Plasma Membrane Mechanical Stress Activates TRPC5 Channels

Mechanical forces exerted on cells impose stress on the plasma membrane. Cells sense this stress and elicit a mechanoelectric transduction cascade that initiates compensatory mechanisms. Mechanosensitive ion channels in the plasma membrane are responsible for transducing the mechanical signals to electrical signals. However, the mechanisms underlying channel activation in response to mechanical stress remain incompletely understood. Transient Receptor Potential (TRP) channels serve essential functions in several sensory modalities. These channels can also participate in mechanotransduction by either being autonomously sensitive to mechanical perturbation or by coupling to other mechanosensory components of the cell. Here, we investigated the response of a TRP family member, TRPC5, to mechanical stress. Hypoosmolarity triggers Ca²⁺ influx and cationic conductance through TRPC5. Importantly, for the first time we were able to record the stretch-activated TRPC5 current at single-channel level. The activation threshold for TRPC5 was found to be 240 mOsm for hypoosmotic stress and between −20 and −40 mmHg for pressure applied to membrane patch. In addition, we found that disruption of actin filaments suppresses TRPC5 response to hypoosmotic stress and patch pipette pressure, but does not prevent the activation of TRPC5 by stretch-independent mechanisms, indicating that actin cytoskeleton is an essential transduction component that confers mechanosensitivity to TRPC5. In summary, our findings establish that TRPC5 can be activated at the single-channel level when mechanical stress on the cell reaches a certain threshold.     

A scanning and transmission electron microscopic study of the bat lung

The lungs of two adult species of bat Epomophorus wahlbergi and Miniopterus minor fixed with 2.3% glutaraldehyde were processed for SEM (scanning electron microscope) and TEM (transmission electron microscope) examination by the standard procedures. The bat lung comprised a blood and air conducting zone (consisting of bronchi, bronchioles and large blood vessels), the intermediate zone (made up of alveolar ducts), and the respiratory zone, which consisted of alveoli and blood capillaries. The interalveolar septa comprised basically granular pneumocytes (type II cells), squamous pneumocytes (type I cells), endothelial cells, and, in the interstitium, collagen and elastic fibres with occasional fibrocytes. Blood capillaries were interposed in the interalveolar septa, thus bulging into adjacent alveoli. It was noted that grossly, architecturally and structurally, the bat lung was similar to that of a terrestrial mammal. However, in previous morphometric and physiological studies it has been found that bats have a large lung, a thin pulmonary blood-gas barrier, a large pulmonary capillary blood volume, and high haematocrit and haemoglobin concentration. The bat lung, while retaining the basic mammalian pulmonary design, is well adapted to provide the large amount of oxygen demanded by flight. The avian pulmonary design (the lung-air sac system) is thus not a prerequisite to flight.     

Differential responsiveness of late passage C-6 glial cells and advanced passages of astrocytes derived from aged mouse cerebral hemispheres to cytokines and growth factor: Glutamine synthetase activity

In this study, we were interested to compare the responsiveness to growth factors, NGF, b-FGF and EGF and cytokines, IL1β, and TNF-α, in late passages (74–79) C6 glial cells committed astrocytes and astrocytes of advanced passages (26–28) in cultures derived from aged mouse cerebral hemispheres (MACH). Cultures were grown in either DMEM or chemically defined medium (CDM/TIPS) in order to test the effects of growth factors or cytokines. The activity of glutamine synthetase (GS), a marker for astrocytes, was used as a test parameter. We found that treatment with growth factors increased GS activity in both glial cell culture systems with the exception of EGF in C-6 glial cells. Treatment with cytokines markedly decreased GS activity in the late passage C6 glial cells whereas only TNF-α had a similar effect on MACH astrocytes. In view of the generally opposite effects of growth factors and cytokines on GS activity, we-speculate that these molecules which are also endogenously present in glial cells may play a role in the maintenance of cellular homeostasis.     

Spectrum of Microbial Diseases and Resistance Patterns at a Private Teaching Hospital in Kenya: Implications for Clinical Practice

Background

Accurate local prevalence of microbial diseases and microbial resistance data are vital for optimal treatment of patients. However, there are few reports of these data from developing countries, especially from sub-Saharan Africa. The status of Aga Khan University Hospital Nairobi as an internationally accredited hospital and a laboratory with an electronic medical record system has made it possible to analyze local prevalence and antimicrobial susceptibility data and compare it with other published data.

Methods

We have analyzed the spectrum of microbial agents and resistance patterns seen at a 300 bed tertiary private teaching hospital in Kenya using microbial identity and susceptibility data captured in hospital and laboratory electronic records between 2010 and 2014.

Results

For blood isolates, we used culture collection within the first three days of hospitalization as a surrogate for community onset, and within that group, Escherichia coli was the most common, followed by Staphylococcus aureus. In contrast, Candida spp. and Klebsiella pneumoniae were the most common hospital onset causes of bloodstream infection. Antimicrobial resistance rates for the most commonly isolated Gram negative organisms was higher than many recent reports from Europe and North America. In contrast, Gram positive resistance rates were quite low, with 94% of S. aureus being susceptible to oxacillin and only rare isolates of vancomycin-resistant enterococci.

Conclusions

The current report demonstrates high rates of antimicrobial resistance in Gram negative organisms, even in outpatients with urinary tract infections. On the other hand, rates of resistance in Gram positive organisms, notably S. aureus, are remarkably low. A better understanding of the reasons for these trends may contribute to ongoing efforts to combat antimicrobial resistance globally.     

Phenol metabolism, phytoalexins, and respiration in potato tuber tissue treated with Fatty Acid

Potato (solanum tuberosum L. cv Katahdin) tuber discs treated with arachidonic acid become necrotic and accumulate sesquiterpenoid phytoalexins. The arachidonic acid also causes increases in both phenylalanine ammonia lyase and lignin, but no change in total alcohol-soluble phenols. Linoleic acid does not alter any of these parameters. A high concentration of nonanoic acid promotes both necrosis and accumulation of low levels of phytoalexins, but decreased levels of phenols, phenylalanine ammonia lyase, and lignin. The respiration of the control discs and those treated with linoleic acid declines by 24 hours after treatment, but the respiration of arachidonic acid-treated discs remains constant for at least 48 hours.     

Morphometric characterization of the airway and vascular systems of the lung of the domestic pig, Sus scrofa: comparison of the airway, arterial and venous systems

The bronchial system (BS), the pulmonary artery (PA) and the pulmonary vein (PV) of the lung of the domestic pig, Sus scrofa were simultaneously cast with silicone rubber and studied. Asymmetrical dichotomous bifurcation preponderated in the tree-like arrangement of the three conducting systems. Lengths and diameters of the various generations were measured. At the extremities of the BS and the PA, alveoli and blood capillaries related very closely. In the cranial and middle lobes of the right and left lungs, topographically, the PA and the PV closely followed the BS, but in the accessory and the caudal (diaphragmatic) lobes, only the PA accompanied the BS: the PV run intersegmentally. Certain similarities and differences were observed between the diameters and lengths of the various generations of the three conducting systems. The strong correlations between some of the structural parameters indicated a high level of structural optimization. While morphometric variations suggest that the air and the blood flow dynamics may somewhat differ between the three conducting systems, they may also register structural features unique to the lung of the domestic pig, an animal that has been highly genetically exploited for fast growth and now leads an indolent lifestyle in captivity.     

Deconvoluting lung evolution: from phenotypes to gene regulatory networks

Speakers in this symposium presented examples of respiratoryregulation that broadly illustrate principles of evolution fromwhole organ to genes. The swim bladder and lungs of aquaticand terrestrial organisms arose independently from a commonprimordial "respiratory pharynx" but not from each other. Pathwaysof lung evolution are similar between crocodiles and birds buta low compliance of mammalian lung may have driven the developmentof the diaphragm to permit lung inflation during inspiration.To meet the high oxygen demands of flight, bird lungs have evolvedseparate gas exchange and pump components to achieve unidirectionalventilation and minimize dead space. The process of "screening"(removal of oxygen from inspired air prior to entering the terminalunits) reduces effective alveolar oxygen tension and potentiallyexplains why nonathletic large mammals possess greater pulmonarydiffusing capacities than required by their oxygen consumption.The "primitive" central admixture of oxygenated and deoxygenatedblood in the incompletely divided reptilian heart is actuallyco-regulated with other autonomic cardiopulmonary responsesto provide flexible control of arterial oxygen tension independentof ventilation as well as a unique mechanism for adjusting metabolicrate. Some of the most ancient oxygen-sensing molecules, i.e.,hypoxia-inducible factor-1alpha and erythropoietin, are up-regulatedduring mammalian lung development and growth under apparentlynormoxic conditions, suggesting functional evolution. Normalalveolarization requires pleiotropic growth factors acting viahighly conserved cell–cell signal transduction, e.g.,parathyroid hormone-related protein transducing at least partlythrough the Wingless/int pathway. The latter regulates morphogenesisfrom nematode to mammal. If there is commonality among thesediverse respiratory processes, it is that all levels of organization,from molecular signaling to structure to function, co-evolveprogressively, and optimize an existing gas-exchange framework.     

The anatomy, physics, and physiology of gas exchange surfaces: is there a universal function for pulmonary surfactant in animal respiratory structures?

(Orgeig and Daniels) This surfactant symposium reflects theintegrative and multidisciplinary aims of the 1st ICRB, by encompassingin vitro and in vivo research, studies of vertebrates and invertebrates,and research across multiple disciplines. We explore the physicaland structural challenges that face gas exchange surfaces invertebrates and insects, by focusing on the role of the surfactantsystem. Pulmonary surfactant is a complex mixture of lipidsand proteins that lines the air–liquid interface of thelungs of all air-breathing vertebrates, where it functions tovary surface tension with changing lung volume. We begin witha discussion of the extraordinary conservation of the blood–gasbarrier among vertebrate respiratory organs, which has evolvedto be extremely thin, thereby maximizing gas exchange, but simultaneouslystrong enough to withstand significant distension forces. Theprincipal components of pulmonary surfactant are highly conserved,with a mixed phospholipid and neutral lipid interfacial filmthat is established, maintained and dynamically regulated bysurfactant proteins (SP). A wide variation in the concentrationsof individual components exists, however, and highlights lipidomicas well as proteomic adaptations to different physiologicalneeds. As SP-B deficiency in mammals is lethal, oxidative stressto SP-B is detrimental to the biophysical function of pulmonarysurfactant and SP-B is evolutionarily conserved across the vertebrates.It is likely that SP-B was essential for the evolutionary originof pulmonary surfactant. We discuss three specific issues ofthe surfactant system to illustrate the diversity of functionin animal respiratory structures. (1) Temperature: In vitroanalyses of the behavior of different model surfactant filmsunder dynamic conditions of surface tension and temperaturesuggest that, contrary to previous beliefs, the alveolar filmmay not have to be substantially enriched in the disaturatedphospholipid, dipalmitoylphosphatidylcholine (DPPC), but thatsimilar properties of rate of film formation can be achievedwith more fluid films. Using an in vivo model of temperaturechange, a mammal that enters torpor, we show that film structureand function varies between surfactants isolated from torpidand active animals. (2) Spheres versus tubes: Surfactant isessential for lung stabilization in vertebrates, but its functionis not restricted to the spherical alveolus. Instead, surfactantis also important in narrow tubular respiratory structures suchas the terminal airways of mammals and the air capillaries ofbirds. (3). Insect tracheoles: We investigate the structureand function of the insect tracheal system and ask whether pulmonarysurfactant also has a role in stabilizing these minute tubules.Our theoretical analysis suggests that a surfactant system maybe required, in order to cope with surface tension during processes,such as molting, when the tracheae collapse and fill with water.Hence, despite observations by Wigglesworth in the 1930s offluid-filled tracheoles, the challenge persists into the 21stcentury to determine whether this fluid is associated with apulmonary-type surfactant system. Finally, we summarize thecurrent status of the field and provide ideas for future research.     

Aberrant striatal dopamine transmitter dynamics in brain-derived neurotrophic factor-deficient mice

Brain-derived neurotrophic factor (BDNF) modulates the synaptic transmission of several monoaminergic neuronal systems, including forebrain dopamine-containing neurons. Recent evidence shows a strong correlation between neuropsychiatric disorders and BDNF hypofunction. The aim of the present study was to characterize the effect of low endogenous levels of BDNF on dopamine system function in the caudate-putamen using heterozygous BDNF (BDNF(+/-) ) mice. Apparent extracellular dopamine levels in the caudate-putamen, determined by quantitative microdialysis, were significantly elevated in BDNF(+/-) mice compared with wildtype controls (12 vs. 5 nM, respectively). BDNF(+/-) mice also had a potentiated increase in dopamine levels following potassium (120 mM)-stimulation (10-fold) relative to wildtype controls (6-fold). Slice fast-scan cyclic voltammetry revealed that BDNF(+/-) mice had reductions in both electrically evoked dopamine release and dopamine uptake rates in the caudate-putamen. Superfusion of BDNF led to partial recovery of the electrically stimulated dopamine release response in BDNF(+/-) mice. Conversely, tissue accumulation of L-3,4-dihydroxyphenylalanine, extracellular levels of dopamine metabolites, and spontaneous locomotor activity were unaltered. Together, this study indicates that endogenous BDNF influences dopamine system homeostasis by regulating the release and uptake dynamics of pre-synaptic dopamine transmission.