Postnatal maturation of laryngeal chemoreflexes in the preterm lamb.

Laryngeal chemoreflexes (LCR) are triggered by the contact of liquids with the laryngeal mucosa. In the mature organism, LCR trigger lower airway protective responses (coughing, effective swallowing, and arousal) to prevent aspiration. General belief holds that LCR are responsible for apnea and bradycardia in the newborn mammal, including humans. Our laboratory has recently shown that LCR in full-term lambs are consistently analogous to the mature LCR reported in adult mammals, without significant apneas and bradycardias (St-Hilaire M, Nsegbe E, Gagnon-Gervais K, Samson N, Moreau-Bussiere F, Fortier PH, and Praud J-P. J Appl Physiol 98: 2197-2203, 2005). The aim of the present study was to assess LCR in nonsedated, newborn preterm lambs born at 132 days of gestation (term = 147 days). The preterm lambs were instrumented for recording glottal adductor electromyogram, electroencephalogram, eye movements, heart rate, respiration, and oximetry. A chronic supraglottal catheter was used for injecting 0.5 ml of saline, distilled water, and HCl (pH 2) during quiet sleep, active sleep, and wakefulness on postnatal days 7 (D7) and 14 (D14). Laryngeal stimulation by water or HCl on D7 induced significant apneas, bradycardia, and desaturation, which, at times, appeared potentially life-threatening. No significant apneas, bradycardias, or desaturation were observed on D14. No consistent effects of sleep state could be shown in the present study. In conclusion, laryngeal stimulation by liquids triggers potentially dangerous LCR in preterm lambs on D7, but not on D14. It is proposed that maturation of the LCR between D7 and D14 is partly involved in the disappearance of apneas/bradycardias of prematurity with postnatal age.     

Effects of simulated reflux laryngitis on laryngeal chemoreflexes in newborn lambs

It has been suggested that reflux laryngitis (RL) is involved in apneas-bradycardias of the newborn. The aim of the present study was to develop a unique RL model in newborn lambs to test the hypothesis that RL enhances the cardiorespiratory components of the laryngeal chemoreflexes (LCR) in the neonatal period. Gastric juice surrogate (2 ml of normal saline solution with HCl pH 2 + pepsin 300 U/ml) (RL group, n = 6) or normal saline (control group, n = 6) was repeatedly injected onto the posterior aspect of the larynx, 3 times a day for 6 consecutive days, via a retrograde catheter introduced into the cervical esophagus. Lambs instilled with gastric juice surrogate presented clinical signs of RL, as well as moderate laryngitis on histological observation. Laryngeal chemoreflexes were thereafter induced during sleep by injection of 0.5 ml of HCl (pH 2), ewe's milk, distilled water or saline into the laryngeal vestibule via a chronic, transcutaneous supraglottal catheter. Overall, RL led to a significantly greater respiratory inhibition compared with the control group during LCR, including longer apnea duration (P = 0.01), lower minimal respiratory rate (P = 0.002), and a more prominent decrease in arterial hemoglobin saturation (SpO(2)) (P = 0.03). No effects were observed on cardiac variables. In conclusion, 1) our unique neonatal ovine model presents clinical and histological characteristics of RL; and 2) the presence of RL in newborn lambs increases the respiratory inhibition observed with LCR, at times leading to severe apneas and desaturations.     

Inhibitory effects of CO2 on airway defensive reflexes in enflurane-anesthetized humans

We investigated responses of respiration, blood pressure, and heart rate to tracheal mucosa irritation induced by injection of distilled water at three different levels of CO2 ventilatory drive in 11 spontaneously breathing female patients under a constant depth of enflurane anesthesia [1.1 minimum alveolar concentration (MAC)]. The airway irritation at the resting level of spontaneous breathing caused a variety of respiratory responses such as coughing, expiration reflex, apnea, and spasmodic panting, with considerable increases in blood pressure and heart rate. Although the latency of respiratory responses after water injection was much shorter than those of blood pressure and heart rate responses, blood pressure and heart rate responses, once elicited, were prolonged much longer than was the respiratory response. An increase in CO2 ventilatory drive decreased the degree and duration of respiratory, blood pressure, and heart rate responses to the airway irritation, whereas a decrease in CO2 ventilatory drive had the opposite effect on these responses. Our results indicate that changes in CO2 ventilatory drive can modify reflex responses of respiration, blood pressure, and heart rate to airway irritation.     

Modification of cardiac activity in response to dehydration in the terrestrial slug, Limax maximus

The level of body hydration in the terrestrial slug Limax maximus modifies several aspects of behavior such as pneumostome activity, feeding responsiveness, huddling, and contact-rehydration. The relationship between water balance and pneumostome activity and respiratory function suggested that cardiac activity might also be affected. To pursue this possibility, intact slugs and isolated heart-central nervous system (CNS) preparations were used to investigate cardiac responses to the increase in hemolymph osmolality which occurs during dehydration. In intact animals, heart rate increased in response to progressive air-dehydration and to increases in hemolymph osmolality resulting from injections of hyperosmotic solutions of mannitol or NaCl. In isolated preparations, the heart or CNS were separately exposed to hyperosmotic saline. Exposure of the heart alone to hyperosmotic saline resulted in decreased heart rate while exposure of only the CNS resulted in an increase in heart rate. These observations suggest that the increase in heart rate that is observed in intact air-dehydrated slugs is primarily mediated by the CNS.     

Upper airway chemoreflex responses to saline and water in preterm infants

Laryngeal chemoreflex (LCR) responses elicited by fluid irrigation of the larynx have been described repeatedly in animals, whereas evidence for a similar reflex in human infants is extremely limited. Using nasopharyngeal catheters to instill small volumes of warm saline or water into the pharynx, we examined the incidence and characteristics of such a reflex in nine premature infants. Saline and water elicited the same pattern of responses, which frequently included swallows, central apnea, and airway obstruction and less commonly featured coughs, prolonged apnea, and arousal. With the exception of arousal, the incidence of these responses was significantly greater after delivery of water stimuli than after saline bolus administration. We therefore deduce chemoreceptor involvement in generation of these reflex responses and propose a laryngeal site for this sensory system, as in animals. Since greater potency of water compared with saline was demonstrable in all the infants studied, we further conclude that most preterm infants possess an upper airway chemoreflex.     

Pharyngeal fluid clearance and aspiration preventive mechanisms in sleeping infants

We sought to characterize ventilatory and airway protective responses to pharyngeal stimulation in young infants during sleep. We studied the various responses with respect to frequency of occurrence, effect of increased stimulus intensity, and relation of stimulus fluid to laryngeal structures. Two groups of infants were studied: healthy full-term infants (n = 5) and preterm infants with a history of prolonged apnea (n = 9). We used a nasopharyngeal catheter to deliver small boluses of warm saline (0.02-0.35 ml) to the oropharynx. Responses repeatedly observed in both infant groups included swallows, obstructed respiratory efforts, brief apnea, prolonged apnea, and cough. In both infant groups, swallows and obstructed breaths occurred frequently and cough and prolonged apnea infrequently. The functional significance of some response patterns was clear, whereas that of others was obscure. Larger stimulus volumes yielded more frequent responses (P less than 0.01), and preterm infants responded much more frequently than full-term infants (P less than 0.01). Prolonged apnea was a composite of the other responses and was much more common in preterm than full-term infants (P less than 0.01). The stimulus technique was performed under direct visualization of the airway in two deceased infants. The findings suggested that the relation of the piriform fossae to the interarytenoid notch is important in determining response frequency. Implications for regulation of the removal of upper airway secretions during sleep are discussed.     

Heart rate and respiratory patterns in mild hypoxia in unanaesthetized newborn mammals

The heart rate and respiratory patterns in hypoxia are not well documented in unanaesthetized intact newborn animals. We studied heart rate and respiratory patterns during quiet sleep in 17% inspired O2 in 31 unanaesthetized newborns of five species: lamb, piglet, puppy, kitten, and rabbit. There was no significant change in mean heart rate and respiratory rate with hypoxia for any species. Brief apneas greater than 5 s were frequent (5-8/h), both in 21 and 17% O2 only in lambs and puppies. No sustained periodic breathing was induced by hypoxia. Thus, mild hypoxia has little steady-state effect on heart rate and respiratory rate and pattern in these unanaesthetized newborns. These findings are compatible with depressed chemoreceptor threshold, but indicate a remarkably mature respiratory pattern in full-term newborns of these species.     

Effect of ultrasonically nebulized distilled water on airway epithelial cell swelling in guinea pigs.

To investigate the pathogenesis of ultrasonically nebulized distilled water-induced airway narrowing, we studied the role of airway epithelial cells during a distilled water-inhalation challenge in an animal model of airway inflammation. Guinea pigs were divided into four groups: 1) a sham/saline (S/S) group: sham ozone followed by saline inhalation; 2) a sham/water (S/W) group: sham ozone followed by water inhalation; 3) an ozone/saline (O/S) group: ozone followed by saline inhalation; and 4) an ozone/water (O/W) group: ozone followed by water inhalation. After exposure to either 3.0 parts/million ozone or air at the same flow rate for 2 h, guinea pigs were anesthetized and tracheostomized, and then lung resistance (RL) was measured. For morphometric assessment, tissues were fixed with formaldehyde, stained with hematoxylin and eosin, and cut into transverse sections. Airway dimensions were either measured directly or calculated from the internal perimeter, the external perimeter, and airway wall area. There were no statistical differences in the values of RL before distilled water inhalation between the sham groups and the ozone groups. RL increased significantly after 10 min of distilled water inhalation in both the S/W group and the O/W group. In the S/W group, epithelial cells were swollen, and intercellular spaces were wider, resulting in significant increase in epithelial wall thickness, but there was no significant infiltration by inflammatory cells. In the O/S group, the epithelium showed infiltration by inflammatory cells without change in cell volume. In the O/W group, the epithelium showed both infiltration and a greater increase in epithelial wall thickness compared with the S/W group. These results suggest that airway epithelial cell swelling, induced by inhaled distilled water, increases with RL in guinea pigs and that this reaction may be accelerated by airway inflammation.     

Effect of nasal continuous or intermittent positive airway pressure on nonnutritive swallowing in the newborn lamb.

The present study was aimed at investigating the effects of nasal continuous positive airway pressure (nCPAP; 6 cmH2O) or intermittent positive pressure ventilation (nIPPV; 10/4 cmH2O) on nonnutritive swallowing (NNS) and on the coordination between NNS and phases of the respiratory cycle, while taking into account the potential effects of states of alertness. Twelve full-term lambs were chronically instrumented at 48 h after birth for polysomnographic recordings, including NNS, diaphragm electromyographic activity, respiratory movements, pulse oximetry, and states of alertness. Studies in control conditions, with nCPAP and nIPPV, were performed in random order in nonsedated lambs at 4, 5, and 6 days of life. Results demonstrate that nCPAP significantly decreased overall NNS frequency, more specifically isolated NNS during quiet sleep and bursts of NNS in active sleep. In comparison, the effects of nIPPV on NNS frequency were more variable, with an inhibition of NNS only in wakefulness and an increase in isolated NNS frequency in active sleep. In addition, neither nCPAP nor nIPPV disrupted the coordination between NNS and phases of the respiratory cycle. In conclusion, nCPAP inhibits NNS occurrence in newborn lambs. Clinical relevance of this novel finding is related to the importance of NNS for clearing the upper airways from secretions and gastric content frequently regurgitated in the neonatal period.     

Respiratory syncytial virus infection is associated with an altered innate immunity and a heightened pro-inflammatory response in the lungs of preterm lambs

Introduction

Factors explaining the greater susceptibility of preterm infants to severe lower respiratory infections with respiratory syncytial virus (RSV) remain poorly understood. Fetal/newborn lambs are increasingly appreciated as a model to study key elements of RSV infection in newborn infants due to similarities in lung alveolar development, immune response, and susceptibility to RSV. Previously, our laboratory demonstrated that preterm lambs had elevated viral antigen and developed more severe lesions compared to full-term lambs at seven days post-infection. Here, we compared the pathogenesis and immunological response to RSV infection in lungs of preterm and full-term lambs.

Methods

Lambs were delivered preterm by Caesarian section or full-term by natural birth, then inoculated with bovine RSV (bRSV) via the intratracheal route. Seven days post-infection, lungs were collected for evaluation of cytokine production, histopathology and cellular infiltration.

Results

Compared to full-term lambs, lungs of preterm lambs had a heightened pro-inflammatory response after infection, with significantly increased MCP-1, MIP-1α, IFN-γ, TNF-α and PD-L1 mRNA. RSV infection in the preterm lung was characterized by increased epithelial thickening and periodic acid-Schiff staining, indicative of glycogen retention. Nitric oxide levels were decreased in lungs of infected preterm lambs compared to full-term lambs, indicating alternative macrophage activation. Although infection induced significant neutrophil recruitment into the lungs of preterm lambs, neutrophils produced less myeloperoxidase than those of full-term lambs, suggesting decreased functional activation.

Conclusions

Taken together, our data suggest that increased RSV load and inadequate immune response may contribute to the enhanced disease severity observed in the lungs of preterm lambs.     

Low-molecular-weight heparin reduces hyperoxia-augmented ventilator-induced lung injury via serine/threonine kinase-protein kinase B

Introduction

Factors explaining the greater susceptibility of preterm infants to severe lower respiratory infections with respiratory syncytial virus (RSV) remain poorly understood. Fetal/newborn lambs are increasingly appreciated as a model to study key elements of RSV infection in newborn infants due to similarities in lung alveolar development, immune response, and susceptibility to RSV. Previously, our laboratory demonstrated that preterm lambs had elevated viral antigen and developed more severe lesions compared to full-term lambs at seven days post-infection. Here, we compared the pathogenesis and immunological response to RSV infection in lungs of preterm and full-term lambs.

Methods

Lambs were delivered preterm by Caesarian section or full-term by natural birth, then inoculated with bovine RSV (bRSV) via the intratracheal route. Seven days post-infection, lungs were collected for evaluation of cytokine production, histopathology and cellular infiltration.

Results

Compared to full-term lambs, lungs of preterm lambs had a heightened pro-inflammatory response after infection, with significantly increased MCP-1, MIP-1α, IFN-γ, TNF-α and PD-L1 mRNA. RSV infection in the preterm lung was characterized by increased epithelial thickening and periodic acid-Schiff staining, indicative of glycogen retention. Nitric oxide levels were decreased in lungs of infected preterm lambs compared to full-term lambs, indicating alternative macrophage activation. Although infection induced significant neutrophil recruitment into the lungs of preterm lambs, neutrophils produced less myeloperoxidase than those of full-term lambs, suggesting decreased functional activation.

Conclusions

Taken together, our data suggest that increased RSV load and inadequate immune response may contribute to the enhanced disease severity observed in the lungs of preterm lambs.     

Effect of artificial surfactant on pulmonary function in preterm and full-term lambs

We hypothesized that agents very different from surfactant may still support lung function. To test this hypothesis, we instilled FC-100, a fluorocarbon, and Tween 20, a detergent, which have higher minimum surface tensions and less hysteresis than surfactant, into 15 full-term and 14 preterm lambs. FC-100 and Tween 20 were as efficient as natural surfactant in improving gas exchange and compliance in preterm lambs with respiratory failure. Dynamic compliance correlated with the equilibrium surface tension of the alveolar wash in both full-term (P less than 0.02) and preterm (P less than 0.008) lambs. Functional residual capacity in full-term and preterm lambs was lower after treatment with the two test agents than with surfactant, findings consistent with qualitative histology. Oxygenation in full-term lambs correlated with mean lung volumes (P less than 0.003), suggesting that the hysteresis and/or low minimum surface tension of surfactant may improve mean lung volume, and hence oxygenation, by maintaining functional residual capacity. The effects of the test agents suggest that agents with biophysical properties different from surfactant may still aid lung expansion.     

Preterm birth in lambs: sleep patterns and cardio-respiratory changes.

Cardio-respiratory physiology in sleep was examined in eight preterm lambs born at 133-135 (134 +/- 1, mean SEM) days of gestation after 3-5 days of pulsatile ACTH/TRH infusion, and contrasted with eight lambs born at term (147 +/- 1 days). Lambs were instrumented with electrodes for recording electrocorticogram, electro-oculogram and nuchal electromyogram to define behavioural states, as well as carotid arterial catheters for determination of arterial pressure, heart rate and arterial blood gases. Compared to full-term lambs, the preterm lambs exhibited extended active sleep times, elevated PaCO2 and faster heart rate in all behavioural states than full-term lambs; with increasing postnatal age, sleep times and heart rate declined. As similar differences are found in preterm human infants, the preterm lamb will be a useful model to study the underlying physiology of these cardio-respiratory alterations.     

Effects of phrenic nerve section on the respiratory system of fetal lambs

We investigated the effects of phrenic nerve section (PNS) on the respiratory system of fetal lambs. Seven ewes, three of which had twin fetuses, were given a general anesthetic. The thoracic phrenic nerves were cut in two singleton fetuses and in one fetus in each set of twins (116-121 days); two singleton fetuses and one fetus in each set of twins underwent the same procedure except for PNS. Fetal arterial blood pressure, heart rate, and arterial pH and blood gas tensions were the same in both groups. Phrenic nerve section eliminated fetal breathing movements and decreased airway fluid volume, lung weight, and total lung DNA (P less than 0.05). However, PNS did not affect production of tracheal fluid or percent dry weight of the lungs. Furthermore, PNS did not affect the concentration of saturated phosphatidylcholine in the lung or its flux in tracheal fluid. We conclude that PNS in fetal lambs retards lung growth but does not affect tracheal fluid production or formation and release of surfactant.     

Relationship between respiratory pauses and heart rate during sleep in normal premature and full-term newborns

To investigate the relationship between central respiratory pauses and heart rate, we performed polygraphic recordings in 23 normal newborns (35 to 41 weeks conceptional age). We monitored the electroencephalogram, rapid eye movements, movements of the upper and lower limbs, chin and diaphragmatic electromyogram, electrocardiogram, thoracic and abdominal respiratory movements, air flow and transcutaneous PO2. Heart rate changes were analysed by computer measurement of R-R intervals and by cardiotachography. Respiratory pauses occurring after body movements and those not preceded by movements were studied separately. We analysed 1128 respiratory pauses greater than 3 s duration. No respiratory pause lasted more than 12 s. Independently of age, sleep state and respiratory pause duration, heart rate was significantly lower at the onset of respiratory pause, compared to control periods (selected away from the pause: 10 s before its onset and 20 s after its end). Heart rate slowed still further through the respiratory pause and reverted toward the baseline level after its end. When no movements preceded the respiratory pause, heart rate just before the pause was lower compared to control periods. These findings suggest the existence of simultaneous central commands responsible for both respiratory pause and heart rate deceleration.     

The interaction of the upper airway and thermo-metabolism on respiratory rhythm during non-REM sleep in the developing lamb.

This study investigates the role of metabolic rate and of vagal airway mechanisms in sustaining rhythmic breathing in the developing lamb. Fifteen lambs were prepared, at 2 days of age under fluothane anaesthesia, for sequential studies at 4, 14, 30, 45, and 55 days of age. At each age they were maintained at an ambient temperature of 5, 10, 15, 20, 25 and 30 degrees C for at least one hour before measurements were made during N-REM sleep. In 6 lambs at 4 days and in all lambs at older ages the upper airway was by-passed (by opening a tracheal window) for 10-15 minutes at each ambient temperature. Oxygen consumption was unaffected by upper airway by-pass and there were no consistent changes in mean breathing frequency or amplitude, with the exception of shifts to panting at warm ambient temperatures. Breathing pattern was unaffected by upper airway by-pass in lambs at 4 days of age, but at older ages loss of regularity of breathing frequently occurred (up to 47% of 30 days-old lambs at 25 degrees C). This was related to the fall in oxygen consumption with age and to basal values at thermoneutrality, and coincided with lower respiratory rates and increased use of expiratory laryngeal braking. Periodic breathing (and apnea) of a fixed cycle length (9.3 +/- 0.36 s) was a common feature (62%) of the observed breathing dysrhythmia. In young lambs high metabolic rate sustained high frequency rhythmic breathing which was unaffected by upper airway by-pass.(ABSTRACT TRUNCATED AT 250 WORDS)     

Elevated body temperature enhances the laryngeal chemoreflex in decerebrate piglets.

Hyperthermia and reflex apnea may both contribute to sudden infant death syndrome (SIDS). Therefore, we investigated the effect of increased body temperature on the inhibition of breathing produced by water injected into the larynx, which elicits the laryngeal chemoreflex (LCR). We studied decerebrated, vagotomized, neonatal piglets aged 3-15 days. Blood pressure, end-tidal CO(2), body temperature, and phrenic nerve activity were recorded. To elicit the LCR, we infused 0.1 ml of distilled water through a polyethylene tube passed through the nose and positioned just rostral to the larynx. Three to five LCR trials were performed with the piglet at normal body temperature. The animal's core body temperature was raised by approximately 2.5 degrees C, and three to five LCR trials were performed before the animal was cooled, and three to five LCR trials were repeated. The respiratory inhibition associated with the LCR was substantially prolonged when body temperature was elevated. Thus elevated body temperature may contribute to the pathogenesis of SIDS by increasing the inhibitory effects of the LCR.     

Coordination of breathing, sucking, and swallowing during bottle feedings in human infants

Incoordination of sucking, swallowing, and breathing might lead to the decreased ventilation that accompanies bottle feeding in infants, but the precise temporal relationship between these events has not been established. Therefore, we studied the coordination of sucks, swallows, and breaths in healthy infants (8 full-term and 5 preterm). Respiratory movements and airflow were recorded as were sucks and swallows (intraoral and intrapharyngeal pressure). Sucks did not interrupt breathing or decrease minute ventilation during nonnutritive sucking. Minute ventilation during bottle feedings was inversely related to swallow frequency, with elimination of ventilation as the swallowing frequency approached 1.4/s. Swallows were associated with a 600-ms period of decreased respiratory initiation and with a period of airway closure lasting 530 +/- 9.8 (SE) ms. Occasional periods of prolonged airway closure were observed in all infants during feedings. Respiratory efforts during airway closure (obstructed breaths) were common. The present findings indicate that the decreased ventilation observed during bottle feedings is primarily a consequence of airway closure associated with the act of swallowing, whereas the decreased ventilatory efforts result from respiratory inhibition during swallows.     

Influence of CPAP on reflex responses to tracheal irritation in anesthetized humans

We investigated the effects of lung inflation during continuous positive airway pressure breathing (CPAP) on airway defensive reflexes in 10 enflurane-anesthetized spontaneously breathing humans. The airway defensive reflexes were induced by instillation into the trachea of 0.5 ml of distilled water at two different levels of end-expiratory pressure (0 and 10 cmH2O CPAP). The tracheal irritation at an end-expiratory pressure of 0 cmH2O caused a variety of reflex responses including apnea, spasmodic panting, expiration reflex, cough reflex, an increase in heart rate, and an increase in blood pressure. Lung inflation during CPAP of 10 cmH2O did not exert any influence on these reflex responses in terms of the types, latencies, and durations of reflex responses although the intensity of the expiration reflex and cough reflex was augmented by lung inflation. Our results suggest that the pulmonary stretch receptors do not play an important role in the mechanisms of airway defensive reflexes in humans.     

Brain water and electrolytes in response to respiratory acidosis and brain edema in the mutant mouse,Jimpy

Compared to littermate controls, unstressed Jimpy mice have higher brain water, sodium, potassium and chloride contents and lower carbonic anhydrase activity. When stressed by CO₂ to produce a respiratory acidosis or by injection of distilled water to produce brain edema, the Jimpy mouse brain has water and ionic responses essentially like those in controls.