Central Respiratory Neurons Research Articles

Mechanisms of Middle ear aeration: Anatomic and physiologic evidence in primates

Proper aeration is a prerequisite for normal middle ear function in all terrestrial mammals. Our previous studies in primates provided anatomic evidence of neural circuits between the middle ear, brain, and eustachian tube by which central respiratory neurons can control middle ear aeration. Yet mechanisms that regulate middle ear aeration remain poorly understood. This study extends our research by examining maturation of these neural circuits, and investigating their underlying physiology. Ultrastructural examination of tympanic nerves, the afferent limb of the neural circuit, in an age-graded series of cynomolgus monkeys (Macaca fascicularis) showed substantial differences between newborn, young, and adult animals. These included a twofold increase in average myelin thickness, and greater than threefold increase in the ratio of myelinated to unmyelinated fibers from newborn to adult animals. These marked developmental changes may translate into functional differences in regulation of middle ear aeration in young animals, and possibly explain the extraordinarily high incidence of middle ear disease in early childhood. In physiologic experiments, bilateral electromyographic responses were recorded from eustachian tube muscles, the efferent limb of the neural circuit, in adult monkeys after ipsilateral stimulation of the tympanic nerve. Response latencies were 9 to 28 msec, similar to those of other multisynaptic bilateral brainstem reflexes. These physiologic data strongly suggest a concept of active control of middle ear aeration by respiratory neurons in the brain.

CO2 sensitivity of cat phrenic neurogram during hypoxic respiratory depression

The CO2 response of the phrenic neurogram before and during CO-induced isocapnic brain hypoxia was studied in peripherally chemodenervated, vagotomized, paralyzed, ventilated cats with blood pressure held constant. During inhalation of 0.5% CO in 40% O2, arterial O2 content (CaO2) was reduced to 40% and minute phrenic activity to 38.4 +/- 9.4% (SE; n = 9) of prehypoxic levels, primarily due to depression of peak phrenic amplitude (PP). CO2 response, defined as the slope of the plot of PP vs. end-tidal PCO2 during CO2 rebreathing, was unaffected by phrenic depression even to the point of total suppression of phrenic activity in two cats. The effect of the tissue metabolic acidosis associated with hypoxia on phrenic CO2 sensitivity was assessed in a separate group of cats by blocking lactate formation during hypoxia with dichloroacetate (DCA). Preventing lactic acidosis during hypoxia did not affect the CO2 response of the phrenic activity during hypoxia. We conclude that 1) hypoxic depression does not limit the ability of central respiratory neurons to respond to CO2, and 2) the failure of DCA to affect the CO2 response of the phrenic neurogram suggests that brain intracellular lactic acidosis does not modify the phrenic response to hypercapnia.

Inspiratory on-switch evoked by stimulation of the mesencephalon: activity of phrenic and laryngeal motoneurones.

In anaesthetized cats (chloralose-urethan) the effects of brief tetanic electrical stimulation (50 to 100 ms) of the mesencephalic central gray matter and reticular formation on the inspiratory on-switch were studied during the expiratory (E) phase on the gross and unitary activities of phrenic, laryngeal inspiratory and laryngeal expiratory nerves. On the inspiratory laryngeal and phrenic nerves, stimulation elicited a short latency gross response concomitant with the train: the inspiratory Primary Response (Prim.R.) which is followed by an inspiratory Patterned Response (Patt.R.) of longer duration which corresponded to the inspiratory on-switch. The Patt.R. generally appeared from the Prim.R. within a latent period (Silent Phase: Sil.P.) as long as 100 ms. On the expiratory laryngeal nerve, stimulation elicited a brief activation (expiratory Prim.R.) concomitant with the beginning of the inspiratory laryngeal Prim.R. and which rapidly stopped as the latter continued during the stimulus train. The inspiratory Prim.R. corresponded to a simultaneous activation of both early and late (so defined during their spontaneous discharge) inspiratory motoneurones. The laryngeal motoneurones were more strongly activated than the phrenic ones. During the inspiratory Patt.R. all the phrenic motoneurones presented a recruitment delay earlier, compared with the spontaneous one, whereas the recruitment drastically changed from an inspiratory laryngeal motoneurones to another. Thus, the two pools of motoneurones presented different properties of activation. During the inspiratory Sil.P. no concomitant expiratory laryngeal activation was observed when most of the inspiratory motoneurones were inactive. As some inspiratory laryngeal motoneurones did not stop firing, the existence of some central respiratory neurones exhibiting a similar persistent activity and subserving the inspiratory on-switch mechanisms may be hypothesized.

Modifiability of spinal synapses.

Modifiability of spinal synapses.

Neurophysiological studies on superficial medullary chemosensitive area for respiration

In cats anesthetized with chloralose-urethane (40 mg/kg chloralose; 200 mg/kg urethane) pH sensitivity of neurons in the caudal chemosensitive area on the ventrolateral surface of the medulla oblongata was examined while monitoring phrenic nerve activity simultaneously. pH was varied by superfusion of the ventral medullary surface with mock cerebrospinal fluid (CSF) of different pH (pH 7.4 control, 7.0 and 7.8). A total of 130 units from 21 cats changed their firing rate in response to CSF-pH changes. These were subdivided into 3 groups. In Group I, 31 respiratory pH sensitivie units increased their firing rate in response to decreased mock CSF-pH, noxious pinch, joint movement in contralateral forelimb, and increased inspired CO 2. These responses may have originated from respiratory center neurons. Group II consisted of 59 non-respiratory pH sensitive units whose firing rate changed in an inverse manner with CSF-pH changes. Of these, 30 responded to contralateral distal forelimb movement, 15 to hair manipulation, 9 to heavy pressure and 5 to noxious pinch. Increased inspired CO 2 (rebreathing) did not modify activity. The response to pH is believed to be from non-specific neurons. Group III consisted of 40 non-respiratory pH sensitive units responding to CSF-pH changes and to increased inspired CO 2. The firing rate was irregular, the interval distribution approaching an exponential function. It may be postulated that the impulse frequency of chemosensitive impulses may be irregular at the site of impulse generation, the irregularity decreasing by convergence during transmission to the respiratory centers. The time course of Group III chemosensitive units was similar to phrenic nerve responses.

On the cardioinhibitory role of water immersion per se in ducklings (Anas platyrhynchos) during enforced submersion

Experiments were done to clarify the contribution of water immersion per se to the cardiac chronotropic response to enforced submergence in ducklings. Four protocols were used in which one or more of the factors claimed to affect diving heart rate were controlled. Heart rate fell rapidly in the first few seconds of submergence in all protocols. This initial fall was unaffected by changes in head position or by breathing 100% oxygen for 5 min before diving. Also, sudden withdrawal of input from lung receptors or cessation of activity in central respiratory neurones did not affect onset of diving bradycardia. We conclude that about one-third of diving bradycardia in ducklings is caused by water immersion per se.

Activity of the respiratory center as a paired structure during stimulation of the anterior gyrus cinguli in rats

A total of 65 acute experiments on rats anesthetized with urethane were made to study bioelectrical activity of the external intercostal muscles on the right and left sides of the chest and neuronal activity of both halves of the respiratory center during electrical stimulation (4--15 v, 60 Hz) of the right or left anterior gyrus cinguli. The data obtained showed varied asymmetrical and asynchronous changes in the electromyogram of the respiratory muscles and disclosed some features of the effect of the left and right anterior gyrus cinguli on the electromyogram of the respiratory muscles and diverse reactions of respiratory center neurons to unilateral stimulation of the gyrus cinguli. The presence of functional asymmetry of the right and left anterior gyrus cinguli and their effect on the respiratory center are suggested.

Anatomical organization of central respiratory neurons.

Most of our currently held views on the organization of central mechanisms are based on electrophysiological studies of respiratory neu­ rons, i.e. neurons that Salmoiraghi & Burns (66, 67) described as produc­ ing periodic bursts of activity which are locked constantly to some phase of activity of the diaphragm. Neurons that show no sign of rhythmicity or have irregular bursting patterns that may play a part in the genesis of rhythm are excluded by such a definition. Recently, the search for these related units-RRUs (15)-has been intense and much of the conceptual framework of the organization of the respira­ tory centers is derived from these studies. Electrophysiological studies, however, have limitations. For instance, during extracellular stimulation and recording it is not possible to know the exact morphology or location of the neuron being studied. Furthermore, since only spontaneously active units are sampled, it is possible to miss neurons that are silent under the condition of the experiment and that could have been activated under other conditions. Combinations of electrophysio­ logical and neuroanatomical mapping techniques might result in a more meaningful map of the centers. However, it is rarely possible to do definitive anatomical mapping of control mechanisms be­ cause of heterogeneity in the neuronal populations involved.

The progressive onset of spontaneous and induced fetal breathing

Intratracheal pressure (ITP max) and respiratory drive (dITP/dt) from the occluded, liquid-filled trachea of term fetal lambs in utero were measured for each breath at the onset of fetal breathing (1) during spontaneous breathing, (2) during sciatic nerve stimulation, (3) during induced hypercapnia, and (4) following naloxone administration. These responses were characterized by linear increase of ITP max and dITP/dt following which these parameters became stable. The rate of rise of ITP max and dITP/dt was lowest and similar during spontaneous breathing and sciatic stimulation, but increased incrementally with hypercapnia and naloxone. Mechanical factors could not account for these responses in the liquid-filled lung, nor did appreciable chemical changes occur during this period. These results suggest that progressive breathing responses at the onset of fetal breathing may stem from gradual recruitment of central respiratory neurons, and that the rate of rise of such recruitment depends on facilitation by natural arousal, somatosensory stimulation and hypercapnia as well as on release from natural endorphin inhibition.

1701 PROGRESSIVE ONSET OF SPONTANEOUS AND INDUCED FETAL BREATHING

Intratracheal pressure (Pmax) and respiratory drive (dP/dt) from the occluded, liquid filled trachea of term fetal lambs in utero were measured for each breath during the onset of fetal breathing. Progressive breathing responses at the onset of fetal breathing were observed (1) during spontaneous breathing, (2) during sciatic nerve stimulation, (3) during induced hypercapnia by Fetal CO2 Tests (Moss and Scarpelli, J. Appl. Physiol. 47:527, 1979) and (4) following naloxone administration. These responses were characterized by linear increase of both Pmax and dP/dt for 6.8 ± 0.3 breaths (x ± SEM) over 13.9 ± 1.7 seconds, following which these parameters became stable. The rate of rise of Pmax and dP/dt versus both breath number and absolute time was lowest and similar during spontaneous breathing and sciatic stimulation, but increased incrementally with hypercapnia and naloxone. Mechanical factors could not account for these responses in the liquid filled lung, nor did appreciable chemical changes occur during this period. These results suggest that progressive breathing responses at the onset of fetal breathing may stem from gradual recruitment of central respiratory neurons, and that the rate of rise of such recruitment depends on facilitation by natural or somatosensory induced “arousal” and by chemical stimulation, as well as on release from natural (endorphin) inhibition. (Supported by NIH HL 00688 (RCDA) and HL 23995).

The interaction between reflex apnoea and bradycardia produced by injecting 5-HT into the nodose ganglion of the cat.

The apnoea, accompanied by bradycardia and hypotension caused by injection of 5-Hydroxytryptamine (5-HT) into the vascularly isolated nodose ganglion is characterised by intense electrical activity in the internal intercostal muscles. No such activity was recorded when apnoea was provoked by electrical stimulation of the central end of the superior laryngeal nerve. The cardiac slowing induced by 5-HT was less marked when the superior laryngeal nerve was concomitantly stimulated with an intensity sufficient to inhibit the internal intercostal activity. When central respiratory drive was depressed by hypocapnic ventilation nodose ganglion-induced bradycardia was comparable to that provoked during electrical stimulation of the superior laryngeal nerve in eupnoea. Unlike chemoreceptor-induced bradycardia the cardiac slowing caused by nodose ganglion stimulation was not reduced by "ramp" inflation of the lungs. The results were similar during hypocapnic depression of the "Respiratory Centre". These findings suggest that: (a) central expiratory activity contributes to the bradycardia induced by nodose ganglion stimulation, and (b) the inhibition of bradycardia by lung expansion occurs before the neural impulses reach the nucleus ambiguus but does not involve the central respiratory neurones.

Hydrogen sulfide: Effects on avian respiratory control and intrapulmonary CO 2 receptors

The respiratory response to acute inhalation of hydrogen sulfide (H 2S) and the response of intrapulmonary CO 2 receptors to this gas were studied in male White Leghorn chickens. Inhaling low concentrations of H 2S (0.05%) for 30 min had no effect on ventilation, but respiratory frequency and tidal volume became irregular and variable in birds that inhaled 0.2% and 0.3% H 2S for that period. All birds that inhaled 0.4% H 2S died within 15 min. H 2S, presented in the gas stream omnidirectionally ventilated birds, caused an increase in the discharge frequency of intrapulmonary CO 2 receptors and an increase in the amplitude of sternal movements. Because an increase in the discharge of these receptors normally inhibits the central respiratory neurons and may lead to apnea, it is clear that H 2S also has actions that increase the output from these central neurons. The possibility that H 2S affects the intrapulmonary CO 2 receptors by inhibiting carbonic anhydrase in them is discussed.

Somatic-respiratory reflex and onset of regular breathing movements in the lamb fetus in utero.

Breathing activity of six mature lamb fetuses (greater than 135 days of gestation) in utero was monitored from recordings of intraesophageal pressure, intratracheal pressure, and tracheal circumference from a mercury strain gauge before, during, and after stimulation of the central end of a cut sciatic nerve. Stimuli were either low (0.5-2.0 cps) or high (66 cps) frequency, 6-15-V square wave pulses of 0.6-1.25 msec duration. The fetuses remained in utero throughout the experiments in which ambient temperature, paO2, paCO2, arterial pH, mechanical stimulation, and spontaneous respiratory center activity could be ruled out as primary stimuli of the breathing movements observed. In one-third of the trials a "somatic-respiratory reflex" was elicited in which breathing coincided with the period of stimulation: in over 85% of these trials with low frequency stimulation, breathing movements were synchronous with the stimuli; in the rest the synchrony was broken during the period of stimulation. In two-thirds of the trials the "reflex" response was followed by spontaneous regular breathing movements ("onset of regular breathing") which continued for 1 min to 2 hr 30 min after the stimulation was stopped. Thiopental administration to the ewe (5 mg/kg) seemed to depress respiratory responsivity for about 60 min. Changes of tracheal circumference reflected both transmural pressure gradients and possibly also rhythmic vagal activity associated with breathing.

Effect of body temperature and hypoxia on the ventilatory CO 2 response in man

The ventilatory CO 2-response and the effect of hypoxia on this relationship were studied m four healthy subjects at normal and at elevated (+ 1.5 °C) body temperature. The experiments were carried out in a climatic chamber. Expired minute ventilation, rectal temperature and arterial tensions of oxygen and carbon dioxide were measured. Breathing air all subjects hyperventilated at elevated body temperature. The ventilatory CO 2-response line in high oxygen was displaced to the left in all subjects, and in three subjects a potentiated ventilatory response to hypoxia in addition to CO 2 was found during hyperthermia. In three subjects a significant increase in slope of the e t line in a Hey-plot was found in hyperthermia. The results seem to be evidence of a specific action of hyperthermia in the respiratory regulation apart from the drives related to discomfort of the experimental situation. Whether the effect of hyperthermia in our hyperoxic experiments is a direct temperature effect on central respiratory neurones or an indirect one is unknown. The potentiation of the hypoxic stimulus during hyperthermia, however, might be mediated through the arterial chemoreceptors.

Role of the medial zone of the medulla in rhythmic activity of respiratory center neurons

It was concluded from experiments with injection of cocaine and dihydroergotoxin that the medial zone of the medulla contains a system of neurons controlling the rhythmic firing pattern of neurons in the lateral zone of the respiratory center.

Neurophysiological investigations of medullary chemosensitive areas of respiration

The ventrolateral surface of the medulla has been designated as the location of the central chemoreceptors for respiration by Mitchell et al. (1963a, b; 1965). This area is presumably sensitive to the [H +] of CSF. Microelectrode recordings of unitary neuronal activity in the area during changes in pH of the CSF perfusing the area failed to show changes in firing rates correlated with changes in pH of perfusing CSF. Electrical stimulation of the ventral surface also failed to effect any changes in respiration. Recordings of unit activity of respiratory center neurons showed that respiratory center activity increased rapidly with application of acid CSF to the ventral surface and decreased with application of local anesthetic. It was found that respiration and respiratory center activity persisted in spite of the fact that the super-ficial “chemoreceptor areas” were anesthetized by local anesthetic. Bilateral lesioning of the superficial area of the medulla designated as chemosensitive resulted in apnea; however, it also resulted in ischemia of the lateral portions of the medulla and respiratory centers. By calculation it was shown that substances applied to the ventrolateral surfaces of the medulla would diffuse into the small arterioles of the subarachnoid space and thusly, be rapidly transported deep into the medulla and respiratory centers. It is considered feasible to suppose that substances which stimulate or depress respiration when introduced into the CSF produce their effects more directly at deep neurons after diffusion into the arterial supply to the medulla.

The importance of timing on the respiratory effects of intermittent carotid sinus nerve stimulation.

1. The respiratory response, measured directly as tidal volume or indirectly by using integrated peak phrenic activity, to intermittent electrical stimulation of the carotid sinus nerve was determined in anaesthetized cats.2. Stimulation at rates of 20-25 Hz for 0.5 sec had a rapid effect, increasing inspiratory airflow and phrenic discharge, but only if applied during inspiration. An increase in tidal volume or peak level of integrated phrenic discharge occurred only if the stimulus was exhibited during the second half of inspiration. Continuous stimulation had no greater effect on size or frequency of breathing than did intermittent inspiratory stimuli alone. Stimulation during expiration had no effect on the form or magnitude of subsequent breaths.3. Stimuli in expiration led to a prolongation of expiration. Stimuli in late inspiration caused a prolongation of both inspiration and expiration. Because of these effects, the respiratory rate could be changed by stimulation; in some instances entrainment of respiration by the intermittent carotid sinus nerve stimuli occurred.4. The findings are attributable to modulation of incoming carotid sinus nerve information by the central respiratory neurones, which use primarily that which arrives during inspiration. They show a possible mechanism by which oscillating signals may have a different effect than their mean level would indicate.

The importance of timing on the respiratory effects of intermittent carotid body chemoreceptor stimulation.

1. The respiratory response, measured directly as tidal volume or indirectly by using integrated peak phrenic activity, to brief intermittent chemical stimulation or depression of the carotid body was determined in anaesthetized cats. Recordings of carotid sinus nerve impulses allowed precise timing of the stimulus.2. Stimulation of the carotid body had a rapid effect on air flow, tidal volume and phrenic discharge rate only if given during inspiration. Increases in tidal volume and peak phrenic discharge occurred only if stimulation was applied during the last half of inspiration. Stimulation during expiration had no effect on the form or magnitude of subsequent breaths.3. Depression of the carotid body by NH(4)OH led to decreased tidal volume and phrenic discharge if it occurred during inspiration but had no effect if it occurred during expiration.4. Stimuli in expiration led to a prolongation of expiration. Stimuli in late inspiration caused prolongation of both inspiration and expiration.5. All of the effects noted were eliminated by bilateral carotid body denervation.6. The findings are similar to those following electrical stimulation of the carotid sinus nerve and are attributable to modulation of carotid body signals by the central respiratory neurones.