The trigeminal subnuclei interpolaris/caudalis transition zones (Vi/Vc) play an important role in orofacial deep pain, however, the role of primary afferent projections to the Vi/Vc remains to be determined. This study investigated the functional significance of hyperalgesia to the brain-derived neurotrophic factor (BDNF)-tyrosine kinase B (trkB) signaling system in trigeminal ganglion (TRG) neurons projecting to the Vi/Vc transition zone following masseter muscle (MM) inflammation.

In order to evaluate the neuronal mechanisms underlying functional abnormalities of swallowing in orofacial pain patients, this study investigated the effects of noxious orofacial stimulation on the swallowing reflex, phosphorylated extracellular signal-regulated kinase (pERK) and gamma-aminobutyric acid (GABA) immunohistochemical features in brainstem neurons, and also analysed the effects of brainstem lesioning and of microinjection of GABA receptor agonist or antagonist into the nucleus tractus solitarii (NTS) on the swallowing reflex in anaesthetized rats. The swallowing reflex elicited by topical administration of distilled water to the pharyngolaryngeal region was inhibited after capsaicin injection into the facial (whisker pad) skin or lingual muscle. The capsaicin-induced inhibitory effect on the swallowing reflex was itself depressed after the intrathecal administration of MAPK kinase (MEK) inhibitor. No change in the capsaicin-induced inhibitory effect was observed after trigeminal spinal subnucleus caudalis lesioning, but the inhibitory effect was diminished by paratrigeminal nucleus (Pa5) lesioning. Many pERK-like immunoreactive neurons in the NTS showed GABA immunoreactivity. The local microinjection of the GABA(A) receptor agonist muscimol into the NTS produced a significant reduction in swallowing reflex, and the capsaicin-induced depression of the swallowing reflex was abolished by microinjection of the GABA(A) receptor antagonist bicuculline into the NTS. The present findings suggest that facial skin-NTS, lingual muscle-NTS and lingual muscle-Pa5-NTS pathways are involved in the modulation of swallowing reflex by facial and lingual pain, respectively, and that the activation of GABAergic NTS neurons is involved in the inhibition of the swallowing reflex following noxious stimulation of facial and intraoral structures.

The aim of the present study was to determine whether acetazolamide (AZ) contributes to the inhibition of the fast inactivating transient K(+) current (I(A) ) in adult rat nodose ganglion (NG) neurons. We have previously shown that pretreatment with either AZ or 4-AP attenuated or blocked the CO(2) -induced inhibition of slowly adapting pulmonary stretch receptors in in vivo experiments. The patch-clamp experiments were performed by using the isolated NG neurons. In addition to this, the RT-PCR of mRNA and the expression of voltage-gated K(+) (Kv) 1.4, Kv 4.1, Kv 4.2, and Kv 4.3 channel proteins from nodose ganglia were examined. We used NG neurons sensitive to the 1 mM AZ application. The application of 1 mM AZ inhibited the I(A) by approximately 27% and the additional application of 4-AP (1 mM) further inhibited I(A) by 48%. The application of 0.1 μM α-dendrotoxin (α-DTX), a slow inactivating transient K(+) current (I(D) ) blocker, inhibited the baseline I(A) by approximately 27%, and the additional application of 1 mM AZ further decreased the I(A) by 51%. In current clamp experiments, AZ application (1 mM) increased the number of action potentials due to the decreased duration of the depolarizing phase of action potentials and/or due to a reduction in the resting membrane potential. Four voltage-gated K(+) channel proteins were present, and most (80-90%) of the four Kv channels immunoreactive neurons showed the co-expression of carbonic anhydrase-II (CA-II) immunoreactivity. These results indicate that the application of AZ causes the reduction in I(A) via the inhibition of four voltage-gated K(+) channel (Kv) proteins without affecting I(D).

To define the somatotopic arrangement of neurons in the trigeminal spinal subnucleus caudalis and upper cervical cord activated by acute noxious stimulation of various orofacial sites, pERK expression was analyzed in these neurons. After capsaicin injection into the tongue, lower gum, upper and lower lips, or mental region, pERK-like immunoreactive (pERK-LI) cells were distributed mainly in the dorsal half of the trigeminal spinal nucleus interporalis (Vi) and caudalis (Vc) transition zone (Vi/Vc zone), middle Vc, and Vc and upper cervical cord transition zone (Vc/C2 zone). pERK-LI cells were distributed throughout the dorsal to ventral portion of the Vi/Vc zone, middle Vc, and Vc/C2 zone following capsaicin injection into the anterior hard palate, upper gum, buccal mucosa, or vibrissal pad and in the ventral portion of the Vi/Vc zone, middle Vc, and Vc/C2 zone following snout, ophthalmic, or ocular injection of capsaicin. The rostrocaudal distribution area of pERK-LI cells was more extensive from the Vi/Vc zone to the Vc/C2 zone after intraoral injection than that after facial injection, and the rostrocaudal distribution of pERK-LI cells from the Vi/Vc zone to the Vc/C2 zone had a somatotopic arrangement, with the snout being represented most rostrally and ophthalmic, ocular, or mental regions represented most caudally. These findings suggest that the pERK-LI cells expressed from the Vi/Vc zone to the Vc/C2 zone following injection of capsaicin in facial and intraoral structures may be differentially involved in pain perception in facial and intraoral sites.

We examined the role of TRPA1s in triggering the swallowing reflex. TRPA1s predominantly localized on thin nerve fibers and fibroblast-like cells in swallowing-related regions and on small to medium-sized superior laryngeal nerve-afferents in the nodose-petrosal-jugular ganglionic complex. Topical application of a TRPA1 agonist, allyl isothiocyanate (AITC), dose-dependently triggered swallowing reflexes. Prior topical application of a TRPA1 antagonist significantly attenuated the AITC-induced reflexes. Application of cold AITC (4 °C) very briefly reduced the on-site temperature to < 17 °C (temperature at which TRPA1s can be activated), but had no effect on triggering of the reflex. By contrast, reducing the on-site temperature to < 17 °C for a longer time by continuous flow of cold AITC or by application of iced AITC paradoxically delayed/prevented the triggering of AITC-induced reflexes. Prior application of the TRPA1 antagonist had no effect on the threshold for the punctate mechanical stimuli-induced reflex or the number of low-force or high-force continuous mechanical pressure stimuli-induced reflexes. TRPA1s are functional and act as chemosensors, but not as cold sensors or mechanosensors, for triggering of the swallowing reflex. A brief cold stimulus has no effect on triggering of the reflex. However, a longer cold stimulus delays/prevents triggering of the reflex because of cold anesthesia.

The swallowing reflex is an essential physiological reflex that allows food or liquid to pass into the esophagus from the oral cavity. Delayed triggering of this reflex is a significant health problem in patients with oropharyngeal dysphagia for which no pharmacological treatments exist. Transient receptor potential channels have recently been discovered as potential targets to facilitate triggering of the swallowing reflex. However, the ability of transient receptor potential vanilloid 4 (TRPV4) to trigger the swallowing reflex has not been studied. Here, we demonstrate the involvement of TRPV4 in triggering the swallowing reflex in rats. TRPV4 immunoreactive nerve fibers were observed in the superior laryngeal nerve (SLN)-innervated swallowing-related regions. Retrograde tracing with fluorogold revealed localization of TRPV4 on approximately 25% of SLN-afferent neurons in the nodose-petrosal-jugular ganglionic complex. Among them, approximately 49% were large, 35% medium, and 15% small-sized SLN-afferent neurons. Topical application of a TRPV4 agonist (GSK1016790A) to the SLN-innervated regions dose-dependently facilitated triggering of the swallowing reflex, with the highest number of reflexes triggered at a concentration of 250 μM. The number of agonist-induced swallowing reflexes was significantly reduced by prior topical application of a TRPV4 antagonist. These findings indicate that TRPV4 is expressed on sensory nerves innervating the swallowing-related regions, and that its activation by an agonist can facilitate swallowing. TRPV4 is a potential pharmacological target for the management of oropharyngeal dysphagia.

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.

Difficulty swallowing represents a major health problem. Swallowing function is improved by incorporating weak acids in suspensions/food boluses, implicating acid-sensing ion channels (ASICs) in the swallowing reflex. However, the functional involvement of ASICs in the swallowing reflex has not been fully elucidated.

In order to evaluate mechanisms that may underlie the sensitization of trigeminal spinal subnucleus caudalis (Vc; the medullary dorsal horn) and upper cervical spinal cord (C1-C2) nociceptive neurons to heat, cold and mechanical stimuli following topical capsaicin treatment of the facial skin, nocifensive behaviors as well as phosphorylation of extracellular regulated-kinase (pERK) in Vc and C1-C2 neurons were studied in rats.

To determine whether there is a difference between a transient K(+) current (I(A)) and a sustained K(+) current (I(K)) regarding the neuronal function in small-diameter adult rat trigeminal ganglion (TG) neurons, which were insensitive to tetrodotoxin (TTX, 1 microM), we performed two different types of experiments. Primary cultures of dissociated TG neurons were prepared, and electrophysiological recordings were performed with the whole-cell configuration using the patch-clamp technique. In the voltage-clamp mode, two distinct K(+) current components, (I(A)) and (I(K)), were identified, and two different components (59.5% and 96.3%) of I(K) to the total K(+) current were observed at a +50 mV step-pulse. The IC(50) value for 4-aminoprydine (4-AP, 0.05-50 mM), which inhibited the I(A) by 50%, was 0.7 mM. That for tetraethylammonium (TEA, 0.02-20 mM) to inhibit 50% of I(K) was 1.5 mM. In the current-clamp mode, we used 0.5 mM 4-AP and 2 mM TEA at each concentration nearly equal to the IC(50) value. Irrespective of the absence or presence of TEA (2 mM), 0.5 mM 4-AP application increased the number of action potentials due to the decreased duration of the depolarization phase (DDP). TEA in the presence and absence of 4-AP prolonged the duration of action potentials as well as the duration of repolarization phase (DRP). These results suggested that I(A) and I(K) had independent effects regulating the intrinsic firing properties of the action potential number and timing, respectively, in adult rat TTX-R TG neurons.

Endocannabinoids have an important role for the regulation of neuropathic pain. In our previous study, we observed that preventing the degradation of a endocannabinoid, 2-arachidonoylglycerol (2-AG), using an inhibitor of monoacylglycerol lipase (JZL184), attenuated neuropathic orofacial pain (NOP). The present study aimed to investigate mechanisms underlying JZL184-induced attenuation of NOP. We hypothesized that JZL184 may suppress microglial reactivity in the trigeminal spinal subnucleus caudalis (Vc) under NOP. The infraorbital nerve (ION) was hemisected to model NOP in mice, resulting in a significant reduction of mechanical head-withdrawal threshold (MHWT) on day 4 following the ION hemisection. Chronic systemic application of JZL184 at a concentration of 8 or 16 mg/kg/day for 4 days significantly attenuated the reduction of MHWT in mice exposed to NOP. Administering JZL184 at 4 mg/kg/day or its vehicle, however, did not attenuate the MHWT of mice with NOP. The reactivity of microglial cells in the Vc increased in mice with NOP compared to sham-operated controls. The application of JZL184 at 8 or 16 mg/kg/day for 4 days significantly reduced the increased microglial reactivity in the Vc. The changes of microglia under NOP were, by contrast, not reduced by application of the drug at 4 mg/kg/day or its vehicle. The results indicate that preventing 2-AG degradation may increase its accumulation in the Vc and normalize microglial reactivity under NOP, which may contribute to suppressing NOP.

The larynx and associated laryngopharyngeal regions are innervated by the superior laryngeal nerve (SLN) and are highly reflexogenic. Transient receptor potential (TRP) channels have recently been detected in SLN innervated regions; however, their involvement in the swallowing reflex has not been fully elucidated. Here, we explore the contribution of two TRP channels, TRPV1 and TRPM8, located in SLN-innervated regions to the swallowing reflex. Immunohistochemistry identified TRPV1 and TRPM8 on cell bodies of SLN afferents located in the nodose-petrosal-jugular ganglionic complex. The majority of TRPV1 and TRPM8 immunoreactivity was located on unmyelinated neurons. Topical application of different concentrations of TRPV1 and TRPM8 agonists modulated SLN activity. Application of the agonists evoked a significantly greater number of swallowing reflexes compared with the number evoked by distilled water. The interval between the reflexes evoked by the agonists was shorter than that produced by distilled water. Prior topical application of respective TRPV1 or TRPM8 antagonists significantly reduced the number of agonist-evoked reflexes. The findings suggest that the activation of TRPV1 and TRPM8 channels present in the swallowing-related regions can facilitate the evoking of swallowing reflex. Targeting the TRP channels could be a potential therapeutic strategy for the management of dysphagia.

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.

In order to evaluate the neural mechanisms underlying the abnormal facial pain that may develop following regeneration of the injured inferior alveolar nerve (IAN), the properties of the IAN innervated in the mental region were analyzed.

Sarcopenia is a poor prognosis factor in some cancer patients, but little is known about the mechanisms by which malignant tumors cause skeletal muscle atrophy. Tryptophan metabolism mediated by indoleamine 2,3-dioxygenase is one of the most important amino acid changes associated with cancer progression. Herein, we demonstrate the relationship between skeletal muscles and low levels of tryptophan. A positive correlation was observed between the volume of skeletal muscles and serum tryptophan levels in patients with diffuse large B-cell lymphoma. Low levels of tryptophan reduced C2C12 myoblast cell proliferation and differentiation. Fiber diameters in the tibialis anterior of C57BL/6 mice fed a tryptophan-deficient diet were smaller than those in mice fed a standard diet. Metabolomics analysis revealed that tryptophan-deficient diet downregulated glycolysis in the gastrocnemius and upregulated the concentrations of amino acids associated with the tricarboxylic acid cycle. The weights and muscle fiber diameters of mice fed the tryptophan-deficient diet recovered after switching to the standard diet. Our data showed a critical role for tryptophan in regulating skeletal muscle mass. Thus, the tryptophan metabolism pathway may be a promising target for preventing or treating skeletal muscle atrophies.