Recovery In Room Air Research Articles

Recovery of Ventilatory and Metabolic Responses to Hypoxia after Chronic Hypoxia in Neonatal Rats

Chronic hypoxia (CH) from birth blunts the hypoxic ventilatory response (HVR) in newborn rats, but there is conflicting evidence as to whether (or how quickly) the HVR recovers after these rats are returned to room air. In this study, Sprague-Dawley rats were exposed to either 21% O2 (Control) or 12% O2 (CH) for the first 3 postnatal days. Breathing and metabolism were studied using head-body plethysmography and respirometry immediately after the 3 days of CH (i.e., at P3) or after 1, 4-5, or 7 days of recovery in room air (i.e., at P4, P7-8, or P10). The early phase of the HVR was blunted in CH rats at P3 (13.7± 2.3 vs. 30.6±3.4 % increase from baseline during 12% O2), and the HVR was more biphasic in CH rats after 1 day of recovery. However, no significant differences in the HVR were observed between treatment groups after 3-4 or 7 days of recovery. Males and females showed similar patterns. Changes in ventilation during hypoxia were strongly biphasic in the younger age groups: ventilation increased initially, but this was followed by a gradual decline in ventilation towards or below baseline levels. However, when accounting for metabolic depression, all age groups continued to hyperventilate during the late phase of the HVR [i.e., the ratio of ventilation to CO2 production (CO2 convection requirement) remained elevated relative to baseline values]. The CO2 convection requirement response to hypoxia did not differ between treatment groups during the late phase of the HVR at any age, but metabolism and CO2 convection requirement could not be determined during the early phase of the HVR due to non-steady state conditions. In conclusion, 3 days of CH produces only a transient blunting of the HVR in newborn rats. Longer CH exposures may be required to elicit persistent changes in the HVR or, alternatively, this plasticity may not be revealed until after puberty. Supported by NIH grant P20 GM-103423 (Maine INBRE). This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Chronic hyperoxia augments the hypoxic ventilatory response in adult rats

Chronic hyperoxia during postnatal development causes long-lasting attenuation of the hypoxic ventilatory response (HVR) in rats, but similar exposures during adulthood do not attenuate the HVR (Ling et al. J Physiol 405: 561-571, 1996). However, the HVR has not been assessed in adult-exposed rats immediately after chronic hyperoxia (i.e., without several months of recovery in room air). This is important since chronic hyperoxia depresses breathing and blunts the HVR in newborn rats, and only some of these effects persist after return to room air. Accordingly, adult rats were studied after 4 and 14 days of exposure to either 60% O 2 (Hyperoxia) or 21% O 2 (Control). Ventilation was measured by whole-body plethysmography; metabolic O 2 consumption and CO 2 production were also measured in a subset of rats. Normoxic ventilation was unchanged, but Hyperoxia rats exhibited a slight increase in their acute HVR after both durations of chronic hyperoxia. Overall, minute ventilation was approximately 10% greater in Hyperoxia rats while breathing 12% O 2 and 10% O 2 than in Control rats; this effect did not differ significantly between female and male rats. The HVR was also enhanced after 4 days of hyperoxia after controlling for metabolic rate, but the results were less clear after 14 days. After the 14-day exposure, the HVR tended to be enhanced when tested at 12% O 2 but not at 10% O 2 . These data demonstrate that chronic hyperoxia has different effects on the control of breathing in neonatal and adult rats when assessed immediately after the exposure. Supported by NIH grant P20 GM-103423 (Maine INBRE). This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Protective Effect of Dexmedetomidine against Hyperoxia-Damaged Cerebellar Neurodevelopment in the Juvenile Rat.

Impaired cerebellar development of premature infants and the associated impairment of cerebellar functions in cognitive development could be crucial factors for neurodevelopmental disorders. Anesthetic- and hyperoxia-induced neurotoxicity of the immature brain can lead to learning and behavioral disorders. Dexmedetomidine (DEX), which is associated with neuroprotective properties, is increasingly being studied for off-label use in the NICU. For this purpose, six-day-old Wistar rats (P6) were exposed to hyperoxia (80% O2) or normoxia (21% O2) for 24 h after DEX (5 µg/kg, i.p.) or vehicle (0.9% NaCl) application. An initial detection in the immature rat cerebellum was performed after the termination of hyperoxia at P7 and then after recovery in room air at P9, P11, and P14. Hyperoxia reduced the proportion of Calb1+-Purkinje cells and affected the dendrite length at P7 and/or P9/P11. Proliferating Pax6+-granule progenitors remained reduced after hyperoxia and until P14. The expression of neurotrophins and neuronal transcription factors/markers of proliferation, migration, and survival were also reduced by oxidative stress in different manners. DEX demonstrated protective effects on hyperoxia-injured Purkinje cells, and DEX without hyperoxia modulated neuronal transcription in the short term without any effects at the cellular level. DEX protects hyperoxia-damaged Purkinje cells and appears to differentially affect cerebellar granular cell neurogenesis following oxidative stress.

Coenzyme Q10 and Fish Oil Supplementation for Reducing Retinal Oxidative Stress in a Rat Model

Extremely low gestational-age neonates requiring supplemental oxygen experience intermittent hypoxia (IH) episodes, which predispose them to oxidative stress and retinopathy of prematurity. We tested the hypothesis that early supplementation with fish oil or CoQ10 confers benefits reducing the severity of IH-induced retinopathy. At birth, rat pups were exposed to two clinically relevant neonatal IH paradigms with recovery in either hyperoxia (50% O2) or room air (RA) between episodes for 14 days, during which they received daily oral fish oil, coenzyme Q10 (CoQ10) in olive oil (OO), or OO only (vehicle). At postnatal day 14 (P14), pups were allowed to recover in RA with no further treatment until P21. Retinas were examined at P14 and at P21. Both IH paradigms resulted in severe ocular oxidative stress and retinopathy regardless of recovery in hyperoxia or RA in the vehicle groups. Although early supplementation with fish oil was beneficial, CoQ10 provided superior benefits for reducing IH-induced oxidative stress and retinopathy. These effects were associated with lower retinal antioxidants and biomarkers of angiogenesis. The therapeutic benefits of CoQ10 suggest a potential treatment for IH-induced retinopathies. Further studies are needed to establish appropriate, safe, and effective doses for use in preterm infants.

Abstract 9727: Reduced Bmpr2 Causes a Decrease in Foxf1 That Links Unrepaired DNA Damage to Persistent Pulmonary Hypertension

Introduction: B MPR2 mutation is the most common genetic cause of pulmonary arterial hypertension (PAH). Loss of BMPR2 causes pulmonary arterial (PA) endothelial cell (EC) dysfunction and impairs EC-mediated suppression of smooth muscle cells proliferation. These are characteristic features of PAH vascular cells as is the presence of DNA damage that can be induced by oxidative stress. We previously showed that EC- Bmpr2 -/- mice have persistent pulmonary hypertension (PH) with reoxygenation after hypoxia. However, the role of DNA damage in causing persistent PH related to loss of BMPR2 and oxidative stress is unknown. Hypothesis: We hypothesize that loss of BMPR2 causes unrepaired DNA damage and persistent PH by altering the expression of genes that maintain EC homeostasis. Methods and Results: DNA damage assessed by γH2AX foci was observed in PAEC of EC- Bmpr2 -/- mice in room air, that was more extensive after reoxygenation (10% hypoxia for 3 weeks followed by recovery in room-air for 4 weeks). Deletion of the DNA damage sensor ataxia-telangiectasia mutated (ATM) in EC of mice (EC- Atm -/- ) also resulted in persistent DNA damage and PH. RNA sequencing of PAEC after reoxygenation in EC- Bmpr2 -/- and EC- Atm -/- mice vs. controls showed downregulated EC differentiation and development genes enriched in biding motifs for Foxf1, a transcription factor associated with DNA repair and angiogenesis. Foxf1 expression was reduced in PAEC with loss of BMPR2 or ATM. Loss of Foxf1 decreased angiogenic genes Cldn5 and Vegfr2 , and DNA damage response genes Atm and p53 , and impaired PAEC migration and tube formation. Foxf1 overexpression increased these genes in PAEC. Immunohistochemistry showed decreased FOXF1 in PAEC of human pulmonary vascular lesions. Delivery of Foxf1 to EC- Bmpr2 -/- mice with an AAV2-peptide targeting the PA endothelium attenuated DNA damage in PAEC, restored angiogenesis genes and completely reversed persistent PH returning right ventricular systolic pressure to normal levels. Conclusions: We link loss of BMPR2 and DNA damage sensing in PAEC to persistent DNA damage and PAH and show that both are mediated by reduced FOXF1 . In EC- Bmpr2 -/- mice, Foxf1 gene therapy attenuated DNA damage restored angiogenesis genes, and prevented persistent PH.

Maladaptive functional changes in alveolar fibroblasts due to perinatal hyperoxia impair epithelial differentiation.

Infants born prematurely worldwide have up to a 50% chance of developing bronchopulmonary dysplasia (BPD), a clinical morbidity characterized by dysregulated lung alveolarization and microvascular development. It is known that PDGFR alpha–positive (PDGFRA+) fibroblasts are critical for alveolarization and that PDGFRA+ fibroblasts are reduced in BPD. A better understanding of fibroblast heterogeneity and functional activation status during pathogenesis is required to develop mesenchymal population–targeted therapies for BPD. In this study, we utilized a neonatal hyperoxia mouse model (90% O2 postnatal days 0–7, PN0–PN7) and performed studies on sorted PDGFRA+ cells during injury and room air recovery. After hyperoxia injury, PDGFRA+ matrix and myofibroblasts decreased and PDGFRA+ lipofibroblasts increased by transcriptional signature and population size. PDGFRA+ matrix and myofibroblasts recovered during repair (PN10). After 7 days of in vivo hyperoxia, PDGFRA+ sorted fibroblasts had reduced contractility in vitro, reflecting loss of myofibroblast commitment. Organoids made with PN7 PDGFRA+ fibroblasts from hyperoxia in mice exhibited reduced alveolar type 1 cell differentiation, suggesting reduced alveolar niche-supporting PDGFRA+ matrix fibroblast function. Pathway analysis predicted reduced WNT signaling in hyperoxia fibroblasts. In alveolar organoids from hyperoxia-exposed fibroblasts, WNT activation by CHIR increased the size and number of alveolar organoids and enhanced alveolar type 2 cell differentiation.

Dynamic Regulation of GH-IGF1 Signaling in Injury and Recovery in Hyperoxia-Induced Neonatal Lung Injury.

Prematurely born infants often require supplemental oxygen that impairs lung growth and results in arrest of alveolarization and bronchopulmonary dysplasia (BPD). The growth hormone (GH)- and insulin-like growth factor (IGF)1 systems regulate cell homeostasis and organ development. Since IGF1 is decreased in preterm infants, we investigated the GH- and IGF1 signaling (1) in newborn mice with acute and prolonged exposure to hyperoxia as well as after recovery in room air; and (2) in cultured murine lung epithelial cells (MLE-12) and primary neonatal lung fibroblasts (pLFs) after treatment with GH, IGF1, and IGF1-receptor (IGF1-R) inhibitor or silencing of GH-receptor (Ghr) and Igf1r using the siRNA technique. We found that (1) early postnatal hyperoxia caused an arrest of alveolarization that persisted until adulthood. Both short-term and prolonged hyperoxia reduced GH-receptor expression and STAT5 signaling, whereas Igf1 mRNA and pAKT signaling were increased. These findings were related to a loss of epithelial cell markers (SFTPC, AQP5) and proliferation of myofibroblasts (αSMA+ cells). After recovery, GH-R-expression and STAT5 signaling were activated, Igf1r mRNA reduced, and SFTPC protein significantly increased. Cell culture studies showed that IGF1 induced expression of mesenchymal (e.g., Col1a1, Col4a4) and alveolar epithelial cell type I (Hopx, Igfbp2) markers, whereas inhibition of IGF1 increased SFTPC and reduced AQP5 in MLE-12. GH increased Il6 mRNA and reduced proliferation of pLFs, whereas IGF1 exhibited the opposite effect. In summary, our data demonstrate an opposite regulation of GH- and IGF1- signaling during short-term/prolonged hyperoxia-induced lung injury and recovery, affecting alveolar epithelial cell differentiation, inflammatory activation of fibroblasts, and a possible uncoupling of the GH-IGF1 axis in lungs after hyperoxia.

Prenatal Maternal Lipopolysaccharide and Mild Newborn Hyperoxia Increase Intrapulmonary Airway but Not Vessel Reactivity in a Mouse Model.

Maternal infection is a risk for preterm delivery. Preterm newborns often require supplemental oxygen to treat neonatal respiratory distress. Newborn hyperoxia exposure is associated with airway and vascular hyperreactivity, while the complications of maternal infection are variable. In a mouse model of prenatal maternal intraperitoneal lipopolysaccharide (LPS, embryonic day 18) with subsequent newborn hyperoxia (40% oxygen × 7 days) precision-cut living lung slices were used to measure intrapulmonary airway and vascular reactivity at 21 days of age. Hyperoxia increased airway reactivity to methacholine compared to room air controls. Prenatal maternal LPS did not alter airway reactivity in room air. Combined maternal LPS and hyperoxia exposures increased airway reactivity vs. controls, although maximal responses were diminished compared to hyperoxia alone. Vessel reactivity to serotonin did not significantly differ in hyperoxia or room air; however, prenatal maternal LPS appeared to attenuate vessel reactivity in room air. Following room air recovery, LPS with hyperoxia lungs displayed upregulated inflammatory and fibrosis genes compared to room air saline controls (TNFαR1, iNOS, and TGFβ). In this model, mild newborn hyperoxia increases airway but not vessel reactivity. Prenatal maternal LPS did not further increase hyperoxic airway reactivity. However, inflammatory genes remain upregulated weeks after recovery from maternal LPS and newborn hyperoxia exposures.

Club Cell Heme Oxygenase-1 Deletion: Effects in Hyperoxia-Exposed Adult Mice.

Thioredoxin reductase-1 (TXNRD1) inhibition activates nuclear factor (erythroid-derived 2)-like 2 (Nrf2) responses and prevents acute lung injury (ALI). Heme oxygenase-1 (HO-1) induction following TXNRD1 inhibition is Nrf2-dependent in airway epithelial (club) cells in vitro. The influence of club cell HO-1 on lung development and lung injury responses is poorly understood. The present studies characterized the effects of hyperoxia on club cell-specific HO-1 knockout (KO) mice. These mice were generated by crossing Hmox1 flox mice with transgenic mice expressing cre recombinase under control of the club cell-specific Scgb1a1 promoter. Baseline analyses of lung architecture and function performed in age-matched adult wild-type and KO mice indicated an increased alveolar size and airway resistance in HO-1 KO mice. In subsequent experiments, adult wild-type and HO-1 KO mice were either continuously exposed to >95% hyperoxia or room air for 72 h or exposed to >95 hyperoxia for 48 h followed by recovery in room air for 48 h. Injury was quantitatively assessed by calculating right lung/body weight ratios (g/kg). Analyses indicated an independent effect of hyperoxia but not genotype on right lung/body weight ratios in both wild-type and HO-1 KO mice. The magnitude of increases in right lung/body weight ratios was similar in mice of both genotypes. In the recovery model, an independent effect of hyperoxia but not genotype was also detected. In contrast to the continuous exposure model, right lung/body weight ratio mice were significantly elevated in HO-1 KO but not wild-type mice. Though club cell HO-1 does not alter hyperoxic sensitivity in adult mice, it significantly influences lung development and resolution of lung injury following acute hyperoxic exposure.

Tracheomalacia in bronchopulmonary dysplasia: Trachealis hyper-relaxant responses to S-nitrosoglutathione in a hyperoxic murine model.

Bronchopulmonary dysplasia (BPD) with airway hyperreactivity is a long-term pulmonary complication of prematurity. The endogenous nonadrenergic, noncholinergic signaling molecule, S-nitrosoglutathione (GSNO) and its catabolism by GSNO reductase (GSNOR) modulate airway reactivity. Tracheomalacia is a major, underinvestigated complication of BPD. We studied trachealis, left main bronchus (LB), and intrapulmonary bronchiolar (IPB) relaxant responses to GSNO in a murine hyperoxic BPD model. Wild-type (WT) or GSNOR knockout (KO) newborn mice were raised in 60% (BPD) or 21% (control) oxygen during the first 3 weeks of life. After room air recovery, adult trachealis, LB, and IPB smooth muscle relaxant responses to GSNO (after methacholine preconstriction) were studied using wire myographs. Studies were repeated after GSNOR inhibitor (GSNORi) pretreatment and in KO mice. GSNO relaxed all airway preparations. GSNO relaxed WT BPD trachealis substantially more than WT controls (P < .05). Pharmacologic or genetic ablation of GSNOR abolished the exaggerated BPD tracheal relaxation to GSNO and also augmented BPD IPB relaxation to GSNO. LB ring contractility was not significantly different between groups or conditions. Additionally, GSNORi treatment induced relaxation of WT IPBs but not trachealis or LB. GSNO dramatically relaxed the trachealis in our BPD model, an effect paradoxically reversed by loss of GSNOR. Conversely, GSNOR inhibition augmented IBP relaxation. These data suggest that GSNOR inhibition could benefit both the BPD trachealis and distal airways, restoring relaxant responses to those of room air controls. Because therapeutic options are limited in this high-risk population, future studies of GSNOR inhibition are needed.

Respiratory Plasticity in Adult Rats after Exposure to Chronic Hyperoxia

Neonatal rats reared in moderate hyperoxia (60% O2) from birth hypoventilate when acutely returned to room air (e.g., Bavis et al., Respir. Physiol. Neurobiol. 198: 1–12, 2014); this hypoventilation reflects reductions in both respiratory frequency and tidal volume. Since many forms of respiratory plasticity are specific to development, we questioned whether chronic hyperoxia would have similar effects on normoxic ventilation in adult rats. Adult Sprague‐Dawley rats (3–5 months old) were randomly assigned to Control (chronic 21% O2; n=11) or Hyperoxia (chronic 60% O2; n=10) treatment groups. Normoxic ventilation was measured by whole‐body plethysmography prior to exposure, after 4 and 14 days of exposure, and after 4 days of recovery in room air. Chronic hyperoxia altered minute ventilation in adult rats (Treatment × Day, P=0.002), but surprisingly this effect was not apparent during the hyperoxic exposure itself. Instead, Hyperoxia rats exhibited a modest increase in normoxic ventilation when measured during the recovery period (39±2 vs. 33±1 ml min−1 100g−1; P=0.003). Tidal volumes and respiratory frequencies were similar between groups before and during the chronic exposures, but Hyperoxia rats exhibited greater tidal volumes and respiratory frequencies during the recovery period (Treatment × Day, P=0.047 and 0.038, respectively). Neither metabolism nor blood gases were measured during these experiments, so it is unclear whether the increase in ventilation constitutes a true hyperventilation. We conclude that the effects of chronic hyperoxia on normoxic ventilation are age‐specific.Support or Funding InformationSupported by NIH grant P20 GM‐103423 (Maine INBRE).This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Persistent HIF‐1 Activation by Long‐Term Intermittent Hypoxia

Long‐term (30 days) exposure of rats to intermittent hypoxia (LT‐IH) results in persistent elevation of reactive oxygen species levels in central and peripheral nervous system (J. Nanduri et al., J. Physiol.2016,2018). HIF‐ regulates pro‐oxidant enzyme (POE) genes during IH. Present study examined the effect of LT‐IH on HIF‐1α protein and NADPH oxidase (Nox) 4 mRNA abundances in the carotid body (CB) and adrenal medullae (AM) of LT‐IH treated rats. CB and AM of LT‐IH treated rats exhibited robust increase in HIF‐1α protein, Nox4 mRNA and ROS levels. Remarkably, these effects persisted after recovery in room air for 30d. The effects of LT‐IH on HIF‐1α, Nox 4 mRNA, and ROS abundance were absent in rats treated with digoxin, an inhibitor of HIF‐1α during LT‐IH as well as in mice with partial deficiency of HIF‐1a (HIF‐1α heterozygote mice). These findings suggest that HIF‐1 dependent upregulation of POE genes contributes to LT‐IH induced persistent oxidative stress in the CB and AM.Support or Funding InformationSupported by NIH‐ HL‐90554.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Duration‐dependent effects of IH‐stimulated Hif1a signaling on adult hippocampal neurogenesis

Intermittent hypoxia (IH), is a hallmark consequence of sleep apnea, a common untreated disorder associated with impaired neurological health. We have recently demonstrated that thirty days of IH impairs long term potentiation and reduced the proportion of neurons generated during adult neurogenesis. Hypoxia Inducible Factor 1a (HIF1a) is a transcription factor important to stem cell development that is regulated by the state of oxygenation. The objective of this on‐going study is to characterize how the duration of IH exposure influences the generation of adult‐born neurons and to determine the role of IH‐dependent HIF1a signaling in the generation of adult‐born neurons in the hippocampus. We hypothesize that IH has duration dependent influence over neurons generated through adult neurogenesis and involves HIF1a dependent signaling. We used a cre‐loxP transgenic strategy to birth label a discrete cohort of stem cells that experience room air for 30 days (control), 10 days of IH +20 days of recovery in room air (IH10+20RA), or 30 days of IH (IH30) in wild‐type reporter mice (Ai27) and in Ai27‐HIF1a+/flox mice. In wild‐type mice, exposure to IH10+20RA led to higher proportion of birth‐labelled neurons compared to the control (+19% ± 5%, n=6) while the proportion of neurons following IH30 decreased (−40% ± 10%, n=4). In Ai27‐HIF1a+/− mice, the proportion of birth‐labeled neurons was similar between control and IH10+20RA groups. However, following IH30 the proportion of birth‐labeled neurons was markedly decreased when compared to control in Ai27‐Hif1a+/− and to the IH30 wild‐type group. Dendritic complexity, as determined by Sholl analysis, was similar among labeled neurons in both wild‐type and Ai27‐HIF1a+/− groups suggesting that neither IH nor IH‐dependent HIF1a signaling influences dendritic complexity. These findings indicate that exposure duration is factor impacting how IH affects the generation of adult‐born neurons through a HIF1a signaling shaping how IH influences, associated with sleep apnea affects the generation of adult‐born neurons.Support or Funding InformationThis work was supported by NIH R01 NS10742101 (AJG).This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Neonatal hyperoxia promotes asthma-like features through IL-33–dependent ILC2 responses

Premature infants often require oxygen supplementation and, therefore, are exposed to oxidative stress. Following oxygen exposure, preterm infants frequently develop chronic lung disease and have a significantly increased risk of asthma. We sought to identify the underlying mechanisms by which neonatal hyperoxia promotes asthma development. Mice were exposed to neonatal hyperoxia followed by a period of room air recovery. Agroup of mice was also intranasally exposed to house dust mite antigen. Assessments were performed at various time points for evaluation of airway hyperresponsiveness, eosinophilia, mucus production, inflammatory gene expression, and TH and group 2 innate lymphoid cell (ILC2) responses. Sera from term- and preterm-born infants were also collected and levels of IL-33 and type 2 cytokines were measured. Neonatal hyperoxia induced asthma-like features including airway hyperresponsiveness, mucus hyperplasia, airway eosinophilia, and type 2 pulmonary inflammation. In addition, neonatal hyperoxia promoted allergic TH responses to house dust mite exposure. Elevated IL-33 levels and ILC2 responses were observed in the lungs most likely due to oxidative stress caused by neonatal hyperoxia. IL-33 receptor signaling and ILC2s were vital for the induction of asthma-like features following neonatal hyperoxia. Serum IL-33 levels correlated significantly with serum levels of IL-5 and IL-13 but not IL-4 in preterm infants. These data demonstrate that an axis involving IL-33 and ILC2s is important for the development of asthma-like features following neonatal hyperoxia and suggest therapeutic potential for targeting IL-33, ILC2s, and oxidative stress to prevent and/or treat asthma development related to prematurity.

Hemodynamic and Pathologic Characterization of the TASK-1-/- Mouse Does Not Demonstrate Pulmonary Hypertension.

Pulmonary hypertension (PH) carries significant associated morbidity and mortality and the underlying molecular mechanisms of PH are not well understood. Loss-of-function mutations in TASK-1 potassium channels are associated with PH in humans. Although TASK-1 has been considered in the development of PH for over a decade, characterization of TASK-1 knockout mice has been limited to in vitro studies or in vivo studies in room air at isolated time points. The purpose of this study was twofold. First, we sought to determine if TASK-/- male and female mice developed PH over the span of one year. Second, we sought to determine the effect of chronic hypoxia, a stimulus for PH, and its recovery on PH in TASK-1-/- mice. We measured right ventricular systolic pressure (RVSP) and vascular remodeling in male and female C57BL/6 WT and TASK-1-/- mice at separate time points: 20-24 weeks and 1 year of age. Additionally, we measured RVSP and vascular remodeling in TASK-1-/- and wild-type mice between 13 and 16 weeks of age exposed to 10% hypoxia for 3 weeks followed by recovery to room air conditions for an additional 6 weeks. RVSP was similar between WT and TASK-/- mice. Male and female WT and TASK-1-/- mice all demonstrated age-related increases in RVSP, which correlated to age-related vascular remodeling in male mice but not in female mice. Male TASK-1-/- and WT mice exposed to chronic hypoxia demonstrated increased RVSP, which decreased following room air recovery. WT and TASK-1-/- male mice demonstrated vascular remodeling upon exposure to hypoxia that persisted in room air recovery. Female and male TASK-1-/- mice do not develop hemodynamic or vascular evidence for PH, but RVSP rises in an age-dependent manner independent of genotype. TASK-1-/- and WT male mice develop hypoxia-induced elevations in RVSP that decrease to baseline after recovery in room air. TASK-1-/- and WT male mice demonstrate vascular remodeling after exposure to hypoxia that persists despite recovery to room air conditions and does not correlate with RVSP normalization.

Aberrant cGMP signaling persists during recovery in mice with oxygen-induced pulmonary hypertension.

Bronchopulmonary dysplasia (BPD), a common complication of preterm birth, is associated with pulmonary hypertension (PH) in 25% of infants with moderate to severe BPD. Neonatal mice exposed to hyperoxia for 14d develop lung disease similar to BPD, with evidence of associated PH. The cyclic guanosine monophosphate (cGMP) signaling pathway has not been well studied in BPD-associated PH. In addition, there is little data about the natural history of hyperoxia-induced PH in mice or the utility of phosphodiesterase-5 (PDE5) inhibition in established disease. C57BL/6 mice were placed in room air or 75% O2 within 24h of birth for 14d, followed by recovery in room air for an additional 7 days (21d). Additional pups were treated with either vehicle or sildenafil for 7d during room air recovery. Mean alveolar area, pulmonary artery (PA) medial wall thickness (MWT), RVH, and vessel density were evaluated at 21d. PA protein from 21d animals was analyzed for soluble guanylate cyclase (sGC) activity, PDE5 activity, and cGMP levels. Neonatal hyperoxia exposure results in persistent alveolar simplification, RVH, decreased vessel density, increased MWT, and disrupted cGMP signaling despite a period of room air recovery. Delayed treatment with sildenafil during room air recovery is associated with improved RVH and decreased PA PDE5 activity, but does not have significant effects on alveolar simplification, PA remodeling, or vessel density. These data are consistent with clinical studies suggesting inconsistent effects of sildenafil treatment in infants with BPD-associated PH.

Intermittent hypercapnic hypoxia during sleep does not induce ventilatory long-term facilitation in healthy males.

Intermittent hypoxia-induced ventilatory neuroplasticity is likely important in obstructive sleep apnea pathophysiology. Although concomitant CO2 levels and arousal state critically influence neuroplastic effects of intermittent hypoxia, no studies have investigated intermittent hypercapnic hypoxia effects during sleep in humans. Thus the purpose of this study was to investigate if intermittent hypercapnic hypoxia during sleep induces neuroplasticity (ventilatory long-term facilitation and increased chemoreflex responsiveness) in humans. Twelve healthy males were exposed to intermittent hypercapnic hypoxia (24 × 30 s episodes of 3% CO2 and 3.0 ± 0.2% O2) and intermittent medical air during sleep after 2 wk washout period in a randomized crossover study design. Minute ventilation, end-tidal CO2, O2 saturation, breath timing, upper airway resistance, and genioglossal and diaphragm electromyograms were examined during 10 min of stable stage 2 sleep preceding gas exposure, during gas and intervening room air periods, and throughout 1 h of room air recovery. There were no significant differences between conditions across time to indicate long-term facilitation of ventilation, genioglossal or diaphragm electromyogram activity, and no change in ventilatory response from the first to last gas exposure to suggest any change in chemoreflex responsiveness. These findings contrast with previous intermittent hypoxia studies without intermittent hypercapnia and suggest that the more relevant gas disturbance stimulus of concomitant intermittent hypercapnia frequently occurring in sleep apnea influences acute neuroplastic effects of intermittent hypoxia. These findings highlight the need for further studies of intermittent hypercapnic hypoxia during sleep to clarify the role of ventilatory neuroplasticity in the pathophysiology of sleep apnea.NEW & NOTEWORTHY Both arousal state and concomitant CO2 levels are known modulators of the effects of intermittent hypoxia on ventilatory neuroplasticity. This is the first study to investigate the effects of combined intermittent hypercapnic hypoxia during sleep in humans. The lack of neuroplastic effects suggests a need for further studies more closely replicating obstructive sleep apnea to determine the pathophysiological relevance of intermittent hypoxia-induced ventilatory neuroplasticity.

Neonatal Hyperoxia Perturbs Neuronal Development in the Cerebellum.

Impaired postnatal brain development of preterm infants often results in neurological deficits. Besides pathologies of the forebrain, maldeveolopment of the cerebellum is increasingly recognized to contribute to psychomotor impairments of many former preterm infants. However, causes are poorly defined. We used a hyperoxia model to define neonatal damage in cerebellar granule cell precursors (GCPs) and in Purkinje cells (PCs) known to be essential for interaction with GCPs during development. We exposed newborn rats to 24h 80% O2 from age P6 to P7 to identify postnatal and long-term damage in cerebellar GCPs at age P7 after hyperoxia and also after recovery in room air thereafter until P11 and P30. We determined proliferation and apoptosis of GCPs and immature neurons by immunohistochemistry, quantified neuronal damage by qPCR and Western blots for neuronal markers, and measured dendrite outgrowth of PCs by CALB1 immunostainings and by Sholl analysis of Golgi stainings. After hyperoxia, proliferation of PAX6+ GCPs was decreased at P7, while DCX+CASP3+ cells were increased at P11. Neuronal markers Pax6, Tbr2, and Prox1 were downregulated at P11 and P30. Neuronal damage was confirmed by reduced NeuN protein expression at P30. Sonic hedgehog (SHH) was significantly decreased at P7 and P11 after hyperoxia and coincided with lower CyclinD2 and Hes1 expression at P7. The granule cell injury was accompanied by hampered PC maturation with delayed dendrite formation and impaired branching. Neonatal injury induced by hyperoxia inhibits PC functioning and impairs granule cell development. As a conclusion, maldevelopment of the cerebellar neurons found in preterm infants could be caused by postnatal oxygen toxicity.

Effects of nitrous oxide sedation on sympathetic baroreflex sensitivity

The number of patients with cardiovascular diseases is still growing worldwide, which may increase the risk for cardiovascular events associated with pain stimulation during medical and dental treatments. Nitrous oxide provides minimal and moderate sedation and suppresses pressor responses to pain. We found that an increase in heart rate (HR) in response to pain stimulation remained unchanged by nitrous oxide inhalation, while an increase in muscle sympathetic nerve activity (MSNA) was attenuated, suggesting that blood pressure (BP) control during nitrous oxide sedation may be modified via MSNA. We evaluated whether sympathetic baroreflex sensitivity (BRS) would be decreased by nitrous oxide sedation. Arm cuff and beat‐to‐beat BP (finger photoplethysmography), HR, MSNA (microneurography), cardiac output (Qc) and stroke volume (SV) were measured during inhalation of room air, 100% oxygen, 30%, 40% nitrous oxide, and room air (recovery) for 20 min through a mask, respectively, in 9 young men [age 31±2 (SD) yrs]. Sympathetic BRS was analyzed from the spontaneous changes in beat‐by‐beat diastolic BP and corresponding MSNA (burst incidence and total MSNA) during the last 5 min of each condition. Systolic and diastolic BP did not differ among the inhalation conditions. HR, Qc, and SV were also similar between any conditions. Sympathetic BRS was remained unchanged by inhalation sedation with any nitrous oxide concentration (burst incidence : −3.16 ±1.77, −2.98 ± .02, −2.86 ± 2.05, −2.19 ± 1.95, and−2.21 ± 1.34; P= 0.204 total MSNA : −6.56 ± 2.89, −6.79 ± 4.89, −6.89 ± 5.25, −4.75 ± 3.86, and −5.14 ± 3.44; P= 0.161). These results suggest that baroreflex function is maintained during nitrous oxide sedation and nitrous oxide itself does not directly affect BP regulations.Support or Funding InformationSupported by JSPSKAKENHI 16K20705.

Adult Lysophosphatidic Acid Receptor 1-Deficient Rats with Hyperoxia-Induced Neonatal Chronic Lung Disease Are Protected against Lipopolysaccharide-Induced Acute Lung Injury.

Aim: Survivors of neonatal chronic lung disease or bronchopulmonary dysplasia (BPD) suffer from compromised lung function and are at high risk for developing lung injury by multiple insults later in life. Because neonatal lysophosphatidic acid receptor-1 (LPAR1)-deficient rats are protected against hyperoxia-induced lung injury, we hypothesize that LPAR1-deficiency may protect adult survivors of BPD from a second hit response against lipopolysaccharides (LPS)-induced lung injury.Methods: Directly after birth, Wistar control and LPAR1-deficient rat pups were exposed to hyperoxia (90%) for 8 days followed by recovery in room air. After 7 weeks, male rats received either LPS (2 mg kg−1) or 0.9% NaCl by intraperitoneal injection. Alveolar development and lung inflammation were investigated by morphometric analysis, IL-6 production, and mRNA expression of cytokines, chemokines, coagulation factors, and an indicator of oxidative stress.Results: LPAR1-deficient and control rats developed hyperoxia-induced neonatal emphysema, which persisted into adulthood, as demonstrated by alveolar enlargement and decreased vessel density. LPAR1-deficiency protected against LPS-induced lung injury. Adult controls with BPD exhibited an exacerbated response toward LPS with an increased expression of pro-inflammatory mRNAs, whereas LPAR1-deficient rats with BPD were less sensitive to this “second hit” with a decreased pulmonary influx of macrophages and neutrophils, interleukin-6 (IL-6) production, and mRNA expression of IL-6, monocyte chemoattractant protein-1, cytokine-induced neutrophil chemoattractant 1, plasminogen activator inhibitor-1, and tissue factor.Conclusion: LPAR1-deficient rats have increased hyperoxia-induced BPD survival rates and, despite the presence of neonatal emphysema, are less sensitive to an aggravated “second hit” than Wistar controls with BPD. Intervening in LPA-LPAR1-dependent signaling may not only have therapeutic potential for neonatal chronic lung disease, but may also protect adult survivors of BPD from sequelae later in life.