Takanori Tsujimura

Involvement of ERK phosphorylation in brainstem neurons in modulation of swallowing reflex in rats

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 γ‐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 GABAA 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 GABAA 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.

Individual-dependent effects of pharyngeal electrical stimulation on swallowing in healthy humans.

The present study tested whether electrical stimulation increases the number of voluntary repetitive swallows in humans. In addition, the potential of initiating both voluntary and involuntary swallows was compared using electrical stimulation. Fourteen normal male adults were instructed to perform repetitive voluntary swallowing as quickly as possible over 30 s, and the number of swallows was counted with and without repetitive electrical stimulation (80% of maximal tolerated intensity; 0.1 ms in pulse duration; 30 Hz) of the nasal cavity, nasopharynx, oropharynx, or laryngopharynx. Although the number of swallows was significantly increased during electrical stimulation of the laryngopharynx, oropharynx, and nasopharynx, there was a wide variation in the number among subjects. The number of reflexively evoked swallows (i.e., involuntary swallows) by pharyngeal stimulation also varied greatly, and there was a significant linear correlation in the number of swallows between voluntary and involuntary swallows. The present study demonstrated the facilitatory effects of pharyngeal electrical stimulation on voluntary swallowing in humans. Furthermore, the potentials of initiation of voluntary swallows are identical to those of involuntary swallows, which suggests that the swallowing central pattern generator is a common component of both neuronal networks and therefore is responsible for inter-individual variations.

Differential involvement of two cortical masticatory areas in modulation of the swallowing reflex in rats

To clarify the functional role of cortical descending inputs involved in the swallowing reflex, the effect of electrical stimulation of two cortical masticatory areas (CMAs: A- and P-area) on rhythmic jaw movements (RJMs) and superior laryngeal nerve (SLN)-evoked swallows were studied. RJMs and swallowing reflex were elicited by repetitive electrical stimulation of CMAs and the SLN, respectively. The electromyographic activities of jaw-closer (masseter), jaw-opener (digastric), and laryngeal-elevator (thyrohyoid) muscles were recorded to identify the RJMs and swallowing reflex. The number of evoked swallows was significantly lower, and swallowing interval was significantly longer during A-area stimulation compared with those without stimulation. Conversely, these parameters were not significantly altered during P-area stimulation. The inhibition of swallows by A-area stimulation was not affected by an increase in sensory input by wooden stick application between upper and lower teeth, or A-area stimulation preceding SLN stimulation. The present findings suggest that the swallowing reflex is inhibited by activation of the A-area, but not the P-area. Since no changes in swallows were seen after the increase in intraoral sensory input and prior activation of masticatory central pattern generator (CPG), swallowing inhibition may be mediated by direct inputs from the A-area or inputs via the masticatory CPG into the swallowing CPG.

Effects of electrical stimulation of the superior laryngeal nerve on the jaw-opening reflex

The present study aimed to examine whether the jaw-opening reflex (JOR) is modulated during swallowing, and if so, to compare the modulation between the low- and high-threshold afferent-evoked reflex responses. Experiments were carried out on 11 anesthetized rabbits. The inferior alveolar nerve was stimulated to evoke the JOR in the digastric muscle. The stimulus intensity was either 1.5 (low threshold) or 4.0 (high threshold) times the threshold for eliciting the JOR. As a conditioning stimulation, the superior laryngeal nerve (SLN) was repetitively stimulated to evoke the swallowing reflex. The stimulus intensity ranged from 0.6 to 8.0 times the threshold to evoke the swallowing reflex during SLN stimulation over 20s. Electromyographic (EMG) activities of the digastric and mylohyoid muscles were recorded, and the peak-to-peak EMG amplitude of the digastric muscle was measured and compared with and without SLN stimulation, as well as with and without swallowing. Comparisons were also made between low- and high-threshold afferent-evoked JORs. The JOR was strongly suppressed during SLN stimulation. The degree of suppression increased and the latency for the JOR was delayed when the stimulus current applied to the SLN was increased. Such modulation was apparent when the low-threshold afferent-evoked JOR was recorded. Effects of motor outputs of swallowing events and those of single-pulse stimulation of SLN on the inhibition of the JOR were not noted. These results suggest that the JOR evoked by both the low- and high-threshold afferents was inhibited during laryngeal sensory input and following swallowing, probably to prevent opposing jaw movements evoked by oral sensory input during swallowing.

Changes in jaw muscle activity and the physical properties of foods with different textures during chewing behaviors

This study aimed to investigate how the activity of the masseter (Mas) and suprahyoid (Hyoid) muscles is influenced by the physical properties of food, how changes in the rheological properties of food differ between different foods during the process of food reduction, and how different salivary flow rates affect bolus-making capability during masticatory behavior in healthy humans. Ten healthy adults participated in this study. Electromyographic (EMG) recordings were obtained from the Mas and Hyoid muscles, and 15 g of steamed rice and rice cake was prepared as test foods. In the ingestion test, the subjects were asked to eat each food in their usual manner. The chewing duration, number of chewing cycles before the first swallow, Mas and Hyoid EMG activity, and chewing cycle time were compared between the foods. Total chewing duration was divided into three substages: early, middle, and late; chewing cycle time and EMG activity per chewing cycle of each substage were compared between the foods and among the substages. In the spitting test, the rheological properties of the bolus at the end of each substage were compared between the foods and among the substages. Finally, stimulated salivary flow rates were measured and the relationships between salivary flow rate and chewing duration, EMG activity, and changes in physical food characteristics were investigated. There were significant differences in total chewing duration and the number of chewing cycles, but not in chewing cycle time, between the foods, which had similar hardness values. The EMG activity levels of the Mas and Hyoid per chewing cycle for the rice cake were significantly greater than for the steamed rice throughout the recording periods. While Mas activity did not change among the substages during chewing, Hyoid EMG activity decreased as chewing progressed. Chewing cycle time also gradually decreased as chewing progressed. The hardness of both foods initially increased, then gradually decreased back to baseline. The adhesiveness of the rice cake initially increased, and did not fall throughout the recording period; the adhesiveness of the steamed rice did not significantly change. Cohesiveness barely changed in either of the two foods during chewing, but was significantly greater for the rice cake than for the steamed rice. Finally, a correlation between the stimulated salivary flow rate and chewing performance was evident only in a change in Mas EMG activity. The current results demonstrate that the activities of the Mas and Hyoid muscles changed as chewing progressed, and were affected by hardness, adhesiveness, and cohesiveness. Salivary flow rate may affect the changes in Mas activity during the process of bolus formation.

Changes in the frequency of swallowing during electrical stimulation of superior laryngeal nerve in rats.

The aim of the present study was to investigate the adaptation of the swallowing reflex in terms of reduced swallowing reflex initiation following continuous superior laryngeal nerve stimulation. Forty-four male Sprague Dawley rats were anesthetized with urethane. To identify swallowing, electromyographic activity of the left mylohyoid and thyrohyoid muscles was recorded. To evoke the swallowing response, the superior laryngeal nerve (SLN), recurrent laryngeal nerve, or cortical swallowing area was electrically stimulated. Repetitive swallowing evoked by continuous SLN stimulation was gradually reduced, and this reduction was dependent on the resting time duration between stimulations. Prior SLN stimulation also suppressed subsequent swallowing initiation. The reduction in evoked swallows induced by recurrent laryngeal nerve or cortical swallowing area stimulation was less than that following superior laryngeal nerve stimulation. Decerebration had no effect on the reduction in evoked swallows. Prior subthreshold stimulation reduced subsequent initiation of swallowing, suggesting that there was no relationship between swallowing movement evoked by prior stimulation and the subsequent reduction in swallowing initiation. Overall, these data suggest that reduced sensory afferent nerve firing and/or trans-synaptic responses, as well as part of the brainstem central pattern generator, are involved in adaptation of the swallowing reflex following continuous stimulation of swallow-inducing peripheral nerves and cortical areas.

How do tablet properties influence swallowing behaviours

Behavioural performance of tablet swallowing was evaluated with different tablet conditions in terms of size, number and surface coating.

Inhibition of swallowing reflex following phosphorylation of extracellular signal-regulated kinase in nucleus tractus solitarii neurons in rats with masseter muscle nociception

Pain is associated with swallowing abnormalities in dysphagic patients. Understanding neuronal mechanisms underlying the swallowing abnormalities associated with orofacial abnormal pain is crucial for developing new methods to treat dysphagic patients. However, how the orofacial abnormal pain is involved in the swallowing abnormalities is not known. In order to evaluate neuronal mechanisms of modulation of the swallows by masticatory muscle pain, here we first induced swallows by topical administration of distilled water to the pharyngolaryngeal region. The swallowing reflex was significantly inhibited after capsaicin (10, 30mM) injection into the masseter muscle compared to vehicle injection. Moreover the number of phosphorylated extracellular signal-regulated kinase-like immunoreactive (pERK-LI) neurons in the nucleus tractus solitarii (NTS) was significantly increased in the rats with capsaicin injection into the masseter muscle compared to that with vehicle injection. Rostro-caudal distribution of pERK-LI neurons in the NTS was peaked at the obex level. The capsaicin-induced inhibitory effect on swallowing reflex was reversed after intrathecal administration of mitogen-activated protein kinase (MAPK) kinase (MEK) inhibitor, PD98059. The present findings suggest that phosphorylation of ERK in NTS neurons may be involved in capsaicin-induced inhibition of swallowing reflex.

Organization of pERK-immunoreactive cells in trigeminal spinal nucleus caudalis, upper cervical cord, NTS and Pa5 following capsaicin injection into masticatory and swallowing-related muscles in rats

Many phosphorylated extracellular signal-regulated kinase (pERK)-immunoreactive (IR) cells are expressed in the trigeminal spinal subnucleus caudalis (Vc), upper cervical spinal cord (C1-C2), nucleus tractus solitarii (NTS) and paratrigeminal nucleus (Pa5) after capsaicin injection into the whisker pad (WP), masseter muscle (MM), digastric muscle (DM) or sternohyoideus muscle (SM). The pERK-IR cells also showed NeuN immunoreactivity, indicating that ERK phosphorylation occurs in neurons. The pERK-IR cells were significantly reduced after intrathecal injection of MEK 1/2 inhibitor PD98059. The pERK-IR cells expressed bilaterally in the Vc and C1-C2 after capsaicin injection into the unilateral DM or SM, whereas unilaterally in the Vc and C1-C2 after unilateral WP or MM injection. After capsaicin injection into the WP or MM, the pERK-IR cell expression in the Vc was restricted rostrocaudally within a narrow area. However, the distribution of pERK-IR cells was more wide spread without a clear peak in the Vc and C1-C2 after capsaicin injection into the DM or SM. In the NTS, the unimodal pERK-IR cell expression peaked at 0-720μm rostral from the obex following capsaicin injection into WP, MM, DM or SM. In the ipsilateral Pa5, many pERK-IR cells were observed following capsaicin injection into the SM. The number of swallows elicited by distilled water administration was significantly smaller after capsaicin injection into the WP, MM or DM but not SM compared to that of vehicle-injected rats. Various noxious inputs due to the masticatory or swallowing-related muscle inflammation may be differentially involved in muscle pain and swallowing reflex activity.

Peripheral and central control of swallowing initiation in healthy humans.

We investigated (1) how peripheral inputs might assist central inputs in the control of voluntary evoked swallowing, (2) inter-individual variation in involuntary and voluntary swallowing initiation, and (3) whether natural chewing behavior affects the initiation of involuntary swallowing in healthy humans. Eleven participants completed a repetitive saliva swallowing test (RSST), chewing test (CHEW), and rest period (REST). In RSST, participants repetitively swallowed as quickly as possible. In CHEW, subjects chewed gum freely. We delivered pharyngeal electrical stimulation (PEStim) to the laryngopharynx and compared the number of swallows that occurred with and without PEStim. PEStim significantly increased the number of voluntary evoked swallows in RSST, as well as the number of swallows in CHEW and REST trials, although this facilitatory effect was larger in REST trials. We found a positive correlation between the number of swallows at RSST without PEStim and that at REST with PEStim within individuals. Additionally, we found a significant positive correlation between the number of swallows at RSST with PEStim and the sum of that at RSST without PEStim and at REST with PES. Based on the current results, we suggest that (1) peripheral inputs within a certain range appear to facilitate the central inputs that control voluntary swallowing, (2) inter-individual variations in swallowing initiation may arise from differences in the excitability of the common neural network in the lower brainstem, and (3) during chewing, food reduction in the oral cavity is prioritized, such that the neural network associated with chewing may regulate swallowing initiation.