Expiratory Activity Research Articles

A Physiological Point of View on Expiratory (Re)action during Mechanical Ventilation

A Physiological Point of View on Expiratory (Re)action during Mechanical Ventilation

GABAergic neurons of the medullary raphe regulate active expiration during hypercapnia.

The parafacial respiratory group (pFRG), located in the lateral aspect of the rostroventral lateral medulla, has been described as a conditional expiratory oscillator that emerges mainly in conditions of high metabolic challenges to increase breathing. The convergence of inhibitory and excitatory inputs to pFRG and the generation of active expiration may be more complex than previously thought. We hypothesized that the medullary raphe, a region that has long been described to be involved in breathing activity, is also responsible for the expiratory activity under hypercapnic condition. To test this hypothesis, we performed anatomical and physiological experiments in urethane-anesthetized adult male Wistar rats. Our data showed anatomical projections from serotonergic (5-HT-ergic) and GABAergic neurons of raphe magnus (RMg) and obscurus (ROb) to the pFRG region. Pharmacological inhibition of RMg or ROb with muscimol (60 pmol/30 nL) did not change the frequency or amplitude of diaphragm activity and did not generate active expiration. However, under hypercapnia (9-10% CO2), the inhibition of RMg or ROb increased the amplitude of abdominal activity, without changing the increased amplitude of diaphragm activity. Depletion of serotonergic neurons with saporin anti-SERT injections into ROb and RMg did not increase the amplitude of abdominal activity during hypercapnia. These results show that the presumably GABAergic neurons within the RMg and ROb may be the inhibitory source to modulate the activity of pFRG during hypercapnia condition.NEW & NOTEWORTHY Medullary raphe has been involved in the inspiratory response to central chemoreflex; however, these reports have never addressed the role of raphe neurons on active expiration induced by hypercapnia. Here, we showed that a subset of GABA cells within the medullary raphe directly project to the parafacial respiratory region, modulating active expiration under high levels of CO2.

Retrograde projections to the expiratory oscillator, the parafacial respiratory group, in adult male rats

The parafacial respiratory group (pFRG) is a conditional oscillator known to drive active expiration and enhance ventilation during high metabolic and respiratory demands, through the recruitment of abdominal (ABD) expiratory muscles. At rest and under anesthesia, pFRG neurons appear to be silent, but when disinhibited or activated, they trigger active expiration. Furthermore pFRG modulation appears to be critical for recruitment of expiratory ABD activity during REM sleep. There is extensive evidence for functional modulation of pFRG and its role in active expiration, although no systematic analysis of neuroanatomical projections to the pFRG is currently available. Thus, the objectives of this study are: 1) to identify brainstem regions that directly project to pFRG; and 2) define phenotypes of these projections, mainly excitatory and/or inhibitory.To test the objective 1, we used a retrograde virus (herpes simplex virus) injected unilaterally in the pFRG area in adult male rats that expressed the reporter protein eYFP (enhanced yellow fluorescent protein) driven by the neuronal promoter synapsin in order to track neurons projecting to the pFRG. To determine the phenotype of these projections (objective 2), we combined immunofluorescent labeling for eYFP with different neuronal markers: somatostatin (SST), choline acetyltransferase (CHAT), tyrosine hydroxylase (TH), paired Like Homeobox 2 (PHOX2B) and 5‐hydroxytryptamine (5HT); and we used RNAScope assay to target mRNA for Vesicular Glutamate Transporter (Vglut2); Glutamate Decarboxylase 1 (Gad1) and Glycine Transporter 2 (Glyt2).We identified several brainstem structures that directly project to pFRG ipsilaterally and contralaterally: in the medulla, projecting neurons were located in the ventral respiratory column (Bötzinger Complex, preBötzinger Complex, rostral ventral respiratory group, retrotrapezoid nucleus ‐ RTN and pFRG), the nucleus of the solitary tract (NTS), the reticular formation (RF), the vestibular nuclei (extended to the pons and midbrain), the medullary raphe and the nucleus of Roller. Several projecting neurons were also located rostrally in the pons and midbrain within the Kölliker‐Fuse nucleus, the parabrachial nucleus, the periaqueductal grey, the pedunculopontine nucleus (PPT), the laterodorsal tegmental (LDT) and the A5 area. Overall, the ipsilateral distribution was more extensive than in the contralateral side of the virus injection. Phenotype analysis demonstrated the presence of both excitatory (Vglut2+) and inhibitory (Gad1/Glyt2+) cells projecting to pFRG deriving from all the above regions, in addition to PHOX2B+ cells located in the RTN, NTS and part of the RF. Moreover, we observed lack of direct projections to the pFRG area from TH+ cells in the NTS, A5 and Locus coeruleus; 5HT+ cells in the raphe nuclei; SST+ cells in the preBötzinger Complex and CHAT+ cells in the PPT and LDT.The present study provides evidence that pFRG neurons receive excitatory and inhibitory inputs from several breathing‐ and non‐breathing related brainstem regions that could be involved with their conditional activity and the modulation of active expiration in various physiological conditions.Support or Funding InformationCIHR 156338, NSERC 435843; WCHRI Fellowship (VB)

Selective optogenetic stimulation of RTN neurons drives ventilation and active expiration in unanesthetized rats

BackgroundThe retrotrapezoid nucleus (RTN) consists of neurons that are chemosensitive (i.e., CO2/pH sensitive), glutamatergic, Phox2b+ and Neuromedin B+ (Nmb+). Lentiviral vectors currently used to transduce RTN neurons with various opto‐ or pharmacogenetic actuators contain a Phox2b‐activated promoter, PRSx8. When injected into the RTN, these vectors also transduce substantial numbers of nearby C1 and catecholaminergic neurons. The purpose of the present study was to determine the effects on breathing elicited by selectively activating RTN neurons.MethodsTo selectively transduce RTN neurons we microinjected a mixture of Lenti‐PRSx8‐ChR2‐mCherry and AAV5‐FLEX‐taCasP3‐TEVp into the RTN of adult TH‐Cre rats. One month after the surgery, RTN neurons were stimulated with blue light (473 nm, 10 ms pulse, 20 Hz, for 20 s). All the measurements were conducted inside a plethysmography chamber. Diaphragm (DIA) and abdominal (ABD) muscle activity were recorded in conscious conditions. Results (mean ± SD), were analyzed using one‐ or two‐way ANOVA as required and differences were considered significant at P<0.05.ResultsRTN neurons (Nmb+, TH−) were selectively transduced (96.8 % ± 4) with the PRSx8 vector, the FLEX‐caspase AAV5 having caused the apoptotic death of C1 neurons (TH+) in the area where neurons were transduced with ChR2. Selective stimulation of RTN (NMB+ neurons; n=6) increased respiratory rate during wake and NREM sleep but not during REM sleep (Δ 30 ± 7 vs. 22.7 ± 6 vs. 4 ± 4 bpm from baseline), increased tidal volume also during quiet waking and non‐REM but not REM sleep (Δ 0.15 ± 0.07 vs. 0.13 ± 0.04 vs. 0.02 ± 0.007 mL/100g) and VE also during wake and non‐REM sleep but not REM (27.8 ± 11 vs. 21.4 ± 8 vs. 3.1 ±1 mL/100g/min). Selective RTN stimulation in rats breathing room air elicited active expiration (from 8.4 ± 7 to 15.9 ± 8 % of normalized abdominal EMG recorded at 9% FiCO2, n=3). RTN stimulation produced greater active expiration in rats exposed to 6% FiCO2 (from 56.3 ± 32 to 125.3 ± 104 % of normalized abdominal EMG).ConclusionsSelective RTN neuron stimulation drives breathing amplitude and frequency during wake, NREM but not in REM sleep, and elicits active expiration, especially at elevated levels of FiCO2.Support or Funding InformationSupported by grants from the National Institutes of Health (HL28785, 18 HL074011) and the Congenital Central Hypoventilation Syndrome Foundation to PGG.

Model‐Based Analysis of Lower Thoracic High‐Frequency Spinal Cord Stimulation (HF‐SCS) to Restore Effective Cough

BackgroundSpinal cord stimulation (SCS, 50 Hz) is a useful method to restore an effective cough in persons with spinal cord injury (SCI). In animal models, dorsal HF‐SCS (~500 Hz) produces similar level of expiratory muscle activation with much lower stimulus amplitudes. Currently, the mechanism of this behavior is unknown.ObjectiveThe aim of the present study was to develop a computational model of lower thoracic SCS to investigate the mechanisms of the expiratory muscle activation via HF‐SCS.MethodsWe created a computer model of an anatomically‐accurate finite element model (FEM) of the lower thoracic (T9) canine spinal cord and surrounding anatomy with an implanted SCS electrode array. This model consisted of two main components: 1) multicompartment cable models of dorsal afferent fibers and 2) ventral motor axons. Model analysis allowed calculation of the extracellular voltages generated during SCS and the corresponding activation thresholds of each fiber type.ResultsIn our simulations, we calculated the activation thresholds for several electrode combinations and stimulation frequencies (50 Hz, 500 Hz). As expected, the activation thresholds of the ventral motor axons were significantly higher than the activation thresholds of dorsal root axons. Therefore, low‐frequency SCS likely functions through the direct activation of ventral motor axons, while high‐frequency SCS operates via recruitment of myelinated afferent fibers at significantly lower stimulation amplitudes. In our simulations, the stimulation frequency did not significantly alter activation thresholds in dorsal or ventral axons.ConclusionsOur results suggest that both high‐ and low‐frequency SCS directly activate dorsal rootlets at low amplitudes (~1 mA); however, the trans‐synaptic spinal circuits responsible for eliciting cough are only effective at higher stimulation frequencies. Consequently, low‐frequency stimulation results in effective cough by bypassing this circuitry entirely to directly activate the ventral motor axons at the cost of requiring higher stimulation amplitudes. In future work, we will perform simulations directly assessing the ability of SCS to engage these spinal circuits. We will compare our model results to experimental measurements in an animal model of SCS to produce cough.DisclosureDr. DiMarco holds two U.S. patents for technology related to the content of this abstract: Method and Apparatus for Electrical Activation of the Expiratory Muscles to Restore Cough (5,999,855) and Bipolar Spinal Cord Stimulation to Activate the Expiratory Muscles to Restore Cough (8,751,004).Support or Funding InformationMerit Review Award I01‐RX‐001488 from the U.S. Department of Veterans Affairs, Rehabilitation Research and Development Service.

Effects of Different Systemic Opioid Doses on Subareas of the Ventral Respiratory Column

BackgroundThe Parabrachial Nucleus (PBN)/Kölliker‐Fuse (KF) complex are important subareas of the ventral respiratory column (VRC) that regulate inspiratory on‐ ( 1) and off‐switch ( 2). We have shown in vagus‐intact rabbits that respiratory rate depression caused by clinical opioid concentrations was in part mediated by the PBN ( 3), and that systemic opioid effects on respiratory phase timing could be partially reversed in the preBötzinger complex (preBC) albeit without reversing respiratory rate depression ( 4). The present study was to determine how much the combination of local microinjections of naloxone into the KF, PBN and preBC could reverse respiratory depression from intravenous remifentanil (REMI).MethodsThe study was approved by the local Animal Care Committee and conformed to NIH standards. Adult New Zealand White rabbits (3–4kg) were anesthetized, tracheotomized, ventilated, decerebrated and vagotomized. Phrenic nerve activity was recorded from the c5 rootlet, time averaged and used to calculate inspiratory and expiratory duration and peak phrenic activity (PPA). Gridwise localized pressure microinjections of α‐amino‐3‐hydroxy‐5‐methyl‐4‐isoxazolepropionic acid (AMPA, 50 μM, 70 nl) utilizing a multibarrel glass pipette into the VRC caudal to the inferior collicle identified the PBN where AMPA injection caused significant tachypnea and the KF where AMPA caused transient bradypnea and a decrease in PPA. The tachypneic response to AMPA injection in the medulla was used to identify the preBC area. After functional identification of the PBN, KF and preBC areas, we injected a bolus of 10mcg REMI IV, which resulted in apnea. On return of the breathing pattern, a REMI infusion was started to achieve a steady‐state respiratory rate depression of 50% of control rate. After reaching steady‐state, we injected naloxone (1mM, 700nl) into the bilateral KF, PBN and preBC. We then repeated the REMI bolus injection with the apneic dose of 10mcg. In a subset of animals, the 10 mcg REMI bolus was repeated after naloxone injection into the KF/PBN and after injection into the preBC. Residual respiratory depression was reversed at the end of the protocol with IV naloxone (300μM). Drug effects were compared with one‐way RM ANOVA. Data are expressed as mean±SD.ResultsIn 11 animals, IV REMI depressed respiratory rate from 39±7 to 18±6 breaths per minute (BPM). Naloxone injection recovered respiratory rate to 20±9 in the KF (p=0.36), to 26±9 after additional injection into the PBN (p<0.001), and to 28±7 bpm after injection into the preBC area (p=0.54). The IV REMI bolus caused apnea >30s at control but only depressed respiratory rate to 12±6 bpm after naloxone injection into the KF/PBN (p<0.001) and to 16±7 after additional naloxone reversal in the preBC area (p=0.21, n=5).ConclusionRespiratory rate depression from IV opioids is mostly mediated by the PBN, however, higher opioid doses may also affect other brainstem areas.Support or Funding InformationSupported by NIH R01GM112960701A1 (Dr. Stucke) and VA merit grant I01BX000721 (Dr. Zuperku)Figure 1

Water Deprivation Enhances the Late Expiratory Activity of Abdominal Nerve During Hypercapnia and Hypoxia in Rats

It is known that osmolarity challenges produce reflex adjustments in cardiorespiratory function. Herein, we investigated whether 48 h of water deprivation (WD) modifies respiratory pattern under room air conditions and in response to either hypercapnia or hypoxia using male juvenile (60–90 g) and adults (280–320 g) Holtzman rats. Control rats (n=6–7/group) had continuous access to tap water and food, whereas water‐deprived (WD; n=6–8/group) rats had water, but not food removed for 48 h before the experiments. Adult unanesthetized rats were placed in whole‐body plethysmographic chamber (5 L) for recording oxygen consumption (VO2), carbon dioxide production (VCO2) and ventilatory parameters. VO2 and VCO2 were evaluated under baseline conditions (room air), whereas respiratory frequency (fR) and tidal volume (VT) were measured during baseline, hypercapnia (7% CO2 balanced in 93% O2, for 5 min) or hypoxia (7% O2 balanced in N2, for 5 min). Decorticated, arterially‐perfused in situ preparations of juvenile rats were also used for recording respiratory motor outputs during normocapnia (5% CO2), hypercapnia (8% CO2) and while potassium cyanide (KCN; 25 μg in 50 μL) was intra‐arterially infused. WD rats presented a reduction in VO2 (18 ± 0.5 mL.kg−1.min−1 STPD, vs. control: 23 ± 1.2 mL.kg−1.min−1 STPD; p<0.05) and VCO2 (17 ± 0.5 mL.kg−1.min−1 STPD, vs. control: 26 ± 1.5 mL.kg−1.min−1 STPD; p<0.05). At baseline, WD rats exhibited increased VT (9.7 ± 0.5 mL.kg−1, vs. control: 7.7 ± 0.3 mL.kg−1; p<0.05), but reduced fR (76 ± 2 breaths.min−1, vs. control: 103 ± 7 breaths.min−1; p<0.05), resulting in similar ventilation (VE) between groups (746 ± 54 mL.kg−1.min−1, vs. control: 799 ± 64 mL.kg−1.min−1). During hypercapnia, WD rats had a higher increase in VT (15.3 ± 1.1 mL.kg−1, vs. control: 10.2 ± 0.3 mL.kg−1; p<0.05) and VE (1745 ± 202 mL.kg−1.min−1, vs. control: 1239 ± 49 mL.kg−1.min−1; p<0.05) compared to control rats, with similar increase in fR (112 ± 5 breaths.min−1, vs. control: 122 ± 6 breaths.min−1). Under hypoxia, WD rats presented a greater increase in VT (12.2 ± 0.4 mL.kg−1, vs. control: 8.1 ± 0.8 mL.kg−1; p<0.05) and VE (1247 ± 71 mL.kg−1.min−1, vs. control: 991 ± 84 mL.kg−1.min−1; p<0.05) than control rats, with a lower increase in fR (102 ± 4 breaths.min−1, vs. control: 122 ± 3 breaths.min−1; p<0.05). In in situ preparations, the respiratory motor outputs at baseline were not modified by WD, however, WD produced a greater increase in late expiratory activity of abdominal (ABD) nerve during hypercapnia (22 ± 2 μV, vs. control: 15 ± 2 μV; p<0.05) and hypoxia (14 ± 2 μV, vs. control: 7 ± 1 μV; p<0.05). The results show that 48 h of WD produces a hypometabolic stated, modifies baseline VE and enhances the ventilatory response to hypercapnia and hypoxia in conscious rats. The increased ABD nerve activity might contribute to ventilatory response after WD.Support or Funding InformationCapes, CNPq, FAPESP (grant numbers: 2015/23467‐7; 2017/04649‐2)

C1 neurons are part of the circuitry that recruits active expiration in response to the activation of peripheral chemoreceptors.

Breathing results from the interaction of two distinct oscillators: the pre-Bötzinger Complex (preBötC), which drives inspiration; and the lateral parafacial region (pFRG), which drives active expiration. The pFRG is silent at rest and becomes rhythmically active during the stimulation of peripheral chemoreceptors, which also activates adrenergic C1 cells. We postulated that the C1 cells and the pFRG may constitute functionally distinct but interacting populations for controlling expiratory activity during hypoxia. We found in rats that: a) C1 neurons are activated by hypoxia and project to the pFRG region; b) active expiration elicited by hypoxia was blunted after blockade of ionotropic glutamatergic receptors at the level of the pFRG; and c) selective depletion of C1 neurons eliminated the active expiration elicited by hypoxia. These results suggest that C1 cells may regulate the respiratory cycle, including active expiration, under hypoxic conditions.

Heart to breathe: partial ablation of rostral ventrolateral medulla catecholaminergic neurons mediates disordered breathing in volume overload heart failure rats

Heart to breathe: partial ablation of rostral ventrolateral medulla catecholaminergic neurons mediates disordered breathing in volume overload heart failure rats

Active expiratory oscillator regulates nasofacial and oral motor activities in rats.

What is the central question of this study? Does the parafacial respiratory group (pFRG), which mediates active expiration, recruit nasofacial and oral motoneurons to coordinate motor activities that engage muscles controlling airways in rats during active expiration. What is the main finding and its importance? Hypercapnia/acidosis or pFRG activation evoked active expiration and stimulated the motoneurons and nerves responsible for the control of nasofacial and oral airways patency simultaneously. Bilateral pFRG inhibition abolished active expiration and the simultaneous nasofacial and oral motor activities induced by hypercapnia/acidosis. The pFRG is more than a rhythmic oscillator for expiratory pump muscles: it also coordinates nasofacial and oral motor commands that engage muscles controlling airways. Active expiration is mediated by an expiratory oscillator located in the parafacial respiratory group (pFRG). Active expiration requires more than contracting expiratory muscles as multiple cranial nerves are recruited to stabilize the naso- and oropharyngeal airways. We tested the hypothesis that activation of the pFRG recruits facial and trigeminal motoneurons to coordinate nasofacial and oral motor activities that engage muscles controlling airways in rats during active expiration. Using a combination of electrophysiological and pharmacological approaches, we identified brainstem circuits that phase-lock active expiration, nasofacial and oral motor outputs in an in situ preparation of rat. We found that either high chemical drive (hypercapnia/acidosis) or unilateral excitation (glutamate microinjection) of the pFRG evoked active expiration and stimulated motoneurons (facial and trigeminal) and motor nerves responsible for the control of nasofacial (buccal and zygomatic branches of the facial nerve) and oral (mylohyoid nerve) motor outputs simultaneously. Bilateral pharmacological inhibition (GABAergic and glycinergic receptor activation) of the pFRG abolished active expiration and the simultaneous nasofacial and oral motor activities induced by hypercapnia/acidosis. We conclude that the pFRG provides the excitatory drive to phase-lock rhythmic nasofacial and oral motor circuits during active expiration in rats. Therefore, the pFRG is more than a rhythmic oscillator for expiratory pump muscles: it also coordinates nasofacial and oral motor commands that engage muscles controlling airways in rats during active expiration.

Urgent problems of employment and promotion of competitiveness for discharged military personnel in Ukraine

The article is devoted to the investigation of urgent problems concerning the employment and promotion of competitiveness for discharged military personnel in Ukraine. According to the results of sociological survey conducted by means of the author's tools, it is determined that significant number of respondents are characterized by recognition of risks of non-employment after active service expiration, lack of awareness of existing rights and guarantees in the labor market, low efficiency of public services in the sphere of employment and adaptation. It is determined that increase of competitiveness in Ukraine labor market for discharged military personnel requires modernization of technologies of state institutions activity related to the employment of the investigated category of citizens, search of additional funding sources, adaptation of discharged military personnel, providing them with free legal services on employment, active cooperation of the employment service with educational institutions, employers and other labor market entities, etc. The structural analysis of employment services in Ukraine, the United States and the Kingdom of Norway aimed at the improvement of discharged military personnel employment and increase of their competitiveness in the labor market of Ukraine, showed the need to create the department of social protection and employment of military personnel in the State Employment Service of Ukraine and to provide this department with the following functions: social adaptation, training and retraining in nondefense occupations and employment of discharged military personnel increasing competitiveness in the labor market of Ukraine and providing the opportunity for fast readjustment to civil life. This department will also provide better employment service work with discharged military personnel and reduce the risk of their unemployment, as well as decrease the percentage of unemployed in the labor market of Ukraine. The analysis showed that it is necessary to provide new functions to the State Employment Service in order to improve the work with job hunters, change the required control functions for the functions of employment assistance.

Expiratory Resistances Prevent Expiratory Diaphragm Contraction, Flow Limitation, and Lung Collapse.

Rationale: Tidal expiratory flow limitation (tidal-EFL) is not completely avoidable by applying positive end-expiratory pressure and may cause respiratory and hemodynamic complications in ventilated patients with lungs prone to collapse. During spontaneous breathing, expiratory diaphragmatic contraction counteracts tidal-EFL. We hypothesized that during both spontaneous breathing and controlled mechanical ventilation, external expiratory resistances reduce tidal-EFL.Objectives: To assess whether external expiratory resistances 1) affect expiratory diaphragmatic contraction during spontaneous breathing, 2) reduce expiratory flow and make lung compartments more homogeneous with more similar expiratory time constants, and 3) reduce tidal atelectasis, preventing hyperinflation.Methods: Three positive end-expiratory pressure levels and four external expiratory resistances were tested in 10 pigs after lung lavage. We analyzed expiratory diaphragmatic electric activity and respiratory mechanics. On the basis of computed tomography scans, four lung compartments-not inflated (atelectasis), poorly inflated, normally inflated, and hyperinflated-were defined.Measurements and Main Results: Consequently to additional external expiratory resistances, and mainly in lungs prone to collapse (at low positive end-expiratory pressure), 1) the expiratory transdiaphragmatic pressure decreased during spontaneous breathing by >10%, 2) expiratory flow was reduced and the expiratory time constants became more homogeneous, and 3) the amount of atelectasis at end-expiration decreased from 24% to 16% during spontaneous breathing and from 32% to 18% during controlled mechanical ventilation, without increasing hyperinflation.Conclusions: The expiratory modulation induced by external expiratory resistances preserves the positive effects of the expiratory brake while minimizing expiratory diaphragmatic contraction. External expiratory resistances optimize lung mechanics and limit tidal-EFL and tidal atelectasis, without increasing hyperinflation.

Epigenetic silencing by early-life hypoxic stress programmes respiratory motor control.

Epigenetic silencing by early-life hypoxic stress programmes respiratory motor control.

A6 neurons simultaneously modulate active expiration and upper airway resistance in rats.

What is the central question of this study? Do A6 neurons modulate active expiratory and airway responses evoked by hypercapnia/acidosis? What is the main finding and its importance? Acute inhibition of A6 neurons reduced active expiratory, inspiratory and the associated oropharyngeal and laryngeal motor responses to hypercapnia/acidosis. A6 neurons provide excitatory synaptic drive contributing to the central generation of inspiratory and expiratory motor activity as well as the control of upper airway resistance. During rest, inspiration is an active phenomenon, whereas expiration is passive. Under conditions of high chemical drive, such as hypercapnia/acidosis, there is an increase in inspiratory activity, expiration becomes active and upper airway resistance is reduced. The locus coeruleus noradrenergic neurons (A6 neurons) are activated when exposed to elevated CO2 /[H+ ] levels and modulate respiratory brainstem neurons regulating ventilation. However, the role of A6 neurons in the control of upper airway resistance is not fully understood. We tested the hypothesis that A6 neurons contribute to the central generation of active inspiratory and expiratory responses and the associated changes in the motor nerves controlling upper airway resistance during hypercapnia/acidosis in rats. Using a perfused brainstem-spinal cord preparation, we inhibited A6 neurons using pharmacogenetics and evaluated the active expiratory (abdominal nerve), laryngeal (cervical vagus nerve), oropharyngeal (hypoglossal nerve) and inspiratory (phrenic nerve) motor nerve responses to hypercapnia/acidosis. Acute inhibition of A6 neurons did not produce significant changes in the respiratory pattern in normocapnia. However, the hypercapnia/acidosis-induced active expiratory response and the associated changes in the motor nerves responsible for control of oropharyngeal and laryngeal resistance, as well as the inspiratory response were all reduced after inhibition of A6 neurons. Our data demonstrate that A6 neurons exert an important excitatory synaptic drive to the central generation of both active inspiratory and expiratory activities and modulate the control of upper airway resistance during hypercapnia/acidosis.

Chemogenetic modulation of the parafacial respiratory group influences the recruitment of abdominal activity during REM sleep.

Current theories on respiratory control postulate that the respiratory rhythm is generated by oscillatory networks in the medulla: preBötzinger complex (preBötC) is the master oscillator responsible for generating inspiration, while parafacial respiratory group (pFRG) drives active expiration through recruitment of expiratory abdominal (ABD) muscle activity. Research addressing the role of pFRG in ventilation and rhythm generation across sleep states is limited. We recently reported the occurrence of ABD recruitment occurring despite the induction of muscle paralysis during REM sleep. This ABD recruitment was associated with increased tidal volume and regularization of the respiratory period in rats. As pFRG generates active expiration through the engagement of ABD muscles, we hypothesized that the expiratory oscillator is also responsible for the ABD recruitment observed during REM sleep. To test this hypothesis, we inhibited and activated pFRG using chemogenetics (i.e. designer receptors exclusively activated by designer drugs) while recording EEG and respiratory muscle EMG activities across sleep-wake cycles in male Sprague-Dawley rats. Our results suggest that inhibition of pFRG reduced the number of REM events expressing ABD recruitment, in addition to the intensity and prevalence of these events. Conversely, activation of pFRG resulted in an increase in the number of REM events in which ABD recruitment was observed, as well as the intensity and prevalence of ABD recruitment. Interestingly, modulation of pFRG activity did not affect ABD recruitment during NREM sleep or wakefulness. These results suggest that the occurrence of ABD recruitment during sleep is dependent on pFRG activity and is state dependent.

Feedforward discharges couple the singing central pattern generator and ventilation central pattern generator in the cricket abdominal central nervous system

We investigated the central nervous coordination between singing motor activity and abdominal ventilatory pumping in crickets. Fictive singing, with sensory feedback removed, was elicited by eserine-microinjection into the brain, and the motor activity underlying singing and abdominal ventilation was recorded with extracellular electrodes. During singing, expiratory abdominal muscle activity is tightly phase coupled to the chirping pattern. Occasional temporary desynchronization of the two motor patterns indicate discrete central pattern generator (CPG) networks that can operate independently. Intracellular recordings revealed a sub-threshold depolarization in phase with the ventilatory cycle in a singing-CPG interneuron, and in a ventilation-CPG interneuron an excitatory input in phase with each syllable of the chirps. Inhibitory synaptic inputs coupled to the syllables of the singing motor pattern were present in another ventilatory interneuron, which is not part of the ventilation-CPG. Our recordings suggest that the two centrally generated motor patterns are coordinated by reciprocal feedforward discharges from the singing-CPG to the ventilation-CPG and vice versa. Consequently, expiratory contraction of the abdomen usually occurs in phase with the chirps and ventilation accelerates during singing due to entrainment by the faster chirp cycle.

Tracheobronchomalacia in the Adult: Is Imaging Helpful?

The aim of this article is to: 1) Review the imaging features of tracheobronchomalacia and excessive dynamic airway collapse 2) To highlight the recent advances in imaging of the central airways. Imaging of the central airways requires dedicated techniques optimized to evaluate the transient collapse of the central airways either due to excessive movement of the posterior membrane and/or secondary to weakness of the tracheal cartilages. Routine imaging of the chest is limited in its ability to demonstrate the true extent of collapsibility of the airways. The more recently introduced imaging protocols utilize dynamic acquisition during active expiration which demonstrate the transient but true extent of the airway collapse. CT also has potential applications in treatment in the form of 3D-printed splints. Dedicated imaging protocols introduced with recent advances in CT technology allow diagnosis of tracheobronchomalacia (TBM) noninvasively. However, there is a current lack of clear threshold for diagnosis which has led to a wide range of reported prevalence of TBM and excessive dynamic airway collapse.

Distinct pathways to the parafacial respiratory group to trigger active expiration in adult rats.

Active expiration (AE) is part of the breathing phase; it is conditional and occurs when we increase our metabolic demand, such as during hypercapnia, hypoxia, or exercise. The parafacial respiratory group (pFRG) is involved in AE. Data from the literature suggest that excitatory and the absence of inhibitory inputs to the pFRG are necessary to determine AE. However, the source of the inputs to the pFRG that trigger AE remains unclear. We show in adult urethane-anesthetized Wistar rats that the pharmacological inhibition of the medial aspect of the nucleus of the solitary tract (mNTS) or the rostral aspect of the pedunculopontine tegmental nucleus (rPPTg) is able to generate AE. In addition, direct inhibitory projection from the mNTS or indirect cholinergic projection from the rPPTg is able to contact pFRG to trigger AE. The inhibition of the mNTS or the rPPTg under conditions of high metabolic demand, such as hypercapnia (9-10% CO2), did not affect the AE. The present results suggest for the first time that inhibitory sources from the mNTS and a cholinergic pathway from the rPPTg, involving M2/M4 muscarinic receptors, could be important sources to modulate and sustain AE.

Cardiovascular and respiratory profiles during the sleep-wake cycle of rats previously submitted to chronic intermittent hypoxia.

What is the central question of this study? Chronic intermittent hypoxia (CIH) causes increased arterial pressure (AP), sympathetic overactivity and changes in expiratory modulation of sympathetic activity. However, changes in the short-term sleep-wake cycle pattern after CIH and their potential impact on cardiorespiratory parameters have not been reported previously. What is the main finding and its importance? Exposure to CIH for 10 days elevates AP in wakefulness and sleep but does not cause major changes in short-term sleep-wake cycle pattern. A higher incidence of muscular expiratory activity was observed in rats exposed to CIH only during wakefulness, indicating that active expiration is not required for the increase in AP in rats submitted to CIH. Chronic intermittent hypoxia (CIH) increases arterial pressure (AP) and changes sympathetic-respiratory coupling. However, the alterations in the sleep-wake cycle after CIH and their potential impact on cardiorespiratory parameters remain unknown. Here, we evaluated whether CIH-exposed rats present changes in their short-term sleep-wake cycle pattern and in cardiorespiratory parameters. Male Wistar rats (∼250g) were divided into CIH and control groups. The CIH rats were exposed to 8hday-1 of cycles of normoxia (fraction of inspired O2 =0.208, 5min) followed by hypoxia (fraction of inspired O2 =0.06, 30-40 s) for 10 days. One day after CIH, electrocorticographic activity, cervical EMG, AP and heart rate were recorded for 3h. Plethysmographic recordings were collected for 2h. A subgroup of control and CIH rats also had the diaphragm and oblique abdominal muscle activities recorded. Chronic intermittent hypoxia did not alter the time for sleep onset, total time awake, durations of rapid eye movement (REM) and non-REM (NREM) sleep and number of REM episodes in the 3h recordings. However, a significant increase in the duration of REM episodes was observed. The AP and heart rate were increased in all phases of the cycle in rats exposed to CIH. Respiratory frequency and ventilation were similar between groups in all phases, but tidal volume was increased during NREM and REM sleep in rats exposed to CIH. An increase in the incidence of active expiration during wakefulness was observed in rats exposed to CIH. The data show that CIH-related hypertension is not caused by changes in the sleep-wake cycle and suggest that active expiration is not required for the increase in AP in freely moving rats exposed to CIH.

Brainstem pre‐sympathetic neurons contribute to irregular breathing patterns in volume overload heart failure

Chronic heart failure (CHF) is characterized by sympathoexcitation and a high prevalence of breathing disorders. It has been shown that rostral ventrolateral medulla catecholaminergic neurons (RVLM‐C1) contributes to sympathetic outflow regulation in CHF and that RVLM‐C1 neurons are hyperactivated in experimental CHF. Recently, it has been shown that acute activation of RVLM‐C1 neurons also stimulates breathing in healthy rodents. However, there is no evidence showing the contribution of RVLM‐C1 neurons, if any, on breathing disorders in CHF. Therefore, we aimed to determine the role of RVLM‐C1 neurons on disordered breathing in CHF. Sprague‐Dawley rats underwent volume overload to induce CHF. Stereotaxic bilateral injections of anti‐dopamine β‐hydroxylase‐saporin toxin (DβH–SAP: 5ng/150nl) were used to selectively eliminate RVLM‐C1 neurons. Whole‐body plethysmography, blood pressure radiotelemetry and pression‐volume conductance catheter were used to study breathing stability, chemoreflex function, cardiac autonomic control, respiratory‐cardiovascular coupling (RCC) and cardiac function. Depletion of RVLM‐C1 neurons (~70%) in CHF rats (CHF+Veh vs. CHF+DβH+SAP, respectively) restored regular breathing pattern (Breathing irregularity score: 25.5 ± 3.0 vs. 11.9 ± 1.2 %; p<0.05), decreased apnea/hypopnea incidence (11.0 ± 0.9 vs. 6.5 ± 0.8 events/hr; p<0.05) and improves cardiac autonomic control as evidenced by a decrease in the low/high frequency component of spectral analysis of heart rate variability (HRV) (LF/HFHRV 2.7 ± 0.3 vs. 1.5 ± 0.1; p<0.05). In addition, CHF rats displayed active expiration compared to Sham rats, evidenced by the increase in the early/late expiration ratio (E2/E1: 0.51 ± 0.02 vs. 1.07 ± 0.07 Sham+Veh vs. CHF+Veh, p<0.05) and DβH–SAP toxin treatment decrease active expiration in CHF rats (E2/E1: 1.07 ± 0.07 vs. 0.81 ± 0.01 ml, CHF+Veh vs. CHF+DβH‐SAP; p<0.05). Accordingly, DβH–SAP toxin treatment eliminate RCC in CHF rats (Coherencepeak: 0.74 ± 0.04 vs. 0.59 ± 0.05, CHF+Veh vs. HF+DβH–SAP; p<0.05). In addition, the deleterious effects of central chemoreflex activation on diastolic cardiac function and cardiac autonomic control were blunted after RVLM‐C1 neuron ablation (EDPVR: 0.023 ± 0.008 vs. 0.005 ± 0.001 mmHg/μl; LF/HFHRV 2.19 ± 0.15 vs. 1.09 ± 0.23; CHF+Veh+Hypercapnia vs. CHF+DβH‐SAP+ Hypercapnia; p<0.05). No changes in central chemoreflex drive and/or in the number of central chemoreceptor neurons (from the retrotrapezoid nucleus) were found in CHF+DβH‐SAP rats compared to CHF+Veh treated rats. Our results showed that RVLM‐C1 neurons play a critical role in the maintenance of altered breathing patterns in CHF rats and highlighted their contribution to the worsening of cardiac function during central chemoreflex activation.Support or Funding InformationFondecyt 1180172; The basal Center of Excellence in Aging and Regeneration (AFB 170005)This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.