Inhibitory and excitatory effects on respiration by phrenic nerve afferent stimulation in cats

Respiratory effects of electrical stimulation of phrenic nerve afferents were studied in anesthetized cats, either spontaneously breathing or paralyzed and ventilated. The type of phrenic afferent fibers activated was controlled by recording the evoked action potentials from dorsal root fibers. In both preparations, stimulation at a strength sufficient to activate small diameter myelinated phrenic nerve afferents induced a biphasic response. The first phase lasted a few respiratory cycles and was inhibitory and consisted of a decrease in tidal volume (VT) or phrenic activity (NA), inspiratory time (TI), respiratory duty cycle (TI/Ttot) and instantaneous ventilation (VE) or minute phrenic activity (NMA). Expiratory time (TE) increased and breathing frequency (f) and mean inspiratory flow (VT/TI) or mean inspiratory neural activity (NA/TI) did not change. This short-term response was suppressed in animals pretreated with bicuculline. The second phase was a long-term excitation in which VT or NA, f, VE or NMA and VT/TI increased whereas both TI and TI/Ttot decreased and TE did not change. Unlike published data, our results suggest that small-diameter myelinated phrenic nerve afferents are involved in these responses. These phrenic fibers, like afferents from other muscles, affect respiratory output and may play a role in the control of breathing.     

Phrenic afferent input to the lateral medullary reticular formation of the cat

The afferent inputs from phrenic nerve stimulation to the lateral reticular formation of the lower brain stem were studied in anesthetized spontaneously breathing cats. The activity of reticular neurons was recorded by means of extracellular tungsten microelectrodes. Electrical stimulation of the central end of the right phrenic nerve evoked excitatory or inhibitory responses in the lateral reticular nucleus (LRN), in the nucleus ambiguus (AMB) and in a region dorsal to the AMB of ipsi- and contralateral sides. Phrenic afferents belonging to the flexor reflex afferent group were involved in these responses. The discharge pattern of the respiratory related units (RRU) of the AMB were exceptionally affected by phrenic nerve stimulations. It is concluded that high threshold phrenic afferents relay in the LRN before projecting to the cerebellar cortex. The overlapping of respiratory and non-respiratory afferents in the reticular formation may participate to the adaptations of respiratory and somatomotor functions during specific behaviors.     

Influence of lung volume on respiratory responses to spontaneous bladder contractions

Spontaneous bladder contractions (SBCs) in decerebrate, vagotomized, paralyzed, ventilated cats have been shown to decrease phrenic and hypoglossal inspiratory nerve activities, as well as the activities of other respiratory motor nerves. To determine whether vagal afferents from the lung influence the respiratory inhibition associated with SBCs, we recorded phrenic and hypoglossal nerve activities in decerebrate, paralyzed, vagally intact cats. The animals were ventilated by a servo-respirator, which inflated the lungs in accordance with integrated phrenic nerve activity. Maintained increases in end-expiratory lung volume were produced by the application of 2–10 cm H₂O positive end-expiratory pressure (PEEP). SBCs were accompanied by decreases in both phrenic and hypoglossal peak integrated nerve activities, as well as by marked decreases in respiratory frequency. The reduction of respiratory frequency was greater with higher levels of PEEP, a few animals becoming apneic during SBCs. After bilateral vagotomy, SBCs continued to decrease phrenic and hypoglossal peak integrated nerve activities as previously reported, but the reduction of respiratory frequency was much less striking than when the vagi were intact. These results indicate that activity of vagal afferents from the lung augments the respiratory influence of SBCs. Furthermore, SBCs in vagally intact animals can induce periodic breathing.     

Selective depression by ethanol of upper airway respiratory motor activity in cats

We studied the effects of systemically administered ethyl alcohol on the respiratory motor activity of the phrenic, hypoglossal and recurrent laryngeal nerves in unanesthetized, decerebrate cats. Some of the cats were studied after carotid sinus nerve section. In addition, parallel studies were done in intact, awake cats with chronic electromyographic electrodes in the diaphragm, genioglossus, and posterior cricoarytenoid (PCA) muscles. In decerebrate animals, alcohol induced a significant reduction of hypoglossal and recurrent laryngeal nerve activities at doses that had little or no effect on the phrenic nerve discharge. Similar changes were observed in chemodenervated cats. In awake animals, genioglossal and PCA muscle activities were depressed by alcohol, whereas diaphragm activity showed no consistent change. Alcohol caused a significant increase in respiratory frequency in awake cats and reduced the responses of genioglossal and PCA muscle activities to hypercapnia and normocapnic hypoxia. We conclude that alcohol induces a selective reduction in upper airway respiratory motor activity by an action that does not require intact suprapontile structures, vagal afferents, or peripheral chemoreceptors. This reduction may contribute to the alcohol-induced exacerbation of obstructive sleep apnea.     

Thoracic dorsal rhizotomy in the anesthetized cat: maintenance of eupnic breathing

In order to assess the level of participation of thoracic afferents in the determination of the eupnic breathing pattern, thoracic dorsal rhizotomies from T1 through T13 were performed in cats anesthetized with alpha-chloralose or sodium pentobarbital, with or without intact vagi. While recording several respiratory parameters, no consistent changes in the spontaneous breathing pattern could be demonstrated either 10--20 min or 90--100 min following disruption of thoracic afferent input. Thus, respiratory frequency, tidal volume and total ventilation were unaffected by rhizotomy. Inspiratory (TI) and expiratory (TE) durations and TI/TE ratios remained constant. Tracheal air flows during inspiration and expiration and intrapleural pressure were not altered. Finally, end-expiratory %CO2 and %O2 did not change after section of the thoracic dorsal roots. Only arterial blood pressure was found to fall as a function of time into the experiment. Since the results were identical in vagotomized and in vagi intact preparations, thoracic reflexes were not being masked by vagal dominance in eupnea. Rather, thoracic afferents, including intercostal muscle spindle afferents, appear to be ineffective in shaping the pattern of quiet breathing in anesthetized cats. This is unlike situations of loaded or stressed breathing where these afferents assume a more active role in the modulation of rate and depth mechanisms.     

Phrenic afferents and ventilatory control

It has been understood since the late 1800s that the diaphragm has significant sensory innervation. The role of phrenic afferents in the control of breathing, however, has been obscure. The phrenic nerve has been shown to contain a full array of afferent fibers. However, proprioceptive (group 1 fibers) afferents are few compared to postural muscles or the intercostals. The diaphragm, unlike the inspiratory intercostal muscles, has a small complement of spindle afferents and not all of these spindles are supplied with fusorial fibers. Reduced spindle afferents under gamma control help to explain previous studies of the diaphragm that have failed to reveal autogenic facilitation, that is, a reflex-mediated increase in drive during inspiratory loading. Nevertheless, some clinical studies have revealed increased activation of the diaphragm when its length is reduced. Group 1 fibers, which are predominantly tendon organ afferents in the diaphragm, have been shown to have a phasic inhibitory function. A reduction in this inhibition brought about by a reduction in diaphragmatic length during lung inflation may explain the increased diaphragmatic activation reported in clinical studies. Phrenic afferents have been shown to have multiple spinal and supraspinal projections. Recent studies have explored the ventilatory effects of thin fiber afferents (group III and IV fibers) in the phrenic nerve. Stimulation of these afferents has been shown both to inhibit and excite ventilation. These afferents arise from polymodal receptors that respond to both mechanical and chemical stimulation. Activation of these receptors may occur in a variety of conditions and the ventilatory response may be determined by the specific receptor activated.     

Phrenic afferents and ventilatory control

It has been understood since the late 1800s that the diaphragm has significant sensory innervation. The role of phrenic afferents in the control of breathing, however, has been obscure. The phrenic nerve has been shown to contain a full array of afferent fibers. However, proprioceptive (group 1 fibers) afferents are few compared to postural muscles or the intercostals. The diaphragm, unlike the inspiratory intercostal muscles, has a small complement of spindle afferents and not all of these spindles are supplied with fusorial fibers. Reduced spindle afferents under gamma control help to explain previous studies of the diaphragm that have failed to reveal autogenic facilitation, that is, a reflex-mediated increase in drive during inspiratory loading. Nevertheless, some clinical studies have revealed increased activation of the diaphragm when its length is reduced. Group 1 fibers, which are predominantly tendon organ afferents in the diaphragm, have been shown to have a phasic inhibitory function. A reduction in this inhibition brought about by a reduction in diaphragmatic length during lung inflation may explain the increased diaphragmatic activation reported in clinical studies. Phrenic afferents have been shown to have multiple spinal and supraspinal projections. Recent studies have explored the ventilatory effects of thin fiber afferents (group III and IV fibers) in the phrenic nerve. Stimulation of these afferents has been shown both to inhibit and excite ventilation. These afferents arise from polymodal receptors that respond to both mechanical and chemical stimulation. Activation of these receptors may occur in a variety of conditions and the ventilatory response may be determined by the specific receptor activated.     

Properties of inspiratory termination by superior laryngeal and vagal stimulation.

Electrical stimulation of two respiratory afferent nerves, the vagus and the internal branch of the superior laryngeal, was used to terminate inspiration. The short latency responses of phrenic motoneurones to these stimuli were studied to determine if inspiratory termination was preceded by a characteristic phrenic motoneurone discharge pattern, reflecting changes in brainstem inspiratory neurone discharge and inspiratory terminating mechanisms. Stimulus trains of sufficient intensity delivered to the superior laryngeal nerve terminated inspiration within 50 ms and were preceded by a stereotyped pattern of phrenic motoneurone discharge. This consisted of a short latency (disynaptic), predominantly contralateral excitation in response to the first shock of the train, followed by a marked and long lasting inhibition. In contrast, vagal stimulation typically terminated inspiration hundreds of milliseconds after the onset of the stimulus train and was not preceded by a stereotyped pattern of phrenic motoneurone responses to single shocks. Transient short latency responses were obtained but were extremely small, requiring considerable excitation followed by a moderate bilateral depression of activity. Inspiration could be terminated with or without the presence of these short latency responses. These results indicate that superior laryngeal and vagal (presumably pulmonary stretch receptor) afferents have different projections to brainstem inspiratory neurones and may exert their effects on inspiratory duration through different, but as yet undefined, neural mechanisms.     

Functions of the retrofacial nucleus in chemosensitivity and ventilatory neurogenesis

The hypothesis was evaluated that neurons within the retrofacial nucleus of medulla integrate afferent stimuli from the central chemoreceptors. In decerebrate, vagotomized, paralyzed and ventilated cats, activity of the phrenic nerve was monitored. Peak integrated phrenic activity increased in hypercapnia; the frequency of phrenic bursts typically declined slightly. The retrofacial nucleus was ablated by radio-frequency lesions or neurons within this nucleus were destroyed by microinjections of kainic acid. Results were similar following lesions or injections. Following unilateral ablations, peak phrenic activity was greatly reduced at normocapnia and hypercapnia; the frequency of phrenic bursts typically rose. Both frequency and peak phrenic activity fell further after the contralateral destruction with a cessation of all phasic phrenic discharge being observed in most animals. Injections of kainic acid in regions rostral, caudal or medial to the retrofacial nucleus produced no consistent changes in phrenic activity. We conclude that neuronal activities in the region of the retrofacial nucleus are important both in the integration of stimuli from the central chemoreceptors and in defining the discharge patterns of respiratory neurons.     

Influence of extreme hypercapnia on respiratory motor nerve activity in cats

Sedative drugs have been found to depress the respiratory activity of upper airway muscles more than that of the diaphragm. To determine whether CO2 at narcotic levels has a similar action, we recorded phrenic and hypoglossal nerve activities in decerebrate, vagotomized, paralyzed cats. T5 or T6 external intercostal nerve activity was also recorded in some animals. End-tidal CO2 concentration was raised progressively to over 30% or until depression of nerve activity was apparent. Respiratory frequency was reduced by severe hypercapnia in most cats. Hypoglossal nerve activity was consistently decreased more than that of the phrenic nerve. In most cases intercostal nerve activity was also more susceptible than phrenic nerve activity to hypercapnic depression. The results indicate that CO2 at narcotic levels interferes both with the central pattern generator for breathing movements and with the expression of the pattern in specific motor nerves.     

Respiratory resetting induced by activation of inspiratory bulbo-spinal neurons

The respiratory effects elicited by spinal (C2-C3) stimulation at the level of descending inspiratory axons were studied in paralysed, non-vagotomized and artificially ventilated cats anaesthetized with urethane-chloralose. The activation of inspiratory bulbospinal axons in the ventrolateral quadrant was confirmed by recording the ipsilateral phrenic excitation following a single pulse. Brief stimulus trains delivered at the same locus during expiration elicited short- and long-term phrenic activations. The short-term activation consisted of a tetanic orthodromic response. The long-term activation, of central origin, exhibited the same pattern as a spontaneous inspiration and consisted of an inspiratory resetting which necessitated weak anaesthesia and light hypocapnia. Control experiments (restricted lesions of the medulla and the cervical cord, recording of afferent activity in thalamic sensory nuclei, medullary stimulation) revealed that this inspiratory resetting could not be related to appreciable activation of either non-respiratory efferents or spinal afferent pathways studied but was likely to depend on the activation of the descending inspiratory axons. We conclude that the respiratory resetting obtained by spinal stimulation resulted from mass antidromic activation of the inspiratory bulbospinal neurons which thus appear to be involved in the generation of the respiratory rhythm.     

Activation of visceral thin-fiber afferents increases respiratory output in cats

Respiratory responses to chemical activation of thin-fiber afferents from the stomach and the gallbladder were measured in anesthetized cats. Capsaicin or bradykinin applied to the serosal surface of either the stomach or the gallbladder elicited increases in breathing and phrenic nerve activity. Transection of the cervical vagi or the carotid sinus nerves had no effect on these responses. However, the respiratory responses to visceral stimulation were abolished by bilateral transection of the splanchnic nerves. We conclude that activation of thin-fiber afferents from the stomach and gallbladder causes a reflex increase in respiratory output. The initial afferent limb of this reflex is via the splanchnic nerves.     

Reflex inhibition of crural diaphragmatic activity by esophageal distention in cats

Distention of the esophagus has been shown to result in selective inhibition of phasic inspiratory activity in the crural portion of the diaphragm, with no effect on costal diaphragmatic activity. The purpose of this study was to determine rigourously the afferent pathways that mediate this response. Bipolar EMG electrodes were placed in the costal and crural portions of the diaphragm in decerebrate, spontaneously breathing cats. Distention of the esophagus by inflation of a Foley catheter balloon with 20 ml of air resulted in a selective inhibition of crural hiatal EMG activity, while costal EMG activity was maintained at predistention levels. The distention was accompanied by a reduction in respiratory frequency. Transection of the spinal cord at the C8-T1 level did not obliterate the crural inhibition produced by inflation. Section of the C4-C8 dorsal roots also failed to abolish the response. However, after bilateral cervical vagotomy, esophageal distention no longer influenced diaphragmatic EMG activity. These results indicate that the crural inhibition observed with esophageal distention is vagally mediated and is not influenced importantly by intercostal or phrenic afferents. Records of activity of the phrenic nerve branch innervating the crural portion of the diaphragm showed a similar response pattern, confirming that the inhibition is central in origin and that the crural fibers inhibited by distention are only a fraction of the total population of crural phrenic motoneurons.     

Vagal influences on parasternal intercostal muscle inspiratory shortening during hypercapnia and airway occlusion

To determine whether increases in electromyographic activity (EMG) are necessary for respiratory muscle shortening to occur during airway occlusion, respiratory changes in parasternal intercostal muscle length were measured using sonomicrometry in 11 anesthetized dogs during unoccluded (UB) and occluded (OB) breaths before and after vagotomy. During UB the extent of parasternal intercostal inspiratory shortening was greater after than before vagotomy both during oxygen breathing and during progressive hyperoxic hypercapnia. The relation between parasternal shortening, parasternal EMG, and tidal volume was not substantially affected by vagotomy. During OB parasternal intercostal EMG increased significantly compared to UB when the vagi were intact, but airway occlusion did not significantly change EMG activity post-vagotomy. However, both before and after vagotomy the parasternal intercostal shortened during OB in all animals. Parasternal intercostal inspiratory shortening during OB as a % of that during UB was significantly greater before compared to after vagotomy during oxygen breathing and moderate hypercapnia, but vagal integrity made no significant difference at high CO2. These results suggest that (1) pulmonary stretch receptor afferents inhibit parasternal intercostal inspiratory shortening but in proportion to their inhibitory effects on parasternal intercostal EMG and tidal volume, and (2) even when the EMG stays constant the parasternal intercostal muscle does not contract isometrically during occluded breaths.     

Effect of blocking medial area of nucleus retrofacialis on respiratory rhythm

Experiments were performed on anaesthetized, vagotomized rabbits. Respiratory movement and phrenic rhythmical discharge were reversibly abolished by the symmetrical injection of 1% procaine into the medial area of the nucleus retrofacialis (mNRF). Blocking other areas of the medulla had no obvious effect on respiratory rhythm, with the exception of the rostral portion of the ventral respiratory group (VRG), which overlaps with the mNRF. When the mNRF was blocked, most inspiratory and expiratory neurons recorded in the VRG and DRG (dorsal respiratory group) gradually started to fire continuously, and no longer exhibited respiratory rhythm. A minority of respiratory neurons was inactivated during apnea. Stimulation of the caudal portion of the DRG and VRG evoked only a short cluster of phrenic discharges instead of rhythmical firing, indicating that the respiratory neurons situated in these areas cannot generate rhythmic activity by themselves. This suggests that the mNRF plays an important role in the genesis and maintenance of basic respiratory rhythm.     

Determinants of rib motion in flail chest

We have previously developed a canine model of isolated flail chest to assess the effects of this condition on the mechanics of breathing, and these studies have led to the conclusion that the respiratory displacement of the fractured ribs is primarily determined by the fall in pleural pressure (Delta Ppl) and the action of the parasternal intercostal muscles. The present studies were designed to test the validity of this conclusion. A flail was induced in six supine anesthetized animals by fracturing both dorsally and ventrally the second to fifth ribs on the right side of the chest, after which the phrenic nerve roots were bilaterally sectioned in the neck. Sectioning the phrenic nerves caused a 34% decrease in Delta Ppl, associated with a 39% increase in parasternal intercostal inspiratory EMG activity (p < 0.05), and resulted in a marked reduction in the inspiratory inward displacement of the ribs. In three animals, the inward rib displacement was even reversed into a small outward displacement. When the airway was then occluded at end-expiration to increase Delta Ppl during the subsequent inspiration, all animals again showed a clear-cut inward rib displacement. These observations therefore confirm that in dogs with flail chest, the inspiratory displacement of the fractured ribs is set by the balance between the force related to pleural pressure and that generated by the parasternal intercostals. These observations also point to the critical importance of the pattern of inspiratory muscle activation in determining the magnitude of rib cage paradox in such patients.     

Medullo-Spinal Modulation of Sympathetic Output and Spinal Afferent Input

Associated Inhibition of Sympathetic and Afferent Activities. In intact animals, it is important that most "tonic" sympathetic activity be regulated by brainstem systems that have access to "central commands" from higher centers and to highly specific cranial nerve afferents. The importance of this supraspinal regulation is manifested by severe derangements of sympathetic regulation after spinal cord injury. A major component of these derangements is an exaggerated responsiveness of sympathetic neurons to visceral and somatic afferent activity. Although much is known about medullo-spinal systems that excite sympathetic preganglionic neurons, little is known about those systems that, in intact animals, isolate preganglionic neurons from spinal afferent input. This brief review will consider the evidence that either identical or cospatial medullary systems regulate the output of sympathetic preganglionic neurons and afferent input to the spinal cord. Further, evidence from this laboratory is presented, which indicates that a recently described system in the rostral cervical spinal cord, perhaps in concert with medullary systems, may play an important role in regulating the excitation of sympathetic preganglionic neurons by spinal afferents.     

Effect of chest wall vibration on the canine diaphragm during breathing.

High-frequency mechanical vibration of the ribcage reduces dyspnoea in patients with chronic obstructive pulmonary disease, and the suggestion has been made that this effect might be related to a decrease in central respiratory drive resulting from an increase in afferent inputs from intercostal muscles. In the present studies, the effects of ribcage vibration on central respiratory drive have been assessed without the confounding influence of conscious reactions. The electromyographic (EMG) activity of the diaphragm and the changes in pleural (Ppl) and abdominal (Pab) pressure were measured in six anaesthetized, spontaneously-breathing dogs while the rostral, the middle, or the caudal portion of the ribcage was vibrated at intervals during inspiration. The EMG activity of the external and parasternal intercostals was also measured. Ribcage vibration consistently elicited a marked increase in the inspiratory EMG activity recorded from the external intercostals, thus indicating that the procedure did activate intercostal muscle spindles. However, no alteration in diaphragmatic or parasternal intercostal EMG activity was seen in any animal. Transdiaphragmatic pressure and the relationship between deltaPab and deltaPpl during inspiration were also unaltered. The authors conclude that ribcage vibration and, with it, stimulation of external intercostal muscle spindles has no significant influence on phrenic motoneurones or on medullary inspiratory neurones. It is unlikely, therefore, that the beneficial effect of the procedure on dyspnoea results from a specific reduction in central respiratory drive.     

Fatigue-induced changes in diaphragmatic afferents and cortical activity in the cat.

The rationale for the present study was to test the hypothesis that changes in phrenic sensory activity during diaphragmatic fatigue may modify the transmission of phrenic afferent action potentials to the cortex and also the spontaneous EEG activity. This was performed in anesthetized cats. Diaphragmatic fatigue was produced by intermittent direct muscle stimulation for a 30 min period. Diaphragmatic metaboreceptors (tonically active afferents) and mechanoreceptors (phasic phrenic activity) were identified by their activation by intraarterial lactic acid injection or their discharge in phase with diaphragmatic contraction, respectively. Cortical phrenic evoked potentials (CPEPs) and spontaneous EEG activity were recorded from the left sensorimotor area. Diaphragmatic failure was shown from the 10th minute of stimulation. Then, the activity of tonic phrenic afferents increased markedly whereas, in parallel, the phasic discharge of mechanoreceptors decreased progressively. This was associated with progressive lengthening in onset and peak latencies of CPEPs. The main EEG changes (visual and fast Fourier transform analysis) were characterized by a transient increased energy in the delta frequency band during the first minutes of the fatigue run, followed by decreased energy in the theta frequency band after 11-25 min of stimulation. Denervation of the diaphragm suppressed the EEG changes during the fatigue run. The present observations suggest that the cortical integration of sensory information from the diaphragm may be altered during fatigue.     

Mechanical action of the interosseous intercostal muscles as a function of lung volume

On the basis of local stimulation of individual muscles, it has been suggested that both the external (EI) and internal interosseous intercostal muscles have an inspiratory action at low lung volumes and an expiratory action at high lung volumes. In this study, we assessed the action of the interosseous intercostal muscles at different lung volumes in 19 anesthetized dogs by synchronously activating the intercostal muscles via ventral root stimulation (VRS). An electrode was positioned on the upper thoracic spinal cord according to previously described techniques. The cervical phrenic rootlets were sectioned bilaterally, the accessory muscles were sectioned from the rib cage, and the origins of the abdominal muscles were sectioned from the middle region of the rib cage. Changes in airway pressure (delta P) were monitored during the application of supramaximal stimuli after hyperventilation-induced apnea and during airway occlusion. Animals were passively inflated or deflated with a volume syringe. Precontractile airway pressure was used as an index of lung volume. External and parasternal intercostal muscle (PA) lengths were monitored by sonomicrometry in the third intercostal space. Thoracoabdominal motion was monitored by Respitrace bands. During VRS, both PA and EI shortened at all lung volumes. Mean delta P progressively decreased with increasing lung volume. At precontractile airway pressures of -10, 0, and +30 cm H2O, delta P were -25 +/- 1, -16 +/- 1, and -5 +/- 1 cm H2O, respectively. After section of the internal intercostal nerves lateral to the costochondral junctions from the first through the seventh intercostal spaces to eliminate PA action, EI shortened, whereas PA usually lengthened.(ABSTRACT TRUNCATED AT 250 WORDS)