Menthol in the upper airway depresses ventilation in newborn dogs.

Upper airway cooling depresses ventilation in the newborn dog. Since airway cooling stimulates laryngeal cold receptors and inhibits laryngeal mechanoreceptors, the type of afferent ending responsible for this reflex cannot be easily identified. l-menthol, a specific stimulant of cold receptors in the absence of any cooling, has been used to ascertain the discrete role of upper airway cold receptors in this ventilatory depression. Experiments were carried out in 8 anesthetized 7-14-day-old dogs breathing through a tracheostomy with the upper airway functionally isolated. Constant flows of warm air (37 degrees C), with and without addition of l-menthol, and cold air (25 degrees C) were delivered through the upper airway in the expiratory direction. As compared to warm air trials, cold air and warm air + l-menthol trials greatly reduced ventilation (57.5 +/- 10.7% and 52.8 +/- 11.7% of control, respectively; P less than 0.01) mostly due to a prolongation of Te (291.2 +/- 106.4% and 339.2 +/- 90.0%, respectively, P less than 0.01). Section of the superior laryngeal nerve abolished the response to cold air. However, a residual depressive effect of l-menthol was still present in 3 of 5 animals and was abolished by nasal anesthesia, suggesting the involvement of nasal cold receptors. The results suggest that in the newborn dog stimulation of laryngeal cold receptors, without any concurrent inhibition of laryngeal mechanoreceptors, is a sufficient stimulus to cause respiratory depression.     

Effects of cooling on laryngeal reflexes in the dog

We investigated the reflex effects of laryngeal cooling on posterior cricoarytenoid (PCA) muscle activity, breathing pattern, arterial blood pressure and heart rate. We performed experiments on 9 anesthetized, spontaneously breathing dogs. Laryngeal temperature was decreased by passing cold air through the functionally isolated larynx while the dog was breathing through a tracheostomy. Inspiratory and expiratory durations, esophageal pressure, peak PCA activity, heart rate and blood pressure did not change significantly during laryngeal cooling. Upon interruption of cold airflow, while the laryngeal temperature was returning to control values, we assessed PCA response to upper airway occlusion. At laryngeal temperatures of 20-25 degrees C the peak PCA activity during upper airway occlusion was approximately 2/3 of that observed at control temperature (approximately equal to 33 degrees C). This difference was abolished by topically applied anesthetics or by superior laryngeal nerve section. In addition, we recorded from 4 laryngeal mechanoreceptors stimulated by negative pressure; their response to upper airway occlusion was reduced to 1/2 by laryngeal cooling. These results indicate that laryngeal cooling has a marked depressive effect on the PCA response to collapsing pressure in the larynx, thereby compromising the mechanism subserving upper airway patency.     

Characteristics of laryngeal cold receptors

This study was designed to further characterize the properties of previously described laryngeal cold receptors (Respir. Physiol. 59:35, 1985). Single unit action potentials were recorded from the internal branch of the superior laryngeal nerve (SLN) in anesthetized, spontaneously breathing dogs. The nervous conduction of fibers originating from 12 laryngeal cold receptors was blocked at a mean (+/- SE) temperature of 18.8 +/- 0.7 degrees C. Twelve receptors were localized on the edge of the vocal folds in correspondence of the vocal process of the arytenoid cartilage. Topical anesthesia (2% lidocaine) blocked their activity within 4-18 sec, suggesting a superficial location. Paralysis of the vocal folds during spontaneous breathing through the upper airway did not alter the activity of 9 of 13 cold receptors. On the other hand, 7 of 12 cold receptors tested with constant flow showed respiratory modulation and laryngeal paralysis abolished the modulation of 3 of these tested with a constant flow of air. During progressive cooling in a stepwise fashion, as in frigid air breathing, laryngeal cold receptors maintained a phasic discharge. Our results indicate that these endings are particularly suited for detecting changes in temperature.     

Phrenic and hypoglossal neural responses to cold airflow in the upper airway.

Cold air flowing through the larynx is known to alter the activities of laryngeal receptors with afferents in the superior laryngeal nerves (SLNs) and to induce reflex apnea in neonatal mammals. To examine the ventilatory response in adult animals and to explore associated upper airway motor responses, we recorded phrenic and hypoglossal neural responses to cooling the isolated larynx with cold air in decerebrate, vagotomized, paralyzed, ventilated cats. The most consistent response was phrenic inhibition, which occurred in all animals tested. Either excitation or inhibition of hypoglossal activity was seen consistently in individual cats, with the result that the group response was not statistically significant. All responses to laryngeal cooling were abolished by section of the SLNs. The findings confirm that directing cold air through the larynx causes reflex inhibition of ventilatory (phrenic) activity, but raise new questions as to how the two, directionally opposite hypoglossal responses are mediated.     

Laryngeal receptors responding to transmural pressure, airflow and local muscle activity

The larynx has a rich sensory supply which is the main source of several respiratory reflexes. These reflexes, that influence both the patency of the upper airway and the pattern of breathing, are related to transmural pressure and/or airflow in the upper airway. Yet hardly any information is available on the response of laryngeal mechanoreceptors to transmural pressure and airflow. We recorded action potentials from single fibers separated from the superior laryngeal nerve of anesthetized dogs, breathing spontaneously either through a tracheostomy or the upper airway. The airway could be occluded above or below the larynx. On the basis of their behavior during tracheostomy breathing, upper airway breathing, tracheal occlusion and upper airway occlusion, laryngeal mechanoreceptors were classified as pressure receptors, flow receptors or 'drive' receptors (stimulated by the respiratory activity of upper airway muscles). Pressure receptors were encountered most frequently, representing 63.6% of our sample of 110 receptors, 'drive' receptors constituted 21.8% and flow receptors the remaining 14.6%. Our findings indicate that, even though the three types of receptors differ in sensory modality, they concur in exhibiting a predominant activity during inspiration. In fact, 65% of all receptors are active during eupneic inspiration. Moreover, their activity increases markedly during upper airway obstruction.     

Cooling mediates the ventilatory depression associated with airflow through the larynx

Although constant airflow through the upper airway has been shown to induce ventilatory depression in anesthetized newborn animals, the role of laryngeal temperature in this response has not been studied. Experiments were performed in fourteen 1-5 day-old anesthetized puppies breathing through a tracheostomy. Tidal volume and laryngeal temperature were recorded while a constant stream of air (15-25 ml/sec) at room temperature was passed in the expiratory direction for 20 sec through the isolated upper airway. Warm (35-37 degrees C), humidified air at the same flow served as control. When laryngeal temperature was decreased by 7.5 +/- 0.9 degrees C, a marked change in breathing pattern was observed (VT = 54 +/- 5, TI = 187 +/- 33, TE = 636 +/- 179, VT/TI = 45 +/- 10% of control; n = 9). Warm air at the same flow induced no significant changes. Superior laryngeal nerve section abolished the effects of cooling on breathing pattern. In 5 puppies we compared the effect of 'fast' and 'slow' laryngeal cooling. Fast trials altered breathing pattern earlier than slow trials. We conclude that the depressant effect of airflow through the upper airway is entirely due to a decrease in laryngeal temperature and is mediated by superior laryngeal nerve afferents.     

Laryngeal pressure receptors

We studied the response characteristics of laryngeal pressure receptors in anesthetized dogs, breathing through a tracheal cannula, by recording single unit action potentials from the peripheral cut end of the internal branch of the superior laryngeal nerve. The larynx, with the rest of the upper airway, was isolated and cannulated separately for the application of distending and collapsing pressures. We identified receptors responding to either negative or positive pressure and a few responding to both. All these receptors showed a marked dynamic sensitivity and had the characteristics of slowly adapting mechanoreceptors. The majority of pressure receptors were active at zero transmural pressure and the gain of their response to pressure was higher at lower values, suggesting a role for these receptors in eupnea. Reflex alterations in breathing pattern and upper airway muscle activity during upper airway pressure changes, previously reported, are presumably mediated by the receptors described here. Moreover, these receptors may play a role in certain pathological states, such as obstructive sleep apnea, in which the upper airway is transiently subjected to large collapsing pressure.     

Carbon dioxide-responsive laryngeal receptors in the dog

The purpose of this study was to relate the carbon dioxide (CO2) response of laryngeal receptors to their behavior during the breathing cycle (i.e. their response to transmural pressure changes, laryngeal movement or decreases in temperature) or during exposure to irritant stimuli (water or cigarette smoke). In 9 anesthetized mongrel dogs breathing spontaneously through a tracheostomy, unit activity from the superior laryngeal nerve was recorded while warmed and humidified gas mixtures (air or 10% CO2 in O2) were passed, for 1 min, through the functionally isolated upper airway in the expiratory direction. None of the 10 cold receptors studied were affected by CO2. Eleven of 20 laryngeal non-modulated mechano-receptors were stimulated (from 0.3 to 1.6 imp/sec) by exposure to CO2. These CO2-responsive receptors were also stimulated by known irritant stimuli (cigarette smoke, water), although not all receptors which responded to these irritants were stimulated by CO2. Twelve of 33 respiratory-modulated receptors were affected by CO2; 4 were stimulated and 8 inhibited. Receptors inhibited by CO2 were also inhibited by negative pressure while receptors stimulated by CO2 were also stimulated by negative pressure. These results show that CO2-responsive laryngeal receptors are not specialized endings. Although it is not clear to what extent each separate group of laryngeal receptors is involved, each may contribute to the reflex bradypnea which has been observed during exposure of the upper airway to elevated levels of CO2. However, the importance of CO2-responsive laryngeal receptors in physiological conditions remains unclear.     

Role of intrinsic muscles and tracheal motion in modulating laryngeal receptors

Recording from the superior laryngeal nerve discloses a respiratory modulated activity even in the absence of airflow and pressure changes in the larynx. The present study evaluates the relative contribution of intrinsic laryngeal muscle activity and transmitted tracheal movement on the respiratory modulation of laryngeal mechanoreceptors. Seventy-four receptors were studied in 22 anesthetized spontaneously breathing dogs. The modulation of 31 receptors depended solely on laryngeal muscle activity since it was abolished by cold block of laryngeal nerves. Twelve receptors were primarily activated by tracheal movement since tracheal stabilization alone reduced or abolished their modulation. The respiratory modulation of the remaining 31 receptors was found to be dependent on both laryngeal muscle activity and tracheal movements. Lidocaine (2%) was applied to the receptor field of 13 endings; the results indicate that while some receptors are located superficially (blocked within 1 min) others are located in deeper structures (not affected in 30 min). These receptors may be involved in the precise coordination of laryngeal muscle activity and could play a role in the regulation of breathing pattern and airway patency due to their pressure sensitivity.     

Effects of halothane,enflurane, and isoflurane on laryngeal receptors in dogs

The effects of halothane, enflurane, and isoflurane on laryngeal receptors were investigated in 6 anethetized dogs breathing spontaneously through a tracheostomy. Single unit actiion potentials were recorded from the peripheral cut end of the superior laryngeal nerve (SLN) while different concentrations of volatile anesthetics (1.25, 2.5., 5.0%) were administered in the expiratory direction at a constant air-flow (6 1/min) for 1 min through the functionally isolated upper airway. A total of 21 respiratory-modulated mechanoreceptors, 18 “irritant” receptors, and 7 cold receptors were studied. The overall results obtained from the 16 respiratory-modulated mechanoreceptors challenged with the 3 anesthetic gases disclosed a prevalent inhibitory effect and halothane proved to be the most effective of the 3 gases. The activity during both the inspiratory and expiratory phase was significantly reduced only by halothane (inspiratory phase, P<0.01; expiratory phase, P<0.05), while neither isoflurane nor enflurane caused significant changes in receptor activity. Of the 18 irritant receptors, 14 receptors increased their activity in a dose-related manner in response to one or more of the anesthetics although the effect of halothane was more pronounced than those of enflurane and isoflurane. All of the 7 cold receptos consistently increased their activity in a dose-related manner in response to halothane whereas 3 of 7 receptors were insensitive to enflurane and 4 of 7 receptors were insensitive to isoflurane. Our results indicate that, while all three commonly used anethetics can have an effect on different types of laryngeal receptors, the effects of halothane are more pronounced than those of the other two gases in terms of changes in receptor activity.     

Water-responsive laryngeal receptors in the dog are not specialized endings

The primary purpose of this study was to ascertain whether laryngeal receptors activated by water are specialized endings or whether they also respond to other stimuli, such as pressure, temperature and laryngeal motion as they occur during the breathing cycle. In 35 anesthetized mongrel dogs, breathing spontaneously through a lower cervical tracheostomy, water and other test solutions at approximately 37 degrees C were injected into the functionally isolated larynx with a small catheter. Of the 130 receptors studied, none of the cold receptors (N = 13) responded to water, whereas approximately 60% of all laryngeal mechanoreceptors (72 of 117) responded with either a short delay, short duration or a long delay, long duration response. In general the former pattern of response was exhibited by nonrespiratory-modulated receptors, whereas the latter was typical of respiratory-modulated receptors. The specific nature of the stimulus (hypotonicity or lack of chloride ion) of the water response was further studied in 53 receptors with isoosmotic solutions of dextrose and sodium gluconate. The long delay, long duration response was dependent on a decreased osmolality, while the short delay, short duration response was dependent on the lack of chloride ion of the test solutions. All water-responsive receptors tested (N = 17) were blocked within 50 sec by topically applied 2% lidocaine and thus presumed to be superficial. However, 10 receptors which did not respond to water were also blocked within 50 sec, suggesting that not all superficial receptors are stimulated by water. Based on these observations, we propose that changes in osmolality or ionic composition of the laryngeal surface liquid could play an important role in modifying reflexes involved in the maintenance of upper airway patency.     

Dynamic analysis of airway temperature in dogs breathing cold air]

Exercise in cold, dry air induces bronchoconstriction. However, the time course of airway cooling during the breathing of cold air has not been investigated. In this study in dogs, the temperatures of tracheal gas and the tracheal wall were measured continuously while the animal was breathing cold air at approximately 4.5 degrees C. The temperature of the tracheal gas decreased during the inspiratory phase, increase slightly early in the expiratory phase, and then decreased to the level at end-expiration. The lowest temperature of the tracheal gas decreased significantly, from 29.7 +/- 2.4 degrees C (mean +/- SD) to 25.7 +/- 2.8 degrees C. The highest temperature also changed significantly, but the decrease was less than 1 degree C. The lowest temperature of the tracheal wall decreased from 31.1 +/- 2.6 degrees C to 30.5 +/- 2.5 degrees C during cold-air breathing, but the difference was not significant. When a dog inspired deeply during a cough, the temperature of tracheal gas did not decrease linearly with the progression of inspiration. However, the decrease in the tracheal wall temperature was almost directly proportional to inspiratory volume. We concluded that both the tracheal gas and the tracheal wall temperatures were resistant to cooling with cold-air breathing. It remains uncertain whether rapid, deep breathing during exercise decreased the tracheal wall temperature.     

Respiratory activity in the superior laryngeal nerve of the rabbit

We studied the respiratory modulation of laryngeal afferents and their response to transmural pressure in 24 anesthetized, spontaneously breathing rabbits. Laryngeal afferent activity has a predominant inspiratory augmentation during tracheal breathing or tracheal occlusion that can be accounted for by the respiratory movement transmitted to the larynx through the trachea. During upper airway breathing or upper airway occlusion SLN afferent activity increases in expiration and decreases in inspiration. This respiratory modulation is due to changes in upper airway pressure (Pua). In fact, positive pressure stimulates SLN afferent activity, while negative pressure inhibits it. Mechanical restriction of epiglottal movement reduced the response to Pua changes during upper airway occlusion and application of maintained positive (0.1-0.5 kPa) and negative (-0.1 to -0.5 kPa) pressures (P less than 0.005). Furthermore, surgical removal of epiglottis decreased the baseline activity of SLN to 16.5% of control. These experiments suggest that in the rabbit the epiglottis is the main source of SLN afferent activity and that its displacement, due to changes in Pua, is the most important factor for modulating SLN activity. Most of the laryngeal receptors showed an inspiratory augmentation with tracheal breathing and occlusion, were stimulated by positive pressure and inhibited by negative pressure, reflecting the behavior observed in the whole nerve.     

Laryngeal receptors and their reflex responses

The afferent activity originating from the larynx shows a considerable respiratory modulation. Receptors responding to pressure changes, inspiratory airflow (cold), and laryngeal movements have been identified. In addition, other receptors without a respiratory modulation are also described. Possible reflex effects of these receptors on breathing pattern, upper airway patency, and defense mechanisms in both adults and newborns are discussed.     

Respiratory effects of cold air breathing in anesthetized cats

Respiratory effects of cold air breathing were studied in anesthetized cats. Two different protocols were used: the air temperature was either lowered in an isolated segment, constituted by the larynx and oropharynx or in lower airways, so that the cats inspired the cold air directly. Temperatures ranged between 37 and 8 degrees C (first protocol) or between 37 and 15 degrees C (second protocol). When the temperature fell below 15 degrees C in the upper segment, marked increase in lung resistance occurred, without any significant changes in ventilatory variables nor in diaphragmatic electrical activity. The section of superior laryngeal nerves abolished this bronchomotor effect. In present experimental circumstances, thermal changes measured in lower airways when cats breathed cold air were mainly located in the cervical trachea. An increase in lung resistance and weak but significant changes in the diaphragmatic electromyogram began when the inspired air temperature fell below 25 degrees C. A selective local block of conduction in small vagal fibres by procaine or section of vagus nerves suppressed all these effects. In all cases the cold-induced changes in lung mechanics began very early (less than 10 sec) but continued for few minutes after the physiological temperature range had been restored in airways. The present data strongly suggest that the bronchomotor response to cold air breathing is a reflex, mediated by afferent fibres in the superior laryngeal nerves and in the vagus nerves.     

Laryngeal afferents activated by phenyldiguanide and their response to cold air or helium-oxygen

In anesthetized cats, sensory neurons in the superior laryngeal nerves (SLN) were identified with respect to their response to (1) phenyldiguanide (PDG) i.v., (2) mechanical stimulation and (3) lowering temperature in an isolated tracheolaryngeal segment. The activity originating from 107 SLN afferent units activated by PDG was recorded using glass microelectrodes advanced in the nodose ganglion. All tested afferent units increased their discharge rate during direct touching of the airway mucosa. None showed flow or pressure related activity during abrupt changes in constant laryngeal flow or transmural pressure in the isolated segment. Fifteen units were inhibited by cold air. Sixty-two units significantly increased their firing rate when the temperature approached 18 degrees C, reached a peak discharge near 15 degrees C, then their activity decreased or stopped. The response to cold air was compared to cold heliox (79% He-21% O2), which enhanced the respiratory heat loss by conduction. The peak firing rate was significantly higher with heliox (+356% compared to +246% with air), the temperature threshold higher (25 degrees C +/- 1.0 degree C) and the temperature range broader (25-11.5 degrees C). Present results show that a large proportion (58%) of afferent SLN fibres activated by PDG are likely non-proprioceptive units, which are stimulated by cooling the inspired gas. Thermosensitive units in the upper airways may act as sensors of the thermal flux through the airway wall more than as detectors of the absolute value of temperature in the airway lumen.     

The respiratory activity of the superior laryngeal nerve in the rat.

The aim of this study was to characterize the laryngeal afferent activity of the rat. The animals were anesthetized and breathing spontaneously. Laryngeal afferent activity was recorded from both the whole superior laryngeal nerve (SLN) and from single fibers isolated from this nerve. An overall inspiratory augmenting activity was observed in the whole SLN during tracheostomy breathing, tracheal occlusion and upper airway breathing, but an expiratory augmenting activity was present during upper airway occlusion. The inspiratory modulated activity was abolished by bilateral section of the hypoglossal nerves but not the recurrent laryngeal nerves. A great number of receptors (46/80, 58%) were identified as 'drive' receptors, and others as 'pressure' (22/80, 28%) and 'irritant' type receptors (9/80, 11%). Nineteen pressure receptors were stimulated by positive transmural pressure, while only three stimulated by negative pressure. Nine drive receptors were also stimulated by positive pressure and inhibited by negative pressure. Such response to pressure was further evaluated by applying maintained pressures to the functionally isolated upper airway. These results are essentially consistent with findings obtained in the rabbit, but differ from those reported for the dog.     

Effects of upper airway anaesthesia on respiratory-related evoked potentials in humans.

Cortical potentials evoked by mid-inspiratory occlusion arise from numerous receptors, many of which are probably within the upper airway. Their precise nature is not known. The aim of the current study was to improve knowledge of this by studying the effects of topical upper airway anaesthesia on respiratory-related evoked potentials. Respiratory-related evoked potentials were described through the averaging of electroencephalogram (EEG) epochs following mid-inspiratory occlusions (C3-CZ; C4-CZ). A total of 21 healthy volunteers (13 male, aged 22-52 yrs) were studied during mouth breathing, before and after topical upper airway anaesthesia (lidocaine). Moreover, 15 subjects were studied during nose breathing with and without anaesthesia. Six subjects were studied whilst inhaling L-menthol. Typical potentials were present in all the subjects, their components featuring normal amplitudes and latencies. The route of breathing and upper airway anaesthesia did not modify the EEG responses to inspiratory occlusions, qualitatively or quantitatively, during mouth or nose breathing. L-menthol had no effect. Upper airway receptors sensitive to topical anaesthesia are unlikely to contribute significantly to mid-inspiratory occlusion-evoked potentials. On the contrary, deeper receptors, such as joint and muscle receptors, could contribute dominantly to these potentials.     

Effects of airway cooling on tracheal stretch receptors

We studied the effect of cold air on tracheal slowly adapting stretch receptors (SAR) in 6 anesthetized, spontaneously breathing dogs. Air at constant flow and two different temperatures was passed through an isolated segment of the extrathoracic trachea. We recorded SAR action potentials, esophageal pressure, tracheal pressure and temperature. With a reduction in tracheal temperature of approximately 10 degrees C the steady state response of 30 SARs to a distending pressure of 1.0 kPa decreased to 75% of control (P less than 0.001). At lower distending pressure the inhibitory effect of tracheal cooling decreased: 87% of control at 0.5 kPa (P less than 0.05, n = 8) and 96% of control at 0.2 kPa (P greater than 0.05, n = 8). The response of 13 tracheal SARs to sinusoidal pressure oscillations (0.15 kPa) superimposed on a bias pressure (0.5 kPa) was reduced (P less than 0.01) by local cooling to the same extent at the 2 pressure extremes ('peak' value = 71% of control; 'valley' = 67% of control), resulting in a similar change in receptor discharge within the oscillatory cycle. The inhibitory effect of airway cooling on stretch receptors may play a role in cold-induced bronchoconstriction.     

Ventilation in kittens with chronic section of the superior laryngeal nerves

The breathing pattern of conscious newborn kittens one-to-two weeks old was studied by the barometric method about 5 days after bilateral section of the superior laryngeal nerve (SLN-denervated group) or a sham operation (SLN-sham operated group). None of the ventilatory variables differed between the two groups, whether during normoxia or acute hypoxia (10 min of 10% O2). After anesthesia, delivery of steady airflows in the expiratory direction through the upper airways of the SLN-sham operated had marked inhibitory effects on ventilation which entirely disappeared after SLN section. A small inhibition was still present in the SLN-denervated group, possibly indicating that other non-SLN upper airways receptors developed inhibitory ventilatory effects during the period of chronic denervation. Intermittent expiratory upper airway airflows were much less effective than steady flows and no inhibition was seen with oscillatory flows, indicating that the mode of application of the stimulus to the laryngeal receptors is crucial in determining the magnitude of their reflex response. Under anesthesia, acute bilateral section of the SLN determined a small increase of the integrated peak EMG activity of the diaphragm. We conclude that laryngeal SLN afferents are inhibitory on ventilation in newborn kittens, but this effect is very small during normal conscious conditions. Only under special circumstances, including anesthesia and sustained upper airways flows and pressures, the ventilatory inhibition can be disproportionately magnified.