Intracellular activity of pharyngeal motoneurons during breathing, swallowing, and coughing.

We recorded membrane potentialp changes in 45 pharyngeal motoneurons (PMs) including 33 expiratory modulated and 12 nonrespiratory neurons during breathing, swallowing, and coughing in decerebrate paralyzed cats. Four types of membrane potential changes were observed during swallowing: 1) depolarization during swallowing (n = 27), 2) depolarization preceded by a brief (≤ 0.1 s) hyperpolarization (n = 4), 3) longer term (> 0.3 s) hyperpolarization followed by depolarization (n = 11), and 4) hyperpolarization during the latter period of swallowing (n = 3). During coughing, PMs showed two types of membrane potential changes (n = 10). Nine neurons exhibited a ramp-like depolarization during the expiratory phase of coughing with the potential peak at the end of expiratory phase. This depolarization was interrupted by a transient repolarization just before the potential peak. The membrane potential of the remaining neuron abruptly depolarized at the onset of the expiratory phase and then gradually decreased even after the end of the expiratory phase. Single-shock stimulation of the superior laryngeal nerve (SLN) induced inhibitory postsynaptic potentials in 19 of 21 PMs. Two motoneurons exhibited an SLN-induced excitatory postsynaptic potential. The present study revealed that PMs receive the central drive, consisting of a combination of excitation and inhibition, from the pattern generator circuitry of breathing, swallowing, and coughing, which changes the properties of their membrane potential to generate these motor behaviors of the pharynx. Our data will provide the basis of studies of pharyngeal activity and its control from the medullary neuronal circuitry responsible for the upper airway motor activity.NEW & NOTEWORTHY We have provided the first demonstration of the multifunctional activity of the pharyngeal motoneurons at the level of membrane potential during respiration, swallowing, and coughing.

Role of the retrotrapezoid nucleus/parafacial respiratory group in coughing and swallowing in guinea pigs.

The retrotrapezoid/parafacial respiratory group (RTN/pFRG) located ventral to the facial nucleus plays a key role in regulating breathing, especially enhanced expiratory activity during hypercapnic conditions. To clarify the roles of the RTN/pFRG region in evoking coughing, during which reflexive enhanced expiration is produced, and in swallowing, during which the expiratory activity is consistently halted, we recorded extracellular activity from RTN/pFRG neurons during these fictive behaviors in decerebrate, paralyzed, and artificially ventilated guinea pigs. The activity of the majority of recorded respiratory neurons was changed in synchrony with coughing and swallowing. To further evaluate the contribution of RTN/pFRG neurons to these nonrespiratory behaviors, the motor output patterns during breathing, coughing, and swallowing were compared before and after brain stem transection at the caudal margin of RTN/pFRG region. In addition, the effects of transection at its rostral margin were also investigated to evaluate pontine contribution to these behaviors. During respiration, transection at the rostral margin attenuated the postinspiratory activity of the recurrent laryngeal nerve. Meanwhile, the late expiratory activity of the abdominal nerve was abolished after caudal transection. The caudal transection also decreased the amplitude of the coughing-related abdominal nerve discharge but did not abolish the activity. Swallowing could be elicited even after the caudal end transection. These findings raise the prospect that the RTN/pFRG contributes to expiratory regulation during normal respiration, although this region is not an essential element of the neuronal networks involved in coughing and swallowing.

Activity of respiratory neurons in the rostral medulla during vocalization, swallowing, and coughing in guinea pigs.

To examine the relationship between the neuronal networks underlying respiration and non-respiratory behaviors such as vocalization and airway defensive reflexes, we compared the activity of respiratory neurons in the ventrolateral medulla during breathing with that during non-respiratory behaviors including vocalization, swallowing, and coughing in guinea pigs. During fictive vocalization the activity of augmenting expiratory neurons ceased, whereas the other types of expiratory neurons did not show a consistent tendency of increasing or decreasing activity. All inspiratory neurons discharged in synchrony with the phrenic nerve activity. Most of the phase-spanning neurons were activated throughout the vocal phase. During fictive swallowing, many expiratory and inspiratory neurons were silent, whereas many phase-spanning neurons were activated. During fictive coughing, many expiratory neurons were activated during the expiratory phase of coughing. Most inspiratory neurons discharged in parallel with the phrenic nerve activity during coughing. Many phase-spanning neurons were activated during the expiratory phase of coughing. These findings indicate that the medullary respiratory neurons help shape respiratory muscle nerve activity not only during breathing but also during these non-respiratory behaviors, and thus suggest that at least some of the respiratory neurons are shared among the neuronal circuits underlying the generation of breathing and non-respiratory behaviors.

Glycosylation of Nα-lauryl-O-(ß-D-xylopyranosyl)-L-serinamide as a saccharide primer in cells.

N(α)-Lauryl-O-(ß-D-xylopyranosyl)-L-serinamide (Xyl-Ser-C12) was synthesized as a saccharide primer to obtain oligosaccharides of glycosaminoglycan using the glycan biosynthetic potential of mouse osteosarcoma FBJ-S1 cells and Chinese hamster ovary (CHO) cells. The glycosylated products secreted into the culture medium were collected and analyzed by liquid chromatography-mass spectrometry and glycosidase digestion. The structure of the Xyl-Ser-C12 derivatives was investigated. Several glycosaminoglycan-type oligosaccharides, such as GalNAc-(GlcA-GlcNAc)(n)-GlcA-Gal-Gal-Xyl-Ser-C12, were detected, and identified as intermediates of the biosynthesis of heparan sulfate glycosaminoglycans. Xyl-Ser-C12 exhibited greater acceptor activity for the glycosylation of glycosaminoglycan-type oligosaccharides than p-nitrophenyl-ß-D-xylopyranoside.

Axonal projections of medullary swallowing neurons in guinea pigs.

A central pattern generator (CPG) for swallowing in the medulla oblongata generates spatially and temporally coordinated movements of the upper airway and alimentary tract. To reveal the medullary neuronal network of the swallowing CPG, we examined the cytoarchitecture of the swallowing CPG and axonal projections of its individual neurons by extracellular recording and juxtacellular labeling of swallowing-related neurons (SRNs) in the medulla in urethane-anesthetized and paralyzed guinea pigs. Three major types of neuronal discharge patterns were identified during fictive swallowing induced by stimulation of the superior laryngeal nerve: early (burst-like activation during the pharyngeal stage), late (activation after the pharyngeal stage), and inhibited (inhibition during the pharyngeal stage) types. Sixteen neurons were successfully labeled in the nucleus tractus solitarii (NTS) and in the medullary reticular formation (RF). No motoneuron was labeled. The SRNs in the NTS had axons projecting to the NTS, RF, nucleus ambiguus, nucleus hypoglossus, and dorsal motor nucleus of the vagus on the ipsilateral side. Some NTS SRNs projected only within the NTS. The axons of SRNs in the RF projected also to the NTS, RF, motor nuclei on the ipsilateral side, and to the other side RF. These findings show anatomic substrates for the neuronal network of the CPG for swallowing, which consists of complex neuronal connections among SRNs in the NTS, RF, and motor nuclei.

Swallow-related inhibition in laryngeal motoneurons.

Inhibitory postsynaptic potentials (IPSPs) of laryngeal motoneurons (LMs) are essential for narrowing the glottis at just the right time during swallowing, which prevents aspiration. To examine the property of IPSPs of LMs during swallowing, we monitored the effects of intracellular application of chloride ion and extracellular application of inhibitory neurotransmitter antagonists on the membrane potential trajectories of LMs during fictive swallowing in decerebrate, paralyzed cat. Adductor LMs hyperpolarized briefly at the beginning of the pharyngeal stage of swallowing (PS) and then depolarized explosively during the remaining part of the PS. Abductor LMs exhibited various patterns during swallowing; hyperpolarization during the PS followed by depolarization at the offset of the PS, slight depolarization, or plateau potentials. Chloride-dependent IPSPs were revealed during the initial part of PS in adductor LMs and during the whole PS in abductor LMs. The swallow-related IPSPs were depressed by iontophoretic extracellular application of bicuculline in both adductor and abductor LMs, but they were not modified by strychnine application. It is concluded that the swallow-related inhibition of both adductor and abductor LMs is chloride-dependent IPSPs mediated through GABA(A) receptors, not through glycine receptors.

Brainstem vocalization area in guinea pigs.

The purpose of the present study was to determine whether murines could be substituted for traditional experimental mammals to study the brainstem mechanism of vocalization. We conducted systematic electrical and chemical stimulation of the brainstem in guinea pigs to identify the similarities in the call sites between murines and other mammals. We further examined whether or not fictive vocalization could be induced in paralyzed guinea pigs, an experimental model which facilitates neuronal recording in the brainstem. The sites where electrical stimulation evoked vocalization were distributed continuously from the periaqueductal grey (PAG) to the lower brainstem. This call area usually ended at the most caudal part of the inferior olive and thus did not continuously extend to the nucleus retroambiguus. Microinjections of d,l-homocysteic acid and bicuculline induced vocalization at the PAG, parabrachial nucleus, and the most dorsal part of the pontine reticular formation. The brainstem call areas and vocal motor patterns induced from these areas were approximately consistent with those in other mammals. Fictive vocalization induced by PAG stimulation could be identified from activities of the phrenic, abdominal, and superior laryngeal nerves in paralyzed guinea pigs. We thus concluded that guinea pigs can be utilized in studies of brainstem vocal mechanism.

Multifunctional laryngeal premotor neurons: their activities during breathing, coughing, sneezing, and swallowing.

To examine whether motor commands of two or more distinct laryngeal motor patterns converge onto a common premotor network, we conducted dual recordings from the laryngeal adductor motoneuron and its premotor neuron within the brainstem respiratory circuitry during fictive breathing, coughing, sneezing, and swallowing in decerebrate paralyzed cats. Expiratory neurons with an augmenting firing pattern (EAUG), whose action potentials evoked monosynaptic IPSPs in the adductor motoneurons, sharply fired during the expulsive phases of fictive coughing and sneezing, during which the adductor motoneurons transiently repolarized. In contrast, these premotor neurons were silent during the swallow-related hyperpolarization in adductor motoneurons. These results show that one class of medullary respiratory neuron, EAUG, is multifunctional and shared among the central pattern generators (CPGs) for breathing, coughing, and sneezing. In addition, although the CPGs underlying these three behaviors and the swallowing CPG do overlap, EAUG neurons are not part of the swallowing CPG and, in contrast to the other three behaviors, are not a source of inhibitory input to adductor motoneurons during swallowing.

Paradoxical vocal cord motion: a review focused on multiple system atrophy.

OBJECTIVE: Paradoxical vocal cord motion (PVCM) is a well recognized respiratory condition in which active adduction of the vocal cords during inspiration causes functional airway obstruction. It is considered that laryngeal reflex acceleration underlies the generation of nonorganic PVCM. In various situations producing PVCM, multiple system atrophy (MSA) is a representative neurological disease causing nocturnal laryngeal stridor attributed to PVCM. The purpose of this review is to identify the underlying mechanisms associated with nonorganic and MSA-related PVCM. The following issues are addressed in this review: (1) the pathophysiology of nonorganic and MSA-related PVCM, (2) the relationships between PVCM and airway reflexes, and (3) the treatment for MSA-related PVCM. METHODS: Review. RESULTS AND CONCLUSIONS: An abnormality of the laryngeal output-feedback control underlies nonorganic PVCM, which is usually triggered by an excessive response to external and internal airway stimuli. Similarly, several clinical and experimental evidence suggest that MSA-related PVCM is attributed to the airway reflex as well as to paradoxical central outputs resulting from the MSA-induced damage to the pontomedullary respiratory center. Application of continuous positive airway pressure (CPAP), which suppresses the reflexive inspiratory activation of adductors, is recommended as the treatment for MSA-related PVCM.

Tracheostomy abolishes paradoxical activation of the vocal cord adductor in multiple system atrophy.

OBJECTIVES: Inspiratory activation of the vocal cord adductor, which causes paradoxical vocal cord motion, develops in patients with multiple system atrophy (MSA). To confirm the hypothesis that airway reflexes trigger such paradoxical activation, we investigated the effects of tracheostomy on the adductor activation in a MSA patient. METHODS: We compared the adductor electromyograms before and after breathing was diverted to a tracheostoma under propofol anesthesia. RESULTS: The adductor inspiratory activation disappeared during tracheostoma breathing. CONCLUSION: Airway reflexes as well as MSA-related damage to the respiratory center contribute to the generation of paradoxical adductor activation in MSA patients.

Membrane potential changes in vocal cord tensor motoneurons during breathing, vocalization, coughing and swallowing in decerebrate cats.

We studied the patterns of membrane potential changes in vocal cord tensor motoneurons, i.e. cricothyroid muscle motoneurons (CTMs), during fictive breathing, vocalization, coughing, and swallowing in decerebrate paralyzed cats to determine the nature of central drives to CTMs during these behaviors. CTMs were identified by antidromic activation from the superior laryngeal nerve. During breathing, CTMs always depolarized during the inspiratory phase, and sometimes depolarized during the expiratory phase as well. During vocalization, CTMs strongly depolarized. During coughing, CTMs exhibited depolarizations during both inspiratory and expiratory phases, but it was interrupted by a transient repolarization between the last part of the inspiratory phase and the first part of the abdominal burst during which chloride-dependent inhibitory postsynaptic potentials were revealed. During swallowing, most CTMs hyperpolarized, and this hyperpolarization was sometimes followed by a weak depolarization. We conclude that the main role of the cricothyroid muscle is vocalization but the functional roles in coughing and swallowing are minor, and that the CTM activity during resting breathing and vocalization are primarily controlled by excitatory inputs, while during coughing and swallowing, inhibitory inputs play roles in shaping membrane potential trajectories.

Inspiratory activation of the vocal cord adductor, part I: human study in patients with restricted abduction of the vocal cords.

OBJECTIVES/HYPOTHESIS: In patients with restricted abduction of the vocal cords, it has generally been accepted that glottis narrowing with laryngeal stridor during inspiration is attributed to static and passive obstruction of the glottis. However, active glottis narrowing can also be contributory. We tested the hypothesis that the vocal cord adductor is activated during inspiration in patients with restricted abduction of the vocal cords. STUDY DESIGN: Electromyographic evaluation of vocal cord adductor activity in patients with restricted abduction of the vocal cords. METHODS: Five patients with restricted abduction of the vocal cords who had stridor with mild to severe dyspnea during wakefulness were anesthetized with propofol. We recorded the adductor muscle electromyogram during breathing through a laryngeal mask airway while observing the vocal cord movement endoscopically. In three patients who had undergone tracheostomy, we also recorded adductor firing patterns not only while closing but also while opening the tracheostoma. RESULTS: The adductor was activated during inspiration, and the glottis was narrowed in accordance with inspiratory stridor. This adductor inspiratory activity was abolished by opening the tracheostoma in the tracheostomized patients. CONCLUSION: Not only static or passive glottis narrowing but also active narrowing may contribute to inspiratory flow limitation in patients with restricted abduction of the vocal cords. This active glottis narrowing is probably induced by an airway reflex.

Inspiratory activation of the vocal cord adductor, part II: Animal study in the cat.

OBJECTIVES/HYPOTHESIS: The authors have shown previously that the vocal cord adductor is activated during inspiration in patients with vocal cord abduction impairment and that this adductor inspiratory activity is abolished by relief from inspiratory tracheal negative pressure by opening the tracheostoma. (Shiba K. Isono S, Sekita Y, Tanaka A. Inspiratory activation of the vocal cord adductor, Part I: human study in patients with restricted abduction of the vocal cords. Laryngoscope 2004;114:372-375). The authors hypothesized that insufficient opening of the glottis during inspiration generates strong negative pressure in the trachea and that this negative pressure triggers an airway reflex that activates the adductor. STUDY DESIGN: Experimental study of the mechanism of laryngeal obstruction using an animal model of restricted abduction of the vocal cords. METHODS: To identify such an airway reflex, the authors recorded the adductor electromyogram in anesthetized cats whose vocal cords were mechanically adducted by stitching both cords together. To determine whether this reflex modulation of adductor activity is induced through afferents from the larynx or from the lower airway, the authors applied negative pressure to the subglottic space and lower airway separately. RESULTS: The adductor was activated during inspiration with powerful negative pressure in the trachea. Negative pressure in the subglottic space had a more marked effect on the adductor activity than did pressure in the lower airway. The adductor inspiratory activity was virtually abolished by laryngeal deafferentation. CONCLUSION: Glottal narrowing during inspiration reflexly activates the vocal cord adductor. This paradoxical inspiratory-related adductor activation is induced by an airway reflex triggered mainly through afferents from the larynx and probably contributes to stridor and dyspnea in patients with laryngeal obstruction.

Ipsilateral full-thickness forearm skin graft for covering the radial forearm flap donor site.

OBJECTIVES/HYPOTHESIS: The technique of the ipsilateral full-thickness forearm skin graft for covering the defect of radial forearm free flap (RFFF) improves aesthetic impairment at the recipient and donor sites by split-thickness skin graft repair and omits the need to make an extraoperative site for harvesting the skin graft. However, in this technique, the RFFF is limited in size. In the present study, we considered a model of the forearm and calculated the possible size of the RFFF for using this technique. METHODS: The calculation was conducted under assumptions that the isosceles-triangle skin graft is elevated as its height is twice the RFFF length in the direction of the forearm axis and that the forearm skin defect can be primarily closed with a width shorter than one-fourth of the wrist circumference. The calculation revealed that this technique is feasible when the RFFF width, that is, the length vertical to the forearm axis, is shorter than half of the wrist circumference. We repaired the RFFF defect using this technique in 15 patients with head and neck cancer in whom the RFFF size conformed to the above-mentioned condition. RESULTS: When the RFFF width was shorter than half of the wrist circumference and the isosceles-triangle skin graft was elevated as its height was twice the RFFF length, the RFFF defect could be repaired using this technique in all 15 patients. CONCLUSION: The above-mentioned condition (that the RFFF width is shorter than half of the wrist circumference) is useful for determining whether or not the technique of ipsilateral full-thickness forearm skin graft can be used for covering the RFFF defect.

Hemodynamic changes of the hand after radial forearm flap harvesting.

After radial forearm flap harvesting, there is some risk for hand circulatory disorders. To reveal the changes in circulatory dynamics in the hand after harvesting this flap, the authors compared blood pressure and flow by color Doppler ultrasonography in the donor and nondonor hands, and evaluated the long-term changes in these factors in 40 patients undergoing this operation. Blood pressure and flow of the index finger in the donor hands were lower than those in the nondonor hands during the first 2 months postoperatively, but they virtually returned to the level of those in the nondonor hands within 1 year of the operation. These results suggest that after harvesting the radial artery, collateral circulation in the hand developed during a short postoperative period. Therefore, the authors can predict the long-term safety of forearm flap harvesting by evaluating the hemodynamic changes of the digits caused by acute occlusion of the radial artery preoperatively, which would reflect the hemodynamics at an early postoperative stage.

Firing patterns of micturition-related neurons in the pontine storage centre in cats.

The pontine storage centre (PSC) and the pontine micturition centre (PMC) are known to be critical for urinary filling and emptying, respectively. In the present study, firing patterns of 45 neurons in the PSC area where electrical stimulation induced inhibition of the micturition reflex were analyzed in 20 male decerebrated and paralyzed cats. The electrically determined PSC area was widespread in the dorsolateral pontine reticular formation (P0-P4), ventrolateral to the PMC. Four major types of neurons were detected according to urinary storage/micturition cycles: tonic storage neurons (38%), phasic storage neurons (40%), tonic micturition neurons (9%) and phasic micturition neurons (13%). These four types of neurons were intermingled in the PSC. However, the tonic and phasic micturition neurons tended to be located within a limited area (P2-P3). These neurons were further classified into augmenting, constant and decrementing firing patterns. Some increased their firing prior to the storage/micturition phase initiation. Such preceding pattern was more frequently found in the tonic neurons than in the phasic neurons. In conclusion, the PSC neurons with diverse heterogeneous discharge patterns suggest that these neurons may organize a complex neuronal circuitry, which is critical in the neural control of the urinary continence.

Usefulness of color Doppler sonography for assessing hemodynamics of free flaps for head and neck reconstruction.

This report focuses on the monitoring of intraoperative and postoperative hemodynamics of free flaps for repair of head and neck defects by color Doppler sonography (CDS). The study group included 20 patients with head and neck cancer who underwent resection and reconstruction with free flaps. The hemodynamics in the feeding arteries of the flaps were measured during the following six stages: before surgery, immediately after microvascular anastomosis, and on postoperative days 1, 3, 5, and 7. The pulsatility index (PI) was used as the index for measuring changes in hemodynamics over time. Grafts showed the maximal PI immediately after vascular anastomosis. PI decreased over 3 to 7 days. Of the 20 patients, 1 patient in whom the hypopharynx was reconstructed with the radial forearm flap developed venous occlusion. This was diagnosed during the early stage using CDS, allowing the flap to be saved. CDS proved to be very useful for real-time observation of the hemodynamics in free flaps.