Central Respiratory Rhythm Research Articles

On the dominant rhythm in the discharges of single postganglionic sympathetic neurones innervating the rat tail artery.

1. In anaesthetized rats, using a focal recording technique, activity was recorded from single sympathetic postganglionic neurones innervating the caudal ventral artery of the tail. The following hypotheses were tested: (i) that the frequency of the dominant rhythmic discharge of the neurones can be different from the frequency of the central respiratory rhythm (as indicated by rhythmic phrenic discharge); and (ii) that the dominant sympathetic rhythm is not reliant on afferent feedback carried in aortic, sinus and vagus nerves. 2. Four types of preparation were used: spontaneously breathing (group 1), artificially ventilated (group 2), artificially ventilated with vagi cut (group 3), and artificially ventilated with vagus and sino-aortic denervation (group 4). 3. The frequencies of the dominant sympathetic rhythm under control conditions were: group 1, 0.91 +/- 0.12 Hz (mean +/- S.E.M., n = 5); group 2, 0.81 +/- 0.04 Hz (n = 18); group 3, 0.83 +/- 0.03 Hz (n = 17); group 4, 0.95 +/- 0.06 Hz (n = 11). The frequency of the dominant sympathetic rhythm was different from that of the phrenic rhythm in thirty-five out of fifty-one cases. 4. The mean frequency of the dominant sympathetic rhythm was not influenced significantly by hypocapnic apnoea. 5. Hyperthermia increased the frequency of the phrenic rhythm whilst decreasing that of the dominant sympathetic rhythm. 6. In all cases the frequency of the dominant sympathetic rhythm was different from that of the artificial ventilation cycle. 7. It is concluded that the frequency of the dominant sympathetic rhythm can be different from that of central respiratory drive and that it is not "driven' by afferent feedback relayed via sinus, aortic and vagus nerves. 8. It is proposed that the dominant sympathetic rhythm is unlikely to be generated by a central respiratory oscillator.

Postnatal changes in the mammalian respiratory network as revealed by the transverse brainstem slice of mice.

1. Spontaneous rhythmic activity in hypoglossal (XII) rootlets is generated at all postnatal stages from postnatal day (P) 0 to P22 in the transverse brainstem slice of mice containing the pre-Bötzinger complex (PBC). The PBC is known to be a region essential for respiratory rhythm generation. It contains neurones generating periodic bursts that occur in synchrony with rhythmic XII activity. This synchrony indicates that the rhythmic PBC activity generated by the transverse slice is the central respiratory rhythm. 2. The strength of coupling between XII bursts and PBC bursts decreased during early postnatal development. In younger mice (P0-4) each burst in XII rootlets corresponded to one burst in the PBC. In older mice (P5-18) one burst in XII rootlets occurred only every third to fourth burst in PBC neurones. 3. Cycle length and burst duration of rhythmic XII activity did not change significantly during the first three postnatal weeks. However, the pattern of XII bursts changed from decrementing (P0-7) to bell shaped (P8-18) while the rate of rise of XII bursts decreased significantly. 4. The rate of rise of rhythmic depolarizations in neurones of the PBC discharging in phase with XII bursts ('inspiratory neurones') decreased with postnatal development. During interburst intervals, membrane potentials of neurones of older mice (P6-18) were characterized by waves of synaptic input that were not observed in neonatal animals (P0-5). 5. Blockade of glycine receptors by strychnine increased the frequency of rhythmic XII activity in neonatal and older mice (P0-22). Although in expiratory PBC neurones glycinergic transmission was blocked at 10 microM strychnine, in inspiratory PBC neurones and XII rootlets even higher concentrations of up to 50 microM strychnine failed to abolish rhythmic activity.

A working heart-brainstem preparation of the mouse

An intra-arterially perfused working heart-brainstem preparation (WH-BP) was developed to allow studies into functionally identified cardiovascular and respiratory neurones in an in vitro milieu. This report provides the first description of this preparation. Evidence is presented indicating that the WH-BP: (1) spontaneously generates eupneic-like phrenic nerve activity, indicative of adequate oxygenation of the brainstem; (2) preserves the integrity of central coupling between the central respiratory rhythm generator and cardiac vagal motor neurones and, (3), allows an intracellular analysis of medullary cardio-respiratory neurones. The WH-BP may provide an advantaged environment for analysis of both synaptic and cellular mechanisms within the medulla that regulate cardio-respiratory activity.

Modulation of respiratory rhythm by alpha 2-adrenoceptors in awake and anesthetized goats

The purpose of this study was to examine the effects of alpha 2-adrenoceptor agonists in the control of breathing with goats that were either awake (n = 7) or anesthetized and artificially ventilated (n = 11). Awake goats infused intravenously with either of the alpha 2-agonists clonidine (1.0-6.0 micrograms/kg) or guanabenz (15.0-63.0 micrograms/kg) exhibited two distinct ventilatory patterns. One pattern was characterized by tachypnea in which respiratory frequency and minute ventilation increased to approximately 50% above control values. A second ventilatory pattern consisted of slow breathing with reductions of respiratory frequency and minute ventilation and highly variable expiratory duration intervals. These two patterns were unaffected by bilateral carotid body denervation. In anesthetized goats, alpha 2-agonists also caused an arrhythmia in phrenic nerve activity that was similar to the slow breathing pattern seen in awake goats. Respiratory disturbances were abolished by the selective alpha 2-receptor antagonist SKF-86466 (100-500 micrograms/kg), indicating that the effects are mediated by alpha 2-receptors. The results suggest that stimulation of alpha 2-adrenoceptors generally has an inhibitory effect on breathing in goats. The disruption of ventilation with clonidine or guanabenz suggests that alpha 2-adrenoceptors may play an important role in the control of central respiratory rhythm.

Endogenous serotonin modulates the fetal respiratory rhythm: an in vitro study in the rat

The aim of the present work was to know whether the excitatory modulation of the central respiratory rhythm generator by serotonin (5-HT) previously found to occur in the newborn rat, is already functional during the fetal life. Experiments were performed at embryonic day 18 (D18) and 20–21 (D20–21; full-term day 21) on the fetal rat brainstem-spinal cord preparation in which the ability to generate central respiratory activity in vitro persists. Replacing the normal medium which bathed the preparation by a medium containing 5-HT increased the respiratory frequency (RF) within 2–3 min in a dose-dependent manner in both D18 and D20–21 fetuses but the effect was particularly drastic at D18. Applying a medium containing the 5-HT antagonist, methysergide, to block the effect of endogenous 5-HT, if any, reduced the RF within 2–3 min and the reduction was especially drastic at D18 where respiratory arrests occurred for several minutes in most of the experiments. Applying a medium containing either the 5-HT reuptake inhibitor fluoxetine to potentiate the effect of endogenous 5-HT or the 5-HT precursor, l-tryptophan, to activate 5-HT biosynthesis mechanisms, increased the RF. To define the type of 5-HT receptors involved in the modulation of the RF, experiments were conducted with specific 5-HT agonists and antagonists. Both 5-HT 1 (8-OH-DPAT, buspirone) and 5-HT 2 agonists (DOI, α-methyl-5-HT) increased the RF but only the 5-HT IA agonist 8-OH-DPAT was efficient at submicromolar concentrations. Applying the 5-HT IA antagonist NAN-190 alone decreased the RF and even elicited respiratory arrests while the 5-HT 2 antagonist ketanserine was inefficient. NAN-190 pre-treatment blocked the increase in the RF due to 8-OH-DPAT and 5-HT. Taken as a whole these results clearly indicate that endogenous 5-HT exerts an excitatory modulation on the respiratory rhythm generator via activation of medullary 5-HT IA receptors well before birth, as soon as D18 where the modulation is particularly potent.

Physiological studies of thoracic spinocerebellar tract neurons in relation to respiratory movement

The differential roles of thoracic spinocerebellar tract (SCT) neurons with axons ascending in ipsi- (uncrossed) and contralaterally (crossed) spinal cord and their activities during respiratory movement were examined by extracellular recordings in the T 9–T 12 spinal segments of the anaesthetized cat. A total of 36 uncrossed and 7 crossed SCT neurons showed rhythmic discharges in relation to either spontaneous or artificial respiration. Uncrossed neurons were located in and around Clarke's column and thus are cells of origin of the dorsal spinocerebellar tract (DSCT). Crossed neurons were located in laminae VII and VIII. Almost all DSCT neurons were modulated during artificial respiration. Nineteen DSCT neurons showed high-frequency discharges during chest expansion and 15 DSCT neurons showed high-frequency discharges during the chest retraction phase of artificial respiration. Their respiration-related activity maintained the same phase relation and firing patterns after vagotomy. The phase relationship of neural rhythmicity to chest movement during artificial respiration and spontaneous breathing was the same in 14 neurons examined. Artificially induced pneumothorax caused a marked decrease of respiration-related modulation, and severing of a single nerve to the appropriate muscle caused a marked decrease of modulation, suggesting that respiration-related rhythmic activity of DSCT neurons is induced by the extension of respiratory muscles in the chest wall during both spontaneous and artificial respiration. Crossed SCT neurons showed rhythmic activity in phase with the central respiratory rhythm as indexed by phrenic nerve activity. Two neurons received an excitatory influence and five an inhibitory influence in the inspiratory phase from the centre. Four neurons in the latter group also received excitatory inputs from the periphery during chest expansion. Since inspiration brings chest expansion during spontaneous breathing, the central and peripheral inputs seem antagonistic in function. These neurons seemed to signal a discrepancy between the chest wall movement and the central respiratory rhythm. Functional differences between uncrossed and crossed SCTs in the lower thoracic segments in regard to the central and peripheral inputs as observed here are similar to those observed in the lumbar SCTs during locomotion.

Responses of ventral respiratory neurones in the rat to vagus stimulation and the functional division of expiration.

In anaesthetized rats, extracellular and intracellular recordings were taken from 106 respiratory neurones in the intermediate region of the nucleus ambiguus. We observed unprovoked shortening of expiratory time accompanied, in all classes of respiratory neurone, by the elimination of the changes in membrane potential that were characteristic of stage II expiration. The demonstration of the elimination of stage II expiration in both the rat and cat strongly supports the functional division of expiration into stage I expiration (post-inspiration) and stage II expiration. In order to identify the neurones in the rat that receive inputs from vagal afferents and modulate the central respiratory rhythm, we examined whether any respiratory neurones responded to stimulation of the vagus nerve. Some post-inspiratory and stage II expiratory neurones responded. The short latency (< 2 ms) of four of the responses indicates that some vagal afferents act on post-inspiratory neurones via a disynaptic pathway. While repetitive stimulation of the vagus nerve could inhibit the phrenic rhythm, it appears that most inspiratory neurones in the intermediate region of the nucleus ambiguous complex are not directly involved in integrating the information from vagal afferents with the central respiratory rhythm.

Mechanisms of respiratory rhythm generation change profoundly during early life in mice and rats

To study the ontogenesis of central respiratory rhythm generation, a novel brainstem slice preparation was developed that generates respiratory rhythmic activity spontaneously in mice and rats at all post-natal ages. The slice was made by tilting the brainstem to include both the ventrolateral and dorsomedial medulla. This ‘tilted-sagittal’ slice contained the nucleus ambiguus, the hypoglossal motor nucleus (XII) and the nucleus of the solitary tract which were preserved intact throughout their rostro-caudal extent. Using this rhythmic preparation it has been possible for the first time to directly compare the significance of glycinergic mechanisms for respiratory rhythmogenesis between newborns and mature rodents in vitro. Our findings demonstrate that during the first two weeks of life there are profound changes in both the motor pattern of rhythmic XII neurons and sensitivity of the respiratory rhythm to strychnine blockade of glycine receptors. Thus, developmental changes in strychnine-sensitive receptors are vital for the maturation of the respiratory network and it is suggested that any disturbance in their development may be lethal.

Pontine respiratory neurons in anesthetized cats

The pontine respiratory neurons (PRG) in the ‘pneumotaxic centre’ have been hypothesized to contribute to phase-switching of neural respiratory activity, especially in terminating inspiration. To define the neural elements involved in phase-switching, we recorded respiratory neurons extra- and intracellularly in anesthetized cats with an intact central nervous system. In total, 54 neurons were recorded: 49 neurons with activity modulated by central respiratory rhythm (20 inspiratory, 17 postinspiratory and 12 expiratory) and 5 neurons with activity correlated to tracheal pressure. The recorded neurons were clustered in dorsolateral pontine tegmentum within the Ko¨lliker-Fuse (KF) subnucleus of the parabrachial nuclei. Stable intracellular membrane potential was recorded in 11 of the 49 respiratory neurons (8 postinspiratory, 1 early inspiratory and 2 inspiratory). During continuous injection of chloride ions( n = 6), synaptic noise increased and IPSPs reversed, including a wave of IPSPs during stage-2 expiration in postinspiratory neurons. Further, relative input resistance varied through the respiratory cycle such that the least input resistance occurred during the neuron's( n = 5) quiescent period. No IPSPs nor EPSPs were evoked in pontine respiratory neurons by vagal stimulation. In conclusion, various types of respiratory neurons were recorded in the KF nucleus. Prominent excitatory and inhibitory postsynaptic activities were similar to those described for medullary neurons. These pontine respiratory neurons do not appear to receive a strong afferent input from the vagus. Rather, vagal afferent inputs seem to be directed towards non-respiratory neurons that are located more medially in the dorsal pons.

Interactions between Respiratory, Cardiac and Stepping Rhythms in Decerebrated Cats: Functional Hierarchical Structures of Biological Oscillators

Interactions are described of central origin between respiratory, cardiac and stepping rhythms during fictive locomotion in paralyzed, vagotomized, and decerebrated cats. Fictive locomotion was induced by tonic electrical stimulation of the mesencephalic locomotor region (MLR). The coherence between heart beat fluctuation, the efferent discharges of the phrenic, and the lateral gastrocnemius nerves was used to evaluate the strength of the coupling between those three rhythms. The heart beat rhythm was modulated by the centrally generated respiratory and stepping rhythms. The central respiratory rhythm was modulated by the centrally generated stepping rhythm. Based on the present findings, we have proposed a new model concerning the functional hierarchical structures of the three biological oscillators.

Computational Aspects of the Respiratory Pattern Generator

To help evaluate the hypothesis that the central respiratory rhythm is generated by a network of interacting neurons, a network model of respiratory rhythmogenesis is formulated and examined computationally. The neural elements of the network are driven by tonic inputs and generate a continuous variable representing firing rate. Each neural element in the model can be described by an activation time constant, an adaptation time constant, and a step nonlinearity. Initial network connectivity was based on an earlier proposed model of the central respiratory pattern generator. These connections were adjusted interactively until the model trajectories resembled those observed electrophysiologically. The properties of the resulting network were examined computationally by simulation, determination of the phase resetting behavior of the network oscillator, and examination of the localized eigenstructure of the network. These results demonstrate that the network model can account for a number of diverse physiological observations, and, thus, support the network hypothesis of respiratory rhymogenesis.

Possible involvement of serotonin in obstructive apnea of the newborn.

Experiments were conducted on newborn rats in order to study the serotonergic modulation of the central respiratory activity during the neonatal period. In vitro experiments performed on isolated brainstem spinal cord preparations demonstrated that serotonin exerts a permanent excitatory modulation on the central respiratory rhythm generator but depresses the inspiratory motor output to the genioglossus muscle. In vivo experiments confirmed that increasing endogenous serotonin levels decreases the inspiratory activity of the genioglossus and elicits drastic obstructive apneas.

Centrally driven slow oscillating potential of extrathoracic trachea.

Spontaneous electrical activity of extrathoracic trachea was recorded along with force developed by tracheal smooth muscle and phrenic nerve activity in decerebrated, paralyzed, and artificially ventilated dogs with pneumothorax. The tracheal electrical activity exhibited slow oscillating potentials that were coupled with spontaneous phasic contraction of trachea. Both rhythmic changes were synchronous with central respiratory rhythm represented by phrenic burst, independent of the respirator's rhythm. The dominant component of the slow oscillating potentials consisted of sinusoidal waves with large amplitude that occurred shortly after cessation of phrenic burst, i.e., in the postinspiratory phase. The concomitant small change in the slow oscillating potentials began in the late inspiratory phase just before the initiation of the tracheal contraction. This phase relationship was preserved after removal of intrathoracic vagal afferents from lungs. Such slow oscillating potentials were also observed during lung collapse produced by disconnecting the tube attached to the respirator. Transection of recurrent laryngeal nerves abolished the slow oscillating potentials. These results indicate that the slow oscillating potentials of the extrathoracic trachea are generated by a physiological process associated with the central respiratory rhythm. The dominant component of the slow oscillating potentials occurs in the postinspiratory phase.

Entrainment of respiratory rhythm by periodic lung inflation: effect of airflow rate and duration.

In anesthesized, paralyzed, and mechanically ventilated rabbits, the rate and duration of airflow were varied to investigate the effect on the central respiratory rhythm during 1:1 entrainment (1 phrenic nerve burst for 1 pump inflation). Our results showed that 1) the largest range of 1:1 entrainment was obtained with the longest inflation durations associated with the lowest flow rates and 2) both inspiratory (TI) and expiratory (TE) durations (as estimated by the period of phrenic nerve activity and phrenic nerve silence, respectively) increased when total respiratory duration (TT = TI+TE) increased. The phase relationships we studied were DI (the time elapsing between the onset of phrenic activity and the beginning of pump inflation) and DE (the time between the end of phrenic activity and the end of pump inflation). Both DI and DE increased when the total ventilatory period increased. However, for constant ventilatory period, DI changed with the rate and duration of airflow, whereas DE remained constant. In addition, under these conditions, TI, TE, and the integrated phrenic nerve activity remained constant independently of airflow rate and duration. The present results are discussed with regard to the pulmonary receptors (slowly and rapidly adapting receptors and C fibers) that are stimulated during mechanical ventilation, and hypotheses are drawn regarding their possible contribution to rhythm generation and the mechanism through which they may act.

Possible mechanism for central respiratory rhythm generation: Respiratory cycle duration is determined dependently by inspiratory and expiratory duration

Possible mechanism for central respiratory rhythm generation: Respiratory cycle duration is determined dependently by inspiratory and expiratory duration

Identification of neurons receiving input from pulmonary rapidly adapting receptors in the cat.

1. Extracellular and intracellular recordings were made in the caudal subdivisions of the nucleus tractus solitarii (NTS) to locate and characterize neurons excited by afferents from pulmonary rapidly adapting receptors (RARs) in Nembutal-anaesthetized cats. 2. Neurons identified as second-order cells activated by RARs (RAR-cells) were activated by electrical stimulation of myelinated afferents in the cervical portion of the vagus nerve(s) and by at least two of the following 'physiological' stimuli: (a) collapse of the lungs to atmospheric pressure; (b) hyperinflation of the lungs by either increasing tidal volume or maintained lung inflations; and (c) brief inhalation of ammonia vapour. 3. Of the ninety-nine RAR-cells identified and studied extracellularly, eighty-four were localized within the commissural nucleus of the NTS. Seventy-four cells responded monosynaptically to electrical stimulation of both ipsi- and contralateral vagal stimulation. The remaining ten RAR-cells located in the commissural nucleus and the fifteen located in the caudal portion of the medial subnucleus of the NTS rostral to the obex, responded to the ipsilateral vagus only. 4. Under control ventilatory conditions (bilateral pneumothorax, positive end-expiratory pressure of approx. 2 cmH2O), forty-eight of the ninety-nine RAR-cells showed spontaneous ventilator-related activity, occurring primarily during ventilator-induced deflations (thirty of forty-eight). 'Reversal' of this ventilator-related modulation, from firing predominantly during lung deflations to lung inflations, and vice versa, could be induced in eleven of the RAR-cells by changing the lung volume or by the inhalation of ammonia vapour. 5. Modulation of firing in synchrony with the central respiratory rhythm was observed in the activity of fourteen of the ninety-nine RAR-cells. 6. Intracellular recordings were made from twenty-two NTS neurons caudal to the obex that received monosynaptic excitatory postsynaptic potentials (EPSPs) from both ipsi- and contralateral vagus nerves. Two were positively identified as RAR-cells. 7. It is concluded that the commissural nucleus of the NTS is the major location of RAR-cells, and that the response characteristics of these neurons largely, but not entirely, correspond to the known properties of RARs.

The medullary respiratory network in the rat.

1. In urethane or Nembutal anaesthetized and artificially ventilated Wistar rats, respiratory neurones of the ventrolateral medulla oblongata were analysed in extracellular (n = 74) and intracellular (n = 43) recordings. 2. Some respiratory neurones were identified as bulbospinal by their antidromic response to spinal cord stimulation at the C4 level. The neurones examined were not antidromically excited by vagal nerve stimulation. 3. Based on their discharge pattern in relation to efferent phrenic and vagal nerve activity, six types of respiratory neurones were classified: early-inspiratory, throughout-inspiratory, late-inspiratory, post-inspiratory, expiratory, and phase-spanning expiratory-inspiratory neurones. 4. Analysis of postsynaptic activities and IPSP reversal following chloride injection revealed post-inspiratory and expiratory inhibition in inspiratory neurones a pronounced early-inspiratory and a relatively weak expiratory inhibition in post-inspiratory neurones, and an early-inspiratory and post-inspiratory inhibition in expiratory neurones. 5. In phase-spanning expiratory-inspiratory neurones the post-inspiratory inhibition was strong and effectively blocked action potential discharge. Expiratory-inspiratory neurones were quite similar to the group of inspiratory neurones, but seemed to receive tonic excitatory inputs not shunted by weak expiratory inhibition. This pre-inspiratory discharge was readily blocked by weak negative DC injection. 6. Under conditions of experimental hypoxia, or long lasting lung inflation and non-inflation, post-inspiratory neurones displayed a second burst of discharge at the end of the expiratory phase in addition to their longer lasting post-inspiratory discharge. 7. We conclude that in the rat the central respiratory rhythm is organized in three (inspiratory, post-inspiratory, expiratory) phases, and that synaptic interaction within the medullary respiratory network of the rat occurs similarly to that described for the cat.

Activity of crossed spinocerebellar tract neurones in the thoracic spinal cord in relation to the central respiratory rhythm

Correlation between respiratory movement and neural discharges of antidromically identified 20 crossed spinocerebellar tract (SCT) neurones in the T 8–11 segments was examined in the anaesthetized, paralysed cat. Activity of 11 neurones of them was related to phrenic nerve activity; 7 neurons increased and 4 decreased their activity during the active phase of the phrenic nerve. Their rhythmic activity remained even after artificial ventilation was turned off, indicating the central genesis of their rhytmic activity. By comparing discharge patterns with the ventilator on and off, it is suggested that half of these crossed SCT neurones appear to receive additional inputs from the periphery. Neurones which increased their firing rate during phrenic nerve activity tend to respond with decrease discharge to passive chest inflation, and vice versa.

Rostral ventrolateral medulla and sympathorespiratory integration in rats

The respiratory modulation of the lumbar sympathetic nerve discharge (LSND) was examined in halothane-anesthetized, paralyzed, and vagotomized rats by means of phrenic nerve discharge (PND)-triggered histograms. The respiratory modulation was 1) proportional to PND amplitude during chemoreceptor activation with CO2 and 2) reduced at elevated arterial pressure. Bilateral injections of bicuculline [gamma-aminobutyric acid (GABA)A receptor antagonist, n = 5] into the rostral ventrolateral medulla (RVLM), but not into medullary raphe, reversibly increased mean arterial pressure (MAP) and resting LSND, decreased the baroreflex, reduced PND amplitude and central respiratory rate, and greatly magnified the respiratory modulation of LSND. Injections of strychnine (glycine receptor antagonist, n = 5) or phaclofen (GABAB receptor antagonist, n = 2) into RVLM were without effect. Injections of kynurenic acid (excitatory amino acid receptor antagonist) into RVLM (n = 8), but not raphe (n = 3), reduced PND amplitude, increased central respiratory rate, reduced MAP, elevated resting LSND slightly, and greatly reduced the respiratory modulation of LSND. These data suggest that the rostral tip of the ventrolateral medulla represents a critical link between the central respiratory rhythm generator and the vasomotor outflow. Also, it indicates that the respiratory modulation of SND does not involve a gating of the activity of the medullary neurons that convey baroreceptor information to RVLM sympathoexcitatory neurons.

Activity of crossed spinocerebellar tract (SCT) neurones in the thoracic spinal cord synchronized with central respiratory rhythm

Activity of crossed spinocerebellar tract (SCT) neurones in the thoracic spinal cord synchronized with central respiratory rhythm