Eupneic Pattern Research Articles

Endogenous hydrogen sulfide maintains eupnea in an in situ arterially perfused preparation of rats

Hydrogen sulfide (H2S) is constitutively generated in the human body and works as a gasotransmitter in synaptic transmission. In this study, we aimed to evaluate the roles of endogenous H2S in generating eupnea at the respiratory center. We employed an in situ arterially perfused preparation of decerebrated rats and recorded the central respiratory outputs. When the H2S-producing enzyme cystathionine β-synthase (CBS) was inhibited, respiration switched from the 3-phase eupneic pattern, which consists of inspiration, postinspiration, and expiration, to gasping-like respiration, which consists of inspiration only. On the other hand, when H2S synthesis was inhibited via cystathionine γ-lyase (CSE) or when H2S synthesis was activated via CBS, eupnea remained unchanged. These results suggest that H2S produced by CBS has crucial roles in maintaining the neuronal network to generate eupnea. The mechanism of respiratory pattern generation might be switched from a network-based system to a pacemaker cell-based system in low H2S conditions.

Transitioning to eupnea during the first hour after birth in term and preterm mice (1177.6)

We used lipopolysaccharide (LPS) to induce cytokine‐mediated inflammation and preterm labor. Dams injected with LPS gave birth on G18.5, one day early. Saline injected dams either gave birth at term (G19.5) or had a c‐section (C‐S) on G18.5. Breathing and cardiac function were then assessed <10 and >60 min. after birth. C‐S preterm and term animals had high survival rates (~94%). LPS preterm animals had low survival rates (43%). Survival corresponded with a fast transition from a low frequency, high tidal volume, long duration ('large') breath pattern to a eupneic pattern. Non‐survivors failed to make the transition. Heart rate was directly related to breathing rate and correlated with survival. However, cardiac activity continued after non‐survivors took their last breath indicating that respiratory, not cardiac, failure was the primary cause of death. A subset of animals that did not transition to a eupneic pattern were administered caffeine. Caffeine improved survival outcomes, however, after an anoxic bout these animals had delayed heart and breathing rate recovery times. In conclusion, LPS‐mediated preterm birth increases mortality rates. This is due to a failure to transition from an initial 'large' respiratory pattern to a eupneic pattern and may correspond with an inability to alter network dynamics when transitioning from a low to high oxygen environment.Grant Funding Source: Supported by Seattle Children's Hospital Inter‐Center Funds

Respiratory rhythm and pattern after ibotenic acid (A) injections into the pontine respiratory group (PRG) of awake goats

We tested the hypothesis that IA injections into the PRG would disrupt eupneic respiratory rhythm and pattern. Cannula were bilaterally implanted in adult goats into either the lateral (LPBN, n = 4) or medial (MPBN, n = 4) parabrachial nuclei, or the Kölliker‐Fuse nucleus (KFN, n = 4). After recovery from surgery, 1 or 10μl injections of IA were made bilaterally through the implanted cannula during the awake state. Over the first 5 hours after the 10 μl injections, pulmonary ventilation was increased (P<0.05) in LPBN goats, there were no effects (P > 0.05) in MPBN goats, and there was a biphasic response (P < 0.05) in KFN goats. This biphasic response consisted of a tachypneic hyperpnea for 30 minutes followed by a prolonged hypopnea and hypoventilation with marked apneas and apneusis‐like breathing patterns. While awake and during NREM sleep 10–15 hours after the 1 μl and 10 μl injections respectively, there were above control number of apneas and augmented breaths in all groups. However, there were no incidences of terminal apneas. Breathing rhythm and pattern were normal 22 hours after the injections. Subsequent histological analysis showed <10%, 27%, and 62% reduction in number of neurons in the LPBN, MPBN and KFN goats, respectively. The data do not provide evidence that any of the lesioned sites are obligatory for respiratory rhythm generation, but suggest that the KFN contributes to eupneic pattern generation.

Pontomedullary transection in arterially‐perfused adult rat elicits diverse phrenic burst patterns and alters spectral dynamics

It is widely accepted that transection of the brainstem at the pontomedullary (PM) border shifts the eupneic pattern of breathing to gasping in vivo. Recent observations in the decerebrate arterially-perfused adult rat preparation, however, have demonstrated that similar transection may elicit multiple patterns of inspiratory motor output. Here, we examined the temporal and spectral (including time-frequency) characteristics of inspiratory discharge patterns following PM transection as well as transection caudal to the PM border in 6 decerebrate arterially-perfused adult rat preparations. Following PM transection, fractionated, square wave, bell-shaped, augmenting, and decrementing phrenic burst patterns were seen. These modified bursts exhibited increased burst duration (from 0.73±0.08 to 1.17±0.09 s; p=0.037) and reduced burst amplitude. Spectral analysis revealed decreased HFO power with little or no effect on MFO power, resulting in an increase in the MFO/HFO ratio by ~ 3-fold (p=0.009). The frequency of the MFO peak, however, was reduced from 43.8±1.3 to 36.3±2.3 Hz (p=0.047). Spectral activity also exhibited an earlier onset (~10% TI) and prolonged duration (~80% TI) following PM transection. Transection ~1–2 mm caudal to the PM border was ineffective in further altering either burst pattern or spectral content; however, TI was decreased. Subsequent transection at or caudal to the obex abolished all rhythmic output. These results confirm that PM transection elicits multiple patterns of phrenic nerve discharge in the arterially-perfused adult rat preparation, and further demonstrates that these modified burst patterns are associated with alterations in the spectral dynamics underlying the central inspiratory network. Supported by NS045321

A decerebrate, artificially-perfused in situ preparation of rat: Utility for the study of autonomic and nociceptive processing

By extending established technology, we have developed a relatively simple in situ preparation from juvenile rat that overcomes some technical restrictions present in vivo (e.g. need for anaesthesia, mechanical instability for intracellular recording and control over the extracellular milieu). The in situ preparation is decerebrate and artificially perfused via the left ventricle with a colloid containing solution. It exhibits an eupneic pattern of respiratory motor activity and demonstrates numerous somatic and visceral reflexes including those evoked by stimulation of the tail, hindlimbs, bladder, baroreceptors and peripheral chemoreceptors. We have employed this preparation to allow recordings from multiple sympathetic motor outflows such as thoracic and lumbar chain, inferior cardiac, splanchnic, renal and adrenal nerves. We show that the sympathetic motor discharge shows strong respiratory modulation and exhibits pronounced reflex modulation indicating intact communication between the periphery, the brainstem and the spinal cord. Further, we have made extracellular and whole cell recordings from neurones in the spinal cord, demonstrating good mechanical stability. The decerebrate, artificially-perfused rat (DAPR) provides a powerful methodology with which to study peripheral and central control of the autonomic nervous system with many of the benefits of an in vitro environment.

Ionotropic excitatory amino acid receptors in pre-Botzinger complex play a modulatory role in hypoxia-induced gasping in vivo.

Activation of ionotropic excitatory amino acid (EAA) receptors in pre-Bötzinger complex (pre-BötC) not only influences the eupneic pattern of phrenic motor output but also modifies hypoxia-induced gasping in vivo by increasing gasp frequency. Although ionotropic EAA receptor activation in this region appears to be required for the generation of eupneic breathing, it remains to be determined whether similar activation is necessary for the production and/or expression of hypoxia-induced gasping. Therefore, we examined the effects of severe brain hypoxia before and after blockade of ionotropic EAA receptors in the pre-BötC in eight chloralose-anesthetized, deafferented, mechanically ventilated cats. In each experiment, before blockade of ionotropic EAA receptors in the pre-BötC, severe brain hypoxia (6% O2 in a balance of N2 for 3-6 min) produced gasping. Although bilateral microinjection of the broad-spectrum ionotropic EAA receptor antagonist kynurenic acid (20-100 mM; 40 nl) into the pre-BötC eliminated basal phrenic nerve discharge, severe brain hypoxia still produced gasping. Under these conditions, however, the onset latency to gasping was increased (P < 0.05), the number of gasps was reduced for the same duration of hypoxic gas exposure (P < 0.05), the duration of gasps was prolonged (P < 0.05), and the duration between gasps was increased (P < 0.05). These findings demonstrate that hypoxia-induced gasping in vivo does not require activation of ionotropic EAA receptors in the pre-BötC, but ionotropic EAA receptor activation in this region may modify the expression of the hypoxia-induced response. The present findings also provide additional support for the pre-BötC as the primary locus of respiratory rhythm generation.

Defining eupnea.

To describe a pattern of rhythmic activity as "breathing" or "respiration" inevitably leads to the conclusion that this rhythmic activity is "normal" or "eupneic". Initially, it must be noted that, by strictest definition, "eupnea" can only be applied to "breathing" in an unanesthetized preparation. Any experimental perturbation, including anesthesia, changes eupnea, primarily by reducing the frequency of "breathing". However, a "eupneic pattern", in terms of the pattern of airflow of individual breaths, remains. Also remaining are patterns of neural and neuronal activities which are characteristic of individual breaths of eupnea. In this commentary, we consider these patterns of activities, which define a eupneic pattern and contrast these with patterns during apneusis and gasping. It has long been recognized that these three different patterns of "respiratory activity", eupnea, apneusis and gasping, can be generated in preparations in which all of the central nervous system has been removed, exclusive of the brainstem and spinal cord.

Respiratory changes induced by kainic acid lesions in rostral ventral respiratory group of rabbits.

The role played by the Bötzinger complex (BötC), the pre-Bötzinger complex (pre-BötC), and the more rostral extent of the inspiratory portion of the ventral respiratory group (iVRG) in the genesis of the eupneic pattern of breathing was investigated in anesthetized, vagotomized, paralyzed, and artificially ventilated rabbits by means of kainic acid (KA, 4.7 mM) microinjections (20-30 nl). Unilateral KA microinjections into all of the investigated VRG subregions caused increases in respiratory frequency associated with moderate decreases in peak phrenic amplitude in the BötC and pre-BötC regions. Bilateral KA microinjections into either the BötC or pre-BötC transiently eliminated respiratory rhythmicity and caused the appearance of tonic phrenic activity ("tonic apnea"), whereas injections into the rostral iVRG completely suppressed inspiratory activity. Rhythmic activity resumed as low-amplitude, high-frequency oscillations and displayed a progressive, although incomplete, recovery. Combined bilateral KA microinjections (BötC and pre-BötC) caused persistent (>3 h) tonic apnea. Results show that all of the investigated VRG subregions exert a potent control on both the intensity and frequency of inspiratory activity, thus suggesting that these areas play a major role in the genesis of the eupneic pattern of breathing.

Modulation of gasp frequency by activation of pre-Bötzinger complex in vivo.

Under hyperoxic conditions, both chemical stimulation of neurons and focal hypoxia in the pre-Bötzinger complex (pre-BötC) in vivo modify the eupneic pattern of inspiratory motor output by eliciting changes in the patterning and timing of phrenic bursts, which includes both phasic and tonic excitation. The influence of this region on the gasping pattern of phrenic motor output produced during severe brain hypoxia is unknown. We therefore examined the effects of chemical stimulation of neurons (DL-homocysteic acid; DLH; 10 mM; < or =20 nl) and focal hypoxia (sodium cyanide; NaCN; 1 mM; < or =20 nl) in the pre-BötC on hypoxia-induced gasping in chloralose-anesthetized, vagotomized, mechanically ventilated cats. Unilateral microinjection of DLH into the pre-BötC during hypoxia-induced gasping increased phrenic burst frequency by approximately 630% (P < 0.01) over baseline frequency due predominantly to a reduction in T(E) (from 28.9 +/- 6.2 to 5.2 +/- 1.8 s; mean +/- SE; P < 0.01). No significant changes in T(I) or rate of rise between hypoxia-induced gasps and the DLH-induced bursts were observed; the effects on peak amplitude of integrated phrenic nerve discharge were variable. Similar responses were evoked by unilateral microinjection of NaCN into the pre-BötC. These findings demonstrate that both activation of pre-BötC neurons and focal hypoxia in the pre-BötC not only influence the eupneic pattern of phrenic motor output but also modify the expression of hypoxia-induced gasping in vivo. These findings also provide additional support to the concept of intrinsic hypoxic chemosensitivity of the pre-BötC.

Brain stem PO(2) and pH of the working heart-brain stem preparation during vascular perfusion with aqueous medium.

The rat working heart-brain stem preparation (WHBP) is an in situ preparation having many of the advantages associated with in vitro preparations while retaining cardiovascular response functionality and an eupnoeic respiratory motor pattern. The preparation is perfused arterially with an aqueous medium having a much lower oxygen-carrying capacity than blood. To evaluate the efficacy of the artificial perfusion in providing adequate gas exchange within the brain stem, we used polarographic PO(2) and pH microelectrodes to determine the tissue PO(2) and pH of the medulla oblongata at various depths. When the perfusate was equilibrated with 5% CO(2) and 95% O(2), average tissue PO(2) was 294 Torr and no hypoxic areas were encountered. Tissue pH was remarkably uniform throughout the tissue, and on average was only 0.04 +/- 0.02 pH units more acidic than that of the perfusate. Increasing the PCO(2) of the perfusate increased tissue PO(2) and decreased arterial resistance. Decreasing perfusate PCO(2) (while keeping pH constant) decreased tissue PO(2) and reduced the respiratory activity. These results suggest that arterial PCO(2), independent of arterial pH, is an essential variable in determining both respiratory drive and cerebrovascular perfusion. We conclude that the medulla of the WHBP is oxygenated and within a physiological pH, which accounts for the eupneic pattern of respiratory motor activity it generates. Furthermore, this preparation may be a useful model for exploring mechanisms of central chemoreception as well as the dynamics of the cerebral vasculature responses following changes in blood gases.

Ventrolateral medullary respiratory network and a model of cough motor pattern generation.

The primary hypothesis of this study was that the cough motor pattern is produced, at least in part, by the medullary respiratory neuronal network in response to inputs from "cough" and pulmonary stretch receptor relay neurons in the nucleus tractus solitarii. Computer simulations of a distributed network model with proposed connections from the nucleus tractus solitarii to ventrolateral medullary respiratory neurons produced coughlike inspiratory and expiratory motor patterns. Predicted responses of various "types" of neurons (I-DRIVER, I-AUG, I-DEC, E-AUG, and E-DEC) derived from the simulations were tested in vivo. Parallel and sequential responses of functionally characterized respiratory-modulated neurons were monitored during fictive cough in decerebrate, paralyzed, ventilated cats. Coughlike patterns in phrenic and lumbar nerves were elicited by mechanical stimulation of the intrathoracic trachea. Altered discharge patterns were measured in most types of respiratory neurons during fictive cough. The results supported many of the specific predictions of our cough generation model and suggested several revisions. The two main conclusions were as follows: 1) The Bötzinger/rostral ventral respiratory group neurons implicated in the generation of the eupneic pattern of breathing also participate in the configuration of the cough motor pattern. 2) This altered activity of Bötzinger/rostral ventral respiratory group neurons is transmitted to phrenic, intercostal, and abdominal motoneurons via the same bulbospinal neurons that provide descending drive during eupnea.

Simultaneous measurement of the spontaneous eupneic pattern of breathing and pulmonary gas exchanges in normoxic and hypoxic subjects (author's transl)

In 158 normoxic or hypoxic subjects suffering from different bronchopulmonary diseases, we studied relationships between eupneic respiratory frequency (fR) or tidal volume (VT) and (1) ductances of O2 (DuO2) or CO2 (DuCO2); (2) alveolar-arterial difference in PO2, measured from end-tidal value (ET-a) or calculated from mean alveolar gas (A-a). All these variables were simultaneously measured. The individual mean value of PO2 in arterial blood (Pao2) allowed us to separate our subjects in three groups of similar Pao2 ( ± 5%). Negative correlations were found in each group between DuO2 or DuCo2 and fR but the slope of the Duo2 vs. fR relationship decreased with Pao2. A positive correlation, unrelated to Pao2 value, exists between Duo2 and VT. D(ET-a)O2 and D(A-a)O2 were not correlated with fR or VT. A mathematical model was obtained using equations of ductances (Lacoste) and taking into account part of our results, i.e. independency between D(A-a)O2 and breathing pattern; this model allowed us to verify the experimental relationships obtained between Duo2 and fR or VT for different values of Pao2. Studying all the previous relationships in subjects classified by their VT or fR individual values, we showed that the effects of interindividual changes in fR on Duo2 and Duco2 were most potent. It was concluded that: (l)the interindividual variations of eupneic VT and fR were not related to the alveolar-arterial O2 difference; (2) the individual changes in breathing were correlated with Duco2 in all subjects but with Duco2 essentially in normoxic subjects. So the calculation of the theoretical value of Duco2 must consider both components of the individual breathing pattern and measured value of Pao2.