Role In Respiration Research Articles

High-resolution chemical dissection of a model eukaryote reveals targets, pathways and gene functions

Due to evolutionary conservation of biology, experimental knowledge captured from genetic studies in eukaryotic model organisms provides insight into human cellular pathways and ultimately physiology. Yeast chemogenomic profiling is a powerful approach for annotating cellular responses to small molecules. Using an optimized platform, we provide the relative sensitivities of the heterozygous and homozygous deletion collections for nearly 1800 biologically active compounds. The data quality enables unique insights into pathways that are sensitive and resistant to a given perturbation, as demonstrated with both known and novel compounds. We present examples of novel compounds that inhibit the therapeutically relevant fatty acid synthase and desaturase (Fas1p and Ole1p), and demonstrate how the individual profiles facilitate hypothesis-driven experiments to delineate compound mechanism of action. Importantly, the scale and diversity of tested compounds yields a dataset where the number of modulated pathways approaches saturation. This resource can be used to map novel biological connections, and also identify functions for unannotated genes. We validated hypotheses generated by global two-way hierarchical clustering of profiles for (i) novel compounds with a similar mechanism of action acting upon microtubules or vacuolar ATPases, and (ii) an un-annotated ORF, YIL060w, that plays a role in respiration in the mitochondria. Finally, we identify and characterize background mutations in the widely used yeast deletion collection which should improve the interpretation of past and future screens throughout the community. This comprehensive resource of cellular responses enables the expansion of our understanding of eukaryotic pathway biology.

Blockade of Na+/H+ exchanger type 3 causes intracellular acidification and hyperexcitability via inhibition of pH-sensitive K+ channels in chemosensitive respiratory neurons of the dorsal vagal nucleus in rats

Extracellular pH (pHe) and intracellular pH (pHi) are important factors for the excitability of chemosensitive central respiratory neurons that play an important role in respiration and obstructive sleep apnea. It has been proposed that inhibition of central Na(+)/H(+) exchanger 3 (NHE-3), a key pHi regulator in the brainstem, decreases the pHi, leading to membrane depolarization for the maintenance of respiration. However, how intracellular pH affects the neuronal excitability of respiratory neurons remains largely unknown. In this study, we showed that NHE-3 mRNA is widely distributed in respiration-related neurons of the rat brainstem, including the dorsal vagal nucleus (DVN). Whole-cell patch clamp recordings from DVN neurons in brain slices revealed that the standing outward current (Iso) through pH-sensitive K(+) channels was inhibited in the presence of the specific NHE-3 inhibitor AVE0657 that decreased the pHi. Exposure of DVN neurons to an acidified pHe and AVE0657 (5 μmol/L) resulted in a stronger effect on firing rate and Iso than acidified pHe alone. Taken together, our results showed that intracellular acidification by blocking NHE-3 results in inhibition of a pH-sensitive K(+) current, leading to synergistic excitation of chemosensitive DVN neurons for the regulation of respiration.

P1-20. Respiratory changes after lesions of the medullary raphe nuclei

It is known that respiratory movements change after chemical or electrical stimulation of the midline regions of the brain stem, including raphe obscurus, raphe pallidus and raphe magnus. We previously reported that neurons located in these regions had respiratory-related activity and a part of the neurons projected to the phrenic nuclei, suggesting the midline regions are involved in the control of respiration. However, it is unclear whether or not these neurons play a crucial role in respiration. To elucidate the role, we tested effects of chemical lesions of the medullary raphe nuclei on respiration using spontaneously breathing Sprague–Dawley rats which were anesthetized with pentobarbital sodium. Local injection of kainic acid into the medullary midline induced a transient increase in amplitude and frequency of diaphragm activities, and the frequency subsequently decreased. Then, apnea was elicited 20 min after the injection. Injection sites were located 1.5–3.0 mm rostral to the obex. The results suggest that the caudal medullary raphe nuclei may participate in the control of respiratory movements in rats.

Soil microbial carbon turnover decreases with increasing molecular size

It is well established that soil microorganisms play an important role in respiration of newly fixed plant carbon. Recent results show that they also contribute significantly to soil organic matter (SOM) formation. We hypothesized that different molecular size classes of compounds in soil microbial biomass (SMB) have variable turnover time and in consequence influence SOM formation differentially. Here we used natural differences in carbon stable isotope signatures (δ13C values) after C3–C4 vegetation change to track newly fixed C4 plant carbon into SMB molecular size classes. SMB was obtained by chloroform fumigation extraction (SFE) and δ13C values of its size classes were measured using size exclusion chromatography coupled online to liquid chromatography‒isotope ratio mass spectrometry (SEC–LC–IRMS). Resolved SMB was assigned to 5 size classes of 1800–9800, 800–1800, 380–800, 180–380 and 50–180 Da respectively. The contribution of recent C4 plant carbon to size classes of SMB decreased with increasing molecular weight (MW). It ranged from 77 ± 19% in the lowest MW size class size class to 41 ± 14% in the highest MW size class in a sandy soil and from 59 ± 18% in the lowest MW size class to 8 ± 15% in the highest MW size class in a clayey soil. A decreasing carbon turnover of compounds in SMB extracts along a continuum of molecular size from small to large implies that low molecular weight microbial compounds are rapidly metabolized products that link to fast respiratory carbon fluxes, whereas high molecular weight ones could be products of microbial synthesis like structural compounds that have slower turnover rates and link to slower SOM formation. Our methods help avoid contamination of CFE extracts and the results help explain why SMB turnover is faster in CFE extracts when compared to calculations using membrane lipids (e.g. PLFA-based).

The Neural Control of Spinal Stability Muscles during Different Respiratory Patterns

[Purpose] It is well-known that the muscles of spinal stability also play roles in respiration. The spinal stability muscles are divided into two subgroups, the local muscle group and the global muscle group. Appropriately coordinated activation of muscle groups are recommended for more efficient spinal stability. The indirect method of measuring coordination of muscle groups is the synergist ratio of local muscles to global muscles. The purpose of this study was to investigate the synergist ratios of the spinal stability muscles of different breathing patterns. [Subjects and Methods] Forty healthy subjects performed 4 different breathing patterns and 3 synergist ratios calculated from % maximal voluntary isometric contraction of 2 local group muscles and 3 global group muscles were analyzed. [Results] The results of this study show synergist ratios were consistent among the breathing patterns and there was a consistent muscle reliance pattern of synergist ratios during each breathing pattern. The synergist ratio of extensors stayed around 1. The results were consistent with those of previous studies of spinal stability exercises. [Conclusion] We suggest that different breathing patterns could be used as a component of spinal stability exercises, secondary to the similarities of muscle coordination with spinal stability exercises, commonly used in clinics.

The transcriptional response of Arabidopsis leaves to Fe deficiency

Due to its ease to donate or accept electrons, iron (Fe) plays a crucial role in respiration and metabolism, including tetrapyrrole synthesis, in virtually all organisms. In plants, Fe is a component of the photosystems and thus essential for photosynthesis. Fe deficiency compromises chlorophyll (Chl) synthesis, leading to interveinal chlorosis in developing leaves and decreased photosynthetic activity. To gain insights into the responses of photosynthetically active cells to Fe deficiency, we conducted transcriptional profiling experiments on leaves from Fe-sufficient and Fe-deficient plants using the RNA-seq technology. As anticipated, genes associated with photosynthesis and tetrapyrrole metabolism were dramatically down-regulated by Fe deficiency. A sophisticated response comprising the down-regulation of HEMA1 and NYC1, which catalyze the first committed step in tetrapyrrole biosynthesis and the conversion of Chl b to Chl a at the commencement of Chl breakdown, respectively, and the up-regulation of CGLD27, which is conserved in plastid-containing organisms and putatively involved in xanthophyll biosynthesis, indicates a carefully orchestrated balance of potentially toxic tetrapyrrole intermediates and functional end products to avoid photo-oxidative damage. Comparing the responses to Fe deficiency in leaves to that in roots confirmed subgroup 1b bHLH transcription factors and POPEYE/BRUTUS as important regulators of Fe homeostasis in both leaf and root cells, and indicated six novel players with putative roles in Fe homeostasis that were highly expressed in leaves and roots and greatly induced by Fe deficiency. The data further revealed down-regulation of organ-specific subsets of genes encoding ribosomal proteins, which may be indicative of a change in ribosomal composition that could bias translation. It is concluded that Fe deficiency causes a massive reorganization of plastid activity, which is adjusting leaf function to the availability of Fe.

The Story of Oxygen

The history of oxygen from discovery to clinical application for patients with chronic lung disease represents a long and storied journey. Within a relatively short period, early investigators not only discovered oxygen but also recognized its importance to life and its role in respiration. The application of oxygen to chronic lung disease, however, took several centuries. In the modern era, physiologists pursued the chemical nature of oxygen and its physiologic interaction with cellular metabolism and gas transport. It took brazen clinicians, however, to pursue oxygen as a therapeutic resource for patients with chronic lung disease because of the concern in the 20th century of the risks of oxygen toxicity. Application of ambulatory oxygen devices allowed landmark investigations of the long-term effects of continuous oxygen that established its safety and efficacy. Although now well established for hypoxic patients, many questions remain regarding the benefits of oxygen for varying severity and types of chronic lung disease.

The investigation and comparison of respiratory volumes in patients with chronic low back pain and healthy subjects

Introduction: Besides having major effects on the stability of lumbar vertebrae, Diaphragm and abdominal muscles play an important role in respiration. The function of these muscles may gradually change in patients with chronic low back pain leading to an inefficacious respiration. The aim of this study was to evaluate respiratory volumes in a group of patients with chronic low back pain and to compare these volumes with those of healthy subjects. Materials and Methods: Eighty subjects were recruited in this case-control study. Respiratory volumes were measured in 40 patients with chronic low back pain and in 40 sex- and age-matched healthy control subjects. Data on pain severity, previous physiotherapeutic intervention, duration of pain, was recorded. The respiratory volumes were measured by a spirometer. The t test was used for statistical analysis. Results: The results showed that subjects in the case group had lower VC (in), VC (ex), FVC, FEV1, and F1V1 values than those in the control group (P £ 0.5). The two groups had no significant difference in terms of FEF 25%, FEF 50%, FEF 75%, PEF, PIF, and MEF (P > 0.5). Conclusion: A tendency toward altered contration in the diaphragm and abdominal muscles of patients with chronic low back pain may gradually result in muscle weakness and a changed respiratory pattern. Keywords: Diaphragm, Abdominal muscles, Low back pain, Respiratory volumes

Genetic Analysis of the Hox Hydrogenase in the Cyanobacterium Synechocystis sp. PCC 6803 Reveals Subunit Roles in Association, Assembly, Maturation, and Function

Hydrogenases are metalloenzymes that catalyze 2H(+) + 2e(-) ↔ H(2). A multisubunit, bidirectional [NiFe]-hydrogenase has been identified and characterized in a number of bacteria, including cyanobacteria, where it is hypothesized to function as an electron valve, balancing reductant in the cell. In cyanobacteria, this Hox hydrogenase consists of five proteins in two functional moieties: a hydrogenase moiety (HoxYH) with homology to heterodimeric [NiFe]-hydrogenases and a diaphorase moiety (HoxEFU) with homology to NuoEFG of respiratory Complex I, linking NAD(P)H ↔ NAD(P)(+) as a source/sink for electrons. Here, we present an extensive study of Hox hydrogenase in the cyanobacterium Synechocystis sp. PCC 6803. We identify the presence of HoxEFUYH, HoxFUYH, HoxEFU, HoxFU, and HoxYH subcomplexes as well as association of the immature, unprocessed large subunit (HoxH) with other Hox subunits and unidentified factors, providing a basis for understanding Hox maturation and assembly. The analysis of mutants containing individual and combined hox gene deletions in a common parental strain reveals apparent alterations in subunit abundance and highlights an essential role for HoxF and HoxU in complex/subcomplex association. In addition, analysis of individual and combined hox mutant phenotypes in a single strain background provides a clear view of the function of each subunit in hydrogenase activity and presents evidence that its physiological function is more complicated than previously reported, with no outward defects apparent in growth or photosynthesis under various growth conditions.

Mitochondrial and nuclear genomic integrity after oxidative damage in Saccharomyces cerevisiae

All cells have the ability to adjust their metabolism to their changing environment to be able to survive. This adaptation is coordinated by various systems in the cell and mitochondria seem to play a unique and important role. Most endogenous oxidative damage to cells is actually generated as a byproduct of the mitochondrial function, which in turn damages mitochondrial structures more extensively due to their proximity to the source. Excessive damage to mitochondria leads to loss of parts or all of mtDNA, but unlike other organisms, S. cerevisiae cells are able to survive without mtDNA or respiration when grown on fermentative carbon sources. This allows studies of the role of mitochondria in the maintenance of cellular integrity, since lack of mitochondrial DNA frequently leads to genomic instability. Mitochondria are known for their role in respiration, ATP production and apoptosis, but it is now becoming clear that their function is intimately connected to diverse processes such as calcium and iron homeostasis and amino acid metabolism, and thus their dysfunction is not well tolerated. In this review, we discuss the mechanisms by which mitochondrial dysfunction can lead to genomic instability and the effect of the carbon source on this process.

The Plant Mitochondrial Carrier Family: Functional and Evolutionary Aspects

Mitochondria play a key role in respiration and energy production and are involved in multiple eukaryotic but also in several plant specific metabolic pathways. Solute carriers in the inner mitochondrial membrane connect the internal metabolism with that of the surrounding cell. Because of their common basic structure, these transport proteins affiliate to the mitochondrial carrier family (MCF). Generally, MCF proteins consist of six membrane spanning helices, exhibit typical conserved domains and appear as homodimers in the native membrane. Although structurally related, MCF proteins catalyze the specific transport of various substrates, such as nucleotides, amino acids, dicarboxylates, cofactors, phosphate or H+. Recent investigations identified MCF proteins also in several other cellular compartments and therefore their localization and physiological function is not only restricted to mitochondria. MCF proteins are a characteristic feature of eukaryotes and bacterial genomes lack corresponding sequences. Therefore, the evolutionary origin of MCF proteins is most likely associated with the establishment of mitochondria. It is not clear whether the host cell, the symbiont, or the chimerical organism invented the ancient MCF sequence. Here, we try to explain the establishment of different MCF proteins and focus on the characteristics of members from plants, in particular from Arabidopsis thaliana.

Innate immune response to influenza virus

The recent pandemic of a novel H1N1 influenza virus has stressed the importance of effective approaches to prevent viral infection. The innate immune system is our first line of defense against invading viruses. This review aims to give a brief summary of recent findings on the response of the innate immune system to influenza virus. Three families of pattern recognition receptors, toll-like receptors (TLRs), retinoic acid-inducible gene 1 protein like helicases (RLRs) and nucleotide-binding domain and leucine-rich-repeat-containing proteins (NLRs), are involved in recognition of influenza virus and they cooperatively operate to respond to the virus in cell culture or mouse models. Influenza virus mainly induces two types of innate immune cytokine responses: a proinflammatory response and an antiviral response. Recently, the NLRP3 inflammasome has proved to be an essential component in the host defense against influenza infection. The mitochondrion, traditionally recognized for its key role in respiration, metabolism and apoptosis, is becoming recognized as an important organelle for regulation of innate immune responses to influenza virus. The NLRP3 inflammasome is an essential component in the host defense against influenza infection. Further investigations are required to elucidate whether NLRP3 is associated with the adaptive response and to identify the components of influenza virus that activate this important mediator. The role of mitochondria as a potential central platform of innate response is becoming appreciated.

Culture methods and initial characterization of intercostal skeletal fibers isolated from the adult mouse

Primary cultures of single adult skeletal muscle fibers dissociated from locomotor muscles adhered to glass coverslips are routine models in muscle biology. Such models allow the functional and biological properties of live muscle fibers to be probed under carefully controlled conditions. Single fiber models also make muscle fibers highly accessible to various procedures such as the expression of exogenous proteins, electric field stimulation to drive contraction and pharmacological treatment. Such isolated fiber cultures have not been established for the intercostal muscles, despite the critical function of these muscles and the role of respiration in the mortality of many muscular diseases. We have developed an adherent culture system based on single intercostal muscle fibers isolated from the adult mouse for use in physiological and biochemical studies. We also provide initial characterization and demonstrate several common techniques in this new model system with comparisons to the established Flexor Digitorum Brevis muscle primary culture model.

Oxygen's double-edged sword: balancing muscle O2supply and use during exercise

Oxygen's double-edged sword: balancing muscle O₂supply and use during exercise

The physical origin of sigmoidal respiratory pressure–volume curves: Alveolar recruitment and nonlinear elasticity

An important unsolved problem in medical science concerns the physical origin of the sigmoidal shape of pressure–volume curves of healthy (and some unhealthy) lungs. Conventional wisdom holds that linear response, i.e., Hooke’s law, together with alveolar overdistention play a dominant role in respiration, but such assumptions cannot explain the crucial empirical sigmoidal shape of the curves. Here, we propose a theory of alveolar recruitment together with nonlinear elasticity of the alveoli. The proposed model surprisingly and correctly predicts the observed sigmoidal pressure–volume curves. We discuss the importance of this result and its implications for medical practice.

Phylloseptin-1 (PSN-1) from Phyllomedusa sauvagei skin secretion: A novel broad-spectrum antimicrobial peptide with antibiofilm activity

Amphibian skin secretions have proven to be rich sources of antimicrobial peptides that are proposed to be fundamental components of the innate immune system. As amphibian skin is a multi-functional organ playing, among other things, a crucial role in respiration, it has been deemed that a core biological role for such peptides is control of microbial flora on this surface. To date, however, antimicrobial efficacy has been universally determined by means of establishing minimum inhibitory concentrations (MICs) using planktonic organisms rather than those within a biofilm such as would occur on this exposed surface. Here we describe the identification and structural characterisation of a novel 19 amino acid residue antimicrobial peptide of the phylloseptin family, named PSN-1, from the skin secretion of the waxy monkey frog, Phyllomedusa sauvagei. PSN-1 displayed broad-spectrum activity against a range of planktonic organisms with a high potency (MIC 5 μM) against Staphylococcus aureus. In a specific bioassay with the same organism grown as a biofilm, the minimal biofilm eradication concentration (MBEC) was found to be of the same high potency (5 μM). The present data would suggest that evaluation of actions and potency of amphibian skin secretion antimicrobial peptides might best be achieved by evaluating MBEC rather than MIC using planktonic organisms and that data arising from such studies may have more biological relevance in reflecting the purpose for which they have evolved through natural selection.

Effects of ultraviolet radiation on skin structure and ultrastructure in Caspian Sea Salmon, Salmo trutta caspius, during alevin stage

Ozone reduction enhances the amount of ultraviolet (UV) radiation reaching the aquatic ecosystems, which can induce adverse effects on fish. In order to study the UV radiation effects on skin structure and ultrastructure, the alevins of Salmo trutta caspius were exposed to two different doses of UV radiation (low doses of: 60 µW cm−2 UVC; 100 µW cm−2 UVB; and 40 µW cm−2 UVA and high doses of: 90 µW cm−2 UVC; 130 µW cm−2 UVB; and 50 µW cm−2 UVA) for 15 min once a day in dark condition. Alevins with no exposure to either solar light or UV exposure served as the control group. No mortality was observed in the control group, whereas all of the alevins died in the 9th day in both UV exposure groups. Light microscopic and scanning electron microscopic studies showed significant reduction in the epidermis thickness, number of skin mucous cells, and percentage of mucus secreting cells in both UV exposure groups. Sunburn cells, lifting epidermis from basal membrane, and disappearance of pavement cell microridges were also observed. Our results showed that UV radiation caused intensive skin cell lesions in the Caspian salmon alevins. Since the skin cells of trout alevins possesses essential roles in respiration, osmoregulation, excretion and defense, the observed damages to the skin could be attributed to their sudden mortality in the UV exposure.

Synchrony is altered in the respiratory pattern generator following acute hypoxic challenge

Arthur B. Otis stated in 1954 that the work of breathing accounts for a small fraction of basal metabolism and that inspiration, not expiration, consumes energy. In line with Otis’ findings, our hypothesis is that the respiratory control network uses synchrony, in addition to recruitment and firing rate changes, to generate an efficient respiratory pattern. We hypothesize that synchrony may have unique roles in respiration and that this property is malleable, adapting to maintain homeostasis. To test our hypotheses, we used the perfused in situ preparation of rat (SD/Harlan) and recorded (n=7) from the left phrenic nerve and ensembles of respiratory neurons (n=53) in the Bötzinger/pre‐Bötzinger region using a multielectrode array. Preparations were exposed to acute bouts (20–45s) of hypoxia and spike synchronization between neuronal pairs (n=240) was quantified during baseline and post‐hypoxic epochs (80s/epoch). Neuron pairs from populations which appeared homogenous with respect to firing pattern demonstrated consistent changes in coordinated firing after hypoxia with the exception of a subpopulation of expiratory neurons which showed both increasing and decreasing synchrony. We conclude that synchronization is malleable within populations of the respiratory pattern generator and may contribute to patterning changes during post hypoxic frequency decline. Funded by AHA SDG 0735037N and NIH HL090554

Normal Development of the Lung and Premature Birth

The following review focuses on the normal development of the lung from conception to birth. The defined periods of lung development-Embryonic, Pseudoglandular, Canalicular, Saccular and Alveolar-will be explored in detail in relation to gestational age. Cellular differentiation, formation of the conducting airways and respiratory zone and development of the alveoli will be reviewed. Pulmonary vascular development will also be examined within these periods to relate the formation of the blood-air barrier to the lungs for their essential function of gas exchange after birth. The development of the surfactant and cortisol systems will also be discussed as these need to be mature before the lungs are able to take on their role of respiration following birth. It is clear that premature birth interrupts normal lung development so the effect of preterm birth on lung development will be examined and the respiratory consequences of very preterm birth will be briefly explored.

13C/12C isotope labeling to study carbon partitioning and dark respiration in cereals subjected to water stress

Despite the relevance of carbon (C) loss through respiration processes (with its consequent effect on the lower C availability for grain filling), little attention has been given to this topic. Literature data concerning the role of respiration in cereals are scarce and these have been produced using indirect methods based on gas-exchange estimations. We have developed a new method based on the capture of respired CO(2) samples and their analysis by gas chromatography-combustion-isotope ratio mass spectrometry (GC-C-IRMS). In order to analyse the main processes involved in the C balance during grain filling (photosynthesis, respiration, allocation and partitioning) the ambient isotopic (13)C/(12)C composition (delta(13)C) of the growth chamber was modified during this period (delta(13)C ca. -12.8 +/- 0.3 per thousand to ca. -20.0 +/- 0.2 per thousand). The physiological performance, together with the C allocation on total organic matter (TOM) and respiration of wheat (Triticum aestivum L., var. Califa sur) and two hybrids, tritordeum (X Tritordeum Asch. & Graebn line HT 621) and triticale (X Triticosecale Wittmack var. Imperioso), were compared during post-anthesis water stress. In spite of the larger ear DM/total ratio, especially under drought conditions, the grain filling of triticale and wheat was mainly carried out with pre-anthesis C, while the majority of C assimilated during post-anthesis was invested in respiration processes. In the case of wheat and tritordeum, the C balance data suggested a reallocation during grain filling of photoassimilates stored prior to anthesis from shoot to ear. Furthermore, the lower percentage of labeled C on respired CO(2) of droughted tritordeum plants, together with the lower plant biomass, explained the fact that those plants had more C available for grain filling.