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PubMed1999

Detection of glutathione in the human brain in vivo by means of double quantum coherence filtering.

The feasibility of selective in vivo detection of glutathione (L-gamma-glutamyl-L-cysteinyl-glycine, GSH) in the human brain by means of (1)H magnetic resonance spectroscopy (MRS) at 1.5 T is demonstrated. A double quantum coherence (DQC) filtering sequence was used in combination with PRESS volume selection. The strongly coupled cysteinyl CH(2) compound of GSH was found to be the most suitable target for spectral editing. Analytical calculations employing a product operator description of the cysteinyl ABX three-spin system were made in order to optimize the inherent yield of the sequence. A pulse phase calibration procedure, which precedes the spectrum acquisition, secures maximal signal yield independently of the spatial localization of the volume of interest and thus comparability between individual examinations. In vitro tests show that the DQC filtering method provides good discrimination between the GSH signal at 2.9 ppm and the interfering resonances of creatine, gamma-aminobutyric acid (GABA) and aspartate. In measurements in the frontal lobe of 12 healthy volunteers a mean ratio of GSH signal to tissue water signal of 5.7 +/- 2.3 x 10(-5) was found, corresponding to a mean GSH tissue concentration of 2-5 mmol/L. The proposed technique allows for the detection of a biologically highly relevant metabolite at moderate field strength. Magn Reson Med 42:283-289, 1999.

Brain↗
PubMed2000

2D-spatial/2D-spectral spectroscopic imaging of intracerebral gliomas in rat brain.

1H-MR spectroscopy in vivo is often hampered by poor spectral resolution. Spectral overlap can be avoided with two-dimensional spectroscopic techniques. Correlation peak imaging has been implemented to measure unambiguously the distribution of several metabolites in a rat brain glioma model. Acquisition-weighted spectroscopic imaging reduced the experimental time and provided excellent spatial localization. The choice of an appropriate spectral acquisition window granted good sensitivity. Spectroscopic images presenting a full two-dimensional spectrum in every image pixel were acquired in seven rats at 7 Tesla in 195 min, with a nominal voxel volume of 75 microl. Among other metabolites, the distribution of hypotaurine, phosphoethanolamine, alanine, and even glucose could be visualized both in the C6-glioma and in the unaffected brain.

Animals↗
PubMed2000

Scanning electrochemical microscopy of iontophoretic transport in hairless mouse skin. Analysis of the relative contributions of diffusion, migration, and electroosmosis to transport in hair follicles.

Scanning electrochemical microscopy (SECM) is used to measure spatially localized diffusive and iontophoretic transport rates in hairless mouse skin. Molecular fluxes within individual hair follicles are quantified by measuring the rate at which redox-active probe molecules emerge from the follicle. The influence of an applied current on the flux of an anion (ascorbate), a cation (ferrocenylmethyltrimethylammonium), and a neutral molecule (acetaminophen) is used to determine the contributions of diffusion, migration, and electroosmosis to iontophoretic transport. The direction of electroosmotic transport is consistent with hair follicles possessing a net negative charge at neutral pH. Electroosmosis results in a modest increase in the transport rate of the neutral molecule (a factor of approximately 2.4x at an iontophoretic current density of 0.1 mA/cm(2)). Larger enhancements in the flux of the electrically charged species are associated with migration. The electroosmotic flow velocity within hair follicles is established to be 0.5 (+/-0.1) microm/s at 0.1 mA/cm(2), independent of the electrical charge of permeant. The net volume flow rate across skin resulting from electroosmosis in hair follicles is estimated to be 0.3 microL/cm(2)h. The results suggest that hair follicles are a significant pathway for electroosmotic solution flow during iontophoresis. The radius of the hair follicle openings in hairless mouse skin is measured to be 21 +/- 5 microm.

Algorithms↗
PubMed2001

Biopsy needle tip artifact in MR-guided neurosurgery.

A thorough understanding of both the appearance and origin of metallic biopsy needle tip artifact in magnetic resonance imaging (MRI) as well as its interaction with various magnetic resonance (MR) sequence parameters is beneficial for its application in today's MR-guided therapeutic procedures. In a more practical setting, this investigation has focused on the characteristics of MR image artifacts associated with a finite-length metallic needle, specifically at the tip of a biopsy needle when it is approximately parallel to the main magnetic field. The image artifact at needle tip, which exhibits as a blooming ball-shaped signal void, was demonstrated and studied using MR imaging and numerical simulation employing the finite difference method (FDM). In order to understand the origin of this image artifact, a numerical model or simulation software based on the FDM has been developed specifically to solve for the field disturbance to a uniform magnetic field due to a finite-length metallic needle. The solution for magnetic field shows that the field disturbance is spatially localized at the needle tip. From the numerical results, simulated images were generated which were in a very satisfactory agreement MR imaging experiment. Results showed that the MR image artifacts associated with MR-compatible metallic biopsy needles are not only present due to the magnetic susceptibility difference between the needle and its surrounding tissue, but also predictable in routine MR-guided procedures, and the size of the image artifacts could be reduced if optimal imaging parameters were used. J. Magn. Reson. Imaging 2001;13:16-22.

Artifacts↗
PubMed2000

NMR spectroscopy of single neurons.

The first spatially localized NMR spectra of osmolytes and metabolites from single isolated neurons have been obtained using a combination of high magnetic field strengths and NMR radio frequency (RF) microcoils. The proton spectra display peaks at high concentrations (100-300 mM) assigned to betaine and choline, and other metabolite resonances including lactate at lower concentrations in the order of 10s of millimoles. The volumes examined were approximately 10 nl, over two orders of magnitude less than previously possible. In these initial experiments; the cells were unperfused and the signal intensities of the osmolytes decrease with time, a phenomenon consistent with cell swelling. This work demonstrates the technical feasibility of NMR spectroscopy of single cells, further broadening the scope of NMR spectroscopy of living tissues from application to entire living organisms (man and animal models) and isolated tissues (perfused organs and cultured assemblies of cells) and now to single cells. Magn Reson Med 44:19-22, 2000.

Acetates↗
PubMed2000

Spatiotemporal dynamics of the BOLD fMRI signals: toward mapping submillimeter cortical columns using the early negative response.

The existence of the early-negative blood-oxygenation-level-dependent (BOLD) response is controversial and its practical utility for mapping brain functions with columnar spatial specificity remains questionable. To address these issues, gradient-echo BOLD fMRI studies were performed at 4.7 T and 9.4 T using the well-established orientation column model in the cat visual cortex. A robust transient early-negative BOLD response was consistently observed in anesthetized cat (-0.35 +/- 0.09%, mean +/- SD, n = 8 at 2.9 +/- 0.5 sec poststimulus onset for 4.7 T, TE = 31 ms; -0.29 +/- 0.10%, n = 4 at 3.0 +/- 0.8 sec poststimulus onset for 9.4 T, TE = 12 ms). In addition to its temporal evolution, the BOLD response also evolved dynamically in the spatial domain. The initially spatially localized early-negative signal appeared to dynamically drain from the active sites toward large vessels, followed by a wave of the delayed positive signal, which exhibited similar spatiotemporal dynamics. Only the early-negative BOLD response within 2 sec of the stimulus onset (not the entire dip) yielded columnar layouts without differential subtraction. The functional maps of two orthogonal orientations using the first 2-sec dip were indeed complementary. On the other hand, the delayed positive BOLD response appeared diffused and extended beyond the active sites. It was thus less suitable to resolve columnar layouts. These results have implications for the design and interpretation of the BOLD fMRI at columnar resolution. Magn Reson Med 44:231-242, 2000.

Animals↗
PubMed1991

Virtual rigid body dynamics.

An important direction in biological simulations is the development of methods that permit the study of larger systems and/or longer simulation time scales than is currently feasible by molecular dynamics. One such method designed with this objective in mind is stochastic boundary molecular dynamics (SBMD). SBMD was developed for the characterization of spatially localized processes in proteins, and has been shown to successfully reproduce structural and dynamical properties of these macromolecules, as compared to a molecular dynamics control simulation, when concerted or global motions are not present. The virtual rigid body dynamics method presented in this work extends the range of applicability of the SBMD method, by providing a framework to include these important long time scale conformational transitions. In this paper we describe the two-step implementation of the virtual rigid body model: first, the reduction of the full atomic representation to a reduced particle (virtual bond) model, and second, the propagation of the dynamics of flexibly connected rigid bodies containing virtual atom sites.

Biopolymers↗
PubMed2005

An electro-optic monitor of the behavior of Chlamydomonas reinhardtii cilia.

The unicellular green alga Chlamydomonas reinhardtii steers through water with a pair of cilia (eukaryotic flagella). Long-term observation of the beating of its cilia with controlled stimulation is improving our understanding of how a cell responds to sensory inputs. Here we describe how to record ciliary motion continuously for long periods. We also report experiments on the network of intracellular signaling that connects the environment inputs with response outputs. Local spatial changes in ciliary response on the time scale of the underlying biochemical dynamics are observed. Near-infrared light monitors the cells held by a micropipette. This condition is tolerated well for hours, not interfering with ciliary beating or sensory transduction. A computer integrates the light stimulation of the eye of Chlamydomonas with the ciliary motion making possible long-term correlations. Measures of ciliary responses include the beating frequency, stroke velocity, and stroke duration of each cilium, and the relative phase of the cis and trans cilia. The stationarity and dependence of the system on light intensity was investigated. About 150,000,000 total beat cycles and up to 8 h on one cell have been recorded. Each beat cycle is resolved so that each asynchronous beat is detected. Responses extend only a few hundred milliseconds, but there is a persistence of momentary changes that last much longer. Interestingly, we see a response that is linear with absolute light intensity as well as different kinds of response that are clearly nonlinear, implying two signaling pathways from the cell body to the cilia.

Animals↗
PubMed2005

Actin polymerization promotes the reversal of streaming in the apex of pollen tubes.

Actin polymerization is important in the control of pollen tube growth. Thus, treatment of pollen tubes with low concentrations of latrunculin B (Lat-B), which inhibits actin polymerization, permits streaming but reversibly blocks oscillatory growth. In the current study, we employ Jasplakinolide (Jas), a sponge cyclodepsipeptide that stabilizes actin microfilaments and promotes polymerization. Uniquely, Jas (2 microM) blocks streaming in the shank of the tube, but induces the formation of a toroidal-shaped domain in the swollen apex, of which longitudinal optical sections exhibit circles of motion. The polarity of this rotary motion is identical to that of reverse fountain motility in control pollen tubes, with the forward direction occurring at the edge of the cell and the rearward direction in the cell interior. Support for the idea that actin polymerization in the apical domain contributes to the formation of this rotary motility activity derives from the appearance therein of aggregates and flared cables of F-actin, using immunofluorescence, and by the reduction in G-actin as indicated with fluorescent DNAse. In addition, Jas reduces the tip-focused Ca2+ gradient. However, the alkaline band appears in the swollen apex and is spatially localized with the reverse fountain streaming activity. Taken together, our results support the idea that actin polymerization promotes reversal of streaming in the apex of the lily pollen tube.

Actin Cytoskeleton↗
PubMed1991

Cortical connections of the caudal subdivision of the dorsolateral area (V4) in monkeys.

Evidence suggests that all primates have rostral and caudal subdivisions in the region of visual cortex identified as the dorsolateral area (DL) or V4. However, the connections of DL/V4 have not been examined in terms of these subdivisions. To determine the cortical connections of the caudal subdivision of DL (DLC) in squirrel monkeys, injections of the neuroanatomical tracers wheat germ agglutinin conjugated to horseradish peroxidase, Diamidino Yellow, and Fluoro-Gold were made in cortex rostral to V II. To aid in delineating the borders of DLC, cortex was also evaluated architectonically. Based on similar patterns of connections, DLC extends from dorsolateral to ventrolateral cortex. DLC receives strong, feedforward input from V II and projects in a feedforward fashion to the rostral subdivision of DL (DLR) and caudal inferior temporal (IT) cortex, including a separate location in the inferior temporal sulcus. DLC has weaker connections with V I, the middle temporal area (MT), cortex rostral to MT in the location of the fundal superior temporal area (FST), cortex dorsal to DLC, ventral cortex rostral to V II, and cortex in the frontal lobe, lateral to the inferior arcuate sulcus. Only lateral DLC has connections with V I, and only dorsolateral DLC has connections with cortex dorsal to DLC. The topographic organization of DLC was inferred from its connections with V II. Thus, dorsolateral DLC represents the lower field, lateral DLC represents central vision, and ventrolateral DLC represents the upper field. Limited observations were made on DLR. Confirming earlier observations (Cusick and Kaas: Visual Neurosci. 1:211, 1988), DLR is paler than DLC myeloarchitectonically. DLR receives only sparse feedforward input from V II, but stronger input from DLC. DLR has strong connections with cortex just rostral to dorsal V II, ventral posterior parietal cortex in the sylvian fissure, MT, the medial superior temporal area, FST, and the inferior temporal sulcus. DLR also shares connections with IT cortex. Thus, while both DLC and DLR are involved in the pathway relaying visual information to IT cortex, an area specialized for object vision, DLR also projects densely to areas such as MT involved in the pathway relaying to posterior parietal cortex, a region specialized for spatial localization and motion perception.

Amidines↗
PubMed1991

Differential regulation of the low-affinity nerve growth factor receptor during postnatal development of the rat brain.

We studied the temporal and spatial localization of the low-affinity nerve growth factor receptor (LNGF-R) during the early postnatal period in rat brain in order to understand better the relationship between nerve growth factor (NGF)-like responsiveness and the development of specific central neuronal populations. Four different developmental patterns of LNGF-R mRNA hybridization were found in this study. First, some neurons contain high levels of LNGF-R mRNA from postnatal time points into adulthood, as exemplified by neurons of the cholinergic basal forebrain and mesencephalic trigeminal nucleus. Second, several cell groups exhibit robust hybridization during the early postnatal period but contain much reduced levels of LNGF-R mRNA in the adult brain. These include striatal neurons, Purkinje cells of the cerebellum, and several medullary nuclei. A third group of cells produces the LNGF-R transiently during development, including cranial nerve nuclei of the brainstem, the periolivary nuclei complex, the reticular formation, and the deep cerebellar nuclei. Finally, cell populations which may exist only transiently during central nervous system (CNS) development, such as subplate neurons of the cerebral cortex, appear to express the LNGF-R during only a brief period. These results show that the LNGF-R gene is differentially regulated in a cell type-specific manner during development, and suggests that diverse neuronal populations require only transient growth factor sensitivity, while others exhibit NGF-like responsitivity into maturity.

Animals↗
PubMed2005

Efficient evaluation of nonlocal pseudopotentials via Euler exponential spline interpolation.

An Euler exponential spline (EES) based formalism is employed to derive new expressions for the electron-atom nonlocal pseudopotential interaction (NL) in electronic structure calculations performed using a plane wave basis set that can be computed more rapidly than standard techniques. Two methods, one that is evaluated by switching between real and reciprocal space via fast Fourier transforms, and another that is evaluated completely in real space, are described. The reciprocal-space or g-space-based technique, NLEES-G, scales as NMlogM approximately N2logN, where N is the number of electronic orbitals and M is the number of plane waves. The real-space based technique, NLEES-R, scales as N2. The latter can potentially be used within a maximally spatially localized orbital method to yield linear scaling, while the former could be employed within a maximally delocalized orbital method to yield NlogN scaling. This behavior is to be contrasted with standard techniques, which scale as N3. The two new approaches are validated using several examples, including solid silicon and liquid water, and demonstrated to be improvements on other, reduced-order nonlocal techniques. Indeed, the new methods have a low overhead and become more efficient than the standard technique for systems with roughly 20 or more atoms. Both NLEES methods are shown to work stably and efficiently within the Car-Parrinello ab initio molecular dynamics framework, owing to the existence of p-2 continuous derivatives of a pth-order spline.

Journal Article↗
PubMed2006

Compression-inhibited pore formation of polyelectrolyte multilayers containing weak polyanions: a scanning force microscopy study.

Morphological changes of poly(acrylic acid)/poly(diallyldimethylammonium chloride) multilayers induced by low pH were investigated by scanning force microscopy. The weakened interaction between the charged polymer chains in the protonation process is believed to be the reason for this variation. Kinetic studies have shown that during protonation phase separation and dissociation of the multilayers took place successively. The compression of the multilayers, however, caused a transition of the multilayers from a rubbery state to a glassy state. As a result, the closely compacted multilayers lost their sensitivity to pH change. An increase of electrostatic and hydrophobic interactions, can decrease the free energy of the multilayers, and stabilize the films. By compression of the multilayers with a rubber stamp having geometric patterns, films with spatially localized pores were produced.

Journal Article↗
PubMed2006

Functional proteometrics for cell migration.

BACKGROUND: Advances in living cellular fluorescence biosensors and computerized microscopy enable a vision of fully automated high-resolution measurements of the detailed intracellular molecular dynamics directly linked to cellular behaviors. Given the heterogeneity of cell populations, a statistically relevant study of molecular-cellular dynamics is a key motivation for improved automation. METHODS: We explored automating computerized, microscope-based data extraction and analyses that monitor cell locomotion, rates of mitoses, and spatiotemporal activities of intracellular proteins via ratiometric fluorescent biosensors in mouse fibroblasts. Novel image processing methods included K-means clustering segmentation preprocessing followed by modified discrete, normalized cross-correlational alignment of two-color images; ratiometric processing for fluorescence resonance energy transfer (FRET) measurements; and intracellular spatial distribution measurements of RhoA GTPase activity. RESULTS: The interdivision time was 19.4 h (mean) +/- 6.0 h (SD) (n = 7) for the GFP-histone cells in the two-by-two field that was scanned for 72 h. After registration and ratioing of the cells with the RhoA biosensor, increases in both cell protrusion and retraction were coincident with to increases in RhoA activity. CONCLUSIONS: These advances lay the foundation for extracting and correlating measurements characterizing the functional relationships of spatial localization and protein activation with features of cell migration such as velocity, polarization, protrusion, retraction, and mitosis.

Animals↗
PubMed2003

gbx2 Homeobox gene is required for the maintenance of the isthmic region in the zebrafish embryonic brain.

We isolated cDNA clones for the zebrafish gbx2 gene, which is implicated in the establishment of the midbrain-hindbrain boundary (MHB) in other vertebrates. Spatially localized expression of gbx2 was observed at the MHB from 90% epiboly through to the hatching stage. Comparisons with the expression of otx2, wnt1, and krox20 showed that gbx2 is expressed in the anterior hindbrain. Ectopic expression of gbx2 by mRNA injection caused cyclopia or truncation of the fore- and midbrain and severely affected isthmic and cerebellar structures, while hindbrain formation was not significantly affected. At the molecular level, gbx2 suppressed the expression of otx2 in the fore/midbrain, six3 in the anterior forebrain, and MHB-specific genes such as eng2 and wnt1. In contrast, gbx2 did not down-regulate the expression of the hindbrain marker genes. Therefore, gbx2 specifically suppressed the formation of the entire fore/midbrain. Meanwhile, misexpression of otx2 suppressed the expression of gbx2 in the embryonic brain. Abrogation of gbx2 expression with an antisense morpholino oligonucleotide disrupted the midbrain/anterior hindbrain region, and these loss-of-function effects were rescued by activating the Gbx2 protein immediately after the end of gastrulation. Taken together, these results suggest that the zebrafish gbx2 gene is essential for the maintenance of MHB and/or the formation of the isthmic structure during somitogenesis, rather than for the MHB establishment during gastrulation. We also suggest that other factors, including gbx1, is required for the establishment of the MHB during gastrulation.

Amino Acid Sequence↗
PubMed2005

Mph2 expression in germinal zones of the mouse brain.

The telencephalon develops from a seemingly homogeneous population of precursor cells that generates neurons with divergent properties and cell fates. An important question concerns the genetic control of cell states belonging to different generations. Mph2 was identified by serial analysis of gene expression as a differentially expressed gene during cortical development. Belonging to the Polycomb group, Mph2 may be involved in transcriptional repression of cell states from one generation to the next. Here, we use in situ hybridization to determine the spatial localization of Mph2 expression in the developing nervous system. Mph2 expression evident in proliferating precursor cell populations in the developing mouse brain such as the ventricular zones of the cortex and ganglionic eminences as well as in mature structures such as the granule cell layer of the adult dentate gyrus and cerebellum indicates a role for Mph2 in both the developing and adult brain.

Animals↗
PubMed2005

Role of a spatial distribution of IP3 receptors in the Ca2+ dynamics of the Xenopus embryo at the mid-blastula transition stage.

Periodic calcium activity correlates temporally with the onset of gene expression in the embryo, suggesting a causal relation between these two events. Calcium transients are elicited by the action of fibroblast growth factor (FGF) through the activation of phospholipase C. In this work, we present a reaction-diffusion model that extends our previous results on the generation of calcium oscillations for a single and two coupled blastomere cells to a meridian of the Xenopus embryo at the mid-blastula transition. In the model, all cells are subject to the same amount of FGF and contain the same concentration of intracellular components, except for the amount of IP(3) receptors (IP3R). A bell-shaped distribution of IP3R produces the correct shape of the calcium transients experimentally observed in the Xenopus blastula at stage 8 (mid-blastula transition stage). The model is also capable of predicting period and amplitude values close to the experimental values. In our model, calcium transients induce spatially localized ERK periodic transients that could activate specific nuclear genes, allowing for the regional differentiation of the cells in the zone under the influence of the calcium signal.

Animals↗
PubMed2004

Visual encoding mechanisms and their relationship to text presentation preference.

This study was designed to investigate the importance of spatial encoding in reading, with particular emphasis on visuo-spatial encoding mechanisms. Thirty one school children participated in the first study in which they were measured on their ability to solve a centrally presented spatial encoding task, as well as their sensitivity to the frequency doubling illusion across the retina. We found that both spatial frequency doubling sensitivity and performance in the spatial encoding task were correlated with reading, however these tasks were unrelated to each other. Furthermore, frequency doubling sensitivity was correlated with contextual reading, but not single-word reading, while the central spatial encoding task was correlated with both reading tasks. These findings may have functional implications for text presentation preference. Accordingly, in Experiment 2 we demonstrated that children with poor FDT sensitivity read more accurately when words were presented singularly rather than in a whole-text format. In conclusion, we suggest that contextual reading may depend upon two separate and functionally distinct visual encoding mechanisms--one central, important for the spatial discrimination of letters within words, and the other, a spotlighting mechanism important for spatial localization within a body of text. While both mechanisms may constrain reading efficiency, neither mechanism enforces an absolute limit on reading ability.

Child↗