Correlation between electrical direct current resistivity and plasmonic properties of CMOS compatible titanium nitride thin films.

Damping distances of surface plasmon polariton modes sustained by different thin titanium nitride (TiN) films are measured at the telecom wavelength of 1.55 µm. The damping distances are correlated to the electrical direct current resistivity of the films sustaining the surface plasmon modes. It is found that TiN/Air surface plasmon mode damping distances drop non-linearly from 40 to 16µm as the resistivity of the layers increases from 28 to 130µΩ.cm, respectively. The relevance of the direct current (dc) electrical resistivity for the characterization of TiN plasmonic properties is investigated in the framework of the Drude model, on the basis of parameters extracted from spectroscopic ellipsometry experiments. By probing a parametric space of realistic values for parameters of the Drude model, we obtain a nearly univocal dependence of the surface plasmon damping distance on the dc resistivity demonstrating the relevance of dc resistivity for the evaluation of the plasmonic performances of TiN at telecom frequencies. Finally, we show that better plasmonic performances are obtained for TiN films featuring a low content of oxygen. For low oxygen content and corresponding low resistivity, we attribute the increase of the surface plasmon damping distances to a lower confinement of the plasmon field into the metal and not to a decrease of the absorption of TiN.

[HISTOGENESIS AND PECULIARITIES OF STRUCTURAL ORGANIZATION OF THE CARDIAC MUSCLE TISSUE IN THE WALLS OF HUMAN CAVAL AND PULMONARY VEINS].

To study the structural organization and histogenesis of the cardiac muscle tissue in the walls of human caval and pulmonary veins, the heart was examined in 3 human embryos (at weeks 6-7 of development) and 20 fetuses (at weeks 9-10, 16, 19, 22 and 24 of development), as well as segments of caval and pulmonary veins of adult men and women (n = 50) located at various distances from the heart. The methods of light and electron microscopy were used in this work. To obtain the isolated cells from the walls of caval and pulmonary veins, the method of tissue alkaline dissociation was used. An immunohistochemical study with the monoclonal antibodies against cardiac troponin T was performed. It was found that the cardiomyocytes in humans were located in the middle and outer tunics of caval and pulmonary veins, where they formed thick layers. In the pulmonary veins of the adult humans, cardiac muscle fibers did not reach the intrapulmonary areas, in the inferior vena cava their layer did not extend beyond the pericardium, in the superior vena cava, its length was 2.5-3.0 cm. The formation of the pulmonary vein orifices occured by sequential inclusion of the wall of the common pulmonary vein, and later--of the right and left pulmonary veins into the wall of the left atrium. During the formation of the orifices of the caval veins, the gradual inclusion of the wall of the venous sinus in the wall of the right atrium was observed, resulting in caval veins opening directly into the cavity of the right atrium. The veins studied were referred to the veins of the muscular type with the strong development of muscular elements containing the myocardial component.

NGF and neurotrophin-3 both activate TrkA on sympathetic neurons but differentially regulate survival and neuritogenesis.

In this report we examine the biological and molecular basis of the control of sympathetic neuron differentiation and survival by NGF and neurotrophin-3 (NT-3). NT-3 is as efficient as NGF in mediating neuritogenesis and expression of growth-associated genes in NGF-dependent sympathetic neurons, but it is 20-40-fold less efficient in supporting their survival. Both NT-3 and NGF induce similar sustained, long-term activation of TrkA, while NGF is 10-fold more efficient than NT-3 in mediating acute, short-term TrkA activity. At similar acute levels of TrkA activation, NT-3 still mediates neuronal survival two- to threefold less well than NGF. However, a mutant NT-3 that activates TrkC, but not TrkA, is unable to support sympathetic neuron survival or neuritogenesis, indicating that NT-3-mediated TrkA activation is necessary for both of these responses. On the basis of these data, we suggest that NGF and NT-3 differentially regulate the TrkA receptor both with regard to activation time course and downstream targets, leading to selective regulation of neuritogenesis and survival. Such differential responsiveness to two ligands acting through the same Trk receptor has important implications for neurotrophin function throughout the nervous system.

Extracellular Ca2+ depletion contributes to fast activity-dependent modulation of synaptic transmission in the brain.

Synaptic activation is associated with rapid changes in intracellular Ca(2+), while the extracellular Ca(2+) level is generally assumed to be constant. Here, using a novel optical method to measure changes in extracellular Ca(2+) at high spatial and temporal resolution, we find that brief trains of synaptic transmission in hippocampal area CA1 induce transient depletion of extracellular Ca(2+). We show that this depletion, which depends on postsynaptic NMDA receptor activation, decreases the Ca(2+) available to enter individual presynaptic boutons of CA3 pyramidal cells. This in turn reduces the probability of consecutive synaptic releases at CA3-CA1 synapses and therefore contributes to short-term paired-pulse depression of minimal responses. This activity-dependent depletion of extracellular Ca(2+) represents a novel form of fast retrograde synaptic signaling that can modulate glutamatergic information transfer in the brain.

Activation of AMPA, kainate, and metabotropic receptors at hippocampal mossy fiber synapses: role of glutamate diffusion.

Glutamatergic transmission at mossy fiber (MF) synapses on CA3 pyramidal neurons in the hippocampus is mediated by AMPA, kainate, and NMDA receptors and undergoes presynaptic modulation by metabotropic glutamate receptors. The recruitment of different receptors has thus far been studied by altering presynaptic stimulation to modulate glutamate release and interfering pharmacologically with receptors and transporters. Here, we introduce two novel experimental manipulations that alter the fate of glutamate molecules following release. First, an enzymatic glutamate scavenger reduces the postsynaptic response as well as presynaptic modulation by metabotropic receptors. At physiological temperature, however, the scavenger is effective only when glutamate uptake is blocked, revealing a role of active transport in both synaptic and extrasynaptic communication. Second, AMPA and kainate receptor-mediated postsynaptic signals are enhanced when extracellular diffusion is retarded by adding dextran to the perfusion solution, as is feedback modulation by metabotropic receptors, suggesting that the receptors are not saturated under baseline conditions. These results show that manipulating the spatiotemporal profile of glutamate following exocytosis can alter the involvement of different receptors in synaptic transmission.

Hippocampal synapses: do they talk to their neighbours?

Recent experimental findings show that fast synaptic transmission can extend its actions beyond the immediate synaptic cleft. Whether this phenomenon results in significant crosstalk between typical neighbouring synapses remains unclear. This article considers two areas of the hippocampus, the CA1 and dentate gyrus, where important neural processing occurs. The results discussed do not provide a simple answer to the question of whether synapses can 'talk' to their neighbours, but they do reveal crucial physiological constraints that determine the significance of synaptic crosstalk, thus adding considerably to our understanding of chemical synaptic transmission.

[Effect of 1,2,3,4-tetrahydroimidazo[4,5-c]pyridine and 1,2,3,4-tetrahydroisoquinoline derivatives on the behavior and the brain receptor system in rats].

Adducts obtained via the interaction of formaldehyde with histidine (1,2,3,4-tetrahydroimidazo[4, 5-c]pyridine-3-carboxylic acid (I)), tyrosine (7-hydroxy-1,2,3,4-tetrahydroisoquinoline-3-carboxylic acid (II)), and dopamine (6,7-dihydroxy-1,2,3,4-tetrahydroisoquinoline (III)) influence the behavior and the state of the brain receptor system of rats upon chronic administration (10-day treatment at a daily dose of 50 mg/kg, i.p.). All the compounds studied decrease the horizontal and (to a lower extent) vertical motor activity and increase the quiescence period duration. On the other hand, the effects of compounds tested on the vegetative reactions were different: compounds I and III increased, whereas compound II decreased these reactions. Using the radioligand binding method, it was established that the treatment with compound I led to a decrease in the density of benzodiazepine receptors (B max of [3H]-flunitrazepam was 78% of the control level) and to a significant (148%) increase in the density of specific binding sites for [3H]-spiperone (reflecting the total density of dopamine (D2) and serotonin (5-HT2) receptors. The chronic administration of compound III produced a reliable decrease (76% of the control level) only in the density of benzodiazepine receptors. None of the tested compounds influenced the affinity of receptors with respect to the radioactive ligands used.

Hippocampal circuit dysfunction in the Tc1 mouse model of Down syndrome.

Hippocampal pathology is likely to contribute to cognitive disability in Down syndrome, yet the neural network basis of this pathology and its contributions to different facets of cognitive impairment remain unclear. Here we report dysfunctional connectivity between dentate gyrus and CA3 networks in the transchromosomic Tc1 mouse model of Down syndrome, demonstrating that ultrastructural abnormalities and impaired short-term plasticity at dentate gyrus-CA3 excitatory synapses culminate in impaired coding of new spatial information in CA3 and CA1 and disrupted behavior in vivo. These results highlight the vulnerability of dentate gyrus-CA3 networks to aberrant human chromosome 21 gene expression and delineate hippocampal circuit abnormalities likely to contribute to distinct cognitive phenotypes in Down syndrome.

Synapses in hippocampus occupy only 1-2% of cell membranes and are spaced less than half-micron apart: a quantitative ultrastructural analysis with discussion of physiological implications.

Relatively little information exists regarding the spatial structure of synaptic neuropil in the brain. The present electron microscopic study employs unbiased stereological techniques and Monte Carlo simulations to characterise quantitatively the spatial organisation of synaptic circuitry in the dentate gyrus of the hippocampus, an area of particular importance in mechanisms of learning and the subject of a number of experimental neurobiological models of synaptic plasticity such as long-term potentiation. Firstly, tissue shrinkage/expansion resulting from embedding was assessed by imaging 300-microm thick hippocampal slices in the course of the entire embedding protocol, giving a value of 94.3 +/- 1.1% for distance measures and 84.3 +/- 2.8% for volumetric measures. Secondly, numeric synaptic density, Nv, was estimated using the disector. Thirdly, accumulated area of post-synaptic densities (PSDs) per tissue volume, Sv, and the overall cell membrane area per tissue volume, Sv*, were assessed using unbiased stereological rules coupled with image analysis of single sections. Finally, the mean area of individual PSDs was derived as a ratio Sv/Nv giving: 0.0394 microm2 for axo-spinous PSDs (thus representing approximately 1.3% of total cell membranes) and 0.0769 microm2 for dendritic shaft PSDs (approximately 0.25% of total cell membranes). From these data, the mean nearest neighbour distance between synapses was estimated using Monte Carlo simulations of a random 3D arrangement of synapses constrained by PSD sizes (a truncated Poisson process), giving a value of 0.48-0.51 microm. The physiological importance of the morphometric data obtained is discussed in terms of assessing (i) the role of synaptic environment in modifying synaptic efficacy and (ii) the plausibility of cross talk between synapses in relation to extrasynaptic neurotransmitter diffusion and transient depletion of extracellular Ca2+.

Cytoskeleton-mediated, age-dependent lateral topography of lectin-gold-labelled molecules on the plasma membrane of cultured neurons: a statistical view.

In dissociated spinal cord neurons (12-day-old mouse embryo, monolayer culture), an electron microscopic study was carried out to examine quantitatively the rearrangement of wheat-germ agglutinin-gold-labelled molecules on the neuronal somatic surface at two developmental stages (on the fifth and 15th days in vitro), and after cytoskeletal interruptions. In tests, before labelling the cultures were incubated with colchicine or cytochalasin in order to affect microtubules or mostly actin filaments, respectively. Samples of electron micrographs that display soma membrane (profile) fragments were quantified. A set of stochastic geometry approaches was accomplished, which allowed statistical and stereological analysis of labelling. Images that illustrate the lateral (surface) patterns of label were simulated. On the fifth day in vitro, both colchicine and cytochalasin were found to cause an increase in the surface density and aggregation of wheat-germ agglutinin label relative to controls, the effect of cytochalasin being significantly more profound. By the 15th day in vitro, treatment with both drugs led to a similar tendency towards heavy aggregation of wheat-germ agglutinin labels. In contrast, neuron processes showed an opposite tendency of label rearrangement, which suggests lateral migration of labelled molecules, as a result of drug action. Possible molecular mechanisms involved in the phenomena are discussed.

Branching of active dendritic spines as a mechanism for controlling synaptic efficacy.

Recent experimental findings (Yuste R. and Denk W. (1995) Nature 375, 682-684) suggest that dendritic spines possess excitable membranes. Theoretically, it was shown earlier that the shape of active spines can significantly affect somatopetal synaptic signal transfer. Studies of long-term potentiation in the hippocampus have related the increased synaptic efficacy to a number of structural modifications of spines, including an increased number of branched spines [Trommald M. et al. (1990) In Neurotoxicity of Excitatory Amino Acids, pp. 163-174. Raven Press, New York] and a strengthened capability for spines to alter their spatial positions [Hosokawa T. et al. (1995) J. Neurosci. 15, 5560-5573]. In the present simulation study, the potential physiological impact of several types of spine changes was examined in a compartmental neuron model built using the neuromodelling software NEURON [Hines M. (1993) In Neural Systems: Analysis and Modeling, pp. 127-136. Kluwer Academic, Norwell, MA]. The model included 30 complex spines, with dual component synaptic currents and mechanisms of Ca2+ uptake, diffusion, binding and extrusion within spine heads. The results show that local clustering properties of spine distributions along dendrites are unlikely to affect synaptic efficacy. However, partial fusion of active spines, which results in formation of spine branches, or subtle changes in spine branch positions, could alone significantly increase synaptic signal transfer. These data illustrate possible mechanisms whereby subtle morphological changes in dendritic spines (compatible with changes reported in the literature) may be linked to the cellular mechanisms of learning and memory.

Lateral patterns of the neural cell adhesion molecule on the surface of hippocampal cells developing in vitro.

In monolayer cultures of hippocampal neurons from newborn rats, an immunocytochemical quantitative study was carried out to investigate age-dependent arrangement of the neural cell adhesion molecules in different parts of cell membranes. On the fifth and 12th day in vitro, neural cell adhesion molecules were labelled with specific antibodies and protein A conjugated to colloidal gold particles. Samples of randomly selected electron micrographs that displayed labelled membrane fragments of cell bodies, growth cones, and axons were numerically analysed for the five- and 12-day in vitro neurons. Neural cell adhesion molecules surface topography was quantitatively described and compared, using a statistical stereological approach. The mean surface density of labelled neural cell adhesion molecules was found to be approximately 2.5 times higher in growth cone membranes relative to somatic and axonal membranes in five-day in vitro neurons. By the 12th day in vitro, this density decreases in somatic membranes (approximately 18%) and increases in axonal membranes (approximately 60%). Representative spectra of lateral intervals between labels as well as images that show typical topography of label on membrane surfaces were simulated. The results revealed regular patterns of neural cell adhesion molecules on the somatic surface and allowed consideration of neural cell adhesion molecules arrangement in a view of membrane adhesion properties. Participation of cytoskeleton in neural cell adhesion molecules rearrangement is discussed.

Re-structuring of synapses 24 hours after induction of long-term potentiation in the dentate gyrus of the rat hippocampus in vivo.

In male rats, long-term potentiation was induced unilaterally in the dentate gyrus, either by high frequency (200Hz) or theta rhythm stimulation. Structural synaptic changes were examined 24h after induction using quantitative electron microscopy. A disector technique was employed in order to estimate the density of synapses (using 70-80-nm sections) and of granule cell nuclei (using 2-microm sections) in the middle, and inner molecular layer in both hemispheres. Synaptic height and total lateral areas of synaptic active zones per unit tissue volume were assessed via assumption-free stereological techniques coupled with image analysis. The results obtained indicated that both synaptic density and number (corrected per neuron) of axo-spinous, but not axo-dendritic, synapses were approximately 40% higher in the middle, but not inner molecular layer of the potentiated hemisphere compared to the contralateral (control hemisphere). No significant inter-hemispheric difference was found in the volume densities of lateral areas of active zones. These data suggest that 24h after long-term potentiation induction, active zones of existing axo-spinous synapses either split forming separate contacts, or decrease in size while new synapses are formed.

Increased immunogold labelling of neural cell adhesion molecule isoforms in synaptic active zones of the chick striatum 5-6 hours after one-trial passive avoidance training.

An area of the chick striatum, the lobus parolfactorius plays an important role in one-trial passive avoidance learning tasks. In the present study we report evidence that 5-6 h post-training, a significantly higher proportion of synaptic active zones in this area contain labelled epitopes of the neural cell adhesion molecule, with the greatest occurrence of labels at the edges of active zone profiles (in both control and trained groups). This suggests that there is a period after training when expression of the neural cell adhesion molecule in synaptic membranes almost doubles, and that events at active zone edges may play a specific role in mechanisms of synaptic adhesion. Cellular mechanisms of long-term memory formation are believed to include alterations in neural circuitry at the synaptic level. The involvement of the neural cell adhesion molecule (NCAM) in functional synaptic modifications has been demonstrated using a number of physiological models. Performance of rats in the Morris water maze, a spatial learning paradigm which requires the hippocampus, is impaired by either intraventricular injection of NCAM antibodies, or injection into the hippocampus of an enzyme which increases homophilic adhesion of the molecule, due to the removal of polysialic acid residuals from extracellular NCAM domains. In addition, intraventricular injections of anti-NCAM antibodies 6-8 h post-training were shown to impair memory for a one-trial passive avoidance task in the rat. An avoidance training model in the one-day-old chick indicates a similar time window, 5-6 h post-training during which memory for the task can be impaired by intraventricular injection of NCAM antibodies. In the hyperstriatum ventrale, a chick forebrain area involved in the passive avoidance task. subtle changes in the distribution pattern, but not density of NCAM molecules in synaptic membranes were revealed 5-6 h post-training. However, on the basis of studies of synaptic morphometry, a region of striatum, the lobus parolfactorius (LPO), appears to play a more important role in longer term memory storage for the task.

Ultrastructural synaptic correlates of spatial learning in rat hippocampus.

Memory formation is believed to alter neural circuitry at the synaptic level. Although the hippocampus is known to play an important role in spatial learning, no experimental data exist on the synaptic correlates of this process at the ultrastructural level. Here, we have employed quantitative electron microscopy in order to compare the density, size and spatial arrangement of synapses in the dentate gyrus, and in area CA1, of spatially trained (water maze, invisible platform) versus control (visible platform) rats. No training-associated changes of hippocampal volume were found using a stereological estimaion (disector) of the volume density of dentate granule, or CA1 pyramidal cells. Nor were changes found in either density, or sizes of synapses (spinous or dendritic), in CA1 or dentate gyrus. However, analysis of synaptic spatial distribution showed a training-associated increase in the frequency of shorter distances (i.e. clustering) between synaptic active zones in CA1, but not dentate, thus indicating alterations in local neural circuitry. This finding indicates subtle changes in synaptic organization in area CA1 of the hippocampus following a learning experience, suggesting that spatial memory formation in mammalian hippocampus may involve topographical changes in local circuitry without synapse formation de novo.

Interactions between brain mitochondria and cytoskeleton: evidence for specialized outer membrane domains involved in the association of cytoskeleton-associated proteins to mitochondria in situ and in vitro.

The surface distribution of several proteins (porin, hexokinase, and two proteins associated with microtubules or actin filaments) on the outer membrane of brain mitochondria was analyzed by immunogold labelling of purified mitochondria in vitro. The results suggest the existence of specialized domains for the distribution of porin in the outer mitochondrial membrane. Similarities between the distribution of porin and the distribution of microtubule-associated proteins bound in vitro to mitochondria suggested that mitochondria and microtubules interact by binding microtubule-associated proteins to porin-containing domains of the outer membrane. This hypothesis was supported by biochemical studies on outer mitochondrial proteins involved in in vitro binding of cytoskeleton elements. In vitro interactions between mitochondria and microtubules or neurofilaments were analyzed by electron microscopy. These studies revealed cross-bridging between the outer membrane of mitochondria and the two cytoskeleton elements. Cross-bridging was influenced by ATP hydrolysis and by several proteins associated with the surface of mitochondria or with microtubules. In addition, unidentified proteins which were recognized by antibodies to all intermediate filaments subunits were associated either with the mitochondrial surface or with microtubules. This data suggest the participation of additional cytoplasmic proteins in the interactions between cytoskeleton elements and mitochondria.

Training in chicks alters PSA-N-CAM distribution in forebrain cell membranes.

The intermediate and medial hyperstriatum ventrale (IMHV) in the chick is involved in memory formation following one-trial passive avoidance training. Possible links between neural cell adhesion molecule (N-CAM) distribution and memory consolidation were examined in an immunoelectron microscope study of IMHV 6 h after training. An antibody against alpha 2,8 polysialic acid (PSA), characteristic for embryonic N-CAM, and one against the protein backbones of N-CAM were labelled (post-embedding), separately, with 15 nm immunogold, and binding to their epitopes was analysed using the statistics of point processes. No difference in labelling levels between control and trained chicks, for either antibodies, was found, both groups of birds showing that cell membrane regions are 10-30 times more enriched in N-CAM isoforms, and have a two-fold greater proportion of PSA-N-CAM, than tissue as a whole. However, 375-400 nm regular arrays of labelled PSA-N-CAM were revealed statistically in cell membranes of control, but not trained, chicks, which may be related to the possible involvement of such membrane PSA domains in memory-related neuroplasticity.

Dendritic spines form 'collars' in hippocampal granule cells.

A quantitative study of the distribution of dendritic spines was carried out in three orders of dendritic branches of granule cells from the dentate gyrus of the rat hippocampus. Golgi-stained preparations (7-19 neurones in each of seven rats) were analysed using computerized microscopy. Identification of spines and quantification of stem-spine geometry was performed using a segmentation algorithm and a line skeleton transformation of dendritic images. Analysis of data using the statistics of point processes revealed that, in all three branch orders, the distribution of visible spines along dendrites was not evenly random, but included dense clusters of spines surrounding the dendritic stem (spine 'collars'). Three-dimensional reconstructions from serial ultrathin sections have confirmed the presence of such spine groups. We speculate the spine collars represent a functional element in which associative synaptic plasticity is fostered by the proximity of individual synapses.

Quantification of dendritic spine populations using image analysis and a tilting disector.

A series of image analysis routines, stochastic geometry methodology, and a design-based stereological procedure have been developed to quantify objectively the length, layout, and the true density of neuronal dendritic spines observed at the light (or confocal) microscope level. First, the image of a dendritic fragment of interest (in the plane of view) is scaled to a standard brightness scale, and the dendritic profile is separated from the background using a computerized thresholding algorithm that analyzes the histogram of grey levels. Secondly, the resulting binary image of the dendrite is transformed to a midline skeleton that underlies the dendritic geometry. Thirdly, skeletal branch lengths are directly computed (in pixels), thus giving objective measures of visible spine lengths and inter-spine distances along the dendritic stem. These raw data are the basis for (1) an estimation of the distribution of 3D spine lengths, and (2) a nearest neighbour analysis of the spine layout along the dendrite. A design-based stereological routine, the tilting disector, is suggested for unbiased estimation of the true (3D) density of spines along dendrites. The routine involves tilting the dendritic fragment of interest around its longitudinal axis for a known angular sector and scoring the number of spines seen in one angular position and unseen in the other position. Data from a study of neuronal dendrites in the chick forebrain are presented.

3-dimensional morphometry of intact dendritic spines observed in thick sections using an electron microscope.

An experimental technique is described which allows observation of fixed neuronal dendrites at magnifications from 10-12 K. The method uses 4-7-microns-thick sections of Epon-embedded tissue with nerve cells that are first impregnated by the rapid Golgi technique and then stained with gold particles/aggregates using a modified gold-toning procedure. A relatively high acceleration voltage (200 kV) is employed to observe in fine detail the dendritic fragments of interest at different angular positions in space, by using a eucentric goniometer stage with a tilt angle of +/- 45 degrees. Image analysis methodology is proposed which permits estimation of 3-dimensional (3D) lengths and of the volume of observed intact dendritic spines. The advantages of the technique with respect to 3D reconstruction methodology are discussed.