether the clusters interact, multiple-unit recordings were carried out simultaneously from different clusters, with or without electrical stimulation. Clusters with rhythmic burst activity exhibit highly synchronized firing and electrical stimulation of one cluster elicits an immediate response in another one, apparently involving synapses but not gap junctions. It is hypothesized that the interacting clusters form a network. As the firing is affected by norepinephrine, acetylcholine and Ca2+, the network may monitor the interstitial concentrations of these substances and spread this information in the gland, to modulate melatonin secretion.     

Distribution of thalamic nociceptive neurons activated from the tail of the rat

The purpose of the present study was to map systematically in the thalamus the distribution of neurons processing nociceptive information from the tail of the rat. Pentobarbital-anesthetized and gallamine-paralyzed rats were used. Glass microelectrodes were used to record extracellularly from thalamic neurons. Noxious radiant heat stimuli were applied to the tail with a tail-flick apparatus, and the recorded neurons were localized with horseradish peroxidase deposits or by marking electrodes left in situ. A number of 121 neurons were tested of which 45 responded. Of these, 13 were located in the ventrobasal complex (VB), 17 were located in the central lateral nucleus and the parafascicular nucleus of the intralaminar nuclei (ILN). The rest of the responding neurons were located in the posterior group, the reticular thalamic nucleus, and the zona incerta. The nucleus submedius was not examined specifically. It is concluded that the VB and the ILN are two of the most important thalamic nuclei for processing nociceptive information from the tail of the rat.     

Resveratrol alleviates temporomandibular joint inflammatory pain by recovering disturbed gut microbiota

Patients with temporomandibular disorders (TMDs) often experience persistent facial pain. However, the treatment of TMD pain is still inadequate. In recent years, the disturbance of gut microbiota has been shown to play an important role in the pathogenesis of different neurological diseases including chronic pain. In the present study, we investigated the involvement of gut microbiota in the development of temporomandibular joint (TMJ) inflammation. Intra-temporomandibular joint injection of complete Freund's adjuvant (CFA) was employed to induce TMJ inflammation. Resveratrol (RSV), a natural bioactive compound with anti-inflammatory property, was used to treat the CFA-induced TMJ inflammation. We observed that CFA injection not only induces persistent joint pain, but also causes the reduction of short-chain fatty acids (SCFAs, including acetic acid, propionic acid and butyric acid) in the gut as well as decreases relevant gut bacteria Bacteroidetes and Lachnospiraceae. Interestingly, systemic administration of RSV (i.p.) dose-dependently inhibits CFA-induced TMJ inflammation, reverses CFA-caused reduction of SCFAs and these gut bacteria. Moreover, CFA injection causes blood–brain barrier (BBB) leakage, activates microglia and enhances tumor necrosis factor alpha (TNFα) release in the spinal trigeminal nucleus caudalis (Sp5C). The RSV treatment restores the BBB integrity, inhibits microglial activation and decreases the release of TNFα in the Sp5C. Furthermore, fecal microbiota transplantation with feces from RSV-treated mice significantly diminishes the CFA-induced TMJ inflammation. Taken together, our results suggest that gut microbiome perturbation is critical for the development of TMJ inflammation and that recovering gut microbiome to normal levels could be a new therapeutic approach for treating such pain.     

Intrathecal administration of 5-HT(3) receptor agonist modulates jaw muscle activity evoked by injection of mustard oil into the temporomandibular joint in the rat

The effect of intrathecal administration of the 5-HT(3) receptor agonist 2-methyl-5-hydroxytryptamine (2m-5HT) on jaw muscle activity evoked by mustard oil (MO) injection into the temporomandibular joint of anesthetized rats was examined. One microgram or 100 microg of 2m-5HT significantly enhanced or suppressed jaw muscle responses, respectively. Pre-administration of tropisetron, a 5-HT(3) receptor antagonist, attenuated the effect of 2m-5HT. These results indicate that activation of 5-HT(3) receptors can modulate trigeminal nociceptive responses.     

Modulation of paratrigeminal nociceptive neurons following temporomandibular joint inflammation in rats

To evaluate the involvement of paratrigeminal nucleus (Pa5) nociceptive neurons in temporomandibular joint (TMJ) inflammation-induced pain and its autonomic correlates, we conducted behavioral, single unit recording and Fos immunohistochemical studies in anesthetized rats. Nocifensive behaviors to mechanical, heat or cold stimulation of the lateral face over the TMJ region were significantly enhanced in the TMJ-inflamed rats for 10–14 days after injection of complete Freund's adjuvant (CFA) into the TMJ and gradually decreased at the end of the 14-day observation period. Lowering of the nocifensive threshold in TMJ-inflamed rats lasted longer in vagus nerve-transected rats than vagus nerve-intact rats. A large number of Fos-like immunoreactive (LI) cells were observed in the Pa5, and half of them were retrogradely labeled with Fluorogold (FG) injected into the parabrachial nucleus. Background activity of Pa5 wide dynamic range and nociceptive specific neurons was significantly higher in the TMJ-inflamed rats when compared with controls. Responses to mechanical stimuli were significantly higher in NS neurons in the TMJ-inflamed rats. All thermal responsive Pa5 neurons were exclusively sensitive to cold and the response to cold was significantly higher in the TMJ-inflamed rats compared with control rats. Vagus nerve stimulation significantly decreased responses to mechanical and cold stimuli as well as the background activity in TMJ-treated rats but not in TMJ-untreated rats. The present findings suggest that populations of Pa5 neurons are nociceptive and involved in TMJ inflammation-induced pain as well as in autonomic processes related to TMJ pain.     

Unmyelinated nociceptive units in two skin areas of the rat

Responses of unmyelinated afferent fibres were investigated in two skin nerves of Sprague-Dawley rats. The units were tested as to whether they responded to mechanical probing of the skin, to controlled radiant heat stimulation, and/or to cooling of the skin(to 5°C). Ninety-six units in a n. saphenus and 129 units in a n. coccygealis werre studied, which were identified as afferents by means of the above-mentioned stimuli. In both nerves mechano- and heat-sensitive units (CMHs) were most frequent (56% in n. saphenus and 74% in n. coccygealis). There were, however, significantly more purely mechanoceptive units (CMs) in n. saphenus than in n. coccygealis (30% vs 5%). In contrast cold-sensitive units (CCs) were more frequent in n. coccygealis. They constituted 25% of the afferent C-fibres in this nerve. When testing heat sensitivity of CMHs with ramp stimuli raising the temperature to 55°C at a rate of 0.8°C/s, heat thresholds had a wide range of between 30 and 55°C. Since CMHs with low heat thresholds had the highest discharge rates and the greatest dynamic sensitivity in the range of noxious temperatures, they most probably also had nociceptive functions. It was shown that the low heat thresholds of some CMHs were not due to sensitization by preceding heat stimuli. It is argued that low-frequency discharges ( 2Hz) observed in some nociceptive CMHs of the rat at nonnoxious temperatures are insignificant for nociception. When comparing discharges during a first ramp heat stimulus to 50°C (rise time 1°C/s) with those during a second stimulus of identical time course delivered 5–10 min later, 44% of the CMHs were sensitized, 24% were desensitized and the remainder were not clearly influeced. We did not find a significant correlation between initial heat thresholds and tendency to sensitization or desensitization.     

Effects of chronic sepsis on rat motor units: experimental study of critical illness polyneuromyopathy

Critical illness polyneuromyopathy (CIP) leads to major muscle weakness correlated with peripheral nerve and/or muscle alterations. Because sepsis seems to be the main factor, we used an experimental model of chronic sepsis in rats to study the localization of the first alterations on isolated motor units of soleus muscle. Seven days of chronic sepsis leads to a decrease in muscle force and an increase in muscle fatigability. Muscle twitch contraction time is also slower and all the motor units exhibit a slow profile in septic rats. Motor axon conduction velocity remains normal. We observed a significant increase in the latency between nerve and muscle action potentials but no modifications in the electromechanical delay. The first action of sepsis on motor units seems to be a delayed trigger of muscle action potential along with a muscle weakness but without nerve conduction impairment.     

The effect of systemic cocaine on spinal nociceptive reflex activity in the rat

In the anesthetized rat, cocaine (25 mg/kg i.p.), enhanced the frequency potentiation of nociceptively evoked polysynaptic discharges but did not affect the polysynaptic reflex discharge to single nociceptive stimuli or the habituation of this reflex to repetitive pinch stimuli. The non-nociceptive, short-latency reflex discharge was suppressed for 10-15 min after cocaine administration. The neurogenic extravasation response to antidromic cutaneous C-fiber stimulation was unaffected by cocaine. These findings suggest that systemic cocaine, in doses analgesic for the rat, does not suppress spinal nociceptive reflexes.     

Putative nociceptive modulatory neurons in the dorsolateral pontomesencephalic reticular formation

The present study was undertaken to confirm the dorsolateral pontomesencephalic reticular formation (an area that includes the nucleus cuneiformis (NCF) and nucleus parabrachialis (NPB)) as a sensitive site for stimulation-produced antinociception and to investigate the possibility that there are cells in this region that show a change in activity that can be correlated with the occurrence of a nocifensive reflex (the tail-flick withdrawal response from noxious heat (TF)). Such cells have been previously demonstrated in the rostral ventromedial medulla (RVM). Extracellular single unit recording studies were made at sites from which it was possible to inhibit the TF with currents of 10 microA or less. Cellular response to 3 TF trials at 5 min intervals, and spontaneous activity over a 10 min period, were monitored for each unit. Of the cells encountered, 17% displayed an increase (on-cells) and 4% a decrease (off-cells) in activity that preceded the TF. The remaining cells were unaffected by the TF (neutral cells). On- and off-cells were found throughout the region from which TF suppression was observed but were most concentrated in the areas of the NCF and NPB. The presence of on- and off-cells in the NCF/NPB region as well as the RVM suggests that the input to RVM from NCF/NPB may be important in descending nociceptive modulation.     

Modulation of visually evoked responses in units of the ventral lateral geniculate nucleus of the rat by somatic stimuli

Single unit activity was recorded from the ventral part of the lateral geniculate nucleus (vLGN) in rats anaesthetized with urethane. Most of the cells located laterally in the nucleus were excited by light. The studied vLGN neurones did not respond to electrical stimulation of the tail, but about half of them changed their response to light significantly when the light flash was paired with the electrical stimulation. When the tail stimulus preceded the light, the changes consisted in a pronounced facilitation of flash-evoked activity. When the electrical stimulus was applied after the flash in a forward conditioning paradigm, facilitations were less pronounced and responses of some neurones were suppressed. These results are in contrast to those of similar experiments on the dorsal LGN, neurones of which were mainly facilitated by the conditioning paradigm. Thus, light-evoked activity of ventral geniculate cells can be enhanced by arousal-related processes.     

Maturation of cultured hippocampal slices results in increased excitability in granule cells

The preparation of hippocampal slices results in loss of input neurons to dentate granule cells, which leads to the reorganization of their axons, the mossy fibers, and alters their functional properties in long-term cultures, but its temporal aspects in the immature hippocampus are not known. In this study, we have focused on the early phase of this plastic reorganization process by analyzing granule cell function with field potential and whole cell recordings during the in vitro maturation of hippocampal slices (from 1 to 17 days in vitro, prepared from 6 to 7-day-old rats), and their morphology using extracellular biocytin labelling technique. Acute slices from postnatal 14-22-day-old rats were analyzed to detect any differences in the functional properties of granule cells in these two preparations. In field potential recordings, small synaptically-evoked responses were detected at 2 days in vitro, and their amplitude increased during the culture time. Whole cell voltage clamp recordings revealed intensive spontaneous excitatory postsynaptic currents, and the susceptibility to stimulus-evoked bursting increased with culture time. In acutely prepared slices, neither synaptically-evoked responses in field potential recordings nor any bursting in whole cell recordings were detected. The excitatory activity was under the inhibitory control of gamma-aminobutyric acid type A receptor. Extracellularily applied biocytin labelled dentate granule cells, and revealed sprouting and aberrant targeting of mossy fibers in cultured slices. Our results suggest that reorganization of granule cell axons takes place during the early in vitro maturation of hippocampal slices, and contributes to their increased excitatory activity resembling that in the epileptic hippocampus. Cultured immature hippocampal slices could thus serve as an additional in vitro model to elucidate mechanisms of synaptic plasticity and cellular reactivity in response to external damage in the developing hippocampus.     

Enhanced nociceptive behaviour following conditioning injection of formalin in the perioral area of the rat

The possible existence of long-term modifications in response to a transient nociceptive conditioning stimulation was investigated in the rat in three experiments. (1) A nociceptive conditioning stimulus was delivered in the form of a s.c. formalin injection (conditioning injection) in the left upper lip. Evaluation of the nociceptive behaviour triggered by another formalin injection (testing injection) made in the controlateral right upper lip was carried out in distinct groups of rats 7, 14 or 28 days after the conditioning. An enhanced nociceptive response at days 7 and 14 and a return to the baseline at day 28 were observed. (2) A similar protocol was developed with formalin used for both conditioning and testing but an anaesthetic blockade of the infraorbital nerve was performed just before the conditioning injection to suppress the initial barrage. The change observed at day 7 was suppressed by the nerve block. (3) A conditioning nociceptive stimulus was applied either ipsilaterally to the right lower lip or to the tail. An increased nociceptive response was observed when the conditioning stimulus was applied to the same side as the test stimulus but no increase in the formalin test response was detected when the conditioning stimulus was applied to the tail. These results indicated that, after a single formalin injection in the left upper lip, a hyperexcitability developed that depended on the initial barrage, lasted for at least 2 weeks, was no longer present at 4 weeks and might rely on a segmental mechanism. The hypothesis of a central sensitization triggered by an initial barrage and maintained by an ongoing input induced from the periphery is discussed.     

Evidence for 5-HT autoreceptor-mediated,nerve impulse-independent,control of 5-HT synthesis in the rat brain

To gain further insight into the operation of 5-HT autoreceptor-mediated feedback control of 5-HT biosynthesis in serotonergic nerve terminal areas, the effect of the 5-HT_1B and the 5-HT_1A receptor agonists, TFMPP and 8-OH-DPAT, respectively, were investigated in the rat central nervous system (CNS) using in vivo and in vitro neurochemical approaches. TFMPP suppressed 5-HT synthesis (5-HTP accumulation after decarboxylase inhibition) both in vivo and in vitro. In vivo, the 5-HT synthesissuppressing effect of the drug (3.0 mg/kg, s.c.) proved resistant to either acute hemitransection or reserpine (5 mg/kg, i.p.; 90 min before) pretreatment. In vitro, in cortical, hippocampal and striatal slice preparations, TFMPP (0.1–10 μM) decreased 5-HT synthesis under basal and stimulated (30 mM K⁺) conditions, an effect which was unaltered by prior in vivo reserpine-induced 5-HT depletion but was attenuated in the presence of 5-HT_1B receptor antagonists such as methiothepin, cyanopindolol or propranolol. The 8-OH-DPAT (0.1 mg/kg, s.c.)-induced decrease of 5-HT synthesis in vivo was abolished by hemitransection but resistant to acute reserpine pretreatment; 8-OH-DPAT (10 μM) did not decrease 5-HT synthesis in vitro. In conclusion, the present study confirms the importance of 5-HT autoreceptors in the feedback control of nerve terminal 5-HT biosynthesis. Specifically, our data indicate: (1) that the reduction of rat brain 5-HT synthesis after TFMPP is mediated by 5-HT_1B autoreceptors located on the serotonergic axon terminals, and (2) that the effect is directly mediated and occurs independently of 5-HT neuronal firing and intact monoamine stores. © 1995 Wiley-Liss, Inc.     

Characteristics of nociceptors in the periodontium--an in vitro study in rats

Recent studies have indicated that nociceptors can be classified into various types according to their physiological properties. These studies have clarified that the frequency distribution of various nociceptor types is different among body sites and animal species. In the present study, we investigated the physiological properties of rat's periodontal nociceptors in an in vitro jaw-nerve preparation. Responses were recorded from functional single filaments in the inferior alveolar nerve. To determine the nociceptor type, calibrated von Frey filaments, heat, and bradykinin (BK) stimuli were used. We found five subtypes of nociceptors in the periodontal ligaments of the lower incisor: Adelta-high threshold mechanonociceptors (Adelta-HTM, n=28), Adelta-mechanoheat nociceptors (Adelta-MH, n=6), Adelta-polymodal nociceptors (Adelta-POLY, n=26), C-high threshold mechanonociceptors (C-HTM, n=3) and C-polymodal nociceptors (C-POLY, n=4). Most nociceptors were Adelta-innervated, while only a small number of C-innervated nociceptors were found. The present results suggest that periodontal nociceptors transmit mainly fast pain, and may thus play a role in rapid detection of injure-related stimuli during mastication.