Comparison of phrenic motoneuron activity during eupnea and gasping

Single-fiber phrenic nerve action potentials were recorded together with activity of contralateral whole phrenic nerve rootlets during eupnea and gasping in decerebrate, cerebellectomized, vagotomized, paralyzed, and ventilated cats. Gasping was reversibly produced by cooling a fork thermode positioned through the pontomedullary junction. In eupnea, phrenic motoneurons were distributed into "early" and "late" populations relative to their onset of activity during inspiration. During gasping, however, both fiber types typically commenced activity at the beginning of the phrenic nerve burst. Moreover, late fibers, but not early units, exhibited an augmentation of discharge frequency with the onset of gasping. The concentration of activity of all phrenic motoneurons at the beginning of inspiration and the increase in late-unit discharge frequency account for the faster rise of the gasp as compared with the eupneic breath. It is concluded that the pattern of phrenic nerve activation during gasping differs fundamentally from that during eupnea. These results support the concept that mechanisms underlying the neurogenesis of gasping and eupnea may not be identical.     

Time-frequency coherence analysis of phrenic and hypoglossal activity in the decerebrate rat during eupnea, hyperpnea, and gasping

Fast respiratory rhythms include medium- (MFO) and high-frequency oscillations (HFO), which are much faster than the fundamental breathing rhythm. According to previous studies, HFO is characterized by high coherence (Coh) in phrenic (Ph) nerve activity, thereby providing a means of distinguishing between these two types of oscillations. Changes in Coh between the Ph and hypoglossal (XII) nerves during the transition from normal eupnic breathing to gasping have not been characterized. Experiments were performed on nine unanesthetized, chemo- and barodenervated, decerebrate adult rats, in which sustained asphyxia elicited hyperpnea and gasping. A gated time-frequency Coh analysis was developed and applied to whole Ph and medial XII nerve recordings. The results showed dynamic Ph-Ph Coh during eupnea, including MFO and HFO. XII-XII Coh during eupnea was broadband and included four distinct peaks, with low-frequency Coh dominating the epochs preceding the onset of Ph activity. During gasping, only MFO-peaks were present in Ph-Ph Coh. Bilateral XII activity showed a significant reduction in Coh and a shift toward lower frequencies during gasping. In contrast, contralateral Ph-XII Coh progressively increased during state changes from eupnea to gasping, a tendency mirrored in the startup part of the Ph activity. These data suggest significant hypoxia/hypercapnia-induced alterations in synchronization between respiratory outputs during the transition from eupnea to gasping, reflecting a reconfiguration of the respiratory network and/or alterations in the circuitry associated with the motor pools, including dynamic coupling between outputs.     

High-frequency oscillations of phrenic activity in eupnea and gasping of in situ rat: influence of temperature

We hypothesized that the in situ perfused preparation of the juvenile rat exhibits patterns of ventilatory activity comparable to eupnea and gasping in vivo. To evaluate this hypothesis, we examined high-frequency oscillations of activity of the phrenic nerve at 27-34 degrees C. The peak frequency of these high-frequency oscillations was defined from power spectral analysis. In situ, recordings were obtained in hyperoxic normocapnia, during ventilatory cycles in which the peak of integrated phrenic activity was achieved late in the burst, as in eupnea in vivo. Recordings were also obtained in hypoxic hypercapnia, when the peak of integrated phrenic activity occurred in the first half of the burst, as in gasping in vivo. In situ, peak frequencies in the power spectra were significantly higher in gasping than during eupnea. Frequencies during eupnea and gasping were progressively elevated as the temperature of the in situ preparation was increased. The shift in peak frequencies between eupnea and gasping and the temperature sensitivity of frequencies in situ were the same as in vivo. Results provide additional support for the conclusion that the in situ preparation demonstrates distinctly different patterns of automatic ventilatory activity, comparable to eupnea and gasping in vivo.     

Selective loss of high-frequency oscillations in phrenic and hypoglossal activity in the decerebrate rat during gasping

Respiratory motor outputs contain medium-(MFO) and high-frequency oscillations (HFO) that are much faster than the fundamental breathing rhythm. However, the associated changes in power spectral characteristics of the major respiratory outputs in unanesthetized animals during the transition from normal eupneic breathing to hypoxic gasping have not been well characterized. Experiments were performed on nine unanesthetized, chemo- and barodenervated, decerebrate adult rats, in which asphyxia elicited hyperpnea, followed by apnea and gasping. A gated fast Fourier transform (FFT) analysis and a novel time-frequency representation (TFR) analysis were developed and applied to whole phrenic and to medial branch hypoglossal nerve recordings. Our results revealed one MFO and one HFO peak in the phrenic output during eupnea, where HFO was prominent in the first two-thirds of the burst and MFO was prominent in the latter two-thirds of the burst. The hypoglossal activity contained broadband power distribution with several distinct peaks. During gasping, two high-amplitude MFO peaks were present in phrenic activity, and this state was characterized by a conspicuous loss in HFO power. Hypoglossal activity showed a significant reduction in power and a shift in its distribution toward lower frequencies during gasping. TFR analysis of phrenic activity revealed the increasing importance of an initial low-frequency "start-up" burst that grew in relative intensity as hypoxic conditions persisted. Significant changes in MFO and HFO rhythm generation during the transition from eupnea to gasping presumably reflect a reconfiguration of the respiratory network and/or alterations in signal processing by the circuitry associated with the two motor pools.     

Differing activities of medullary respiratory neurons in eupnea and gasping

Our purpose was to compare further eupneic ventilatory activity with that of gasping. Decerebrate, paralyzed, and ventilated cats were used; the vagi were sectioned within the thorax caudal to the laryngeal branches. Activities of the phrenic nerve and medullary respiratory neurons were recorded. Antidromic invasion was used to define bulbospinal, laryngeal, or not antidromically activated units. The ventilatory pattern was reversibly altered to gasping by exposure to 1% carbon monoxide in air. In eupnea, activities of inspiratory neurons commenced at various times during inspiration, and for most the discharge frequency gradually increased. In gasping, the peak discharge frequency of inspiratory neurons was unaltered. However, all commenced activities at the start of the phrenic burst and reached peak discharge almost immediately. The discharge frequencies of all groups of expiratory neurons fell in gasping, with many neurons ceasing activity entirely. These data are consistent with the hypothesis that brain stem mechanisms controlling eupnea and gasping differ fundamentally.     

Potential switch from eupnea to fictive gasping after blockade of glycine transmission and potassium channels

This study evaluated possible neuronal mechanisms responsible for the transition from normal breathing (eupnea) to gasping. We hypothesized that a blockade of both inhibitory glycinergic synaptic transmission and potassium channels, combined with an increase in extracellular concentration of potassium, would induce a switch from an eupneic respiratory pattern to gasping. Efferent activities of the phrenic, vagal, and hypoglossal nerves were recorded during eupnea and ischemia-induced gasping in a perfused in situ preparation of the juvenile rat (4-6 wk of age). To block potassium channels, 4-aminopyridine (4-AP, 1-10 microM) was administered. Strychnine (0.2-0.6 microM) was used to block glycinergic neurotransmission. After administrations of 4-AP, excess extracellular potassium (10.25-17.25 mM), and strychnine, the incrementing pattern of eupneic phrenic activity was altered to a decrementing discharge. Hypoglossal and vagal activities became concentrated to the period of the phrenic burst with expiratory activity being reduced or eliminated. These changes in neural activities were similar to those in ischemia-induced gasping. Results are consistent with the concept that the elicitation of gasping represents a switch from a network-based rhythmogenesis for eupnea to a pacemaker-driven mechanism.     

Key aspects of phrenic motoneuron and diaphragm muscle development during the perinatal period.

At the time of birth, respiratory muscles must be activated to sustain ventilation. The perinatal development of respiratory motor units (comprising an individual motoneuron and the muscle fibers it innervates) shows remarkable features that enable mammals to transition from in utero conditions to the air environment in which the remainder of their life will occur. In addition, significant postnatal maturation is necessary to provide for the range of motor behaviors necessary during breathing, swallowing, and speech. As the main inspiratory muscle, the diaphragm muscle (and the phrenic motoneurons that innervate it) plays a key role in accomplishing these behaviors. Considerable diversity exists across diaphragm motor units, but the determinant factors for this diversity are unknown. In recent years, the mechanisms underlying the development of respiratory motor units have received great attention, and this knowledge may provide the opportunity to design appropriate interventions for the treatment of respiratory disease not only in the perinatal period but likely also in the adult.     

Tonic pulmonary stretch receptor feedback modulates both eupnea and gasping in an in situ rat preparation

The perfused in situ juvenile rat preparation produces phrenic discharge patterns comparable to eupnea and gasping in vivo. These ventilatory patterns of eupnea and gasping differ in multiple aspects, including most prominently the rate of rise of inspiratory activity. Because gasping, but not eupnea, appeared similar after vagotomy in spontaneous breathing preparations, it has been assumed that gasping was unresponsive to afferent stimuli from pulmonary stretch receptors. In the present study, efferent activity of the phrenic nerve was recorded during eupnea and gasping in the in situ juvenile rat preparation. Gasping was induced in hypoxic-hypercapnia or ischemia. An increase in the pressure of tonic lung inflation from 1 to 10 cmH2O caused a prolongation of the duration between phrenic bursts in both eupnea or gasping. Bilateral vagotomy eliminated these changes. We conclude that the neural substrate mediating the Hering-Breuer reflex is retained in the in situ preparation and that the brain stem circuitry generating the respiratory patterns responds to tonic activation of pulmonary stretch receptors in a similar manner in eupnea and gasping. These findings support the homology of eupnea-like phrenic discharge patterns in the reduced in situ preparation and eupnea in vivo and disprove the common supposition that gasping is insensitive to vagal afferent feedback from pulmonary stretch receptor mechanisms.     

Phasic pulmonary stretch receptor feedback modulates both eupnea and gasping in an in situ rat preparation

The perfused in situ juvenile rat preparation produces patterns of phrenic discharge comparable to eupnea and gasping in vivo. These ventilatory patterns differ in multiple aspects, including most prominently the rate of rise of inspiratory activity. Although we have recently demonstrated that both eupnea and gasping are similarly modulated by a Hering-Breuer expiratory-promoting reflex to tonic pulmonary stretch, it has generally been assumed that gasping was unresponsive to afferent stimuli from pulmonary stretch receptors. In the present study, we recorded eupneic and gasplike efferent activity of the phrenic nerve in the in situ juvenile rat perfused brain stem preparation, with and without phrenic-triggered phasic pulmonary inflation. We tested the hypothesis that phasic pulmonary inflation produces reflex responses in situ akin to those in vivo and that both eupnea and gasping are similarly modulated by phasic pulmonary stretch. In eupnea, we found that phasic pulmonary inflation decreases inspiratory burst duration and the period of expiration, thus increasing burst frequency of the phrenic neurogram. Phasic pulmonary inflation also decreases the duration of expiration and increases the burst frequency during gasping. Bilateral vagotomy eliminated these changes. We conclude that the neural substrate mediating the Hering-Breuer reflex is retained in the in situ preparation and that the brain stem circuitry generating the respiratory patterns respond to phasic activation of pulmonary stretch receptors in both eupnea and gasping. These findings support the homology of eupneic phrenic discharge patterns in the reduced in situ preparation and eupnea in vivo and disprove the common supposition that gasping is insensitive to vagal afferent feedback from pulmonary stretch receptor mechanisms.     

Voltage-sensitive calcium currents and their role in regulating phrenic motoneuron electrical excitability during the perinatal period

This study examined the ontogeny of voltage-sensitive calcium conductances in rat phrenic motoneurons (PMNs) and their role in regulating electrical excitability during the perinatal period. Specifically, we studied the period spanning from embryonic day (E)16 through postnatal day (P)1, when PMNs undergo fundamental transformation in their morphology, passive properties, ionic channel composition, synaptic inputs, and electrical excitability. Low voltage-activated (LVA) and high voltage-activated (HVA) conductances were measured using whole cell patch recordings utilizing a cervical slice-phrenic nerve preparation from perinatal rats. Changes between E16 and P0-1 included the following: an approximately 2-fold increase in the density of total calcium conductances, an approximately 2-fold decrease in the density of LVA calcium conductances, and an approximately 3-fold increase in the density of HVA conductances. The elevated expression of T-type calcium channels during the embryonic period lengthened the action potential and enhanced electrical excitability as evidenced by a hyperpolarization-evoked rebound depolarization. The reduction of LVA current density coupled to the presence of a hyperpolarizing outward A-type potassium current had a critical effect in diminishing the rebound depolarization in neonatal PMNs. The increase in HVA current density was concomitant with the emergence of a calcium-dependent "hump-like" afterdepolarization (ADP) and burst-like firing. Neonatal PMNs develop a prominent medium-duration afterhyperpolarization (mAHP) as the result of coupling between N-type calcium channels and small conductance, calcium-activated potassium channels. These data demonstrate that changes in calcium channel expression contribute to the maturation of PMN electrophysiological properties during the time from the commencement of fetal inspiratory drive to the onset of continuous breathing at birth.     

Phrenic and external intercostal motoneuron activity during progressive asphyxia

Phrenic and external intercostal motoneuron activities were compared during progressive asphyxia induced by the interruption of artificial ventilation in the pentobarbital-urethan-anesthetized, gallamine-paralysed rabbit. The relative augmentation of inspiratory activity of the T1-T4 external intercostal nerves was significantly greater than that of the phrenic nerve during asphyxic hyperpnea. This was associated with a greater recruitment of intercostal than of phrenic motoneurons, particularly late in the hyperpneic phase immediately before the period of asphyxic apnea. However, peak and average discharge frequencies developed by intercostal motoneurons (n = 20) were only approximately 60% of those of the phrenic motoneurons (n = 28). Gasping respiration terminated the apneic period and was associated with a further intense recruitment of intercostal though not of phrenic motoneurons, but discharge frequencies developed by the intercostal motoneurons remained approximately 60% of those of the phrenic motoneurons. The instantaneous frequency profiles generated by the motoneurons often exhibited progressive changes during the terminal stages of hyperpnea (reduction in inspiratory duration and duty cycle and increases in inspiratory slope and discharge frequencies) such that much of the character of gasping respiration became evident before the apnea. Such smooth transitional sequences do not obviate the existence of an "independent gasping center" but do require that such a proposed center at least possess the capacity for interaction with those sites responsible for the generation of eupneic and hyperpneic respiration.