Spontaneous and reflexly evoked activity in pharyngeal, laryngeal, and phrenic motoneurons of cat

Experiments were performed on spontaneously breathing cats that were anesthetized with chloralose-urethane in order to study changing excitability in pharyngeal, laryngeal, and phrenic motoneurons. Pharyngeal constrictor motoneurons chiefly were spontaneously active during expiration; the stylopharyngeus and certain motoneurons serving the superior pharyngeal constrictors were inactive during either phase of tidal respiration. While confirming work of previous investigators that feeble glossopharyngeal (epipharyngeal branch) and superior laryngeal nerve stimulation transiently attenuated on-going phrenic nerve activity, such evoked afferent volleys excited laryngeal motoneurons independent of phases of respiration. On-going pharyngeal constrictor nerve activity (expiration) was attenuated for 20–50 msec by superior laryngeal nerve stimulation and for 100–300 msec by glosso-pharyngeal nerve stimulation. Intercostal nerve stimulation evoked occasional pharyngeal constrictor nerve discharges but more commonly attenuated its spontaneous nerve activity. Stylopharyngeus and an unknown portion of the superior pharyngeal constrictor motoneurons were excited by afferent volleys evoked in glossopharyngeal and superior laryngeal nerves. These aforementioned changes in excitability were unaffected by total neuromuscular paralysis while maintaining artificial respiration. It is concluded that certain pharyngeal constrictor motoneurons of the cat are spontaneously active during tidal expiration, are under central respiratory control, and that their intermittence along with those serving the diaphragm and larynx by modest afferent volleys in the ninth and tenth cranial nerves stage the act of swallow or expulsive reflexes, or both.     

Electrophysiological and functional identification of different neuronal types within the nucleus ambiguus in the cat

Extracellular single neuron recording in nucleus ambiguus demonstrates the existence of different neuronal classes within its electrophysiological limits. Laryngeal motoneurons were identified by their antidromic activation from the recurrent laryngeal and vagus nerves. Interneurons were identified by their antidromic activation from the contralateral cervical spinal cord. Stimulation of the ipsilateral superior laryngeal nerve activated synaptically all laryngeal motoneurons recorded but not interneurons. Most motoneurons and all interneurons showed spontaneous discharges during the inspiratory phase of the respiratory cycle. A few expiratory motoneurons were also recorded. In addition to laryngeal motoneurons and interneurons vagal stimulation also activated a population of efferent neurons located in the nucleus ambiguus with firing patterns not related to respiration. Functional implications of described findings are discussed.     

Efferent activity in the phrenic nerve during the startle reflex in chloralose-anesthetized cats

Reflex activity in the phrenic nerve was studied in chloralose anesthetized cats during development of somatic startle reflexes in limb and lower intercostal nerves. It was shown that the main component of this activity during low-threshold reflexes evoked by acoustic, tactile and low-threshold somatic afferent stimulation was depression of phrenic inspiratory activity. The following reflex discharges were prevalent components of phrenic responses to high-threshold afferent stimulation: early, propriospinal (intercostal-to-phrenic reflex) and late, suprasegmental ones. The latter were of two types: inspiratory (observed mainly during inspiration in about 75% of experiments) and expiratory (observed during expiration in 25% of experiments) which could be classified as "phrenic startle reflexes". Modulation of all responses during the respiratory cycle was described. Structural characteristics of reflex responses evoked in the phrenic nerve by stimulation of various respiratory and nonrespiratory bulbar sites as well as their respiratory modulation have been analyzed. Organization of possible neurophysiological mechanisms of phrenic responses during startle reflexes is discussed.     

Split medulla preparation in the cat: arterial chemoreceptor reflex and respiratory modulation of the renal sympathetic nerve activity

The study was undertaken in order to assess the changes in sympathetic output in a split medulla preparation of the cat which, as shown earlier, has impaired respiratory rhythm generation. The effects of medullary midsagittal sections on renal sympathetic nerve firing were investigated in chloralose anesthetized, paralyzed and artificially ventilated cats. Recordings of phrenic and recurrent laryngeal nerve activity served as indices of central respiratory rhythm generation. Sections, 5 mm deep from the dorsal medullary surface and extending 6 mm rostrally and 3 mm caudally to the obex, did not produce any significant changes in heart rate, blood pressure or tonic renal sympathetic nerve firing levels. They decreased or abolished, however, the respiratory rhythmicity in renal sympathetic nerve which paralleled the reduction of inspiratory discharges in phrenic and recurrent laryngeal nerves, and abolished the carotid body chemoreceptor-sympathetic reflex. The inspiratory activity remaining after the sections could still be enhanced by chemoreceptor stimulation. The inhibitory baroreceptor and pulmonary stretch receptor sympathetic reflexes, and the central excitatory effect of CO2 on renal sympathetic nerve firing were not altered. The effects of electrical stimulation within the midsagittal plane of the medulla have shown that descending pathways from the medullary inspiratory neurons (or their medullary collaterals) do not participate in the facilitation of spinal preganglionic neurons during inspiration and in relaying the pulmonary stretch receptor inhibitory sympathetic reflex. A region located close to the obex was identified from which excitatory responses in renal sympathetic nerves, compatible with the response obtained by carotid sinus nerve stimulation, could be evoked. It is concluded that a lesion in the midline of the lower medulla at the level of the obex selectively destroys cells or pathways which relay the carotid body chemoreceptor-sympathetic reflex.     

Power spectral analysis of inspiratory nerve activity in the decerebrate cat

To investigate the high frequency oscillations observed in the inspiratory activity of respiratory motor nerves of decerebrate cats, we applied a signal processing technique, power spectral analysis, to the electrical activity of the phrenic and recurrent laryngeal nerves. We found two peaks in the phrenic nerve power spectral densities, one at 88.1 ± 6.4 Hz (mean ± S.D.) and the other at 37.1 ± 9.7 Hz, and two peaks for the recurrent laryngeal nerve, at 87.4 ± 10.1 Hz and at 55.4 ± 5.1 Hz. We identified 3 factors affecting the peaks. Anesthetics reduced or eliminated the 88 Hz peak and produced new low frequency peaks in the phrenic and recurrent laryngeal nerves. Increasing end-tidal CO₂ decreased the bandwidth of the 88 Hz peak and increased its amplitude relative to that of the low frequency peak. Decreasing body temperature from 38 to 30°C reduced the frequency of the 88 Hz peak by 5.0 Hz/°C. The power spectral density of the phrenic nerve activity differed from that of the recurrent laryngeal nerve activity because the single fibers in each nerve had different power spectral densities. About 70% of the fibers recorded in a nerve had power spectral densities similar to that of the whole nerve. A minority of the phrenic nerve fibers had the same low spectral peak as the recurrent laryngeal fibers had the same low spectral peak as the a minority of the recurrent laryngeal fibers had the same low spectral peak as the phrenic nerve. Bilateral removal of the dorsal respiratory group eliminated the high frequency peak in the power spectral density of the phrenic nerve and the peripheral reflexes, but rhythmic bursts of inspiratory activity remained. From these findings we hypothesized that there are two central respiratory pattern generators in the brain stem with parallel pathways to the respiratory motoneurons.     

Spontaneous and reflexly evoked laryngeal abductor and adductor muscle activity of cat

A study of spontaneous and reflexly evoked activity of laryngeal abductor (posterior cricoarytenoid) and adductor (thyroarytenoid and lateral cricoarytenoid) muscles was carried out in cats anesthetized with chloraloseurethane or made decerebrate and supplemented with ketamine HCl. The posterior cricoarytenoid muscle was active largely during inspiration but showed tonic activity throughout the respiratory cycle; the thyroarytenoid and lateral cricoarytenoid muscles were rhythmically active during expiration. Anesthetic amounts of pentobarbital abolished adductor rhythmicity and enhanced cyclic inspiratory activity of the abductor muscle. Hyperventilation increased the tonic adductor muscle activity while diminishing abductor muscle activity prior to resolution of apnea. Glossopharyngeal (epipharyngeal branch) and superior laryngeal nerve stimulation evoked chiefly excitatory effects on adductors and largely and attenuating effect on the abductor during inspiration. Stimulation of caudal intercostal nerves caused similar effects but to a lesser degree. Peripheral phrenic nerve stimulation during inspiration facilitated reflex abductor muscle activity whereas such stimulation during expiration facilitated reflex adductor muscle activity. The collective evidence further supports the conclusion that the larynx has a dual function: that of a respiratory organ (widening of the glottis during inspiration and its narrowing during expiration) and of a guardian of the lower respiratory tract from invading foreign matter (reflex sphincter action of contracting adductor muscles with relaxation of the abductor).     

Medium-frequency oscillations dominate the inspiratory nerve discharge of anesthetized newborn rats

In this study we examined the synchronization of the discharge of phrenic and recurrent laryngeal motoneurons in anesthetized rat pups 14 to 36 days of age and kittens, 14–15 days old. We found that the inspiratory nerve activity consisted of synchronized bursts separated by 20–35 ms, corresponding to medium-frequency oscillations (MFO). Accordingly, the autospectra of the neurograms had two peaks, one at the respiratory rate and the other between 22.8–43.0 Hz. No significant coherence was found between MFOs in the discharges of different nerves. High-frequency oscillations (HFO) characteristic for the adult inspiratory nerve activity were not present in the newborn rats. These findings demonstrate that phrenic nerve discharge of rat pups, like that of kittens and piglets, is in the MFO range, and suggest that MFO activity is an index of an early developmental stage of the respiratory system.     

Lesions of the rostral dorsolateral pons have no effect on afferent-evoked inhibition of inspiration.

This study investigated a possible role of the rostral dorsolateral pons (including nucleus parabrachialis medialis and K?lliker-Fuse nucleus) in mediating several inspiratory inhibitions. These inhibitions included the transient inhibition of phrenic inspiratory motor output produced by stimulation of the superior laryngeal nerve (SLN), the intercostal nerve (ICN) or the phrenic nerve (PN), as well as the inspiratory termination produced by trains of stimuli delivered to the SLN or ICN. In decerebrate, paralyzed, and artificially ventilated cats, the inhibitions produced by stimulation of these nerves were observed before and after lesioning (either radiofrequency, n = 8, or electrolytic, n = 9) the dorsolateral pons. Delivery of stimulus trains to the SLN or the ICN continued to elicit inspiratory termination following pontine lesions with no significant change in the threshold. There were no significant effects of bilateral dorsolateral pontine lesions on the threshold, onset latency, or duration of the short-latency, transient inhibitions produced by SLN, ICN or PN stimulation. From these data, we conclude that the rostral dorsolateral pons is not required in the production of any of these inhibitory reflexes.     

Effects of intravenous bicuculline and strychnine on inspiratory inhibitory responses in the cat

Single shock stimulation of the superior laryngeal nerve (SLN), intercostal nerve (ICN), phrenic nerve (PN) or within the medullary respiratory groups (DRG-VRG) produces a transient, short-latency attenuation of inspiratory motor activity. Trains of stimuli delivered to SLN and ICN cause premature termination of inspiration. This study examined involvement of glycine and GABAA receptors in these reflex inhibitions. Experiments were conducted in decerebrate, vagotomized, and paralyzed cats. Control responses of left PN activity to threshold single shock stimulation of SLN, PN, ICN and the DRG-VRG were recorded and the thresholds for SLN- and ICN-evoked inspiratory termination were determined. Five min after intravenous injection of bicuculline (1 mg/kg) or strychnine (50 micrograms/kg), the responses to stimulation were again recorded. This procedure was reiterated until the cumulative dose elicited marked convulsions. Neither drug affected the inspiratory terminating reflexes. Systemic bicuculline had no effect on transient inspiratory inhibition. However strychnine prolonged the onset latency and the duration of all four inhibitory responses. Since the degree of transient inhibition was not lessened (only delayed), it appears that these inspiratory inhibitory reflexes do not rely exclusively on actions of glycine or GABAA receptors.     

The participation of bulbar "respiratory" and "nonrespiratory" neurons in the development of the expiration reflex

The patterns of impulse activity of medullar neurons were investigated in nembutal-anaesthetized cats during the expiration reflex elicited by electrical stimulation of the internal branch of superior laryngeal nerve. It was shown that low- and high-threshold superior laryngeal afferents caused excitatory reflex reactions of different complexity in significant number of reticular nonrespiratory neurons. Respiratory neurons exhibited systemic changes of spontaneous activity, but in 22.4 per cent of them reflex responses were recorded. These responses occurred during activation of low-threshold laryngeal afferents. Oligo- and polysynaptic excitatory connections of low-threshold laryngeal afferents are found with inspiratory beta neurons, P-cells and laryngeal motoneurons, but inhibitory ones-with inspiratory gamma neurons. Participation of investigated neurons in the mechanisms of inhibition of inspiration, vocal cords closure and rate of breathing adaptive decrease during expiration reflex is discussed.     

Somatosplanchnic reflex discharges in rats

Splanchnic efferent reflex discharges caused by electrical stimulation of limb afferent nerves or intercostal afferent nerves were studied in chloralose-urethane anesthetized rats. Stimulation of the limb afferent nerve produced late supraspinal reflex discharges via group II and III afferent excitation. Stimulation of the intercostal afferent nerve produced early spinal reflex discharges via group II and III afferent excitation and also late spinal reflex discharges via group IV afferent excitation. Intercostal afferent nerve stimulation seemed to strongly depress the splanchnic late supraspinal reflex discharges.     

Evaluation of role of upper cervical inspiratory neurons in respiration, emesis and cough

Upper cervical (C₁₋₃) inspiratory (UCI) propriospinal neurons project to spinal segments containing respiratory motoneurons, but their functional significance is unknown. Bilateral kainic acid injections into this cell column in 12 decerebrate cats (11 paralyzed and artificially ventilated) had no major effect on phrenic, intercostal, and abdominal nerve discharge or EMG activity during (fictive) respiration, vomiting and coughing. Thus, UCI neurons are unessential for activation of major respiratory muscles during these behaviors.     

Behavior of VRG neurons during the atonia of REM sleep induced by pontine carbachol in decerebrate cats.

The microinjection of carbachol into the pons of acute decerebrate cats elicits a REM sleep-like atonia and a profound suppression of respiratory motoneuronal activity (J. Appl. Physiol., 69 (1990) 2280-2289). To assess whether this suppression is mediated by medullary neurons that provide respiratory drive to motoneurons of the respiratory pump muscles (diaphragm and intercostals), we studied the effect of pontine carbachol on the activity of neurons of the ventral respiratory group (VRG) in decerebrate, vagotomized, paralyzed and artificially ventilated cats. VRG neurons were recorded extracellularly along with the activity of phrenic and intercostal (external and internal) nerves. Both inspiratory (I) and expiratory (E) VRG neurons had incrementing, ramp-like bursts of activity during their firing periods and were not vagal motoneurons. Carbachol produced a depression of the peak firing rate in most (42/57) neurons studied. However, five cells showed no change and ten had an increase in activity in spite of consistent depression at the motoneuronal level. For the total population of cells (34 I and 23 E), the peak firing was reduced to 88.5% +/- 16.3 (S.D.) of control. The simultaneously recorded phrenic activity was reduced to 77.9% +/- 11.5, while inspiratory intercostal activity fell to 63.4% +/- 21.6 and expiratory to 23.2% +/- 21.2 of control. The carbachol-induced changes in peak firing of both I and E cells were quantitatively similar, and positively correlated to changes in peak phrenic activity. Analysis of this correlation suggested that phrenic and intercostal activities will be depressed to some degree by carbachol even when the average VRG cell activity remains unchanged. In addition, our data show that VRG cells may receive a combination of inhibitory and excitatory inputs during the carbachol-induced depression of respiratory motoneurons. Thus, although some disfacilitation from VRG cells may occur, there must be additional inhibitory or disfacilitatory pathways that mediate the decrease in activity of both phrenic and intercostal motoneurons that accompanies the REM sleep-like atonia.     

Dorsal and ventral respiratory groups of neurons in the medulla of the rat

The aim of the present work was to identify and localize in rat the medullary neurons involved in respiration. Neural activity was recorded in ketamine-anesthetized, paralyzed and artificially ventilated rats. Active sites were marked by electrocoagulation. Neurons firing in relation to phrenic nerve activity were located between 0.5 and 2 mm lateral to the midline, extending from 0.5 mm caudal to 2 mm rostral to the posterior end of the area postrema. Two groups of respiratory neurons were found: a dorsal group located ventrolateral to the tractus solitarius and a ventral group located in the ventrolateral reticular formation close to the nucleus ambiguus. Neurons were classified as bulbospinal or laryngeal if stimulation of the spinal cord or the vagus nerve, respectively, elicited antidromic action potentials, or as propriobulbar if they were not activated. Neurons firing synchronously with lung inflation were termed pump (P) cells. The dorsal respiratory group includes inspiratory (I) bulbospinal and propriobulbar neurons, P cells, but few expiratory (E) propriobulbar neurons. The ventral respiratory group includes bulbospinal, laryngeal and propriobulbar I and E neurons. Laryngeal motoneurons project ipsilaterally whereas bulbospinal neurons project contralaterally. Cross-correlations between inspiratory bulbospinal neuronal activity and phrenic discharge suggest that bulbospinal I neurons of dorsal and ventral groups project monosynaptically to contralateral phrenic motoneurons. These results indicate a similarity of the medullary respiratory centers of rats and cats, suggesting that rats may profitably be used in studies of respiratory rhythmogenesis.     

Diaphragm recovery by laryngeal innervation after bilateral phrenicotomy or complete C2 spinal section in rats

This study aimed to highlight the functional aspects of diaphragm reinnervation by laryngeal motoneurons after bilateral phrenicotomy or complete cervical transection. The left recurrent laryngeal nerve was connected to the left phrenic nerve in 14 rats. Five months later, all bridged rats presented a substantial ipsilateral diaphragm recovery (74.2 +/- 10% of contralateral activity) whereas the diaphragm remained paralysed in non-bridged rats (n = 5/5). After additional right phrenicotomy, functional breathing persisted in bridged rats whereas all non-bridged died. After complete C2 spinal transection, diaphragm respiratory discharges persisted in bridged rats. The reinnnervation by laryngeal motoneurons was confirmed by retrograde labeling, stimulus-elicited diaphragm response by vagal stimulation and diaphragm inactivation after vagotomy. In conclusion, the recurrent-phrenic nerve anastomosis induces a reliable functional diaphragm outcome even after contralateral diaphragm denervation or complete high cervical spinal cord injury, and could be considered as a clinical repair strategy for re-establishing diaphragm autonomy following spinal cord trauma.     

Analysis of the respiratory role of pharyngeal constrictor motoneurons of cat

The electromyogram of the middle pharyngeal constrictor muscle and the electroneurogram of the phrenic nerve was recorded in parallel with measurements of tidal inspiration and tracheal airflow in chloralose-urethane anesthetized cats. In eupnea, pharyngeal constrictor exhibited tonic and expiratory phasic activity. Lung inflation inhibited phasic pharyngeal constrictor activity to reveal its tonic component; lung deflation eliminated both phasic and tonic components. Vagotomy eliminated these static lung volume effects and increased spontaneous cyclic activity. Hypocapnia abolished phasic activity and unmasked its tonic component in vagotomized cats. Hypercapnia increased cyclic pharyngeal constrictor activity (vagi intact or cut). Weak superior laryngeal or glossopharyngeal nerve stimulation had little or no effect during inspiration; during expiration, superior laryngeal nerve stimulation evoked a short lasting attenuating effect, and glossopharyngeal nerve stimulation exerted a long lasting blocking effect. (vagi intact or cut). Pharyngeal constrictor, like laryngeal adductors, is classified as an expiratory resisting type muscle. Pharyngeal constrictor reduces dead space during hypercapnia thereby promoting the exodus of CO₂. Comparison of vagal and other respitatory motoneurons reveals some special features of neural control of respiration heretofore overlooked.     

Diaphragmatic and abdominal muscle activity during coughing in the decerebrate cat.

We studied electrical activity of the phrenic and abdominal nerves and from the diaphragm and abdominal muscles during coughing elicited by stimulation of the superior laryngeal nerves (SLN) in decerebrate spontaneously breathing cats. This activity was compared with that observed after partial and complete paralysis. During coughing, the duration, rate of rise, and amplitude of inspiratory discharge increased concomitantly with activity of the rectus abdominis and external, but not internal, oblique muscle. After end-inspiration, the abdominal muscles discharged with one or more bursts. Similar patterns of activity were observed in phrenic and abdominal nerves during SLN-induced coughing, first in spontaneously breathing and then in paralyzed, ventilated cats. The paralyzed, ventilated decerebrate cat provides a powerful experimental model for studying coughing.     

Pneumotaxic mechanisms influence phrenic, hypoglossal, and trigeminal activities

The influence of the pneumotaxic center on activities of the phrenic, hypoglossal, and trigeminal nerves was investigated in decerebrate, vagotomized, paralyzed, and ventilated cats. Stimulation of the pneumotaxic region caused a premature termination of activity of the phrenic nerve. Currents required for terminating this activity decreased during inspiration. Inspiratory hypoglossal and trigeminal activities were terminated concomitant with phrenic activity. The expiratory duration following stimulus-induced cessation of inspiration was directly related to inspiratory duration. During expiration, trigeminal activity was greatly augmented; a hypoglossal burst was also recorded in some trials. Stimulations in expiration elicited premature onsets of phrenic, hypoglossal, and trigeminal activities. The currents required declined during expiration. The durations of neural inspiration and expiration and peak phrenic and hypoglossal activities which followed stimulus-induced terminations of expiration approximated prestimulation values. Phasic trigeminal and expiratory hypoglossal activities were greatly augmented. Currents required for terminating either inspiratory or expiratory phases were augmented in hypercapnia. The data support pneumotaxic mechanisms as producing global onsets or terminations of inspiratory activity. Respiratory motoneurons of the brain stem and spinal cord differ in their sensitivities to efferent influences from the pneumotaxic center.     

Synaptic actions of vagal afferents on facial motoneurons in the cat

Synaptic potentials in facial motoneurons of cats were intracellularly recorded on stimulation of the vagal nerve, superior laryngeal nerve, solitary tract nucleus and spinal trigeminal tract nucleus. A possible disynaptic excitation was elicited in the facial motoneurons by stimulation of the vagal nerves and superior laryngeal nerves on both sides. Activation of the neurons in the solitary tract nucleus and/or trigeminal tract nucleus induced monosynaptic excitatory postsynaptic potentials (EPSPs) in the facial motoneurons.     

Respiratory rhythmicity in a split medulla preparation of the cat

Splitting the medulla in the cat resulted in the disappearance of inspiratory activity in the phrenic but not in the recurrent laryngeal nerves with minor changes in respiratory cycle duration. In one animal a complete desynchronization of respiratory rhythms was observed in the opposite recurrent laryngeal nerves. We conclude that each half of the cat's brain stem has its own respiratory phase-switching mechanism. Commissural connections are involved in the bilateral synchronization of activity and the mutual excitatory interactions between medullary respiratory neurons.