SubSol-HIe is an AMPK-dependent hypoxia-responsive subnucleus of the nucleus tractus solitarius that coordinates the hypoxic ventilatory response and protects against apnoea in mice.

Functional magnetic resonance imaging (fMRI) suggests that the hypoxic ventilatory response is facilitated by the AMP-activated protein kinase (AMPK), not at the carotid bodies, but within a subnucleus (Bregma -7.5 to -7.1 mm) of the nucleus tractus solitarius that exhibits right-sided bilateral asymmetry. Here, we map this subnucleus using cFos expression as a surrogate for neuronal activation and mice in which the genes encoding the AMPK-α1 (Prkaa1) and AMPK-α2 (Prkaa2) catalytic subunits were deleted in catecholaminergic cells by Cre expression via the tyrosine hydroxylase promoter. Comparative analysis of brainstem sections, relative to controls, revealed that AMPK-α1/α2 deletion inhibited, with right-sided bilateral asymmetry, cFos expression in and thus activation of a neuronal cluster that partially spanned three interconnected anatomical nuclei adjacent to the area postrema: SolDL (Bregma -7.44 mm to -7.48 mm), SolDM (Bregma -7.44 mm to -7.48 mm) and SubP (Bregma -7.48 mm to -7.56 mm). This approximates the volume identified by fMRI. Moreover, these nuclei are known to be in receipt of carotid body afferent inputs, and catecholaminergic neurons of SubP and SolDL innervate aspects of the ventrolateral medulla responsible for respiratory rhythmogenesis. Accordingly, AMPK-α1/α2 deletion attenuated hypoxia-evoked increases in minute ventilation (normalised to metabolism), reductions in expiration time, and increases sigh frequency, but increased apnoea frequency during hypoxia. The metabolic response to hypoxia in AMPK-α1/α2 knockout mice and the brainstem and spinal cord catecholamine levels were equivalent to controls. We conclude that within the brainstem an AMPK-dependent, hypoxia-responsive subnucleus partially spans SubP, SolDM and SolDL, namely SubSol-HIe, and is critical to coordination of active expiration, the hypoxic ventilatory response and defence against apnoea.

Chronic intermittent hypoxia-induced cardiovascular and renal dysfunction: from adaptation to maladaptation.

Chronic intermittent hypoxia (CIH) is the dominant pathological feature of human obstructive sleep apnoea (OSA), which is highly prevalent and associated with cardiovascular and renal diseases. CIH causes hypertension, centred on sympathetic nervous overactivity, which persists following removal of the CIH stimulus. Molecular mechanisms contributing to CIH-induced hypertension have been carefully delineated. However, there is a dearth of knowledge on the efficacy of interventions to ameliorate high blood pressure in established disease. CIH causes endothelial dysfunction, aberrant structural remodelling of vessels and accelerates atherosclerotic processes. Pro-inflammatory and pro-oxidant pathways converge on disrupted nitric oxide signalling driving vascular dysfunction. In addition, CIH has adverse effects on the myocardium, manifesting atrial fibrillation, and cardiac remodelling progressing to contractile dysfunction. Sympatho-vagal imbalance, oxidative stress, inflammation, dysregulated HIF-1α transcriptional responses and resultant pro-apoptotic ER stress, calcium dysregulation, and mitochondrial dysfunction conspire to drive myocardial injury and failure. CIH elaborates direct and indirect effects in the kidney that initially contribute to the development of hypertension and later to chronic kidney disease. CIH-induced morphological damage of the kidney is dependent on TLR4/NF-κB/NLRP3/caspase-1 inflammasome activation and associated pyroptosis. Emerging potential therapies related to the gut-kidney axis and blockade of aryl hydrocarbon receptors (AhR) are promising. Cardiorenal outcomes in response to intermittent hypoxia present along a continuum from adaptation to maladaptation and are dependent on the intensity and duration of exposure to intermittent hypoxia. This heterogeneity of OSA is relevant to therapeutic treatment options and we argue the need for better stratification of OSA phenotypes.

Loss of compensation afforded by accessory muscles of breathing leads to respiratory system compromise in the mdx mouse model of Duchenne muscular dystrophy.

Despite profound diaphragm weakness, peak inspiratory pressure-generating capacity is preserved in young mdx mice revealing adequate compensation by extra-diaphragmatic muscles of breathing in early dystrophic disease. We hypothesised that loss of compensation gives rise to respiratory system compromise in advanced dystrophic disease. Studies were performed in male wild-type (n = 196) and dystrophin-deficient mdx mice (n = 188) at 1, 4, 8, 12 and 16 months of age. In anaesthetised mice, inspiratory pressure and obligatory and accessory respiratory EMG activities were recorded during baseline and sustained tracheal occlusion for up to 30-40 s to evoke peak system activation to task failure. Obligatory inspiratory EMG activities were lower in mdx mice across the ventilatory range to peak activity, emerging in early dystrophic disease. Early compensation protecting peak inspiratory pressure-generating capacity in mdx mice, which appears to relate to transforming growth factor-ß1-dependent fibrotic remodelling of the diaphragm and preserved accessory muscle function, was lost at 12 and 16 months of age. Denervation and surgical lesion of muscles of breathing in 4-month-old mice revealed a greater dependency on diaphragm for peak inspiratory performance in wild-type mice, whereas mdx mice were heavily dependent upon accessory muscles (including abdominal muscles) for peak performance. Accessory EMG activities were generally preserved or enhanced in young mdx mice, but peak EMG activities were lower than wild-type by 12 months of age. In general, ventilation was reasonably well protected in mdx mice until 16 months of age. Despite the early emergence of impairments in the principal obligatory muscles of breathing, peak inspiratory performance is compensated in early dystrophic disease due to diaphragm remodelling and facilitated contribution by accessory muscles of breathing. Loss of compensation afforded by accessory muscles underpins the emergence of respiratory system morbidity in advanced dystrophic disease. KEY POINTS: Despite diaphragm weakness, peak inspiratory performance is preserved in young dystrophin-deficient mdx mice revealing adequate compensation by extra-diaphragmatic muscles. Peak obligatory muscle (diaphragm, external intercostal, and parasternal intercostal) EMG activities are lower in mdx mice, emerging early in dystrophic disease, before the temporal decline in peak performance. Peak EMG activities of some accessory muscles are lower, whereas others are preserved. There is greater recruitment of the trapezius muscle in mdx mice during peak system activation. In phrenicotomised mice with confirmed diaphragm paralysis, there is a greater contribution made by extra-diaphragmatic muscles to peak inspiratory pressure in mdx compared with wild-type mice. Surgical lesion of accessory (including abdominal) muscles adversely affects peak pressure generation in mdx mice. Diaphragm remodelling leading to stiffening provides a mechanical advantage to peak pressure generation via the facilitated action of extra-diaphragmatic muscles in early dystrophic disease. Peak accessory EMG activities are lower in 12-month-old mdx compared to wild-type mice. Peak inspiratory pressure declines in mdx mice with advanced disease. We conclude that compensation afforded by accessory muscles of breathing declines in advanced dystrophic disease precipitating the emergence of respiratory system dysfunction.

Chronic Intermittent Hypoxia-Induced Diaphragm Muscle Weakness Is NADPH Oxidase-2 Dependent.

Chronic intermittent hypoxia (CIH)-induced redox alterations underlie diaphragm muscle dysfunction. We sought to establish if NADPH oxidase 2 (NOX2)-derived reactive oxygen species (ROS) underpin CIH-induced changes in diaphragm muscle, which manifest as impaired muscle performance. Adult male mice (C57BL/6J) were assigned to one of three groups: normoxic controls (sham); chronic intermittent hypoxia-exposed (CIH, 12 cycles/hour, 8 h/day for 14 days); and CIH + apocynin (NOX2 inhibitor, 2 mM) administered in the drinking water throughout exposure to CIH. In separate studies, we examined sham and CIH-exposed NOX2-null mice (B6.129S-CybbTM1Din/J). Apocynin co-treatment or NOX2 deletion proved efficacious in entirely preventing diaphragm muscle dysfunction following exposure to CIH. Exposure to CIH had no effect on NOX2 expression. However, NOX4 mRNA expression was increased following exposure to CIH in wild-type and NOX2 null mice. There was no evidence of overt CIH-induced oxidative stress. A NOX2-dependent increase in genes related to muscle regeneration, antioxidant capacity, and autophagy and atrophy was evident following exposure to CIH. We suggest that NOX-dependent CIH-induced diaphragm muscle weakness has the potential to affect ventilatory and non-ventilatory performance of the respiratory system. Therapeutic strategies employing NOX2 blockade may function as an adjunct therapy to improve diaphragm muscle performance and reduce disease burden in diseases characterised by exposure to CIH, such as obstructive sleep apnoea.

Microbes, metabolites and muscle: Is the gut-muscle axis a plausible therapeutic target in Duchenne muscular dystrophy?

NEW FINDINGS: What is the topic of this review? The contribution of gut microbial signalling to skeletal muscle maintenance and development and identification of potential therapeutic targets in progressive muscle degenerative diseases such as Duchenne muscular dystrophy. What advances does it highlight? Gut microbe-derived metabolites are multifaceted signalling molecules key to muscle function, modifying pathways contributing to skeletal muscle wasting, making them a plausible target for adjunctive therapy in muscular dystrophy. ABSTRACT: Skeletal muscle is the largest metabolic organ making up ∼50% of body mass. Because skeletal muscle has both metabolic and endocrine properties, it can manipulate the microbial populations within the gut. In return, microbes exert considerable influence on skeletal muscle via numerous signalling pathways. Gut bacteria produce metabolites (i.e., short chain fatty acids, secondary bile acids and neurotransmitter substrates) that act as fuel sources and modulators of inflammation, influencing host muscle development, growth and maintenance. The reciprocal interactions between microbes, metabolites and muscle establish a bidirectional gut-muscle axis. The muscular dystrophies constitute a broad range of disorders with varying disabilities. In the profoundly debilitating monogenic disorder Duchenne muscular dystrophy (DMD), skeletal muscle undergoes a reduction in muscle regenerative capacity leading to progressive muscle wasting, resulting in fibrotic remodelling and adipose infiltration. The loss of respiratory muscle in DMD culminates in respiratory insufficiency and eventually premature death. The pathways contributing to aberrant muscle remodelling are potentially modulated by gut microbial metabolites, thus making them plausible targets for pre- and probiotic supplementation. Prednisone, the gold standard therapy for DMD, drives gut dysbiosis, inducing a pro-inflammatory phenotype and leaky gut barrier contributing to several of the well-known side effects associated with chronic glucocorticoid treatment. Several studies have observed that gut microbial supplementation or transplantation exerts positive effects on muscle, including mitigating the side effects of prednisone. There is growing evidence in support of the potential for an adjunctive microbiota-directed regimen designed to optimise gut-muscle axis signalling, which could alleviate muscle wasting in DMD.

Three Days of Chronic Intermittent Hypoxia Induce ß1-Adrenoceptor Dependent Increases in Left Ventricular Contractility.

Sleep apnea is characterized by bouts of chronic intermittent hypoxia (CIH) that elicit sympathetic hyperactivity resulting in residual hypertension. We previously demonstrated that exposure to CIH increases cardiac output and sought to determine if enhanced cardiac contractility manifests prior to hypertension.Male Wistar rats were exposed to cyclical bouts of hypoxia (FiO2 = 0.05 nadir; 90 s) and normoxia (FiO2 = 0.21; 210 s) 8 h/day for 3 days (CIH; n = 6). Control animals (n = 7) were exposed to room air. Data are presented as mean ± SD and were analyzed using unpaired Student t-tests.Three-day exposure to CIH did not elicit changes in heart rate and blood pressure (p > 0.05). However, baseline left ventricular contractility (dP/dtMAX) was significantly increased in CIH-exposed animals compared with control (15300 ± 2002 vs. 12320 ± 2725 mmHg/s; p = 0.025), despite no difference in catecholamine concentrations. Acute ß1-adrenoceptor inhibition reduced contractility in CIH-exposed animals (-7604 ± 1298 vs. -4747 ± 2080 mmHg/s; p = 0.014), to levels equivalent to control, while preserving cardiovascular parameters. Sympathetic ganglion blockade (hexamethonium 25 mg/kg; i.v.) produced equivalent cardiovascular responses suggesting similar global sympathetic activity between groups. Interestingly, gene expression of the ß1-adrenoceptor pathway in cardiac tissue was unchanged.Our results suggest that CIH increases cardiac contractility via ß1-adrenoceptor dependent mechanisms prior to development of global sympathetic hyperactivity suggesting that positive cardiac inotropy contributes to the development of hypertension in CIH-exposed rats.

Ventilatory Effects of Acute Intermittent Hypoxia in Conscious Dystrophic Mice.

Exposure to acute intermittent hypoxia (AIH) elicits a form of respiratory plasticity known as long-term facilitation (LTF). Interest has grown in developing AIH interventions to treat ventilatory insufficiency, with promising results in spinal cord injury and amyotrophic lateral sclerosis. Therapeutic AIH may have application in neuromuscular disorders including muscular dystrophies. We sought to establish hypoxic ventilatory responsiveness and the expression of ventilatory LTF in X-linked muscular dystrophy (mdx) mice.Experiments were performed in 15 male wild-type (BL10) and 15 male mdx mice at 4 months of age. Ventilation was assessed using whole-body plethysmography. Baseline measures of ventilation and metabolism were established. Mice were exposed to 10 successive bouts of hypoxia, each lasting 5 min, interspersed with 5-min bouts of normoxia. Measurements were taken for 60 min following termination of AIH.In mdx mice, ventilation was significantly increased 60 min post-AIH compared to baseline. However, metabolic CO2 production was also increased. Therefore, ventilatory equivalent was unaffected by AIH exposure, i.e., no ventilatory LTF manifestation. In wild-type mice, ventilation and metabolism were not affected by AIH.Eliciting ventilatory LTF is dependent on many factors and may require concomitant isocapnia or hypercapnia during AIH exposures and/or repeated daily AIH exposures, which is worthy of further pursuit.

Machine Learning Detects Intraventricular Haemorrhage in Extremely Preterm Infants.

OBJECTIVE: To test the potential utility of applying machine learning methods to regional cerebral (rcSO2) and peripheral oxygen saturation (SpO2) signals to detect brain injury in extremely preterm infants. STUDY DESIGN: A subset of infants enrolled in the Management of Hypotension in Preterm infants (HIP) trial were analysed (n = 46). All eligible infants were <28 weeks' gestational age and had continuous rcSO2 measurements performed over the first 72 h and cranial ultrasounds performed during the first week after birth. SpO2 data were available for 32 infants. The rcSO2 and SpO2 signals were preprocessed, and prolonged relative desaturations (PRDs; data-driven desaturation in the 2-to-15-min range) were extracted. Numerous quantitative features were extracted from the biosignals before and after the exclusion of the PRDs within the signals. PRDs were also evaluated as a stand-alone feature. A machine learning model was used to detect brain injury (intraventricular haemorrhage-IVH grade II-IV) using a leave-one-out cross-validation approach. RESULTS: The area under the receiver operating characteristic curve (AUC) for the PRD rcSO2 was 0.846 (95% CI: 0.720-0.948), outperforming the rcSO2 threshold approach (AUC 0.593 95% CI 0.399-0.775). Neither the clinical model nor any of the SpO2 models were significantly associated with brain injury. CONCLUSION: There was a significant association between the data-driven definition of PRDs in rcSO2 and brain injury. Automated analysis of PRDs of the cerebral NIRS signal in extremely preterm infants may aid in better prediction of IVH compared with a threshold-based approach. Further investigation of the definition of the extracted PRDs and an understanding of the physiology underlying these events are required.

Steady-state chemoreflex drive captures ventilatory acclimatization during incremental ascent to high altitude: Effect of acetazolamide.

Ventilatory acclimatization (VA) is important to maintain adequate oxygenation with ascent to high altitude (HA). Transient hypoxic ventilatory response tests lack feasibility and fail to capture the integrated steady-state responses to chronic hypoxic exposure in HA fieldwork. We recently characterized a novel index of steady-state respiratory chemoreflex drive (SSCD), accounting for integrated contributions from central and peripheral respiratory chemoreceptors during steady-state breathing at prevailing chemostimuli. Acetazolamide is often utilized during ascent for prevention or treatment of altitude-related illnesses, eliciting metabolic acidosis and stimulating respiratory chemoreceptors. To determine if SSCD reflects VA during ascent to HA, we characterized SSCD in 25 lowlanders during incremental ascent to 4240 m over 7 days. We subsequently compared two separate subgroups: no acetazolamide (NAz; n = 14) and those taking an oral prophylactic dose of acetazolamide (Az; 125 mg BID; n = 11). At 1130/1400 m (day zero) and 4240 m (day seven), steady-state measurements of resting ventilation (V̇I ; L/min), pressure of end-tidal (PET )CO2 (Torr), and peripheral oxygen saturation (SpO2 ; %) were measured. A stimulus index (SI; PET CO2 /SpO2 ) was calculated, and SSCD was calculated by indexing V̇I against SI. We found that (a) both V̇I and SSCD increased with ascent to 4240 m (day seven; V̇I : +39%, p < 0.0001, Hedges' g = 1.52; SSCD: +56.%, p < 0.0001, Hedges' g = 1.65), (b) and these responses were larger in the Az versus NAz subgroup (V̇I : p = 0.02, Hedges' g = 1.04; SSCD: p = 0.02, Hedges' g = 1.05). The SSCD metric may have utility in assessing VA during prolonged stays at altitude, providing a feasible alternative to transient chemoreflex tests.

Impact of cancer cachexia on respiratory muscle function and the therapeutic potential of exercise.

Cancer cachexia is defined as a multi-factorial syndrome characterised by an ongoing loss of skeletal muscle mass and progressive functional impairment, estimated to affect 50-80% of patients and responsible for 20% of cancer deaths. Elevations in the morbidity and mortality rates of cachectic cancer patients has been linked to respiratory failure due to atrophy and dysfunction of the ventilatory muscles. Despite this, there is a distinct scarcity of research investigating the structural and functional condition of the respiratory musculature in cancer, with the majority of studies exclusively focusing on limb muscle. Treatment strategies are largely ineffective in mitigating the cachectic state. It is now widely accepted that an efficacious intervention will likely combine elements of pharmacology, nutrition and exercise. However, of these approaches, exercise has received comparatively little attention. Therefore, it is unlikely to be implemented optimally, whether in isolation or combination. In consideration of these limitations, the current review describes the mechanistic basis of cancer cachexia and subsequently explores the available respiratory- and exercise-focused literature within this context. The molecular basis of cachexia is thoroughly reviewed. The pivotal role of inflammatory mediators is described. Unravelling the mechanisms of exercise-induced support of muscle via antioxidant and anti-inflammatory effects in addition to promoting efficient energy metabolism via increased mitochondrial biogenesis, mitochondrial function and muscle glucose uptake provide avenues for interventional studies. Currently available pre-clinical mouse models including novel transgenic animals provide a platform for the development of multi-modal therapeutic strategies to protect respiratory muscles in people with cancer.