Inspiratory Off-switch Mechanism Research Articles

Compensatory airway dilation and additive ventilatory augmentation mediated by dorsomedial medullary 5-hydroxytryptamine 2 receptor activity and hypercapnia

5-HT2 receptor activity in the hypoglossal nucleus and hypercapnia is associated with airway dilation. 5-HT neurons in the medullary raphe and hypercapnia are responsible for tidal volume change. In this study, the effects of 5-HT2 receptors in the dorsomedial medulla oblongata (DMM), which receives projections from the medullary raphe, and hypercapnia on airway resistance and respiratory variables were studied in mice while monitoring 5-HT release in the DMM. A microdialysis probe was inserted into the DMM of anesthetized adult mice. Each mouse was placed in a double-chamber plethysmograph. After recovery from anesthesia, the mice were exposed to stepwise increases in CO(2) inhalation (5%, 7%, and 9% CO(2) in O(2)) at 8-min intervals with a selective serotonin reuptake inhibitor, fluoxetine, or fluoxetine plus a 5-HT2 receptor antagonist, LY-53857 in the DMM. In response to fluoxetine plus LY-53857 coperfusion, specific airway resistance was increased, and tidal volume and minute ventilation were decreased. CO(2) inhalation with fluoxetine plus LY-53857 coperfusion in the DMM largely decreased airway resistance and additively increased minute ventilation. Thus, 5-HT2 receptor activity in the DMM increases basal levels of airway dilation and ventilatory volume, dependent on central inspiratory activity and the volume threshold of the inspiratory off-switch mechanism. Hypercapnia with low 5-HT2 receptor activity in the DMM largely recovers airway dilation and additively increases ventilatory volume. Interaction between 5-HT2 receptor activity in the DMM and CO(2) drive may elicit a cycle of hyperventilation with airway dilation and hypoventilation with airway narrowing.

Effects of intertrigeminal region NMDA and non-NMDA receptors on respiratory responses in rats

Respiratory disturbance, including apnea, can be induced by microinjection of glutamate into the intertrigeminal region (ITR) of the lateral pons, a region that is anatomically coupled to both the dorsal and ventral respiratory groups of the medulla. We showed that the ITR plays a functional role in regulating both vagal reflex apnea and spontaneous sleep-related apnea in rats, but the mechanisms have not been determined. This study shows that functional NMDA receptors are expressed in the ITR since the blockade of these receptors by AP5, a specific NMDA receptor antagonist, was fully effective in blocking apnea induced by glutamate injection within this region. Selective blockade of ITR NMDA receptors had no effect on the immediate apnea evoked by an intravenous 5-HT bolus, whereas the nonspecific glutamate receptor antagonist kynurenic acid significantly increased the duration of this vagal reflex apnea. These findings are of interest because pontine NMDA receptors participate in inspiratory off-switch mechanisms and have been implicated in various short- and long-term potentiation and depression phenomena. These data support the involvement of ITR non-NMDA receptors in modulation of reflex apnea per se, whereas NMDA receptors play a role in damping respiratory responses to transient disturbances.

The Kölliker‐Fuse nucleus gates the postinspiratory phase of the respiratory cycle to control inspiratory off‐switch and upper airway resistance in rat

Lesion or pharmacological manipulation of the dorsolateral pons can transform the breathing pattern to apneusis (pathological prolonged inspiration). Apneusis reflects a disturbed inspiratory off-switch mechanism (IOS) leading to a delayed phase transition from inspiration to expiration. Under intact conditions the IOS is irreversibly mediated via activation of postinspiratory (PI) neurons within the respiratory network. In parallel, populations of laryngeal premotoneurons manifest the IOS by a brief glottal constriction during the PI phase. We investigated effects of pontine excitation (glutamate injection) or temporary lesion after injection of a GABA-receptor agonist (isoguvacine) on the strength of PI-pool activity determined from respiratory motor outputs or kinesiological measurements of laryngeal resistance in a perfused brainstem preparation. Glutamate microinjections into distinct parts of the pontine Kölliker-Fuse nucleus (KF) evoked a tonic excitation of PI-motor activity or sustained laryngeal constriction accompanied by prolongation of the expiratory phase. Subsequent isoguvacine microinjections at the same loci abolished PI-motor or laryngeal constrictor activity, triggered apneusis and established a variable and decreased breathing frequency. In summary, we revealed that excitation or inhibition of defined areas within the KF activated and blocked PI activity and, consequently, IOS. Therefore, we conclude, first, that descending KF inputs are essential to gate PI activity required for a proper pattern formation and phase control within the respiratory network, at least during absence of pulmonary stretch receptor activity and, secondly, that the KF contains large numbers of laryngeal PI premotor neurons that might have a key role in the regulation of upper airway resistance during reflex control and vocalization.

Intracellular neuronal recordings form the ventral respiratory group (VRG) and region of the pontine Kölliker‐Fuse nucleus (KF) during vagal entrainment of breathing in rat.

Medullary parts of the respiratory network were associated with respiratory rhythm generation, while pontine parts play a role in secondary modulations of breathing. Recently we identified the KF as key structure that synchronise afferent and network mediated inspiratory off-switch mechanisms (IOS) in response to vagal stimulations (X-stim.). However, potential changes or reconfiguration of the activity patterns of respiratory neurones during synchronisation of IOS are unknown. We performed intracellular recordings to identify changes in neuronal activities during the synchronisation in the perfused brainstem preparation of rats. In the VRG we recorded from 33 respiratory neurones before, during and after repetitive trails of X-stim. Only postinspiratory (PI) and late-I neurones had a clear excitatory response to rhythmic X-stim., while the remaining sub-types of neurones were in summary inhibited. PI and late-I neurones were rhythmically activated by X inputs, but maintained bursting patterns independent of the X-stim. This independent bursting was related to the respiratory rhythm before X-stim. In contrast, recordings of KF neurones (n=12) never showed such stimulus independent bursting. We suggest that the independent bursting of VRG neurones is related to the pontine inputs illustrating the potential interference of the two IOS mechanisms on the cellular level. Funded by BCCN Göttingen

Central histamine contributes to the inspiratory off-switch mechanism via H1 receptors in mice

Central histaminergic neurons are distributed in areas of the medulla and pons concerned with respiratory rhythm generation, but their effects on breathing pattern are unknown. We examined breathing pattern during hypercapnic responses in wild type (WT) and H1 receptor knockout (H1RKO) mice at 9-10 weeks of age before and after vagotomy. Minute ventilation increased with PaCO(2) increase equally in both genotypes; respiratory rate response was lower and tidal volume (V(T)) response higher in H1RKO mice than in WT mice. The V(T)-inspiratory time (T(I)) relation during hypercapnia was hyperbolic in both groups, with the curve in H1RKO mice shifted right-upward. After vagotomy, the V(T)-T(I) relation was a vertical line, which shifted right in H1RKO mice. We conclude that alterations of inspiratory off-switch and respiratory rhythm generation change breathing pattern without affecting central chemosensitivity in H1RKO. Histamine might affect breathing pattern centrally via H1 receptors.

Episodic phrenic-inhibitory vagus nerve stimulation paradoxically induces phrenic long-term facilitation in rats

Accumulating evidence suggests that the respiratory control system exhibits an impressive degree of plasticity, as do many other neural networks in the central nervous system (Eldridge & Millhorn, 1986; McCrimmon et al. 1995; Ling et al. 1997a; Powell et al. 1998; Mitchell et al. 2001). For example, episodic hypoxia induces a persistent augmentation of respiratory activity (Cao et al. 1992; Bach & Mitchell, 1996; Turner & Mitchell, 1997; Olson et al. 2001; McGuire et al. 2002), known as long-term facilitation (LTF). Episodic carotid sinus nerve (CNS) stimulation (CSNS) also elicits phrenic LTF in anaesthetized animals (Millhorn et al. 1980a; Hayashi et al. 1993; Ling et al. 1997b), which is not abolished by decerebration or spinal transection at the C7-T1 level (Eldridge & Millhorn, 1986). These results suggest that LTF is elicited by central mechanisms located in the brainstem and/or cervical spinal cord, and that the carotid body, respiratory mechanics, systemic hypoxia, forebrain and the lower spinal cord are not necessary for its expression (Eldridge & Millhorn, 1986). Both hypoxia- and CSNS-induced LTF are serotonin-dependent (Millhorn et al. 1980b; Bach & Mitchell, 1996). Recent evidence further suggests that spinal serotonin receptors (Baker-Herman & Mitchell, 2002) and the synapses transmitting bulbospinal, inspiratory drive to the phrenic motoneurons (Fuller et al. 2002) play key roles in phrenic LTF. In these experiments, respiratory LTF is always preceded by repeated inspiratory augmentation. This fact suggests that induction of respiratory LTF is somehow related to inspiratory augmentation, and more specifically, that phrenic LTF relies heavily on an activity-dependent Hebbian mechanism (coincident pre- and post-synaptic activity strengthens synapses) in the synapses on the phrenic motoneurons. However, as all LTF to date has been induced by episodic inspiratory-excitatory stimulation, it has been difficult to directly test these hypotheses by separating the LTF from inspiratory augmentation. In contrast, vagus nerve (VN) stimulation (VNS) suppresses inspiration during stimulation, and induces a reduction (< 1 min duration) in phrenic amplitude and frequency after stimulation (0.5 min duration; Eldridge & Millhorn, 1986). We speculate that a relatively longer post-stimulation inhibitory memory is possible if using episodic and longer VNS. VNS has also been used as a tool in brain research (e.g. evoked potentials recorded from the cerebral cortex, hippocampus, thalamus and cerebellum; Rutecki, 1990) and neurophysiological studies of several reflexes (e.g. cough, swallow and Hering-Breuer reflex or inspiratory off-switch mechanisms) because vagal afferents provide an easily accessible, peripheral route by which to modulate the central nervous system function. However, these investigators all used brief VNS and focused mainly on immediate or short-term (< several min) effects, during and/or after VNS. For many years, episodic VNS has been used clinically as a common treatment for patients with medically intractable epilepsy (Schachter, 2002). However, the precise underlying mechanisms and the consequence of long-term VNS remain unclear. The aims of the present study were to (1) examine the long-term effects of episodic phrenic-inhibitory VNS on phrenic nerve activity and compare them with those elicited by phrenic-excitatory CSNS and (2) explore the possibility of using VNS as a tool to suppress phrenic motor neurons during LTF elicitation. We hypothesized that episodic VNS would induce phrenic long-term depression, but that phrenic activity would eventually return to baseline in less than 60 min.

The effects of locomotor-respiratory coupling on the pattern of breathing in horses.

1. To investigate the effect of locomotor activity on the pattern of breathing in quadrupeds, ventilatory response was studied in four healthy horses during horizontal and inclined (7%) treadmill exercise at different velocities (1.4-6.9 m s(-1)) and during chemical stimulation with a rebreathing method. Stride frequency (f(s)) and locomotor-respiratory coupling (LRC) were also simultaneously determined by means of video recordings synchronized with respiratory events. 2. Tidal volume (V(T)) was positively correlated with pulmonary ventilation (V(E)) but significantly different linear regression equations were found between the experimental conditions (P < 0.0001), since the chemical hyperventilation was mainly due to increases in V(T), whereas the major contribution to exercise hyperpnoea came from changes in respiratory frequency (f(R)). 3. The average f(R) at each exercise level was not significantly different from f(S), although there was not always a tight 1:1 LRC. At constant speeds, f(S) was independent of the treadmill slope and hence the greater V(E) during inclined exercise was due to increased V(T). 4. At any ventilatory level, the differences in breathing patterns between locomotion and rebreathing or locomotion at different slopes derived from different set points of the inspiratory off-switch mechanism. 5. The percentage of single breaths entrained with locomotor rhythm rose progressively and significantly with treadmill speed (P < 0.0001) up to a 1:1 LRC and was significantly affected by treadmill slope (P < 0.001). 6. A LRC of 1:1 was systematically observed at canter (10 out of 10 trials) and sometimes at trot (5 out of 14) and it entailed (i) a 4- to 5-fold reduction in both V(T) and f(R) variability, and (ii) a gait-specific phase locking of inspiratory onset during the locomotor cycle. 7. It is concluded that different patterns of breathing are employed during locomotion and rebreathing due to the interference between locomotor and respiratory functions, which may play a role in the optimization and control of exercise ventilation in horses.

Significance of airway resistance for the pattern of breathing and lung volumes in exercising humans

The effects of increased airway resistance on lung volumes and pattern of breathing were studied in eight subjects performing leg exercise on a cycle ergometer. Airway resistance was changed 1) by increasing the density (D) of the respired gas by a factor of 4.2 and changing the inspired gas from O2 at 1.3 bar to air at 6 bar and 2) by increasing airway flow rates by exposing the subjects to incremental work loads of 0-200 W. Increased gas D caused a slower and deeper respiration at rest and during exercise and, at work loads greater than 120 W, depressed the responses of ventilation and mean inspiratory flow. Raised airway resistance induced by increases in D and/or airway flow rates altered respiratory timing by increasing the ratio of inspiratory time (TI) to total breath duration. Furthermore, analyses of the relationships between tidal volume and TI and between end-inspiratory volume and TI revealed elevation of Hering-Breuer inspiratory volume thresholds. We propose that this elevation, and hence exercise-induced increases of tidal volume, can largely be explained by previous observations that the threshold of the inspiratory off-switch mechanisms depends on central inspiratory activity (cf. C. von Euler, J. Appl. Physiol. 55: 1647-1659, 1983), which in turn increases with airway resistance (Acta Physiol. Scand. 120: 557-565, 1984).

The pattern of breathing during hypoxic exercise.

Breathing pattern was studied in six subjects in normoxia (FIO2 = 0.21) and hypoxia (FIO2 = 0.12) at rest and during incremental work-rate exercise. Ventilation (V) as well as mean inspiratory flow (VT/TI) increased with exercise intensity and were augmented in the hypoxic environment, whereas the ratio between inspiratory (TI) and total (Ttot) breath durations increased with exercise intensity but was unaffected by hypoxia. The relationship of tidal volume (VT) and inspiratory time duration (TI) showed linear, coinciding ranges for the normoxic and hypoxic conditions up to VT/TI values of about 2.5 1.s-1. At higher VT/TI values TI continued to decrease, whereas VT tended to level off, an effect which was more evident in the hypoxic condition. The results suggest that the hypoxic augmentation of exercise hyperpnea is primarily brought about by an enhancement of central inspiratory drive, the timing component being largely unaffected by the hypoxic environment, and that at low to moderate levels of exercise hyperpnea inspiratory off-switch mechanisms are essentially unaffected by moderate hypoxia.

Effects of graded focal cold block in the solitary and para-ambigual regions of the medulla in the cat.

Unilateral focal cold blocks in the region of the nucleus tractus solitarius and the dorsal respiratory group of neurons, DRG, of anaesthetized cats consistently caused apneustic-type breathing. There was no concomitant change in the initial rate of rise of inspiratory activity. The apneustic prolongation of inspiratory duration, TI, was most pronounced in, but was not confined to, the DRG. The apneustic effects were more marked after vagotomy. In cats with intact vagus nerves being given artificial ventilation, focal cooling at certain sites of the DRG region could produce 'unlocking' of the respiratory rhythm from that of the respiratory pump. At other sites in this region, focal cooling could selectively block the effects of the inspiration-facilitating reflex induced by deflation without blocking the inspiration-inhibiting Hering-Breuer reflex. Unilateral focal cold blocks in the region of the intermediate part of the ventral respiratory group of neurons, VRG, generally caused depression of the rate of rise of inspiratory activity, but almost never apneustic effects. All effects of unilateral focal cooling both in the DRG and VRG were bilaterally symmetrical. No systematic differences between the effects on phrenic and external intercostal inspiratory activity were found in response to focal cooling either of the DRG or VRG suggesting that differential control of phrenic and external intercostal motoneurons is not exerted mainly at the level of these medullary structures. The results suggest that the DRG and VRG areas exert somewhat different effects on the respiratory pattern: DRG appears to be more concerned with integration of vagal and other inputs contributing to the inspiratory off-switch mechanisms which, however, are not confined only to the DRG. The VRG inspiratory mechanisms, on the other hand, appear to be more involved in the gain control of the inspiratory output intensity.

Control of breathing at elevated lung volumes in anesthetized cats.

We monitored the steady-state ventilatory responses of anesthetized cats to increases in lung volume produced by expiratory threshold loads (ETL) to study the roles of peripheral and central neural mechanisms in controlling respiration at elevated lung volumes. Application of an ETL of 5 cmH2O produced a significant decrease in respiratory frequency (-18%) but no change in minute ventilation (VE) due to a significant increase in tidal volume (VT) (19.3%). The drop in frequency was due solely to an increase in expiratory duration. ETL of 10 cmH2O significantly reduced VE (-17.5%) for the same reason. VT was maintained or increased at elevated lung volumes due to both an increase in the rate of rise of phrenic activity and a maintenance of inspiratory duration (TI) despite increases in both chemical drive and pulmonary stretch receptor (PSR) activity. No PSR adapted completely to the maintained change in lung volume. The sensitivity of the inspiratory off-switch mechanism to increases in lung volume, given by the reciprocal of the VT-TI relationship, decreased significantly during breathing on ETL. The results are consistent with the hypothesis that central habituation, not just peripheral adaptation of PSR, determines breathing pattern at elevated lung volumes.

Breuer-Hering inflation reflex and breathing pattern in anesthetized humans and cats.

In nine cats and nine human subjects anesthetized with alfaxalone, respiratory activity and tracheal pressure were recorded prior to and during occlusion of the airway at end inspiration or end expiration. Lung inflations at the end of expiration were also performed. In addition, the ventilatory pattern was analyzed during hypercapnia. The results show that occlusions at the end of inspiration or inflations provoked an apnea in both cats and humans. However, concomitant with increases in tidal volume during hypercapnia, inspiratory duration decreased in cats and did not change in human subjects. These results indicate that the Breuer-Hering reflex, which delays the onset of inspiration during inflation was equally operative in cats and humans. In contrast, the "Breuer-Hering threshold curve," which accounts for the off-switch" of inspiration was different in cats and humans. Thus, in summary, the Breuer-Hering inflation reflex is operative in human subjects, but it does not seem to be involved in the control of the inspiratory off-switch mechanism during increases respiratory activity resulting from hypercapnia.

Time course of intercostal afferent termination of the inspiratory process

The influence exerted by several somatic nerves on the inspiratory off-switch mechanism has been assessed in decerebrate cats. These animals were paralyzed, artificially ventilated, and bilaterally vagotomized. Inspiratory activity was monitored by a phrenic neurogram. Brief stimulation of either the superficial radial nerve or the sciatic nerve had an inconsistent effect on both the depth of inspiration and the timing of the respiratory cycle. However, stimulation of the T6 intercostal nerve during inspiration elicited a premature phase switch to expiration. Distinct, repeatable thresholds were determined for 10 delays in 100 msec increments after the onset of inspiration. As the delay increased, the threshold current was observed to decrease in all 30 decerebrate cats studied. An increase in the end-expiratory %CO 2 caused an elevation of the stimulus threshold. These results correspond to the known characteristics of the inspiratory off-switch. Also, since the intercostal afferents are not normally a major determinant of respiratory rhythmicity in eupnea, this work establishes intercostal nerve stimulation as a very useful technique in the study of inspiratory to expiratory phase switching mechanisms.

Changes in Ventilatory Pattern Induced by Intravenous Anesthetic Agents in Human Subjects

The tidal volume-inspiratory duration relationship was studied during air breathing and rebreathing in conscious and anesthetized human subjects using three different intravenous agents. The results observed have been compared with similar experiments carried out in cats. Its has been shown that anesthesia provokes an increase in breathing rate associated with a decrease in tidal volume in human subjects; an opposite effect on breathing rate was observed in cats. Thus, the tidal volume-inspiratory duration relationship, although very similar in the conscious cat or human subject, is very different under anesthesia. The results were quite consistent in a given species whatever the nature of the drug used. It is suggested that modifications of the breathing rate by anesthesia, related to animal species, are caused by central effects of the drug. These effects are probably mediated by different actions on inputs to the inspiratory off-switch mechanisms in the two species.

Regulation of ventilatory responses in vagotomized cats by caudal brainstem sites

Respiratory responses to hypercapnia and isocapnic hypoxia were evaluated in vagotomized decerebrate cats prior to and following lesions in the midline at the pontomedullary junction. Subsequent to these lesions, frequency responses to hypercapnia were significantly elevated whereas the concomitantly determined tidal volumes were significantly lowered. In the post-lesion phase, the durations of inspiration, expiration and the total respiratory cycle at maximum hypercapnia-induced tidal volumes were lower than comparable values at maximum tidal volumes in the pre-lesion phase. The same lesions produced no significant alteration of respiratory responses to hypoxia. Mesencephalic or rostral pontile lesions caused on systematic changes in either hypercapnia- or hypoxia-induced responses. It is concluded that the lesions at teh pontomedullary junction produce ventilatory alterations by interrupting a pathway interconnecting the caudal pontile apneustic center with the medullary respiratory complex. The possibility that this interruption produces an alteration in the threshold of the inspiratory off-switch mechanism of the brainstem respiratory controller is considered and discussed.

Properties of apneusis produced by reversible cold block of the rostral pons

Reversible cold block of the rostral pons was used to compare properties of normal and apneustic respiration in anesthetized, vagotomized, artificially ventilated cats. During apneusis we observed frequency oscillations (HFO) in phrenic nerve activity which were reduced in frequency compared with those during a normal inspiration. Apneusis produced by mid-pontine transection or punctate pneumotaxic center (PC) lesion produced similar HFO changes. The minimal intensity of superior laryngeal nerve electrical stimulation needed to terminate a breath was higher early in an apneusis than at the same time during a normal breath. Later in apneusis the intensity required became constant and was approximately the same as that needed to end a normal inspiration at its natural termination. With intact vagi lung inflation produced a greater prolongation of expiration during apneustic respiration than during normal respiration. Apneustic type activity was observed in both phrenic and vagal inspiratory motoneurons. We suggest that: (1) HFO are generated without the PC, but the PC elevates the oscillation frequency; and (2) apneusis may result in part from a delayed activation of the normal inspiratory off-switch mechanism.

Alteration from apneusis to more regular rhythmic respiration in decerebrate Cats

In spontaneously breathing or paralyzed, ventilated, decerebrate cats, bilateral pneumotaxic center lesions and bilateral vagotomy resulted in apneusis characterized by a marked prolongation of inspiratory duration ( Ti) and, concomitantly, an elevation of tidal volume (V t) and reduction of respiratory frequency (f). Subsequent placement of medial or lateral lesions in the caudal pons or medulla reduced T i and V t, increased f, and restored a more regular rhythmic respiratory pattern. Placement of these pontile or medullary lesions in animals with pneumotaxic center lesions prevented the development of a typical apneustic pattern upon vagotomy. Large rostral pontile lesions did not change apneusis in spontaneous breathing animals whereas, in paralyzed, ventilated cats, these lesions resulted in some decrease of T i. It is concluded that apneusis generation is not equa table simply with a summation of caudal pontile respiratory unit activities. Rather, activity inherent to intrapontile and/or pontomedullary feedback circuits is considered as a necessary requisite for apneusis development. Interaction of these circuits with an inspiratory off-switch mechanism is considered probable.

A model of the central and reflex inhibition of inspiration in the cat.

An attempt is made to summarize the results obtained in previous work from this and other laboratories on the steady state and transient relationships between the mechanical and neural events in breathing and their precise timing in the breathing cycle at different blood chemical demands for ventilation and at different body temperatures, and to fit these results into a functional and realistic model of the bulbo-pontine inspiratory off-switch mechanisms. The experimentally based requirements for the model are briefly described and listed. After a presentation of the model in qualitative terms its functional properties are considered quantitatively and compared with the performance of the real, biological system. This has been achieved by assuming some simple mathematical approximations for the activities of the introduced physiological parameters and their chemical “drive” dependence. The gaps in our present knowledge are pointed out and some key experiments suggested. The proposed model is consistent with the main conclusions reached in previous work from this laboratory that there are three neural submechanisms which are mainly responsible for the effects of increased CO2 on ventilation: 1) a rise in the inspiratory off-switch threshold, 2) an increased rate of rise of the centrally generated inspiratory activity that projects to the off-switch mechanism (and to the spinal respiratory motoneurons), and 3) the vagal volume feed-back. Of these 1) and 2) are mainly responsible for the increase in tidal volume, whereas the vagal volume feed-back is mainly responsible for the increase in respiratory rate. The comparison between the model behaviour and experimental data suggest that the slight CO2 sensitivity of the pulmonary stretch receptors recently reported on, has to be taken into account. The model studies have suggested the increase in respiratory rate with increased temperature may be due either to an increased rate of rise of inspiratory activity or to a decreased off-switch threshold, or both in combination. The mechanism controlling the expiratory durations are briefly discussed.