VEGFD Downregulation in Hippocampal Area CA1: Effects on Dendritic Morphology of Pyramidal Neurons and Network Activity

Neurons constitute the cellular basis of the nervous system and their activity generates most functions of the brain. Although, neurons exhibit highly diverse structures, their morphology comprises three main functional compartments. Dendrites are ramified structures and the major input side of neurons. Different input signals converge at the soma in order to generate action potentials. This output is transmitted through the axon to other neurons, thus creating complex neuronal networks. The morphology of dendritic arbors has a high impact on the connectivity and the electrical properties of neurons, determining input signals and their transmission and integration. It is therefore of great importance to study the link between dendritic morphology, electrical properties of single neurons and functions on the network level. By studying neurodegenerative diseases, it becomes clear that particularly the maintenance of the neuronal morphology is crucial for proper cognitive performance. Previous studies demonstrated that the growth factor VEGFD regulates the maintenance of the dendritic length in cultured neurons and of basal dendrites in the mouse hippocampus. Furthermore, VEGFD deficient mice show a long-term memory deficit, suggesting a link between altered dendritic morphology and cognitive function deficits. In this study, the regulatory effect of VEGFD on the branching of distinct dendritic compartments and the functional consequences of VEGFD loss both on a single cell level and on the level of network oscillations were investigated. Following the suppression of VEGFD in CA1 pyramidal neurons in vivo, detailed morphometric analyses, including basal and apical dendrites as well as spine density, confirmed that basal dendrites loose length and overall complexity. Interestingly, the apical dendrites increased in length and size. This elongation occurred in areas with a high density of synaptic inputs. The spine density in all areas remained unaltered, indicating a shift in input ratio between longer apical and smaller basal dendritic trees. Furthermore, the functional consequences of VEGFD loss on single cell level and on network oscillations were investigated. Single cell patch clamp recordings revealed no changes in passive membrane properties and action potential firing properties, apart from an increased input resistance. The in vivo electrical stimulation of two distinct input pathways to CA1 revealed no alterations in the synaptic input strength. The comparison of several network oscillations between knockdown mice and control animals revealed no differences. These surprising results indicate that changed dendritic morphology due to the loss of VEGFD has no impact on network oscillations, such as theta, gamma and SPW R, in different behavioral states like running, immobility and sleep

Microbial Cell Factories / Induction without methanol: novel regulated promoters enable high-level expression in Pichia pastoris

Background: Inducible high-level expression is favoured for recombinant protein production in Pichia pastoris. Therefore, novel regulated promoters are desired, ideally repressing heterologous gene expression during initial growth and enabling it in the production phase. In a typical large scale fed-batch culture repression is desired during the batch phase where cells grow on a surplus of e.g. glycerol, while heterologous gene expression should be active in the feed phase under carbon (e.g. glucose) limitation. Results: DNA microarray analysis of P. pastoris wild type cells growing in glycerol-based batch and glucose-based fed batch was used for the identification of genes with both, strong repression on glycerol and high-level expression in the feed phase. Six novel glucose-limit inducible promoters were successfully applied to express the intracellular reporter eGFP. The highest expression levels together with strong repression in pre-culture were achieved with the novel promoters PG1 and PG6. Human serum albumin (HSA) was used to characterize the promoters with an industrially relevant secreted protein. A PG1 clone with two gene copies reached about 230% of the biomass specific HSA titer in glucose-based fed batch fermentation compared to a PGAP clone with identical gene copy number, while PG6 only achieved 39%. Two clones each carrying eleven gene copies, expressing HSA under control of PG1 and PG6 respectively were generated by post-transformational vector amplification. They produced about 1.0 and 0.7 g L-1 HSA respectively in equal fed batch processes. The suitability in production processes was also verified with HyHEL antibody Fab fragment for PG1 and with porcine carboxypeptidase B for PG6. Moreover, the molecular function of the gene under the control of PG1 was determined to encode a high-affinity glucose transporter and named GTH1. Conclusions: A set of novel regulated promoters, enabling induction without methanol, was successfully identified by using DNA microarrays and shown to be suitable for high level expression of recombinant proteins in glucose-based protein production processes

Missing elimination via membrane vesicle shedding contributes to the diminished calcium sensitivity of listeriolysin O

The lytic capacity of cholesterol-dependent cytolysins is enhanced in the extracellular calcium-free environment through a combination of limited membrane repair and diminished membrane toxin removal. For a typical neurotoxin of the group, pneumolysin, this effect has already been observed at reduced (1 mM) calcium conditions, which are pathophysiologically relevant. Here, we tested another neurotoxin of the group, listeriolysin O from L. monocytogenes, active in the primary vacuole after bacterium phagocytosis in host cells. Reduced calcium did not increase the lytic capacity of listeriolysin (in contrast to pneumolysin), while calcium-free conditions elevated it 2.5 times compared to 10 times for pneumolysin (at equivalent hemolytic capacities). To clarify these differences, we analyzed membrane vesicle shedding, known to be a calcium-dependent process for toxin removal from eukaryotic cell membranes. Both pneumolysin and listeriolysin initiated vesicle shedding, which was completely blocked by the lack of extracellular calcium. Lack of calcium, however, elevated the toxin load per a cell only for pneumolysin and not for listeriolysin. This result indicates that vesicle shedding does not play a role in the membrane removal of listeriolysin and outlines a major difference between it and other members of the CDC group. Furthermore, it provides new tools for studying membrane vesicle shedding

Distinct Neurotoxicity Profile of Listeriolysin O from Listeria monocytogenes

Cholesterol-dependent cytolysins (CDCs) are protein toxins that originate from Gram-positive bacteria and contribute substantially to their pathogenicity. CDCs bind membrane cholesterol and build prepores and lytic pores. Some effects of the toxins are observed in non-lytic concentrations. Two pathogens, Streptococcus pneumoniae and Listeria monocytogenes, cause fatal bacterial meningitis, and both produce toxins of the CDC family—pneumolysin and listeriolysin O, respectively. It has been demonstrated that pneumolysin produces dendritic varicosities (dendrite swellings) and dendritic spine collapse in the mouse neocortex, followed by synaptic loss and astrocyte cell shape remodeling without elevated cell death. We utilized primary glial cultures and acute mouse brain slices to examine the neuropathological effects of listeriolysin O and to compare it to pneumolysin with identical hemolytic activity. In cultures, listeriolysin O permeabilized cells slower than pneumolysin did but still initiated non-lytic astrocytic cell shape changes, just as pneumolysin did. In an acute brain slice culture system, listeriolysin O produced dendritic varicosities in an NMDA-dependent manner but failed to cause dendritic spine collapse and cortical astrocyte reorganization. Thus, listeriolysin O demonstrated slower cell permeabilization and milder glial cell remodeling ability than did pneumolysin and lacked dendritic spine collapse capacity but exhibited equivalent dendritic pathology

Respiratory plasticity in response to changes in oxygen supply and demand

Aerobic organisms maintain O2 homeostasis by responding to changes in O2 supply and demand in both short and long time domains. In this review, we introduce several specific examples of respiratory plasticity induced by chronic changes in O2 supply (environmental hypoxia or hyperoxia) and demand (exercise-induced and temperature-induced changes in aerobic metabolism). These studies reveal that plasticity occurs throughout the respiratory system, including modifications to the gas exchanger, respiratory pigments, respiratory muscles, and the neural control systems responsible for ventilating the gas exchanger. While some of these responses appear appropriate (e.g., increases in lung surface area, blood O2 capacity, and pulmonary ventilation in hypoxia), other responses are potentially harmful (e.g., increased muscle fatigability). Thus, it may be difficult to predict whole-animal performance based on the plasticity of a single system. Moreover, plastic responses may differ quantitatively and qualitatively at different developmental stages. Much of the current research in this field is focused on identifying the cellular and molecular mechanisms underlying respiratory plasticity. These studies suggest that a few key molecules, such as hypoxia inducible factor (HIF) and erythropoietin, may be involved in the expression of diverse forms of plasticity within and across species. Studying the various ways in which animals respond to respiratory challenges will enable a better understanding of the integrative response to chronic changes in O2 supply and deman

Magnetospheric Venus Space Explorers (MVSE) Mission: A Proposal for Understanding the Dynamics of Induced Magnetospheres

Induced magnetospheres form around planetary bodies with atmospheres through the interaction of the solar wind with their ionosphere. Induced magnetospheres are highly dependent on the solar wind conditions and have only been studied with single spacecraft missions in the past. This gap in knowledge could be addressed by a multi-spacecraft plasma mission, optimized for studying global spatial and temporal variations in the magnetospheric system around Venus, which hosts the most prominent example of an induced magnetosphere in our solar system. The MVSE mission comprises four satellites, of which three are identical scientific spacecraft, carrying the same suite of instruments probing different regions of the induced magnetosphere and the solar wind simultaneously. The fourth spacecraft is the transfer vehicle which acts as a relay satellite for communications at Venus. In this way, changes in the solar wind conditions and extreme solar events can be observed, and their effects can be quantified as they propagate through the Venusian induced magnetosphere. Additionally, energy transfer in the Venusian induced magnetosphere can be investigated. The scientific payload includes instrumentation to measure the magnetic field, electric field, and ion-electron velocity distributions. This study presents the scientific motivation for the mission as well as requirements and the resulting mission design. Concretely, a mission timeline along with a complete spacecraft design, including mass, power, communication, propulsion and thermal budgets are given. This mission was initially conceived at the Alpbach Summer School 2022 and refined during a week-long study at ESAs Concurrent Design Facility in Redu, BelgiumComment: 23 pages, 5 figures, Submitted to Acta Astronautic

The epithelial barrier: The gateway to allergic, autoimmune, and metabolic diseases and chronic neuropsychiatric conditions

Since the 1960 s, our health has been compromised by exposure to over 350,000 newly introduced toxic substances, contributing to the current pandemic in allergic, autoimmune and metabolic diseases. The "Epithelial Barrier Theory" postulates that these diseases are exacerbated by persistent periepithelial inflammation (epithelitis) triggered by exposure to a wide range of epithelial barrier-damaging substances as well as genetic susceptibility. The epithelial barrier serves as the body's primary physical, chemical, and immunological barrier against external stimuli. A leaky epithelial barrier facilitates the translocation of the microbiome from the surface of the afflicted tissues to interepithelial and even deeper subepithelial locations. In turn, opportunistic bacterial colonization, microbiota dysbiosis, local inflammation and impaired tissue regeneration and remodelling follow. Migration of inflammatory cells to susceptible tissues contributes to damage and inflammation, initiating and aggravating many chronic inflammatory diseases. The objective of this review is to highlight and evaluate recent studies on epithelial physiology and its role in the pathogenesis of chronic diseases in light of the epithelial barrier theory

Observation of associated near-side and away-side long-range correlations in √sNN=5.02 TeV proton-lead collisions with the ATLAS detector

Two-particle correlations in relative azimuthal angle (Δϕ) and pseudorapidity (Δη) are measured in √sNN=5.02  TeV p+Pb collisions using the ATLAS detector at the LHC. The measurements are performed using approximately 1  μb-1 of data as a function of transverse momentum (pT) and the transverse energy (ΣETPb) summed over 3.1<η<4.9 in the direction of the Pb beam. The correlation function, constructed from charged particles, exhibits a long-range (2<|Δη|<5) “near-side” (Δϕ∼0) correlation that grows rapidly with increasing ΣETPb. A long-range “away-side” (Δϕ∼π) correlation, obtained by subtracting the expected contributions from recoiling dijets and other sources estimated using events with small ΣETPb, is found to match the near-side correlation in magnitude, shape (in Δη and Δϕ) and ΣETPb dependence. The resultant Δϕ correlation is approximately symmetric about π/2, and is consistent with a dominant cos⁡2Δϕ modulation for all ΣETPb ranges and particle pT

Search for direct production of charginos and neutralinos in events with three leptons and missing transverse momentum in √s = 7 TeV pp collisions with the ATLAS detector

A search for the direct production of charginos and neutralinos in final states with three electrons or muons and missing transverse momentum is presented. The analysis is based on 4.7 fb−1 of proton–proton collision data delivered by the Large Hadron Collider and recorded with the ATLAS detector. Observations are consistent with Standard Model expectations in three signal regions that are either depleted or enriched in Z-boson decays. Upper limits at 95% confidence level are set in R-parity conserving phenomenological minimal supersymmetric models and in simplified models, significantly extending previous results

Measurements of fiducial and differential cross sections for Higgs boson production in the diphoton decay channel at s√=8 TeV with ATLAS

Measurements of fiducial and differential cross sections are presented for Higgs boson production in proton-proton collisions at a centre-of-mass energy of s√=8 TeV. The analysis is performed in the H → γγ decay channel using 20.3 fb−1 of data recorded by the ATLAS experiment at the CERN Large Hadron Collider. The signal is extracted using a fit to the diphoton invariant mass spectrum assuming that the width of the resonance is much smaller than the experimental resolution. The signal yields are corrected for the effects of detector inefficiency and resolution. The pp → H → γγ fiducial cross section is measured to be 43.2 ±9.4(stat.) − 2.9 + 3.2 (syst.) ±1.2(lumi)fb for a Higgs boson of mass 125.4GeV decaying to two isolated photons that have transverse momentum greater than 35% and 25% of the diphoton invariant mass and each with absolute pseudorapidity less than 2.37. Four additional fiducial cross sections and two cross-section limits are presented in phase space regions that test the theoretical modelling of different Higgs boson production mechanisms, or are sensitive to physics beyond the Standard Model. Differential cross sections are also presented, as a function of variables related to the diphoton kinematics and the jet activity produced in the Higgs boson events. The observed spectra are statistically limited but broadly in line with the theoretical expectations