Involvement of astroglial glutamate-glutamine shuttle in modulation of the jaw-opening reflex following infraorbital nerve injury.

To evaluate the mechanisms underlying orofacial motor dysfunction associated with trigeminal nerve injury, we studied the astroglial cell activation following chronic constriction injury (CCI) of the infraorbital nerve (ION) immunohistochemically, nocifensive behavior in ION-CCI rats, and the effect of the glutamine synthase (GS) blocker methionine sulfoximine (MSO) on the jaw-opening reflex (JOR), and also studied whether glutamate-glutamine shuttle mechanism is involved in orofacial motor dysfunction. GFAP-immunoreactive (IR) cells were observed in the trigeminal motor nucleus (motV) 3 and 14 days after ION-CCI, and the nocifensive behavior and JOR amplitude were also strongly enhanced at these times. The number of GS- and GFAP-IR cells was also significantly higher in ION-CCI rats on day 7. The amplitude and duration of the JOR were strongly suppressed after MSO microinjection (m.i.) into the motV compared with that before MSO administration in ION-CCI rats. After MSO administration, the JOR amplitude was strongly suppressed, and the duration of the JOR was shortened. Forty minutes after m.i. of glutamine, the JOR amplitude was gradually returned to the control level and the strongest attenuation of the suppressive effect of MSO was observed at 180 min after glutamine m.i. In addition, glutamine also attenuated the MSO effect on the JOR duration, and the JOR duration was extended and returned to the control level thereafter. The present findings suggest that astroglial glutamate-glutamine shuttle in the motV is involved in the modulation of excitability of the trigeminal motoneurons affecting the enhancement of various jaw reflexes associated with trigeminal nerve injury.

Cannabinoids facilitate the swallowing reflex elicited by the superior laryngeal nerve stimulation in rats.

Cannabinoids have been reported to be involved in affecting various biological functions through binding with cannabinoid receptors type 1 (CB1) and 2 (CB2). The present study was designed to investigate whether swallowing, an essential component of feeding behavior, is modulated after the administration of cannabinoid. The swallowing reflex evoked by the repetitive electrical stimulation of the superior laryngeal nerve in rats was recorded before and after the administration of the cannabinoid receptor agonist, WIN 55-212-2 (WIN), with or without CB1 or CB2 antagonist. The onset latency of the first swallow and the time intervals between swallows were analyzed. The onset latency and the intervals between swallows were shorter after the intravenous administration of WIN, and the strength of effect of WIN was dose-dependent. Although the intravenous administration of CB1 antagonist prior to intravenous administration of WIN blocked the effect of WIN, the administration of CB2 antagonist did not block the effect of WIN. The microinjection of the CB1 receptor antagonist directly into the nucleus tractus solitarius (NTS) prior to intravenous administration of WIN also blocked the effect of WIN. Immunofluorescence histochemistry was conducted to assess the co-localization of CB1 receptor immunoreactivity to glutamic acid decarboxylase 67 (GAD67) or glutamate in the NTS. CB1 receptor was co-localized more with GAD67 than glutamate in the NTS. These findings suggest that cannabinoids facilitate the swallowing reflex via CB1 receptors. Cannabinoids may attenuate the tonic inhibitory effect of GABA (gamma-aminobuteric acid) neurons in the central pattern generator for swallowing.

Expression of TRPV1 channels after nerve injury provides an essential delivery tool for neuropathic pain attenuation.

Increased expression of the transient receptor potential vanilloid 1 (TRPV1) channels, following nerve injury, may facilitate the entry of QX-314 into nociceptive neurons in order to achieve effective and selective pain relief. In this study we hypothesized that the level of QX-314/capsaicin (QX-CAP)--induced blockade of nocifensive behavior could be used as an indirect in-vivo measurement of functional expression of TRPV1 channels. We used the QX-CAP combination to monitor the functional expression of TRPV1 in regenerated neurons after inferior alveolar nerve (IAN) transection in rats. We evaluated the effect of this combination on pain threshold at different time points after IAN transection by analyzing the escape thresholds to mechanical stimulation of lateral mental skin. At 2 weeks after IAN transection, there was no QX-CAP mediated block of mechanical hyperalgesia, implying that there was no functional expression of TRPV1 channels. These results were confirmed immunohistochemically by staining of regenerated trigeminal ganglion (TG) neurons. This suggests that TRPV1 channel expression is an essential necessity for the QX-CAP mediated blockade. Furthermore, we show that 3 and 4 weeks after IAN transection, application of QX-CAP produced a gradual increase in escape threshold, which paralleled the increased levels of TRPV1 channels that were detected in regenerated TG neurons. Immunohistochemical analysis also revealed that non-myelinated neurons regenerated slowly compared to myelinated neurons following IAN transection. We also show that TRPV1 expression shifted towards myelinated neurons. Our findings suggest that nerve injury modulates the TRPV1 expression pattern in regenerated neurons and that the effectiveness of QX-CAP induced blockade depends on the availability of functional TRPV1 receptors in regenerated neurons. The results of this study also suggest that the QX-CAP based approach can be used as a new behavioral tool to detect dynamic changes in TRPV1 expression, in various pathological conditions.

The effect of minocycline on the masticatory movements following the inferior alveolar nerve transection in freely moving rats.

BACKGROUND: To determine the effects of inferior alveolar nerve transection (IAN-X) on masticatory movements in freely moving rats and to test if microglial cells in the trigeminal principal sensory nucleus (prV) or motor nucleus (motV) may be involved in modulation of mastication, the effects of microglial cell inhibitor minocycline (MC) on masticatory jaw movements, microglia (Iba1) immunohistochemistry and the masticatory jaw movements and related masticatory muscle EMG activities were studied in IAN-X rats. RESULTS: The number of Iba1-immunoreactive (IR) cells both in prV and motV was significantly larger in IAN-X rats compared with sham rats on day 3 after IAN-X. The intraperitoneal (i.p.) administration of MC caused a significant reduction of the number of Iba1-IR cells both in prV and motV that was evident on day 14 after IAN-X. Furthermore, a significant reduction of the number of Iba1-IR cells could be observed in motV but not in prV after microinjection (m.i.) of MC into the motV of IAN-X rats. The rats also exhibited a significant decrease in the head-withdrawal threshold on the side ipsilateral to the IAN-X compared to the threshold before IAN-X and it lasted to day 14. In addition, IAN-X markedly affected the ability to rat to carry out mastication. The number of complete masticatory sequences was significantly decreased. Furthermore, the total masticatory sequence time and food preparatory (PP) period duration was significantly elongated in compared to sham rats. Although IAN-X significantly affected the total number of chewing cycles within the RC period of a masticatory sequence, it had no effect on the duration of the chewing cycles. On the other hand, systemic administration of MC (both i.p. and m.i.) in IAN-X rats significantly improved decreased head-withdrawal threshold and the impaired masticatory jaw movements. CONCLUSIONS: The present findings reveal that the strong modulation of masticatory jaw movements occurs following microglial cell activation after IAN-X, and the modulation recovers after inhibition of the microglial cell activation by MC, suggesting that microglial cell activation in the motV as well as in the prV has a pivotal role in modulating mastication following trigeminal nerve injury associated with orofacial neuropathic pain.

Induction of the Swallowing Reflex by Electrical Stimulation of the Posterior Oropharyngeal Region in Awake Humans.

We designed an electrical stimulation system to safely and reliably evoke the swallowing reflex in awake humans, and then examined the neural control of reflex swallowing initiated by oropharyngeal stimulation. A custom-made electrode connected to a flexible stainless-steel coil spring tube was introduced into the pharyngeal region through the nasal cavity and placed against the posterior wall of the oropharynx. Surface electrodes placed over the suprahyoid muscles recorded the electromyogram during swallowing. Swallowing reflexes were induced several times by 30 s of repetitive electrical pulse stimulation (intensity: 0.2-1.2 mA, frequency: 10-70 Hz, pulse duration: 1.0 ms). The onset latency of the swallowing reflex was measured over the 10-70 Hz frequency range. In addition, the two time intervals between the first three swallows were measured. The onset latency of the swallowing reflex became shorter as the stimulus frequency increased up to ≤30 Hz. Once the frequency exceeded 30 Hz, there was no further reduction in the latency. This finding was consistent with those of previous studies in anesthetized animals. The time intervals between successive swallowing reflexes did not change with increased stimulus frequencies. Furthermore, prolonged stimulation often failed to elicit multiple swallowing reflexes. The frequency dependence of onset latency suggests that temporal summation of pharyngeal afferents is required to activate the medullary swallowing center. This reliable stimulation method may help in rehabilitation of dysphagic patients without causing aspiration.

Neural mechanisms of swallowing and effects of taste and other stimuli on swallow initiation.

Swallowing involves several motor processes such as bolus formation and intraoral transport of a food bolus (oral stage) and a series of visceral events that occur in a relatively fixed timed sequence but are to some degree modifiable (pharyngeal stage or swallow reflex). Reflecting the progressive aging of society, patients with swallowing disorders (i.e., dysphagia) are increasing. Therefore, there is expanding social demand for the development of better rehabilitation treatment of dysphagic patients. To date, many dysphagia diets have been developed and are available commercially to help bring back the pleasure of mealtimes to dysphagia patients. Texture modification of food to make the food bolus easier to swallow with less risk of aspiration is one of the important elements in dysphagia diets from the viewpoint of safety assurance. However, for the further development of dysphagia diets, new attempts based on new concepts are needed. One of the possible approaches is to develop dysphagia diets that facilitate swallow initiation. For this approach, an understanding of the mechanisms of swallow initiation and identification of factors that facilitate or suppress swallow initiation are important. In this review, we first summarize the neural mechanisms of swallowing and effects of taste and other inputs on swallow initiation based on data mainly obtained from experimental animals. Then we introduce a recently established technique for eliciting swallowing using electrical stimulation in humans and our ongoing studies using this technique.

Modulation of spindle discharge from jaw-closing muscles during chewing foods of different hardness in awake rabbits.

The relationships between jaw-closing muscle spindle unit discharge and the hardness of foods were evaluated during chewing in awake rabbits. Spindle unit discharges recorded from the left mesencephalic trigeminal nucleus were correlated with the simultaneous recording of jaw movements and electromyographic (EMG) activities of the left masseter (jaw-closing) muscle during chewing soft and hard foods. A chewing cycle was divided into the fast-closing (FC), slow-closing (SC) and opening (OP) phases according to jaw movements. The chewing was classified as ipsilateral and contralateral chewing according to ipsilateral and contralateral to the recording side of the neuron, respectively. Spindle unit discharge was significantly higher during the FC and SC phases of the hard food than the soft food during both ipsilateral and contralateral chewing. The discharge was observed to be higher when the masseter muscle activity was higher. A comparison between the chewing sides reveals that the discharge was significantly higher during the slow-closing phase of ipsilateral chewing than contralateral chewing. From the above findings, the relationship of the spindle unit discharge with the hardness of foods was observed. Moreover, this relationship exists even when an animal chews food on the contralateral side suggesting the significance of the muscle spindle information for smooth chewing. In addition, the phase dependent difference of the spindle unit discharge between chewing sides suggests the distinct roles of the spindle information on the chewing and non-chewing sides.