Heme oxygenase-2 gene deletion attenuates oxidative stress in neurons exposed to extracellular hemin

Background

Hemin, the oxidized form of heme, accumulates in intracranial hematomas and is a potent oxidant. Growing evidence suggests that it contributes to delayed injury to surrounding tissue, and that this process is affected by the heme oxygenase enzymes. In a prior study, heme oxygenase-2 gene deletion increased the vulnerability of cultured cortical astrocytes to hemin. The present study tested the effect of HO-2 gene deletion on protein oxidation, reactive oxygen species formation, and cell viability after mixed cortical neuron/astrocyte cultures were incubated with neurotoxic concentrations of hemin.

Results

Continuous exposure of wild-type cultures to 1–10 μM hemin for 14 h produced concentration-dependent neuronal death, as detected by both LDH release and fluorescence intensity after propidium iodide staining, with an EC₅₀ of 1–2 μM; astrocytes were not injured by these low hemin concentrations. Cell death was consistently reduced by at least 60% in knockout cultures. Exposure to hemin for 4 hours, a time point that preceded cell lysis, increased protein oxidation in wild-type cultures, as detected by staining of immunoblots for protein carbonyl groups. At 10 μM hemin, carbonylation was increased 2.3-fold compared with control sister cultures subjected to medium exchanges only; this effect was reduced by about two-thirds in knockout cultures. Cellular reactive oxygen species, detected by fluorescence intensity after dihydrorhodamine 123 (DHR) staining, was markedly increased by hemin in wild-type cultures and was localized to neuronal cell bodies and processes. In contrast, DHR fluorescence intensity in knockout cultures did not differ from that of sham-washed controls. Neuronal death in wild-type cultures was almost completely prevented by the lipid-soluble iron chelator phenanthroline; deferoxamine had a weaker but significant effect.

Conclusions

These results suggest that HO-2 gene deletion protects neurons in mixed neuron-astrocyte cultures from heme-mediated oxidative injury. Selective inhibition of neuronal HO-2 may have a beneficial effect after CNS hemorrhage.     

Neural mechanisms underlying the facilitation of naming in aphasia using a semantic task: an fMRI study

Background

In the fruit fly, Drosophila melanogaster, serotonin functions both as a neurotransmitter to regulate larval feeding, and in the development of the stomatogastric feeding circuit. There is an inverse relationship between neuronal serotonin levels during late embryogenesis and the complexity of the serotonergic fibers projecting from the larval brain to the foregut, which correlate with perturbations in feeding, the functional output of the circuit. Dopamine does not modulate larval feeding, and dopaminergic fibers do not innervate the larval foregut. Since dopamine can function in central nervous system development, separate from its role as a neurotransmitter, the role of neuronal dopamine was assessed on the development, and mature function, of the 5-HT larval feeding circuit.

Results

Both decreased and increased neuronal dopamine levels in late embryogenesis during development of this circuit result in depressed levels of larval feeding. Perturbations in neuronal dopamine during this developmental period also result in greater branch complexity of the serotonergic fibers innervating the gut, as well as increased size and number of the serotonin-containing vesicles along the neurite length. This neurotrophic action for dopamine is modulated by the D₂ dopamine receptor expressed during late embryogenesis in central 5-HT neurons. Animals carrying transgenic RNAi constructs to knock down both dopamine and serotonin synthesis in the central nervous system display normal feeding and fiber architecture. However, disparate levels of neuronal dopamine and serotonin during development of the circuit result in abnormal gut fiber architecture and feeding behavior.

Conclusions

These results suggest that dopamine can exert a direct trophic influence on the development of a specific neural circuit, and that dopamine and serotonin may interact with each other to generate the neural architecture necessary for normal function of the circuit.     

Cingulate seizure-like activity reveals neuronal avalanche regulated by network excitability and thalamic inputs

Background

Cortical neurons display network-level dynamics with unique spatiotemporal patterns that construct the backbone of processing information signals and contribute to higher functions. Recent years have seen a wealth of research on the characteristics of neuronal networks that are sufficient conditions to activate or cease network functions. Local field potentials (LFPs) exhibit a scale-free and unique event size distribution (i.e., a neuronal avalanche) that has been proven in the cortex across species, including mice, rats, and humans, and may be used as an index of cortical excitability. In the present study, we induced seizure activity in the anterior cingulate cortex (ACC) with medial thalamic inputs and evaluated the impact of cortical excitability and thalamic inputs on network-level dynamics. We measured LFPs from multi-electrode recordings in mouse cortical slices and isoflurane-anesthetized rats.

Results

The ACC activity exhibited a neuronal avalanche with regard to avalanche size distribution, and the slope of the power-law distribution of the neuronal avalanche reflected network excitability in vitro and in vivo. We found that the slope of the neuronal avalanche in seizure-like activity significantly correlated with cortical excitability induced by γ-aminobutyric acid system manipulation. The thalamic inputs desynchronized cingulate seizures and affected the level of cortical excitability, the modulation of which could be determined by the slope of the avalanche size.

Conclusions

We propose that the neuronal avalanche may be a tool for analyzing cortical activity through LFPs to determine alterations in network dynamics.     

Niche-dependent development of functional neuronal networks from embryonic stem cell-derived neural populations

Background

Minocycline, a second-generation tetracycline with anti-inflammatory and anti-apoptotic properties, has been shown to promote therapeutic benefits in experimental stroke. However, equally compelling evidence demonstrates that the drug exerts variable and even detrimental effects in many neurological disease models. Assessment of the mechanism underlying minocycline neuroprotection should clarify the drug's clinical value in acute stroke setting.

Results

Here, we demonstrate that minocycline attenuates both in vitro (oxygen glucose deprivation) and in vivo (middle cerebral artery occlusion) experimentally induced ischemic deficits by direct inhibition of apoptotic-like neuronal cell death involving the anti-apoptotic Bcl-2/cytochrome c pathway. Such anti-apoptotic effect of minocycline is seen in neurons, but not apparent in astrocytes. Our data further indicate that the neuroprotection is dose-dependent, in that only low dose minocycline inhibits neuronal cell death cascades at the acute stroke phase, whereas the high dose exacerbates the ischemic injury.

Conclusion

The present study advises our community to proceed with caution to use the minimally invasive intravenous delivery of low dose minocycline in order to afford neuroprotection that is safe for stroke.     

Precision and accuracy of the subjective haptic vertical in the roll plane

Background

Autophagy, an intracellular response to stress, is characterized by double membrane cytosolic vesicles called autophagosomes. Prolonged autophagy is known to result in autophagic (Type II) cell death. This study examined the potential role of an autophagic response in cultured cerebellar granule neurons challenged with excitotoxin N-methyl-D-aspartate (NMDA).

Results

NMDA exposure induced light chain-3 (LC-3)-immunopositive and monodansylcadaverine (MDC) fluorescent dye-labeled autophagosome formation in both cell bodies and neurites as early as 3 hours post-treatment. Elevated levels of Beclin-1 and the autophagosome-targeting LC3-II were also observed following NMDA exposure. Prolonged exposure of the cultures to NMDA (8-24 h) generated MDC-, LC3-positive autophagosomal bodies, concomitant with the neurodegenerative phase of NMDA challenge. Lysosomal inhibition studies also suggest that NMDA-treatment diverted the autophagosome-associated LC3-II from the normal lysosomal degradation pathway. Autophagy inhibitor 3-methyladenine significantly reduced NMDA-induced LC3-II/LC3-I ratio increase, accumulation of autophagosomes, and suppressed NMDA-mediated neuronal death. ATG7 siRNA studies also showed neuroprotective effects following NMDA treatment.

Conclusions

Collectively, this study shows that autophagy machinery is robustly induced in cultured neurons subjected to prolonged exposure to excitotoxin, while autophagosome clearance by lysosomal pathway might be impaired. Our data further show that prolonged autophagy contributes to cell death in NMDA-mediated excitotoxicity.     

Analysis of spatial relationships in three dimensions: tools for the study of nerve cell patterning

Background

Multiple technologies have been brought to bear on understanding the three-dimensional morphology of individual neurons and glia within the brain, but little progress has been made on understanding the rules controlling cellular patterning. We describe new matlab-based software tools, now available to the scientific community, permitting the calculation of spatial statistics associated with 3D point patterns. The analyses are largely derived from the Delaunay tessellation of the field, including the nearest neighbor and Voronoi domain analyses, and from the spatial autocorrelogram.

Results

Our tools enable the analysis of the spatial relationship between neurons within the central nervous system in 3D, and permit the modeling of these fields based on lattice-like simulations, and on simulations of minimal-distance spacing rules. Here we demonstrate the utility of our analysis methods to discriminate between two different simulated neuronal populations.

Conclusion

Together, these tools can be used to reveal the presence of nerve cell patterning and to model its foundation, in turn informing on the potential developmental mechanisms that govern its establishment. Furthermore, in conjunction with analyses of dendritic morphology, they can be used to determine the degree of dendritic coverage within a volume of tissue exhibited by mature nerve cells.     

Annihilation and topological cross sections forpp and\bar p p interactions at 200 GeV/c

The results of an analysis of streamer chamber data from the NA5 experiment are presented. Topological cross sections forpp and \(\bar p\) p interactions and strange neutral particle production at 200 GeV/c have been measured and ( \(\bar p\) p?pp) difference cross sections have been determined. Multiplicity moments were calculated.     

Structure of 1-RSB Asymptotic Gibbs Measures in the Diluted p-Spin Models

In this paper we study asymptotic Gibbs measures in the diluted $p$ -spin models in the so called $1$ -RSB case, when the overlap takes two values $q_*, q^*\in [0,1].$ When the external field is not present and the overlap is not equal to zero, we prove that such asymptotic Gibbs measures are described by the Mézard–Parisi ansatz conjectured in Mézard and Parisi (Eur Phys J B 20(2):217–233 2001). When the external field is present, we prove that the overlap can not be equal to zero and all $1$ -RSB asymptotic Gibbs measures are described by the Mézard–Parisi ansatz. Finally, we give a characterization of the exceptional case when there is no external field and the smallest overlap value $q_*=0$ , although it does not go as far as the Mézard–Parisi ansatz. Our approach is based on the cavity computations combined with the hierarchical exchangeability of pure states.     

Directed neuronal differentiation of human embryonic stem cells

Background

We have developed a culture system for the efficient and directed differentiation of human embryonic stem cells (HESCs) to neural precursors and neurons.HESC were maintained by manual passaging and were differentiated to a morphologically distinct OCT-4⁺/SSEA-4^- monolayer cell type prior to the derivation of embryoid bodies. Embryoid bodies were grown in suspension in serum free conditions, in the presence of 50% conditioned medium from the human hepatocarcinoma cell line HepG2 (MedII).

Results

A neural precursor population was observed within HESC derived serum free embryoid bodies cultured in MedII conditioned medium, around 7–10 days after derivation. The neural precursors were organized into rosettes comprised of a central cavity surrounded by ring of cells, 4 to 8 cells in width. The central cells within rosettes were proliferating, as indicated by the presence of condensed mitotic chromosomes and by phosphoHistone H3 immunostaining. When plated and maintained in adherent culture, the rosettes of neural precursors were surrounded by large interwoven networks of neurites. Immunostaining demonstrated the expression of nestin in rosettes and associated non-neuronal cell types, and a radial expression of Map-2 in rosettes. Differentiated neurons expressed the markers Map-2 and Neurofilament H, and a subpopulation of the neurons expressed tyrosine hydroxylase, a marker for dopaminergic neurons.

Conclusion

This novel directed differentiation approach led to the efficient derivation of neuronal cultures from HESCs, including the differentiation of tyrosine hydroxylase expressing neurons. HESC were morphologically differentiated to a monolayer OCT-4⁺ cell type, which was used to derive embryoid bodies directly into serum free conditions. Exposure to the MedII conditioned medium enhanced the derivation of neural precursors, the first example of the effect of this conditioned medium on HESC.

     

Superintegrability in Two Dimensions and the Racah–Wilson Algebra

The analysis of the most general second-order superintegrable system in two dimensions: the generic 3-parameter model on the 2-sphere is cast in the framework of the Racah problem for the \({\mathfrak{su}(1,1)}\) algebra. The Hamiltonian of the 3-parameter system and the generators of its quadratic symmetry algebra are seen to correspond to the total and intermediate Casimir operators of the combination of three \({\mathfrak{su}(1,1)}\) algebras, respectively. The construction makes explicit the isomorphism between the Racah–Wilson algebra, which is the fundamental algebraic structure behind the Racah problem for \({\mathfrak{su}(1, 1)}\) , and the invariance algebra of the generic 3-parameter system. It also provides an explanation for the occurrence of the Racah polynomials as overlap coefficients in this context. The irreducible representations of the Racah–Wilson algebra are reviewed as well as their connection with the Askey scheme of classical orthogonal polynomials.     

Spherical top-hat collapse of a viscous unified dark fluid

In this paper, we test the spherical collapse of a viscous unified dark fluid (VUDF) which has constant adiabatic sound speed and show the nonlinear collapse for VUDF, baryons, and dark matter, which are important in forming the large-scale structure of our Universe. By varying the values of the model parameters $\alpha $ and $\zeta _{0}$ , we discuss their effects on the nonlinear collapse of the VUDF model, and we compare its result to the $\Lambda $ CDM model. The results of the analysis show that, within the spherical top-hat collapse framework, larger values of $\alpha $ and smaller values of $\zeta _{0}$ make the structure formation earlier and faster, and the other collapse curves are almost distinguished with the curve of $\Lambda $ CDM model if the bulk viscosity coefficient $\zeta _{0}$ is less than $10^{-3}$ .     

Neutral strange particle production in proton-proton collisions at\sqrt s = 63 GeV

The inclusive cross section for the production ofK _s ⁰ mesons, Λ and \(\bar \Lambda\) particles in proton-proton interactions at \(\sqrt s = 63\) GeV is presented. The produced particles have been detected in the full phase space. Behaviour of the longitudinal and transversal depandences of the cross sections are discussed. The total production cross sections for _s ⁰ mesons and Λ particles was determined to \(\sigma _{{\rm K}_S^0 } = (25.5 \pm 1.4)\) mb andσ _Λ =(7.8±1.2) mb respectively. A strong energy dependence of the production cross sections is observed.     

A role for the membrane protein M6 in the Drosophila visual system

Background

Members of the proteolipid protein family, including the four-transmembrane glycoprotein M6a, are involved in neuronal plasticity in mammals. Results from our group previously demonstrated that M6, the only proteolipid protein expressed in Drosophila, localizes to the cell membrane in follicle cells. M6 loss triggers female sterility, which suggests a role for M6 in follicular cell remodeling. These results were the basis of the present study, which focused on the function and requirements of M6 in the fly nervous system.

Results

The present study identified two novel, tissue-regulated M6 isoforms with variable N- and C- termini, and showed that M6 is the functional fly ortholog of Gpm6a. In the adult brain, the protein was localized to several neuropils, such as the optic lobe, the central complex, and the mushroom bodies. Interestingly, although reduced M6 levels triggered a mild rough-eye phenotype, hypomorphic M6 mutants exhibited a defective response to light.

Conclusions

Based on its ability to induce filopodium formation we propose that M6 is key in cell remodeling processes underlying visual system function. These results bring further insight into the role of M6/M6a in biological processes involving neuronal plasticity and behavior in flies and mammals.     

Spin Effects in the Interaction of Antiprotons with the Deuteron at Low and Intermediate Energies

Antiproton-deuteron scattering is analyzed within the Glauber theory, accounting for the full spin dependence of the underlying \({\bar{N}N}\) amplitudes. The latter are taken from the Jülich \({\bar{N}N}\) models and from a recently published new partial-wave analysis of \({\bar{p}p}\) scattering data. Predictions for differential cross sections and the spin observables \({A_y^d}\) , \({A_y^{\bar{p}}}\) , A _xx , A _yy are presented for antiproton beam energies up to about 300 MeV. The efficiency of the polarization buildup for antiprotons in a storage ring is investigated.     

Single-inclusive jet production in polarized pp collisions at RHIC to next-to-leading logarithmic accuracy

We perform the resummation of large logarithmic corrections to the partonic cross sections for single-inclusive jet production in polarized pp collisions. We reach the next-to-leading logarithmic accuracy for this observable with the corresponding matching to the next-to-leading order calculation performed in the small-cone approximation. We present numerical results for the BNL-RHIC collider at $\sqrt{S}=200$  GeV and at $\sqrt{S}=500$  GeV. We find an enhancement of the spin-dependent cross section, specially at high transverse momentum for the jet, resulting in a rather small increase of the double-spin asymmetry $A^{\mathrm{jet}}_{\mathrm{LL}}$ for this process.     

Efficient delivery of Cre-recombinase to neurons in vivo and stable transduction of neurons using adeno-associated and lentiviral vectors

Background

As development proceeds the human embryo attains an ever more complex three dimensional (3D) structure. Analyzing the gene expression patterns that underlie these changes and interpreting their significance depends on identifying the anatomical structures to which they map and following these patterns in developing 3D structures over time. The difficulty of this task greatly increases as more gene expression patterns are added, particularly in organs with complex 3D structures such as the brain. Optical Projection Tomography (OPT) is a new technology which has been developed for rapidly generating digital 3D models of intact specimens. We have assessed the resolution of unstained neuronal structures within a Carnegie Stage (CS)17 OPT model and tested its use as a framework onto which anatomical structures can be defined and gene expression data mapped.

Results

Resolution of the OPT models was assessed by comparison of digital sections with physical sections stained, either with haematoxylin and eosin (H&E) or by immunocytochemistry for GAP43 or PAX6, to identify specific anatomical features. Despite the 3D models being of unstained tissue, peripheral nervous system structures from the trigeminal ganglion (~300 μm by ~150 μm) to the rootlets of cranial nerve XII (~20 μm in diameter) were clearly identifiable, as were structures in the developing neural tube such as the zona limitans intrathalamica (core is ~30 μm thick). Fourteen anatomical domains have been identified and visualised within the CS17 model. Two 3D gene expression domains, known to be defined by Pax6 expression in the mouse, were clearly visible when PAX6 data from 2D sections were mapped to the CS17 model. The feasibility of applying the OPT technology to all stages from CS12 to CS23, which encompasses the major period of organogenesis for the human developing central nervous system, was successfully demonstrated.

Conclusion

In the CS17 model considerable detail is visible within the developing nervous system at a minimum resolution of ~20 μm and 3D anatomical and gene expression domains can be defined and visualised successfully. The OPT models and accompanying technologies for manipulating them provide a powerful approach to visualising and analysing gene expression and morphology during early human brain development.     

H ̄ + production from collisions between positronium and keV antiprotons for GBAR

In the framework of the GBAR (Gravitational Behaviour of Antihydrogen at Rest) experiment, cross sections for antihydrogen ion ( \(\bar {\mathrm {H}}^{+}\) ) production in collisions between antiprotons ( \(\bar {\mathrm {p}}\) ) and excited positronium atoms (Ps), with intermediate production of antihydrogen ( \(\bar {\mathrm {H}}\) ), have been computed using a perturbative theory, namely Continuum Distorted Wave - Final State (CDW-FS). The results suggest to use antiprotons at 1, 2 or 6 keV with, respectively, Ps(3p,3d), Ps(2p) or no Ps excitation. A simulation using these cross sections is under development to investigate the reaction chamber geometry and the parameters of the different beams (positrons, antiprotons and laser). This simulation, focusing on Ps(3d), predicts at least one \(\bar {\mathrm {H}}^{+}\) ion per pulse of 3·10⁶ \(\bar {\mathrm {p}}\) at 1 and 6 keV, and highlights both the interest of positronium excitation and the need for short pulses of particles.