Physiological analyses of swallowing changes due to chronic obstructive pulmonary disease in anesthetized male rats.

Chronic obstructive pulmonary disease (COPD) was previously known as chronic bronchitis and emphysema. It has various main symptoms, such as dyspnea, chronic cough, and sputum, and is often accompanied by dysphagia. Although many published clinical reports have described COPD-related dysphagia, the physiological mechanisms underlying swallowing changes due to COPD remain unclear. Therefore, we analyzed how COPD affects the swallowing reflex using COPD model rats. We performed an electrophysiological study of respiration and swallowing using COPD model induced by intratracheal administration of porcine pancreatic elastase and lipopolysaccharide in Sprague-Dawley male rats. To identify the respiration and swallowing responses, electromyographic activity was recorded from the diaphragm, digastric (Dig), and thyrohyoid (TH) muscles. We confirmed COPD using micro-computed tomography analysis and hematoxylin and eosin staining of the lungs. The duty cycle was defined as the ratio of the inspiration duration to the total respiratory duration. In COPD model rats, the duty cycle was significantly higher than that in control rats. The frequency of the swallowing reflex evoked by electrical stimulation of the superior laryngeal nerve during the inspiration phase was higher in COPD model rats than in control rats. Furthermore, long-term COPD altered Dig and TH muscle activity without pathological muscle change. Our results suggest that COPD increases the frequency of swallowing initiation during the inspiration phase. Furthermore, long-term COPD affects swallowing-related muscle activity without pathological muscle changes. These physiological changes may increase the risk of developing dysphagia. Further studies are necessary to clarify the mechanisms contributing to the functional changes in respiration and swallowing in COPD.

Inhibition of Water-Evoked Swallowing During Noxious Mechanical Stimulation of Tongue in Anesthetized Rats.

Dysphagia is sometimes accompanied by pain. Because orofacial structures subserve mastication and swallowing, orofacial pain might impair both functions. Tongue biting can occur not only accidentally while eating but also in some pathological conditions. However, it remains unclear whether noxious mechanical stimulation of the tongue affects swallowing. To explore this question, we evaluated the effects of lingual pinch stimulation on the initiation of swallowing evoked by distilled water (DW) infusion with a flow rate of 5.0 µL/s for 20 s into the pharyngolaryngeal region in anesthetized rats. The swallowing reflex was identified by electromyographic (EMG) bursts in the suprahyoid muscles which include the anterior belly of the digastric muscle, mylohyoid and geniohyoid muscles, and laryngeal elevation by visual inspection. The number of DW-evoked swallows during pinch stimulation was significantly smaller than that in a control condition or during pressure stimulation. The onset latency of the first swallow during pinch stimulation was significantly longer than that in the control condition. DW-evoked swallowing was almost abolished following bilateral transection of the superior laryngeal nerve (SLN) compared with the control condition, suggesting that the SLN plays a crucial role in the initiation of DW-evoked swallowing. Finally, electrophysiological data indicated that some SLN-responsive neurons in the nucleus tractus solitarii (nTS) exhibited delayed latency from a single SLN stimulation during lingual pinch stimulation. These results suggest that noxious mechanical stimulation of the tongue inhibits the initiation of swallowing and modulates neuronal activity in the nTS.

Coordination of Respiration, Swallowing, and Chewing in Healthy Young Adults.

Examining the coordination of respiration and swallowing is important for elucidating the mechanisms underlying these functions and assessing how respiration is linked to swallowing impairment in dysphagic patients. In this study, we assessed the coordination of respiration and swallowing to clarify how voluntary swallowing is coordinated with respiration and how mastication modulates the coordination of respiration and swallowing in healthy humans. Twenty-one healthy volunteers participated in three experiments. The participants were asked to swallow 3 ml of water with or without a cue, to drink 100 ml of water using a cup without breathing between swallows, and to eat a 4-g portion of corned beef. The major coordination pattern of respiration and swallowing was expiration-swallow-expiration (EE type) while swallowing 3 ml of water either with or without a cue, swallowing 100 ml of water, and chewing. Although cueing did not affect swallowing movements, the expiratory time was lengthened with the cue. During 100-ml water swallowing, the respiratory cycle time and expiratory time immediately before swallowing were significantly shorter compared with during and after swallowing, whereas the inspiratory time did not differ throughout the recording period. During chewing, the respiratory cycle time was decreased in a time-dependent manner, probably because of metabolic demand. The coordination of the two functions is maintained not only in voluntary swallowing but also in involuntary swallowing during chewing. Understanding the mechanisms underlying respiration and swallowing is important for evaluating how coordination affects physiological swallowing in dysphagic patients.

Involvement of capsaicin-sensitive nerves in the initiation of swallowing evoked by carbonated water in anesthetized rats.

Capsaicin powerfully evokes the swallowing reflex and is a known therapeutic agent for improving dysphagia and preventing aspiration pneumonia. However, the role of capsaicin-sensitive nerves in the initiation of swallowing evoked by various natural stimuli remains unclear. To explore this question, we blocked laryngeal capsaicin-sensitive nerves following the coapplication of QX-314 and capsaicin (QX/Cap), and investigated the effects on swallowing evoked by mechanical and chemical stimulation in anesthetized rats. Swallows were evoked by capsaicin, carbonated water (CW), distilled water (DW), and punctate mechanical stimulation using von Frey filaments applied topically to the larynx. Swallows were documented by recording electromyographic activation of the suprahyoid and thyrohyoid muscles. The initiation of swallowing by capsaicin was strongly suppressed at 5 min following QX/Cap treatment and returned in a time-dependent manner. CW-evoked swallows at 5 min following QX/Cap treatment were significantly diminished compared with before and 30 min after treatment. In contrast, DW-evoked and mechanically evoked swallows were unchanged by QX/Cap treatment. Furthermore, CW-evoked swallows were virtually abolished by transection of the superior laryngeal nerves and significantly decreased by the topical application of acid-sensing ion channel-3 (ASIC3) inhibitor APETx2, but they were not affected by the nonselective transient receptor potential channel inhibitor ruthenium red or the ASIC1 inhibitor mambalgin-1. Taken together, we speculate that capsaicin-sensitive nerves play an important role in the initiation of CW-evoked swallows.NEW & NOTEWORTHY The initiation of swallowing evoked by laryngeal capsaicin and carbonated water application was diminished by the coapplication of QX-314 and capsaicin. Carbonated water-evoked swallows were also abolished by transection of the superior laryngeal nerves and were inhibited by the acid-sensing ion channel-3 inhibitor. Capsaicin-sensitive nerves are involved in the initiation of carbonated water-evoked swallows.

Sustained laryngeal transient receptor potential vanilloid 1 activation inhibits mechanically induced swallowing in anesthetized rats.

A major component of gastric acid is hydrochloric acid (HCl), which can activate transient receptor potential vanilloid 1 (TRPV1). In the present study, we investigated how sustained laryngeal TRPV1 activation affects the frequency of the swallowing reflex. Experiments were carried out on 85 male Sprague-Dawley rats. The effects of short and sustained application of chemicals (3 µl of 0.1 N HCl or capsaicin) on the frequency of swallowing and on time-dependent changes in the occurrence of swallowing evoked by supralaryngeal nerve stimulation were determined. To evaluate vascular permeability of the larynx, Evans blue dye was intravenously injected after 5 or 60 min of sustained TRPV1 activation. SB366791 (a TRPV1 inhibitor) and Cap/QX-314 (a TRPV1-expressed neuronal inhibitor) significantly inhibited HCl/capsaicin-induced swallowing, but air flow-induced swallowing was not affected. Although the number of air flow-induced swallows followed by capsaicin stimulation was not affected within 5 min, it was significantly reduced by 60-min capsaicin or HCl application. The swallowing threshold associated with supralaryngeal nerve stimulation did not significantly change throughout the recording period. Evans blue dye concentrations in the larynx were significantly higher at 60 min in the 10-5 M capsaicin group than in the control group. Our results suggest that sustained TPRV1 activation not only desensitizes TRPV1 but also inactivates mechanoreceptors, which may be attributed to increases in vascular permeability and edema, as part of an inflammatory process.NEW & NOTEWORTHY Although a transient receptor potential vanilloid 1 (TRPV1) inhibitor or TRPV1-expressed neuronal inhibitor significantly inhibited HCl/capsaicin-evoked swallowing, air flow-induced swallowing was not affected. The number of air flow-induced swallows was significantly reduced within 60 min of TRPV1 activation. Evans blue dye concentration in the larynx increased 60 min after capsaicin application. TPRV1 activation not only desensitizes TRPV1 but also inactivates mechanoreceptors caused by increases in vascular permeability and edema.

Involvement of the epithelial sodium channel in initiation of mechanically evoked swallows in anaesthetized rats.

KEY POINTS: Afferents carried by the superior laryngeal nerve play a primary role in the initiation of laryngeal mechanically evoked swallows in anaesthetized rats. Amiloride and its analogues inhibit swallowing evoked by mechanical stimulation, but not swallowing evoked by chemical and electrical stimulation. The epithelial sodium channel is probably involved in the initiation of laryngeal mechanically evoked swallows. ABSTRACT: The swallowing reflex plays a critical role in airway protection. Because impaired laryngeal mechanosensation is associated with food bolus aspiration, it is important to know how the laryngeal sensory system regulates swallowing initiation. This study was performed to clarify the neuronal mechanism of mechanically evoked swallows. Urethane-anaesthetized Sprague-Dawley male rats were used. A swallow was identified by activation of the suprahyoid and thyrohyoid muscles on electromyography. The swallowing threshold was measured by von Frey filament and electrical stimulation of the larynx. The number of swallows induced by upper airway distension and capsaicin application (0.03 nmol, 3 µl) to the vocal folds was counted. The effects of topical application (0.3-30 nmol, 3 µl) of the epithelial sodium channel (ENaC) blocker amiloride and its analogues (benzamil and dimethylamiloride), acid-sensing ion channel (ASIC) inhibitors (mambalgine-1 and diminazene) and gadolinium to the laryngeal mucosa on swallowing initiation were evaluated. A nerve transection study indicated that afferents carried by the superior laryngeal nerve play a primary role in the initiation of laryngeal mechanically evoked swallows. The mechanical threshold of swallowing was increased in a dose-dependent manner by amiloride and its analogues and gadolinium, but not by ASIC inhibitors. The number of swallows by upper airway distension was significantly decreased by benzamil application. However, the initiation of swallows evoked by capsaicin and electrical stimulation was not affected by benzamil application. We speculate that the ENaC is involved in the initiation of laryngeal mechanically evoked swallows.

Effect of attention on chewing and swallowing behaviors in healthy humans.

We examined how attention alters chewing and swallowing behaviors. Twenty-one healthy volunteers were asked to freely eat 8 g of steamed rice in three separate trials, and we obtained the average number of chewing cycles (N) and chewing duration (T) prior to the first swallow in each trial. We also conducted an N-limited test, in which participants chewed the food while independently counting the number of chewing cycles and swallowed the food when they reached N, and a T-limited test, in which they chewed the food for T sec and then swallowed. We recorded electromyograms (EMGs) from masseter and suprahyoid muscles and collected videoendoscopic images. In the N-limited test, chewing speed decreased, masseter muscle activity (area under the curve of the rectified EMG burst) per cycle increased, and suprahyoid muscle activity per cycle decreased. In the T-limited test, the chewing speed increased, muscle activities per cycle decreased, and the number of cycles increased. The occurrence frequency of bolus propulsion into the pharynx before swallowing was smaller in the N- and T-limited tests than in the free chewing test. Further, the whiteout time was longer in the T-limited test than in the free chewing test. Attentional chewing changes not only chewing but also swallowing behavior.

Effect of peripherally and cortically evoked swallows on jaw reflex responses in anesthetized rabbits.

This study aimed to investigate whether the jaw-opening (JOR) and jaw-closing reflexes (JCR) are modulated during not only peripherally, but also centrally, evoked swallowing. Experiments were carried out on 24 adult male Japanese white rabbits. JORs were evoked by trigeminal stimulation at 1 Hz for 30 s. In the middle 10 s, either the superior laryngeal nerve (SLN) or cortical swallowing area (Cx) was simultaneously stimulated to evoke swallowing. The peak-to-peak JOR amplitude was reduced during the middle and late 10-s periods (i.e., during and after SLN or Cx stimulation), and the reduction was dependent on the current intensity of SLN/Cx stimulation: greater SLN/Cx stimulus current resulted in greater JOR inhibition. The reduction rate was significantly greater during Cx stimulation than during SLN stimulation. The amplitude returned to baseline 2 min after 10-s SLN/Cx stimulation. The effect of co-stimulation of SLN and Cx was significantly greater than that of SLN stimulation alone. There were no significant differences in any parameters of the JCR between conditions. These results clearly showed that JOR responses were significantly suppressed, not only during peripherally evoked swallowing but also during centrally evoked swallowing, and that the inhibitory effect is likely to be larger during centrally compared with peripherally evoked swallowing. The functional implications of these results are discussed.

Distinctive features of Phox2b-expressing neurons in the rat reticular formation dorsal to the trigeminal motor nucleus.

Phox2b encodes a paired-like homeodomain-containing transcription factor essential for development of the autonomic nervous system. Phox2b-expressing (Phox2b+) neurons are present in the reticular formation dorsal to the trigeminal motor nucleus (RdV) as well as the nucleus of the solitary tract and parafacial respiratory group. However, the nature of Phox2b+ RdV neurons is still unclear. We investigated the physiological and morphological properties of Phox2b+ RdV neurons using postnatal day 2-7 transgenic rats expressing yellow fluorescent protein under the control of Phox2b. Almost all of Phox2b+ RdV neurons were glutamatergic, whereas Phox2b-negative (Phox2b-) RdV neurons consisted of a few glutamatergic, many GABAergic, and many glycinergic neurons. The majority (48/56) of Phox2b+ neurons showed low-frequency firing (LF), while most of Phox2b- neurons (35/42) exhibited high-frequency firing (HF) in response to intracellularly injected currents. All, but one, Phox2b+ neurons (55/56) did not fire spontaneously, whereas three-fourths of the Phox2b- neurons (31/42) were spontaneously active. K+ channel and persistent Na+ current blockers affected the firing of LF and HF neurons. The majority of Phox2b+ (35/46) and half of the Phox2b- neurons (19/40) did not respond to stimulations of the mesencephalic trigeminal nucleus, the trigeminal tract, and the principal sensory trigeminal nucleus. Biocytin labeling revealed that about half of the Phox2b+ (5/12) and Phox2b- RdV neurons (5/10) send their axons to the trigeminal motor nucleus. These results suggest that Phox2b+ RdV neurons have distinct neurotransmitter phenotypes and firing properties from Phox2b- RdV neurons and might play important roles in feeding-related functions including suckling and possibly mastication.