Fourier light scattering model for treating textures deeper than the wavelength.

We apply a Fourier-scattering model to describe light scattering in solar cells with textured surfaces. For the size and inclination angle of typical micro-textures, scattering may occur into large angles. This makes the model prone to paraxial errors. We present a non-paraxial formulation and discuss the transition from the domain of refraction at large facets to scattering at small features.

Search for solar axions by the CERN axion solar telescope with 3He buffer gas: closing the hot dark matter gap.

The CERN Axion Solar Telescope has finished its search for solar axions with (3)He buffer gas, covering the search range 0.64 eV ≲ ma ≲ 1.17 eV. This closes the gap to the cosmological hot dark matter limit and actually overlaps with it. From the absence of excess x rays when the magnet was pointing to the Sun we set a typical upper limit on the axion-photon coupling of gaγ ≲ 3.3 × 10(-10) GeV(-1) at 95% C.L., with the exact value depending on the pressure setting. Future direct solar axion searches will focus on increasing the sensitivity to smaller values of gaγ, for example by the currently discussed next generation helioscope International AXion Observatory.

Search for sub-eV mass solar axions by the CERN Axion Solar Telescope with 3He buffer gas.

The CERN Axion Solar Telescope (CAST) has extended its search for solar axions by using (3)He as a buffer gas. At T=1.8 K this allows for larger pressure settings and hence sensitivity to higher axion masses than our previous measurements with (4)He. With about 1 h of data taking at each of 252 different pressure settings we have scanned the axion mass range 0.39 eV≲m(a)≲0.64 eV. From the absence of excess x rays when the magnet was pointing to the Sun we set a typical upper limit on the axion-photon coupling of g(aγ)≲2.3×10(-10) GeV(-1) at 95% C.L., the exact value depending on the pressure setting. Kim-Shifman-Vainshtein-Zakharov axions are excluded at the upper end of our mass range, the first time ever for any solar axion search. In the future we will extend our search to m(a)≲1.15 eV, comfortably overlapping with cosmological hot dark matter bounds.

Comparison and optimization of randomly textured surfaces in thin-film solar cells.

Using rigorous diffraction theory we investigate the scattering properties of various random textures currently used for photon management in thin-film solar cells. We relate the haze and the angularly resolved scattering function of these cells to the enhancement of light absorption. A simple criterion is derived that provides an explanation why certain textures operate more beneficially than others. Using this criterion we propose a generic surface profile that outperforms the available substrates. This work facilitates the understanding of the effect of randomly textured surfaces and provides guidelines towards their optimization.

Neuronal enriched endosomal protein of 21 kDa colocalizes with glutamate receptor subunit GLUR2/3 at the postsynaptic membrane.

Functional evidence suggests that neuronal enriched endosomal protein of 21 kDa (NEEP21) takes part in facilitating transport of AMPA receptors (AMPAR) in the synapse. To explore the anatomical basis for a role in this synaptic trafficking, we investigated the ultrastructural localization of NEEP21 in rodent brain. Using immunogold electron microscopy, we show that NEEP21 is colocalized with the AMPAR subunits GluR2/3 in postsynaptic spines. Quantitative analysis of gold particle distribution along an axis perpendicular to the postsynaptic specialization indicated that NEEP21 occurs in the postsynaptic membrane but also in the interior of the spines. NEEP21 positive endosomes/multivesicular bodies were found throughout cell bodies and dendrites. In light microscopical preparations, the NEEP21 antibody produced a labeling pattern in the neocortex, hippocampus and cerebellum that mimicked that of GluR2/3 and not that of GluR1 or 4. Our findings are consistent with a role for NEEP21 in facilitating vesicular transport of GluR2 between intracellular compartments and the postsynaptic plasma membrane.

Protein interacting with C kinase 1 (PICK1) and GluR2 are associated with presynaptic plasma membrane and vesicles in hippocampal excitatory synapses.

AMPA receptors have been identified in different populations of presynaptic terminals and found to be involved in the modulation of neurotransmitter release. The mechanisms that govern the expression of presynaptic AMPA receptors are not known. One possibility is that pre- and postsynaptic AMPA receptors are regulated according to the same principles. To address this hypothesis we investigated whether protein interacting with C kinase 1 (PICK1), known to interact with AMPA receptors postsynaptically, also is expressed presynaptically, together with AMPA receptors. Subfractionation and high-resolution immunogold analyses of the rat hippocampus revealed that GluR2 and PICK1 are enriched postsynaptically, but also in presynaptic membrane compartments, including the active zone and vesicular membranes. PICK1 and GluR2 are associated with the same vesicles, which are immunopositive also for synaptophysin and vesicle-associated membrane protein 2. Based on what is known about the function of PICK1 postsynaptically, the present data suggest that PICK1 is involved in the regulation of presynaptic AMPA receptor trafficking and in determining the size of the AMPA receptor pool that modulates presynaptic glutamate release.

Presynaptic PICK1 facilitates trafficking of AMPA-receptors between active zone and synaptic vesicle pool.

Previous studies have indicated that presynaptic α-amino-3-hydroxy-5-methyl-4-isoxazole propionate receptors (AMPARs) contribute to the regulation of neurotransmitter release. In hippocampal synapses, the presynaptic surface expression of several AMPAR subunits, including GluA2, is regulated in a ligand-dependent manner. However, the molecular mechanisms underlying the presynaptic trafficking of AMPARs are still unknown. Here, using bright-field immunocytochemistry, western blots, and quantitative immunogold electron microscopy of the hippocampal CA1 area from intact adult rat brain, we demonstrate the association of AMPA receptors with the presynaptic active zone and with small presynaptic vesicles, in Schaffer collateral synapses in CA1 of the hippocampus. Furthermore, we show that GluA2 and protein interacting with C kinase 1 (PICK1) are colocalized at presynaptic vesicles. Similar to postsynaptic mechanisms, overexpression of either PICK1 or pep2m, which inhibit the N-ethylmaleimide sensitive fusion protein (NSF)-GluA2 interaction, decreases the concentration of GluA2 in the presynaptic active zone membrane. These data suggest that the interacting proteins PICK1 and NSF act as regulators of presynaptic GluA2-containing AMPAR trafficking between the active zone and a vesicle pool that may provide the basis of presynaptic components of synaptic plasticity.

Differential effect of alpha-syntrophin knockout on aquaporin-4 and Kir4.1 expression in retinal macroglial cells in mice.

Aquaporin-4 water channels and the inwardly rectifying potassium channels Kir4.1 are coexpressed in a highly polarized manner at the perivascular and subvitreal endfeet of retinal Müller cells and astrocytes. The present study was aimed at resolving the anchoring mechanisms responsible for the coexpression of these molecules. Both aquaporin-4 and Kir4.1 contain PDZ-domain binding motifs at their C-termini and it was recently shown that mice with targeted disruption of the dystrophin gene display altered distribution of aquaporin-4 and Kir4.1 in the retina. To test our hypothesis that alpha-syntrophin (a PDZ-domain containing protein of the dystrophin associated protein complex) is involved in aquaporin-4 and Kir4.1 anchoring in retinal cells, we studied the expression pattern of these molecules in alpha-syntrophin null mice. Judged by quantitative immunogold cytochemistry, deletion of the alpha-syntrophin gene causes a partial loss (by 70%) of aquaporin-4 labeling at astrocyte and Müller cell endfeet but no decrease in Kir4.1 labeling at these sites. These findings suggest that alpha-syntrophin is not involved in the anchoring of Kir4.1 and only partly responsible for the anchoring of aquaporin-4 in retinal endfeet membranes. Furthermore we show that wild type and alpha-syntrophin null mice exhibit strong beta1 syntrophin labeling at perivascular and subvitreal Müller cell endfeet, raising the possibility that beta1 syntrophin might be involved in the anchoring of Kir4.1 and the alpha-syntrophin independent pool of aquaporin-4.

Neuroprotective effects of inhibiting N-methyl-D-aspartate receptors, P2X receptors and the mitogen-activated protein kinase cascade: a quantitative analysis in organotypical hippocampal slice cultures subjected to oxygen and glucose deprivation.

Cell death was assessed by quantitative analysis of propidium iodide uptake in rat hippocampal slice cultures transiently exposed to oxygen and glucose deprivation, an in vitro model of brain ischemia. The hippocampal subfields CA1 and CA3, and fascia dentata were analyzed at different stages from 0 to 48 h after the insult. Cell death appeared at 3 h and increased steeply toward 12 h. Only a slight additional increase in propidium iodide uptake was seen at later intervals. The mitogen-activated protein kinases extracellular signal-regulated kinase 1 and extracellular signal-regulated kinase 2 were activated immediately after oxygen and glucose deprivation both in CA1 and in CA3/fascia dentata. Inhibition of the specific mitogen-activated protein kinase activator mitogen-activated protein kinase kinase by PD98059 or U0126 offered partial protection against oxygen and glucose deprivation-induced cell damage. The non-selective P2X receptor antagonist suramin gave neuroprotection of the same magnitude as the N-methyl-D-aspartate channel blocker MK-801 (approximately 70%). Neuroprotection was also observed with the P2 receptor blocker PPADS. Immunogold data indicated that hippocampal slice cultures (like intact hippocampi) express several isoforms of P2X receptors at the synaptic level, consistent with the idea that the effects of suramin and PPADS are mediated by P2X receptors. Virtually complete neuroprotection was obtained by combined blockade of N-methyl-D-aspartate receptors, P2X receptors, and mitogen-activated protein kinase kinase. Both P2X receptors and N-methyl-D-aspartate receptors mediate influx of calcium. Our results suggest that inhibition of P2X receptors has a neuroprotective potential similar to that of inhibition of N-methyl-D-aspartate receptors. In contrast, our comparative analysis shows that only partial protection can be achieved by inhibiting the extracellular signal-regulated kinase 1/2 mitogen-activated protein kinase cascade, one of the downstream pathways activated by intracellular calcium overload.

Laminar differentiation of the hippocampus, fascia dentata and subiculum in developing rats, observed with the Timm sulphide silver method.

The laminar staining of the rat hippocampal region with the Timm sulphide silver method is from studies on adult rats known to depend on the various fibersystems terminating in these laminae. In order to illustrate the development of these fibersystems the laminar differentiation of the Timm staining of fascia dentata, hippocampus and subiculum is presented for rats between 1 and 31 days old. Corresponding changes in cytoarchitectonics as revealed by thionin staining are briefly demonstrated. Even on the first postnatal day there are indications of the mature, laminar staining pattern, and between three and nine days all the laminae corresponding to the terminal fields of the major afferent and intrinsic systems appear. After 12 days only minor additions to the laminar pattern develop, but there are adjustments of absolute and relative dimensions of layers and fields, and also the staining densities of individual laminae change. These observations are in good correlation with the available information on both hippocampal neurogenesis and cytodifferentiation, and the few fiber tracing studies performed on the developing hippocampal region. Compared to the latter, which ideally marks only one system or one lamina per animal, the Timm method provides an excellent means for getting an overview of the general developmental situation at the different ages. Thus developmental gradients along septotemporal, medio-lateral and basal-apical axes are found; which should be heeded in future studies on hippocampal synaptogenesis. The observations are discussed in relation to current models for neuronal growth and formation of nervous connections.

BK channel activity determines the extent of cell degeneration after oxygen and glucose deprivation: a study in organotypical hippocampal slice cultures.

BK channels are voltage- and calcium-dependent potassium channels whose activation tends to reduce cellular excitability. In hippocampal pyramidal cells, BK channels repolarize somatic action potentials, and recent immunogold and electrophysiological analyses have revealed a presynaptic pool of BK channels that can regulate glutamate release. Agents that modulate BK channel activity would therefore be expected to affect cell excitability and neurotransmitter release also under pathological conditions. We have investigated the role of BK potassium channels in a model of ischemia-induced nerve cell degeneration. Organotypical slice cultures of rat hippocampus were exposed to oxygen and glucose deprivation (OGD), and cell death was assessed by the fluorescent dye propidium iodide. OGD induced cell death in the CA1 region and to a lesser extent in CA3. Treatment with the BK channel blockers, paxilline and iberiotoxin, during and after OGD induced increased cell death in CA1 and CA3. Both BK channel blockers also sensitized the relatively resistant granule cells in fascia dentata to OGD. The effect of paxilline and iberiotoxin was evident from 3 h after OGD, indicating a role of BK channels early in the post-ischemic phase or during OGD itself. The BK channel opener, NS1619, turned out to be gliotoxic, and this effect was not counteracted by paxilline and iberiotoxin. Our data show that blockade of BK channels aggravates OGD-induced cell damage and suggest that BK channels act as a kind of 'emergency brake' during and/or after ischemia. Accordingly, the BK channel is a potential molecular target for neuroprotective therapy in stroke.

Neuroactive amino acids in organotypic slice cultures of the rat hippocampus: an immunocytochemical study of the distribution of GABA, glutamate, glutamine and taurine.

Antisera raised against protein-glutaraldehyde-amino acid conjugates were used to study the light and electron microscopic distribution of GABA, glutamate, glutamine and taurine in organotypic slice cultures of rat hippocampi. In the stratum oriens and radiatum, glutamate-like immunoreactivity was particularly concentrated in nerve endings establishing asymmetric junctions with dendritic spines. Mossy fiber terminals in CA3 and the dentate hilus were also strongly labeled. A quantitative immunogold analysis of the glutamate-immunolabelled profiles showed a pattern that was highly reminiscent of that previously observed in perfusion-fixed hippocampi, including a correspondingly sparse labeling of glial processes and of presynaptic elements in symmetric synapses. GABA-like immunoreactivity was localized predominantly in interneurons and in presynaptic terminals contacting dendritic shafts and neuronal cell bodies, while immunoreactivities for glutamine and taurine were found mainly in astroglial cells and pyramidal cells, respectively. Our data indicate that the major intrinsic fiber systems of the cultured hippocampi have retained their normal transmitter phenotypes.

Ischemic disruption of glutamate homeostasis in brain: quantitative immunocytochemical analyses.

More than 10 years ago, it was shown by microdialysis that the excitatory transmitter glutamate accumulates in the interstitial space of brain subjected to ischemic insult. This was one of the key observations leading to the formulation of the "glutamate hypothesis' of ischemic cell death. It is now assumed that even a transient glutamate overflow may set in motion a number of events that ultimately cause cell loss in vulnerable neuronal populations. The aim of the present review is to discuss the intracellular changes that underlie the dysregulation of extracellular glutamate during and after ischemia, with emphasis on data obtained by postembedding, electron microscopic immunogold cytochemistry. While the time resolution of this approach is necessarily limited, it can reveal, quantitatively and at a high level of spatial resolution, how the intracellular pools of glutamate and metabolically related amino acids are perturbed during and after an ischemic insult. Moreover, this can be done in animals whose extracellular amino acid levels are monitored by microdialysis, allowing a direct correlation of extra- and intracellular changes. Immunogold analyses of brains subjected to ischemia have identified dendrites and neuronal somata as likely sources of glutamate efflux, probably mediated by reversal of glutamate uptake. The vesicular glutamate pool has been found to be largely unchanged after 20 min of ischemia. Ischemia causes an increased glutamate content and an increased glutamate/glutamine ratio in glial cells, as revealed by double immunogold labelling. This argues against the idea that glial cells contribute to the extracellular overflow of glutamate in the ischemic brain.

A simple in vitro model of ischemia based on hippocampal slice cultures and propidium iodide fluorescence.

This protocol describes a model of cerebral ischemia based on organotypic hippocampal slice cultures and quantitative assessment of cell death by use of propidium iodide and image analysis. The cultures were made from rat hippocampal slices that were obtained at postnatal day 4-7 and allowed to develop for >14 days in vitro. For induction of 'in vitro ischemia', the cultures were washed in glucose free buffer and the culture chamber flooded with a nitrogen/carbon dioxide mixture until the oxygen concentration was <1.0%. The cultures were exposed to this atmosphere for 30-35 min, washed in serum-free medium, and returned to ordinary growth medium. After 24 h, dead cells were quantified by use of propidium iodide. The cell death resulting from the oxygen/glucose deprivation was largely confined to the CA1 region and was blocked by NMDA-receptor antagonists but not by antagonists to AMPA-receptors or metabotropic glutamate receptors. The type of cell death was judged to be necrotic, based on ultrastructural observations. The oxygen/glucose deprived cultures exhibited increased phosphorylation of the MAP kinase cascade. This activation of the MAP kinase cascade was blocked by NMDA-receptor antagonists. The in vitro model described in the present report is simple to use and reproduces many features of in vivo ischemia, including the preferential vulnerability of CA1 cells. The model should be suited to analyses of the mechanisms underlying the regionally selective cell death in the hippocampus and ischemic cell death in general.

Subcellular compartmentation of glutathione and glutathione precursors. A high resolution immunogold analysis of the outer retina of guinea pig.

Selective antibodies were used to assess the cellular and subcellular localization of glutathione, and the glutathione precursors gamma-glutamylcysteine, glutamate, and cysteine, in neuronal (photoreceptors) and non-neuronal (pigment epithelial cells and Müller cells) cell types in the outer retina of the guinea pig. In each cell type the highest level of glutathione immunoreactivity occurred in the mitochondria. The labeling density in the cytoplasmic matrix was higher (and the mitochondrial-cytoplasmic gold particle ratio lower) in pigment epithelial cells than in Müller cells and photoreceptors. The latter two cell types showed a mitochondrial-cytoplasmic gold particle ratio of 15.5 and 21.7, respectively. In contrast to glutathione, gamma-glutamylcysteine seemed to be enriched in the cytoplasmic matrix relative to the mitochondria. The immunogold labeling for this dipeptide was stronger in the pigment epithelial cells than in Müller cells and photoreceptors. Glutamate immunoreactivity was high in photoreceptors, intermediate in pigment epithelial cells, and low in Müller cells, while the cysteine immunogold signal was low in each cell type and cell compartment. The present results suggest that glutathione is concentrated in mitochondria but to different degrees in different cells. The low mitochondrial content of gamma-glutamylcysteine (the direct precursor of glutathione) is consistent with biochemical data indicating that glutathione is synthesized extramitochondrially and transported into the mitochondrial matrix. Judged from the immunocytochemical data, cysteine may be a rate-limiting factor in glutathione synthesis in each cell type while glutamate can be rate limiting only in Müller cells.

Topochemistry of trace metals in nasal mucosa. Potentialities of some histochemical methods and energy dispersive X-ray microanalysis.

Histochemical methods and energy dispersive X-ray micro-analysis (EDX-analysis) were evaluated in model experiments and on tissue sections for their usefulness in detecting traces of metals in biological tissue. The goal for this study was to establish a method for localization of nickel deposits in the nasal mucosa, where it has been found in concentrations between 1 and 40 microgram/g in nickel exposed individuals. The histochemical methods tested were staining with dimethylglyoxime, rubeanic acid and dithizone, the Turnbull and Prussian blue methods and TIMM'S sulphide silver procedure. In model experiments nickel-, cobalt-, copper-, zinc- and ironsalts were applied to thin-layer chromatography sheets (TLC-sheets) and stained by the histochemical methods. Spots containing 500 and 50 ng of these metals represented the smallest amounts that could consistently be detected in these experiments, except for the sulphide silver method which seemed a little more sensitive. With the latter method, moreover, zinc was detected in 40 micrometer thick cryostat sections of gelatine made up with 1 microgram/g of the metal. For nickel the corresponding figure was 10 to 50 microgram/g. On specimens of nasal mucosa from nickel-exposed workers, a faint colour was obtained in 40 micron thick cryostat sections from specimens that had been immersed in dithizone, but the colour was too weak for histological analysis. None of the other coloured chelating agents caused noticeable staining when applied to blocks or to cryostat sections. TIMM'S sulphide silver method caused strong staining of the basal layers of the surface epithelium and of fibroblast-like cells in the underlying connective tissue. This staining pattern is described in more detail in a separate report. Rat liver tissue was analyzed by atomic absorption before and after araldite embedding. Blocks of gelatine made up with nickel, copper, zinc and iron were embedded in epoxy resin and analyzed by atomic absorption. Large changes in the metal concentrations, usually an increase, were found after embedding. Ultrathin sections from this material were used to test the sensitivity of the EDX-equipment. Referring to the concentrations determined by atomic absorption in the embedded material, iron was detected at 1215 microgram/g and 362 microgram/g (gelatine standards) but not at 167 microgram/g (rat liver). Similar values could not be determined for nickel, copper or zinc, because of background radiation resulting from the presence of these metals in the instrument. We did not succeed in establishing a procedure for detecting nickel deposits in nasal mucosa with any of the methods which were tested. The most sensitive but least specific of the tested methods for visualizing heavy metals in the nasal mucosa, was TIMM'S sulphide silver procedure. The preparation of tissue for this method is discussed.

Concentration and distribution of heavy metals in nasal mucosa of nickel-exposed workers and of controls, studied with atomic absorption spectrophotometric analysis and with Timm's sulphide silver method.

Biopsy specimens from nasal mucosa of 30 nickel-exposed individuals and 6 controls were analysed by atomic absorption spectrophotometry to determine the content of nickel, copper, cobalt, zinc and iron. Timm's sulphide silver staining method was used for visualizing heavy metals in cryostat sections of biopsy material from each individual. The purpose of the investigation was to study the sulphide silver staining pattern in nasal mucosa of nickel-exposed individuals and controls, and to establish whether variations in heavy metal concentrations, especially nickel, affect the histochemical pattern in the mucosa. No consistent differences were found in the histochemical pattern between biopsies with high and low concentrations of nickel or any of the other metals. Some difference in epithelial types between specimens from the nickel-exposed group and the control group was seen, but the staining pattern was quite similar for corresponding epithelial types in the two groups. Two nasal carcinomas from nickel workers were virtually unstained with the sulphide silver staining method.