Retrotrapezoid Nucleus/parafacial Respiratory Group Research Articles

Oxidative stress in retrotrapezoid nucleus/parafacial respiratory group and impairment of central chemoreception in rat offspring exposed to maternal cigarette smoke

We have reported that smoking during pregnancy is associated with deficit in neonatal central chemoreception. However, the underlying mechanism is not well clarified. In this study, we developed a rat model of maternal cigarette smoke (CS) exposure. Pregnant rats were exposed to CS during gestational day 1−20. Offspring were studied on postnatal day 2. Reactive oxygen species (ROS) content and expressions of antioxidant proteins in retrotrapezoid nucleus/parafacial respiratory group (RTN/pFRG) were examined by fluorogenic dye MitoSOX™ Red and Western blotting, respectively. The response of hypoglossal rootlets discharge to acidification was also detected with micro-injection of H2O2 into RTN/pFRG of offspring brainstem slices in vitro. Results showed that maternal CS exposure led to an increase in ROS production, and brought about decreases in mitochondrial superoxide dismutase and Kelch-like ECH-associated protein-1, and an increase in NF-E2-related factor 2 in offspring RTN/pFRG. Catalase and glutathione reductase expressions were not significantly changed. Moreover, oxidative stress induced by micro-injection of H2O2 into RTN/pFRG in vitro inhibited the discharge response of hypoglossal rootlets to acidification. These findings suggest that maternal CS exposure results in oxidative stress in RTN/pFRG of rat offspring, which might play a role in the impairment of central chemoreception.

Excitatory Modulation of the preBötzinger Complex Inspiratory Rhythm Generating Network by Endogenous Hydrogen Sulfide.

Hydrogen Sulfide (H2S) is one of three gasotransmitters that modulate excitability in the CNS. Global application of H2S donors or inhibitors of H2S synthesis to the respiratory network has suggested that inspiratory rhythm is modulated by exogenous and endogenous H2S. However, effects have been variable, which may reflect that the RTN/pFRG (retrotrapezoid nucleus, parafacial respiratory group) and the preBötzinger Complex (preBötC, critical for inspiratory rhythm generation) are differentially modulated by exogenous H2S. Importantly, site-specific modulation of respiratory nuclei by H2S means that targeted, rather than global, manipulation of respiratory nuclei is required to understand the role of H2S signaling in respiratory control. Thus, our aim was to test whether endogenous H2S, which is produced by cystathionine-β-synthase (CBS) in the CNS, acts specifically within the preBötC to modulate inspiratory activity under basal (in vitro/in vivo) and hypoxic conditions (in vivo). Inhibition of endogenous H2S production by bath application of the CBS inhibitor, aminooxyacetic acid (AOAA, 0.1–1.0 mM) to rhythmic brainstem spinal cord (BSSC) and medullary slice preparations from newborn rats, or local application of AOAA into the preBötC (slices only) caused a dose-dependent decrease in burst frequency. Unilateral injection of AOAA into the preBötC of anesthetized, paralyzed adult rats decreased basal inspiratory burst frequency, amplitude and ventilatory output. AOAA in vivo did not affect the initial hypoxia-induced (10% O2, 5 min) increase in ventilatory output, but enhanced the secondary hypoxic respiratory depression. These data suggest that the preBötC inspiratory network receives tonic excitatory modulation from the CBS-H2S system, and that endogenous H2S attenuates the secondary hypoxic respiratory depression.

Generation of active expiration by serotoninergic mechanisms of the ventral medulla of rats

Abdominal expiratory activity is absent at rest and is evoked during metabolic challenges, such as hypercapnia and hypoxia, or after the exposure to intermittent hypoxia (IH). The mechanisms engaged during this process are not completely understood. In this study, we hypothesized that serotonin (5-HT), acting in the retrotrapezoid nucleus/parafacial respiratory group (RTN/pFRG), is able to generate active expiration. In anesthetized (urethane, ip), tracheostomized, spontaneously-breathing adult male Holtzman rats we microinjected a serotoninergic agonist and antagonist bilaterally in the RTN/pFRG and recorded diaphragm and abdominal muscle activities. We found that episodic (3 times, 5 min apart), but not single microinjections of 5-HT (1 mM) in the RTN/pFRG elicited an enduring (>30 min) increase in abdominal activity. This response was amplified in vagotomized rats and blocked by previous 5-HT receptor antagonism with ketanserin (10 µM). Episodic 5-HT microinjections in the RTN/pFRG also potentiated the inspiratory and expiratory reflex responses to hypercapnia. The antagonism of 5-HT receptors in the RTN/pFRG also prevented the long-term facilitation (>30 min) of abdominal activity in response to acute IH exposure (10 × 6-7% O for 45 s every 5 min). Our findings indicate the activation of serotoninergic mechanisms in the RTN/pFRG is sufficient to increase abdominal expiratory activity at resting conditions and required for the emergence of active expiration after IH in anesthetized animals.

Long-term facilitation of expiratory and sympathetic activities following acute intermittent hypoxia in rats.

Acute intermittent hypoxia (AIH) promotes persistent increases in ventilation and sympathetic activity, referred as long-term facilitation (LTF). Augmented inspiratory activity is suggested as a major component of respiratory LTF. In this study, we hypothesized that AIH also elicits a sustained increase in expiratory motor activity. We also investigated whether the expiratory LTF contributes to the development of sympathetic LTF after AIH. Rats were exposed to AIH (10×6-7% O2 for 45s, every 5min), and the cardiorespiratory parameters were evaluated during 60min using invivo and insitu approaches. In unanesthetized conditions (n=9), AIH elicited a modest but sustained increase in baseline mean arterial pressure (MAP, 104±2 vs. 111±3mmHg, P<0.05) associated with enhanced sympathetic and respiratory-related variabilities. In the insitu preparations (n=9), AIH evoked LTF in phrenic (33±12%), thoracic sympathetic (75±25%) and abdominal nerve activities (69±14%). The sympathetic overactivity after AIH was phase-locked with the emergence of bursts in abdominal activity during the late-expiratory phase. In anesthetized vagus-intact animals, AIH increased baseline MAP (113±3 vs. 122±2mmHg, P<0.05) and abdominal muscle activity (535±94%), which were eliminated after pharmacological inhibition of the retrotrapezoid nucleus/parafacial respiratory group (RTN/pFRG). These findings indicate that increased expiratory activity is also an important component of AIH-elicited respiratory LTF. Moreover, the development of sympathetic LTF after AIH is linked to the emergence of active expiratory pattern and depends on the integrity of the neurones of the RTN/pFRG.

The peptidergic control circuit for sighing.

Sighs are long, deep breaths expressing sadness, relief or exhaustion. Sighs also occur spontaneously every few minutes to reinflate alveoli, and sighing increases under hypoxia, stress, and certain psychiatric conditions. Here we use molecular, genetic, and pharmacologic approaches to identify a peptidergic sigh control circuit in murine brain. Small neural subpopulations in a key breathing control centre, the retrotrapezoid nucleus/parafacial respiratory group (RTN/pFRG), express bombesin-like neuropeptide genes neuromedin B (Nmb) or gastrin-releasing peptide (Grp). These project to the preBötzinger Complex (preBötC), the respiratory rhythm generator, which expresses NMB and GRP receptors in overlapping subsets of ~200 neurons. Introducing either neuropeptide into preBötC or onto preBötC slices, induced sighing or in vitro sigh activity, whereas elimination or inhibition of either receptor reduced basal sighing, and inhibition of both abolished it. Ablating receptor-expressing neurons eliminated basal and hypoxia-induced sighing, but left breathing otherwise intact initially. We propose that these overlapping peptidergic pathways comprise the core of a sigh control circuit that integrates physiological and perhaps emotional input to transform normal breaths into sighs.

Facilitation of the respiratory rhythm in mice by acute exposure to a progestin, the etonogestrel. Involvement of medullary areas

Background Works have underlined the facilitatory influence of progestins on central respiratory drive (CRD). The congenital central hypoventilation syndrome (CCHS) is a neuro-respiratory disease characterized by an alteration of the CO 2 /H + chemosensitivity due to dysfunction of CO 2 /H + chemosensitive/PHOX2b-positive neurons of the retrotrapezoid nucleus/parafacial respiratory group (RTN/pFRG). In our laboratory, a recovery of the CO 2 /H + chemosensitivity was fortuitously observed in two CCHS women who took a potent synthetic progestin, the desogestrel (Respir Physiol Neurobiol 2010;171:171–4). Objective Our aim was to appreciate the effects of an acute exposure to etonogestrel (ETO), the main active metabolite of the desogestrel, on the CRD and to determine its medullary action mechanisms. Methods We used: – an analysis of the effect of ETO (0.05–2 μM) on the CRD of ex vivo medullary-spinal cord preparations from newborn mice in both normopH and metabolic acidosis; – pharmacological co-application of ETO and muscimol or NMDA, agonists of GABA A or NMDA receptors known to be a target of some steroids; – an analysis of the c-fos expression in medullary respiratory structures. Results First, the respiratory frequency was: – dose-dependently increased in normopH by ETO (≈ +23–47%; P – increase by metabolic acidosis similarly under ETO (≈ +28–37% at 0.05–1 μM, P P P Second, changes in respiratory frequency induced by muscimol or NMDA at their EC 50 were more potent under ETO (≈ −60–63%, P P P P c-fos revealed ETO activated cells in respiratory medullary areas i.e. the ventrolateral medulla, nucleus of the solitary tract, RTN/pFRG and medullary raphe nuclei. Conclusions As a whole, data suggest that, by medullary mechanisms, the progestin ETO exerts a facilitatory influence on the CRD by acting at the level of respiratory structures. Involved mechanisms would implicate an interaction with GABA A and NMDA receptors. Besides, at least in newborn mice, ETO do not potentiate the respiratory response to metabolic acidosis by intrinsic medullary mechanisms.

Nociceptin/orphanin FQ slows inspiratory rhythm via its direct effects on the pre-Bötzinger complex

In a previous study, we showed that in an in vitro en bloc preparation of newborn rats perfused with standard [K+] (6.2mM) and high [K+] (11.2mM) artificial cerebrospinal fluid (aCSF), nociceptin/orphanin FQ (N/OFQ) suppresses bursting of pre-inspiratory neurons with 1:1 coupling to the fictive inspiration. However, it is unclear whether the pre-Bötzinger complex (preBötC) is involved in the N/OFQ-induced slowing. Using in vitro en bloc preparations with and without the retrotrapezoid nucleus/parafacial respiratory group (RTN/pFRG) perfused with high [K+] aCSF, we found the following: (1) there were no differences in the effects of N/OFQ on the inspiratory rhythm between the preparations with and without the RTN/pFRG, (2) N/OFQ decreased the input resistance of inspiratory neurons (Insps) in the preparations without the RTN/pFRG and suppressed their ectopic firing activities, and (3) N/OFQ suppressed the spontaneous firing of Insps under a chemical synaptic transmission blockade. In conclusion, it is possible that the preBötC is involved in N/OFQ-induced inspiratory rhythm slowing.

A closed-loop model of the respiratory system: focus on hypercapnia and active expiration.

Breathing is a vital process providing the exchange of gases between the lungs and atmosphere. During quiet breathing, pumping air from the lungs is mostly performed by contraction of the diaphragm during inspiration, and muscle contraction during expiration does not play a significant role in ventilation. In contrast, during intense exercise or severe hypercapnia forced or active expiration occurs in which the abdominal “expiratory” muscles become actively involved in breathing. The mechanisms of this transition remain unknown. To study these mechanisms, we developed a computational model of the closed-loop respiratory system that describes the brainstem respiratory network controlling the pulmonary subsystem representing lung biomechanics and gas (O2 and CO2) exchange and transport. The lung subsystem provides two types of feedback to the neural subsystem: a mechanical one from pulmonary stretch receptors and a chemical one from central chemoreceptors. The neural component of the model simulates the respiratory network that includes several interacting respiratory neuron types within the Bötzinger and pre-Bötzinger complexes, as well as the retrotrapezoid nucleus/parafacial respiratory group (RTN/pFRG) representing the central chemoreception module targeted by chemical feedback. The RTN/pFRG compartment contains an independent neural generator that is activated at an increased CO2 level and controls the abdominal motor output. The lung volume is controlled by two pumps, a major one driven by the diaphragm and an additional one activated by abdominal muscles and involved in active expiration. The model represents the first attempt to model the transition from quiet breathing to breathing with active expiration. The model suggests that the closed-loop respiratory control system switches to active expiration via a quantal acceleration of expiratory activity, when increases in breathing rate and phrenic amplitude no longer provide sufficient ventilation. The model can be used for simulation of closed-loop control of breathing under different conditions including respiratory disorders.

Leptin into the ventrolateral medulla facilitates chemorespiratory response in leptin-deficient (ob/ob) mice.

Leptin, an adipocyte-derived hormone, is suggested to participate in the central control of breathing. We hypothesized that leptin may facilitate ventilatory responses to chemoreflex activation by acting on respiratory nuclei of the ventrolateral medulla. The baseline ventilation and the ventilatory responses to CO2 were evaluated before and after daily injections of leptin into the retrotrapezoid nucleus/parafacial respiratory group (RTN/pFRG) for 3 days in obese leptin-deficient (ob/ob) mice. Male ob/ob mice (40-45 g, n = 7 per group) received daily microinjections of vehicle or leptin (1 μg per 100 nL) for 3 days into the RTN/pFRG. Respiratory responses to CO2 were measured by whole-body plethysmography. Unilateral microinjection of leptin into the RTN/pFRG in ob/ob mice increased baseline ventilation (VE ) from 1447 ± 96 to 2405 ± 174 mL min(-1) kg(-1) by increasing tidal volume (VT ) from 6.4 ± 0.4 to 9.1 ± 0.8 mL kg(-1) (P < 0.05). Leptin also enhanced ventilatory responses to 7% CO2 (Δ = 2172 ± 218 mL min(-1) kg(-1) , vs. control: Δ = 1255 ± 105 mL min(-1) kg(-1) ), which was also due to increased VT (Δ = 4.71 ± 0.51 mL kg(-1) , vs. control: Δ = 2.27 ± 0.20 mL kg(-1) ), without changes in respiratory frequency. Leptin treatment into the RTN/pFRG or into the surrounding areas decreased food intake (83 and 70%, respectively), without significantly changing body weight. The present results suggest that leptin acting in the respiratory nuclei of the ventrolateral medulla improves baseline VE and VT and facilitates respiratory responses to hypercapnia in ob/ob mice.

Understanding the Rhythm of Breathing: So Near, Yet So Far

Breathing is an essential behavior that presents a unique opportunity to understand how the nervous system functions normally, how it balances inherent robustness with a highly regulated lability, how it adapts to both rapidly and slowly changing conditions, and how particular dysfunctions result in disease. We focus on recent advancements related to two essential sites for respiratory rhythmogenesis: (a) the preBötzinger Complex (preBötC) as the site for the generation of inspiratory rhythm and (b) the retrotrapezoid nucleus/parafacial respiratory group (RTN/pFRG) as the site for the generation of active expiration.

A closed model for the respiratory system in mammals

The interactions between rhythmogenic neuronal networks and physiological systems are crucial components in understanding how a system maintains homeostasis under diverse circumstances. Various aspects of such interactions should be addressed including how rhythms are generated in neuronal networks, how feedback signals interact with neuronal networks to regulate levels of important physiological quantities, and how changes in demands on the system can be handled in a way that restores homeostasis. In this work, we explore these issues in the context of neuronal networks in the mammalian brainstem that generate the respiratory rhythm when isolated from the physiological systems that drive actual breathing [1]. These networks include neuronal components within the Botzinger (BotC) and pre-Botzinger (pre-BotC) complexes and the retrotrapezoid nucleus/parafacial respiratory group (RTN/pFRG). Experimental results on respiratory rhythm generation have suggested that neurons within the BotC and pre-BotC are responsible for the generation of primary respiratory rhythms, which are modulated by oscillations in the RTN/pFRG. Previous computational work has elucidated a possible set of mechanisms through which these brain stem neurons interact and produce a variety of experimental results [2]. Separate past work has developed a physiologically detailed yet tractable model of respiratory physiology [3]. This model is comprised of a very simplified version of a neuronal network oscillator that generates respiratory rhythms, an integrator that delivers respiratory output , and lung components responsible for air exchange. This earlier study considered reasonable ways to link the neuronal oscillator/integrator with these physiological components in a closed loop feedback system to achieve respiratory frequency and amplitude control. In our work, we simulate and analyze a model that links the detailed neuronal rhythm generator with the physiological lung components. We consider the roles of mechanical and chemoreceptive feedback signals in maintaining stable respiratory rhythms and in responding to metabolic or environmental perturbations. In particular, we study the response to hypercapnia, or excessive carbon dioxide [4]. The idea is that excessive exposure to CO2 may lead to CO2 accumulation in the blood, which will recruit certain neuronal elements in the RTN/pFRG and alter the drive to core rhythmogenic elements. Changes in rhythmic output patterns activate certain muscles that alter breathing in a way that restores CO2 homeostasis. We analyze the roles of neurons’ intrinsic dynamic features, the architecture and strengths of synaptic connections among neurons, and the roles of particular feedback systems in achieving baseline rhythms and appropriate responses to perturbations.

Postnatal development of Na+–K+–2Cl− co-transporter 1 and K+–Cl− co-transporter 2 immunoreactivity in multiple brain stem respiratory nuclei of the rat

Previously, we reported that in rats, GABAA and glycine receptor immunoreactivity increased markedly in multiple brain stem respiratory nuclei around postnatal days (P) 12–13, a critical period when abrupt neurochemical, metabolic, ventilatory, and electrophysiological changes occur in the respiratory network and when the system is under greater inhibition than excitation. Since Na+–K+–2Cl− co-transporter 1 (NKCC1) and K+–Cl− co-transporter 2 (KCC2) play pivotal roles in determining the responses of GABAA and glycine receptors, we hypothesized that NKCC1 and KCC2 undergo significant changes during the critical period. An in-depth immunohistochemical and single neuron optical densitometric study of neurons in seven respiratory-related nuclei (the pre-Bötzinger complex [PBC], nucleus ambiguus [Amb], hypoglossal nucleus [XII], ventrolateral subnucleus of solitary tract nucleus [NTSVL], retrotrapezoid nucleus/parafacial respiratory group [retrotrapezoid nucleus/parafacial respiratory group (RTN/pFRG)], dorsal motor nucleus of the vagus nerve [dorsal motor nucleus of the vagus nerve (DMNX)], and inferior olivary nucleus [IO]) and a non-respiratory cuneate nucleus (CN, an internal control) was undertaken in P0–P21 rats. Our data revealed that (1) NKCC1 immunoreactivity exhibited a developmental decrease from P0 to P21 in all eight nuclei examined, being relatively high during the first 1½ postnatal weeks and decreased thereafter. The decrease was abrupt and statistically significant at P12 in the PBC, Amb, and XII; (2) KCC2 immunoreactivity in these eight nuclei showed a developmental increase from P0 to P21; and (3) the significant reduction in NKCC1 and the greater dominance of KCC2 around P12 in multiple respiratory nuclei of the brain stem may form the basis of an enhanced inhibition in the respiratory network during the critical period before the system stabilizes to a more mature state.

State-dependent control of the respiratory pattern and coupled oscillators

The respiratory rhythm and motor pattern are generated by a spatially organized brain stem respiratory network with a rhythmogenic core comprising interacting neural populations within the pre-Botzinger (pre-BotC) and Botzinger (BotC) complexes controlled by drives and inputs from other brain stem compartments. Our previous large-scale computational model [3, 4] reproduced the behavior of this network under multiple natural and experimental conditions, but did not consider neural oscillations that were proposed to emerge within the retrotrapezoid nucleus / parafacial respiratory group (RTN/pFRG) and drive pre-inspiratory (or late-expiratory, late-E) oscillations in the abdominal motor output [2]. These late-E oscillations usually emerge with increasing metabolic demands (e.g., during hypercapnia or hypoxia) and couple with the respiratory oscillations generated by the BotC-pre-BotC core of the respiratory central pattern generator [1, 2]. In our experiments, performed in the arterially perfused in situ rat preparation, the respiratory motor activity was recorded simultaneously from abdominal, phrenic, hypoglossal and central vagus nerves. Under normal conditions (eupnea), the abdominal motor outflow showed only a low-amplitude post-inspiratory (post-I) activity. An expression of the abdominal late-E bursts, preceding and coupled to phrenic discharges, occurred during hypercapnia (7–10% CO2) and could be abolished by pharmacological inactivation of the RTN/pFRG region and by administration of riluzole, a persistent sodium current (I NaP ) blocker, hence confirming that the late-E oscillations originate in RTN/pFRG and involve intrinsic, I NaP -dependent mechanisms. Using these data we extended our computational model by incorporating in the RTN/pFRG compartment an additional late-E population (with neurons containing I NaP ) serving as a source of abdominal late-E activity. The proposed interactions between the BotC’s and pre-BotC’s populations and the late-E population of RTN/pFRG allowed the model to reproduce several experimentally observed behaviors, including quantal acceleration of abdominal late-E oscillations with progressive hypercapnia and quantal slowing of phrenic activity with progressive suppression of pre-BotC excitability (simulating the suppressing effects of opioids), as well as to predict a release of late-E oscillations by disinhibition of RTN/pFRG under normal conditions. The latter prediction has been confirmed in our experimentally studies. Our experimental and modeling studies suggest that under normal metabolic conditions the RTN/pFRG oscillator is inhibited by both the post-I population of BotC during inspiration and early-inspiratory (early-I) population of preBotC during inspiration. Therefore the late-E oscillations can be released by either a hypercapnia-evoked activation of chemosensitive RTN/pFRG neurons overcoming this inhibition or a hypoxia-dependent suppression of RTN/pFRG inhibition by BotC-pre-BotC circuits. Our model proposes mechanistic explanations for the emergence of RTN/pFRG oscillations and their interaction with the brain stem respiratory network. The emerging view is that the brain stem respiratory network has rhythmogenic capabilities at multiple hierarchical levels, which allows flexible, state-dependent expression of different rhythmogenic mechanisms under different physiological and metabolic conditions and enables a wide repertoire of respiratory behaviors.

Intermittent hypoxia-induced sensitization of central chemoreceptors contributes to sympathetic nerve activity during late expiration in rats

Hypertension elicited by chronic intermittent hypoxia (CIH) is associated with elevated activity of the thoracic sympathetic nerve (tSN) that exhibits an enhanced respiratory modulation reflecting a strengthened interaction between respiratory and sympathetic networks within the brain stem. Expiration is a passive process except for special metabolic conditions such as hypercapnia, when it becomes active through phasic excitation of abdominal motor nerves (AbN) in late expiration. An increase in CO(2) evokes late-expiratory (late-E) discharges phase-locked to phrenic bursts with the frequency increasing quantally as hypercapnia increases. In rats exposed to CIH, the late-E discharges synchronized in AbN and tSN emerge in normocapnia. To elucidate the possible neural mechanisms underlying these phenomena, we extended our computational model of the brain stem respiratory network by incorporating a population of presympathetic neurons in the rostral ventrolateral medulla that received inputs from the pons, medullary respiratory compartments, and retrotrapezoid nucleus/parafacial respiratory group (RTN/pFRG). Our simulations proposed that CIH conditioning increases the CO(2) sensitivity of RTN/pFRG neurons, causing a reduction in both the CO(2) threshold for emerging the late-E activity in AbN and tSN and the hypocapnic threshold for apnea. Using the in situ rat preparation, we have confirmed that CIH-conditioned rats under normal conditions exhibit synchronized late-E discharges in AbN and tSN similar to those observed in control rats during hypercapnia. Moreover, the hypocapnic threshold for apnea was significantly lowered in CIH-conditioned rats relative to that in control rats. We conclude that CIH may sensitize central chemoreception and that this significantly contributes to the neural impetus for generation of sympathetic activity and hypertension.

Active expiration induced by excitation of ventral medulla in adult anesthetized rats.

Data from perinatal and juvenile rodents support our hypothesis that the preBötzinger complex generates inspiratory rhythm and the retrotrapezoid nucleus-parafacial respiratory group (RTN/pFRG) generates active expiration (AE). Although the role of the RTN/pFRG in adulthood is disputed, we hypothesized that its rhythmogenicity persists but is typically silenced by synaptic inhibition. We show in adult anesthetized rats that local pharmacological disinhibition or optogenetic excitation of the RTN/pFRG can generate AE and transforms previously silent RTN/pFRG neurons into rhythmically active cells whose firing is correlated with late-phase active expiration. Brief excitatory stimuli also reset the respiratory rhythm, indicating strong coupling of AE to inspiration. The AE network location in adult rats overlaps with the perinatal pFRG and appears lateral to the chemosensitive region of adult RTN. We suggest that (1) the RTN/pFRG contains a conditional oscillator that generates AE, and (2) at rest and in anesthesia, synaptic inhibition of RTN/pFRG suppresses AE.

Interacting oscillations in neural control of breathing: modeling and qualitative analysis

In mammalian respiration, late-expiratory (late-E, or pre-inspiratory) oscillations emerge in abdominal motor output with increasing metabolic demands (e.g., during hypercapnia, hypoxia, etc.). These oscillations originate in the retrotrapezoid nucleus/parafacial respiratory group (RTN/pFRG) and couple with the respiratory oscillations generated by the interacting neural populations of the Bötzinger (BötC) and pre-Bötzinger (pre-BötC) complexes, representing the kernel of the respiratory central pattern generator. Recently, we analyzed experimental data on the generation of late-E oscillations and proposed a large-scale computational model that simulates the possible interactions between the BötC/pre-BötC and RTN/pFRG oscillations under different conditions. Here we describe a reduced model that maintains the essential features and architecture of the large-scale model, but relies on simplified activity-based descriptions of neural populations. This simplification allowed us to use methods of dynamical systems theory, such as fast-slow decomposition, bifurcation analysis, and phase plane analysis, to elucidate the mechanisms and dynamics of synchronization between the RTN/pFRG and BötC/pre-BötC oscillations. Three physiologically relevant behaviors have been analyzed: emergence and quantal acceleration of late-E oscillations during hypercapnia, transformation of the late-E activity into a biphasic-E activity during hypercapnic hypoxia, and quantal slowing of BötC/pre-BötC oscillations with the reduction of pre-BötC excitability. Each behavior is elicited by gradual changes in excitatory drives or other model parameters, reflecting specific changes in metabolic and/or physiological conditions. Our results provide important theoretical insights into interactions between RTN/pFRG and BötC/pre-BötC oscillations and the role of these interactions in the control of breathing under different metabolic conditions.

Late-Expiratory Activity: Emergence and Interactions With the Respiratory CPG

The respiratory rhythm and motor pattern are hypothesized to be generated by a brain stem respiratory network with a rhythmogenic core consisting of neural populations interacting within and between the pre-Bötzinger (pre-BötC) and Bötzinger (BötC) complexes and controlled by drives from other brain stem compartments. Our previous large-scale computational model reproduced the behavior of this network under many different conditions but did not consider neural oscillations that were proposed to emerge within the retrotrapezoid nucleus/parafacial respiratory group (RTN/pFRG) and drive preinspiratory (or late-expiratory, late-E) discharges in the abdominal motor output. Here we extend the analysis of our previously published data and consider new data on the generation of abdominal late-E activity as the basis for extending our computational model. The extended model incorporates an additional late-E population in RTN/pFRG, representing a source of late-E oscillatory activity. In the proposed model, under normal metabolic conditions, this RTN/pFRG oscillator is inhibited by BötC/pre-BötC circuits, and the late-E oscillations can be released by either hypercapnia-evoked activation of RTN/pFRG or by hypoxia-dependent suppression of RTN/pFRG inhibition by BötC/pre-BötC. The proposed interactions between BötC/pre-BötC and RTN/pFRG allow the model to reproduce several experimentally observed behaviors, including quantal acceleration of abdominal late-E oscillations with progressive hypercapnia and quantal slowing of phrenic activity with progressive suppression of pre-BötC excitability, as well as to predict a release of late-E oscillations by disinhibition of RTN/pFRG under normal conditions. The extended model proposes mechanistic explanations for the emergence of RTN/pFRG oscillations and their interaction with the brain stem respiratory network.

Hypoxia-sensing properties of the newborn rat ventral medullary surface in vitro.

The ventral medullary surface (VMS) is a region known to exert a respiratory stimulant effect during hypercapnia. Several studies have suggested its involvement in the central inhibition of respiratory rhythm caused by hypoxia. We studied brainstem-spinal cord preparations isolated from newborn rats transiently superfused with a very low O(2) medium, causing reversible respiratory depression, to characterize the participation of the VMS in hypoxic respiratory adaptation. In the presence of 0.8 mM Ca(2+), very low O(2) medium induced an increase in c-fos expression throughout the VMS. The reduction of synaptic transmission and blockade of the respiratory drive by 0.2 mM Ca(2+)-1.6 mM Mg(2+) abolished c-fos expression in the medial VMS (at the lateral edge of the pyramidal tract) but not in the perifacial retrotrapezoid nucleus/parafacial respiratory group (RTN/pFRG) VMS, suggesting the existence of perifacial RTN/pFRG hypoxia-sensing neurons. In the presence of Ca(2+) (0.8 mM), lesioning experiments suggested a physiological difference in perifacial RTN/pFRG VMS between the lateral VMS (beneath the ventrolateral part of the facial nucleus) and the middle VMS (beneath the ventromedial part of the facial nucleus), at least in newborn rats. The lateral VMS lesion, corresponding principally to the most rostral part of the pFRG, produced hypoxia-induced stimulation, whereas the middle VMS lesion, corresponding to the main part of the RTN, abolished hypoxic excitation. This may involve relay via the medial VMS, which is thought to be the parapyramidal group.