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PubMed2003

[Use of three-dimensional fluorescence deconvolution microscopy for study of spatial distribution of secretory vesicles in living cells].

A three-dimensional image of a living cell is helpful for cell secretion study. In this report, the three-dimensional fluorescence deconvolution microscopy for observing living cells was studied, because this technique can obtain a quick three-dimensional imaging with minimal fluorescence quenching and cytotoxicity for living cell observation. The property of three-dimensional point spread function (PSF) of imaging system was analyzed. The relationship between experimental and theoretical PSF was illustrated, and the theoretical PSF was proved that it could reflect the principle of imaging system with NA 1.65 objective in use. Three-dimensional deconvolution algorithm in this report was proved effective by well-defined three-dimensional specimens. Furthermore, the rat pancreatic beta cell secretory vesicles labeled by acridine orange was observed by using this technique. Results showed that the blurring induced by out-of-focus light was removed by the deconvolution algorithm effectively, under current experiment conditions (with NA 1.65 objective) the experimental PSF approximated the theoretical PSF very well, and deconvolved living cell images exhibited the spatial distribution of the secretory vesicles clearly.

Algorithms↗
PubMed2002

Respiration-induced B0 fluctuations and their spatial distribution in the human brain at 7 Tesla.

In functional magnetic resonance imaging (fMRI), it is known that physiological influences such as cardiac pulsation, respiration, and brain motion can induce fluctuations in signal intensity and phase. Some of the mechanisms potentially involved in those phenomena are expected to be amplified at higher magnetic fields. This study addresses the issue of B(0) fluctuations induced by susceptibility changes during respiration attributed to movements of chest and diaphragm, and variations in the oxygen concentration. It is demonstrated that respiration-induced resonance offsets (RIROs) are significant at 7T. Data were acquired with an RF pulse (no gradients), multislice echo-planar imaging (EPI), and dynamic 3D fast low-angle shot (3D- FLASH) imaging. Three main observations summarize the experimental findings. First, in FIDs measured after a single RF pulse, a RIRO with a large amplitude was consistently detected, although the average amplitude varied between subjects from 1.45 Hz to 4 Hz. Second, in transverse EPI images obtained in the occipital lobe, the RIRO amplitude showed a monotonic increase along the z axis toward the lungs. Third, a more detailed spatial analysis with 3D-FLASH phase maps revealed that a previously published analytical model can accurately describe the spatial distribution of RIRO. Consequential apparent motions in the EPI series, as well as the implications of slice orientation for correction strategies are discussed.

Brain↗
PubMed1984

The spatial distribution, intensity and unpleasantness of acute dental pain.

The distribution, intensity and unpleasantness of acute dental pain were studied in 196 patients. Dental pain was classified by source according to 7 anatomical sites: dentinal, pulpal, pulpal and periapical, periapical, pericoronal, papillar, and periodontal. The distribution of dental pain was classified by the frequency of pain spread throughout the sample, by the extent of pain spread for a particular pain source, that is, the number of vectors of spread, and by the pain reference locations in the face and head. Pain-intensity and pain-unpleasantness were assessed on Visual Analogue Scales. The frequency and extent of pain spread varied as a function of source. However, specific patterns of pain spread were not associated with particular pain sources. There was considerable overlap in patterns of spread between maxillary and mandibular pain sources. The frequency and extent of pain spread was correlated positively with both pain intensity and pain unpleasantness. The correlation between pain ratings and pain spread may be attributed to central mechanisms and interactions between trigeminothalamic neurons. Spatial overlap of pain from maxillary and mandibular pain sources may be due to the large receptive fields of wide dynamic range neurons, that can extend beyond one trigeminal division. These observations indicate that the spatial distribution of acute dental pain is not sufficient as a diagnostic tool for indentifying pain source.

Adult↗
PubMedDate not supplied

Microwave-facilitated extraction of furanocoumarins onto paper substrates: an imaging technique to analyse spatial distribution and abundance in leaves.

An impediment to studying the implications of fine-scale variation in the distribution of secondary compounds within plants has been an inability to map their distribution. Conventional approaches require fine-scale dissections, followed by extraction and analysis, a strategy which is often limited by the large number of extractions required and by the difficulty in quantifying compounds in very small samples. A method has been developed which allows the microwave-facilitated transfer of furanocoumarins from fresh leaves to two-dimensional substrates (phase separation paper). The fluorescence of these compounds under UV light is enhanced by treatment with base, which probably opens the lactone ring of the compounds. The developed prints are then photographed under UV light and the images analysed with commonly available software programs. Images so obtained for furanocoumarins in wild parsnip foliage revealed that the total fluorescence from extracted furanocoumarins is correlated with the amount of furanocoumarins remaining in the leaf. Thus, the images provide information not only on spatial distribution but also quantitative data.

Chemistry Techniques, Analytical↗
PubMed1994

Cellular epigenetics: control of the size, shape, and spatial distribution of transformed foci by interactions between the transformed and nontransformed cells.

NIH 3T3 cells that are passaged frequently at low density in high (10%) calf serum lose their original capacity to produce transformed foci on a monolayer of nontransformed cells. They can then be used to form a monolayered background for the assay of the number of focus-forming cells from a transformed population. Continuation of the low-density passages for many weeks gives rise to a population that can suppress the full development of foci by a transformed line. The suppression appears to occur only after the background cells have become confluent and contact inhibited. It can also cause the disappearance of light foci that had developed before suppression began. Another subline of cells that were passaged at cloning density only once a week lose their focus-forming capacity more slowly than those passaged thrice weekly. When used as a background for the assay of a transformed line, they permit continuous expansion of the foci, with no sign of suppression. Not only the number and size of foci but also their detailed morphology is influenced by the background on which they are formed. A suppressive background can also determine the spatial distribution of foci, presumably as a result of gradients in local cell density of the background. The permissiveness of a nontransformed cell population for focus formation by transformed cells appears to be related to the capacity of the nontransformed population itself to undergo transformation when exposed to the constraints used to induce transformation. These findings indicate there are many degrees of capacity to suppress focus formation and to overcome suppression. They have significance for tumor development and for the epigenetic interactions of normal development.

3T3 Cells↗
PubMed1985

Color contrast perimetry. The spatial distribution of color defects in optic nerve and retinal diseases.

Color contrast perimetry was used to evaluate central visual field defects in a group of 28 patients with visual loss resulting from optic nerve or retinal diseases. Kinetic perimetry was performed using colored test objects of constant luminance, equated to a white surround of 10 ft lamberts. Colored test objects were varied in size and in extent of color saturation. Test object color saturation was varied from a white that matched the color and luminance of the adapting background toward either the blue or the red color maxima of a video tangent screen. All central visual field defects that were demonstrable by luminance contrast perimetry were also detected by color contrast testing, and no defects were found for color contrast detection that could not also be demonstrated by conventional luminance increment perimetry. Retinal diseases usually produced scotomas for both color and luminance contrast detection, while optic nerve disorders tended to produce global depressions of both color and luminance contrast sensitivity across the entire visual field in addition to scotomas. There was no systematic difference in visual field defects for either class of disease when comparing color contrast in the blue (tritan) versus the red (protan) axes of color space. The apparent tritan or protan/deutan axes of color confusion found by hue discrimination testing in acquired dyschromatopsias may be determined by the relative spatial distribution of defects in the central visual field rather than by selective impairment of neural mechanisms for color or luminance information processing.

Adult↗
PubMed2005

Spatial distribution and abundance of nonindigenous coral genus Tubastraea (Cnidaria, Scleractinia) around Ilha Grande, Brazil.

The distribution and abundance of azooxanthellate coral Tubastraea Lesson, 1829 were examined at different depths and their slope preference was measured on rocky shores on Ilha Grande, Brazil. Tubastraea is an ahermatypic scleractinian nonindigenous to Brazil, which probably arrived on a ship's hull or oil platform in the late 1980's. The exotic coral was found along a great geographic range of the Canal Central of Ilha Grande, extending over a distance of 25 km. The abundance of Tubastraea was quantified by depth, using three different sampling methods: colony density, visual estimation and intercept points (100) for percentage of cover. Tubastraea showed ample tolerance to temperature and desiccation since it was found more abundantly in very shallow waters (0.1-0.5 m), despite the fact that hard substratum is available at greater depths at all the stations sampled. At most sites, 1 to 5 colonies per 0.25 m2 were found most frequently, but occasionally more than 50 colonies were found per 0.25 m2, indicating a somewhat gregarious spatial distribution for this coral. The coral Tubastraea was found to occupy slopes of every possible angle in the Canal Central of Ilha Grande, but more colonies were found occupying slopes of 80 to 100 degrees. Therefore, its insensitivity to angles of recruitment and its tolerance for different depths makes it an organism with great ecological tolerance, with a potential to colonize new areas and increase its current range in Brazil's coastal waters.

Animals↗
PubMed2002

Afghan refugees and the temporal and spatial distribution of malaria in Pakistan.

Influx of refugees and establishment of camps or settlements in malaria endemic areas can affect the distribution and burden of malaria in the host country. Within a decade of the Soviet invasion of Afghanistan and the arrival of 2.3 million Afghan refugees in Pakistan's North West Frontier Province, the annual burden of malaria among refugees had risen ten fold from 11,200 cases in 1981 to 118,000 cases in 1991, a burden greater than the one reported by the Pakistan Ministry of Health for the entire Pakistani population. Political developments in the 1990s led to over half the refugee population repatriating to Afghanistan, and the Afghan Refugee Health Programme (ARHP) was scaled down proportionately. Districts in which the ARHP recorded a reduced incidence of malaria began to show an increased incidence in the statistics of the Pakistan government health programme. This and other evidence pointed to a change in health seeking practices of the refugees who remained in Pakistan, with many turning from ARHP to Pakistani health services as aid declined. Comparison of the two sources of data produced no evidence for the spatial distribution of malaria in NWFP having changed during the 1990s. Nor was there any evidence for the presence of refugees having increased the malaria burden in the Pakistani population, as is sometimes alleged. This highlights the risk of misinterpreting health trends when parallel health services are operating. Over the decade incidence in the refugee camps decreased by 25% as a result of control activities, and by 1997 the burden among remaining refugees had fallen to 26,856 cases per annum. These trends indicate that the burden would continue to fall if political conditions in Afghanistan were to improve and more refugees returned to their homeland.

Afghanistan↗
PubMed1983

A computer electron microscope plotter for mapping spatial distributions in biological tissues.

We have designed a computer-based electron microscope plotting system which maps the locations of organelles in tissue specimens and analyzes their distribution. The system includes: (1) two optical incremental shaft encoders which translate stage drive rotation to electrical pulses; (2) a Display/Control unit used to convert encoder pulses to binary code for computer input; (3) two 16-bit parallel interfaces for transferring data to the computer; (4) a Hewlett-Packard 9845T microcomputer, used to control data input and to store, graph, and analyze the plots. The software for the plotter is written in enhanced BASIC. The plotter system is driven by 4 programs called Trace, Plot, Analyze, and Density. The Trace program "draws" an outline of the edges of the tissue. The Plot program maps the positions of profiles within the tissue. The Analyze program compares trace and plot data and calculates the depth and medial-lateral distance of each plotted profile from the surfaces of the tissue. The Density program sorts and counts profile types, measures surface areas, and calculates profile densities. Commercial statistical software is used to analyze the data. Our laboratory uses the system to map the spatial distribution of synapses and neurons in the central nervous system. The plotting system will also be of value in other areas of neurobiology research.

Animals↗
PubMed2004

Dynamics and spatial distribution of beta-lactamase expression in Pseudomonas aeruginosa biofilms.

The development of resistance to beta-lactam antibiotics is a problem in the treatment of chronic Pseudomonas aeruginosa infection in the lungs of patients with cystic fibrosis. The main resistance mechanism is high-level expression of the chromosomally encoded AmpC beta-lactamase of P. aeruginosa cells growing in biofilms. Several genes have been shown to influence the level of ampC expression, but little is known about the regulation of ampC expression in P. aeruginosa biofilms. To study the expression of ampC in P. aeruginosa biofilms, we constructed a reporter that consisted of the fusion of the ampC promoter to gfp(ASV) encoding an unstable version of the green fluorescent protein. In vitro biofilms of P. aeruginosa were exposed to the beta-lactam antibiotics imipenem and ceftazidime. Sub-MICs of imipenem significantly induced the monitor system of the biofilm bacteria in the peripheries of the microcolonies, but the centers of the microcolonies remained uninduced. However, the centers of the microcolonies were physiologically active, as shown by experiments with another monitor construction consisting of an arabinose-inducible promoter fused to gfp(ASV). The whole biofilm was induced in the presence of increased imipenem concentrations. Ceftazidime induced the monitor system of the biofilm bacteria as well, but only bacteria in the peripheries of the microcolonies were induced in the presence of even very high concentrations. The experiments illustrate for the first time the dynamic and spatial distributions of beta-lactamase induction in P. aeruginosa cells growing in biofilms. Thus, our experiments show that P. aeruginosa cells growing in biofilms constitute a heterogeneous population unit which may create different antibiotic-selective environments for the bacteria in the biofilm.

Anti-Bacterial Agents↗
PubMed2000

Voronoi tessellation to study the numerical density and the spatial distribution of neurones.

The conditions of regularity and isotropy, required by standard morphometric procedures, are generally not fulfilled in the central nervous system (CNS) where cells are distributed in a highly complex manner. The evaluation of the mean numerical density of neuronal or glial cells does not take into account the topographical heterogeneity and thereby misses the information that it contains. A local measurement of the density can be obtained by evaluating the 'numerical density of one cell', i.e. the ratio 1/(the volume that the cell occupies). This volume is the region of space that is closer to that cell than to any other. It has the shape of a polyhedron, called Voronoi (or Dirichlet) polyhedron. In 2-D, the Voronoi polyhedron is a polygon, the sides of which are located at mid-distance from the neighbouring cells. The Voronoi polygons are contiguous and their set fills the space without interstice or overlap, i.e. they perform a 'tessellation' that may yield a density map when the same colours are used to fill polygons of similar sizes. The use of Voronoi polygons allows computing the confidence interval of a mean numerical density that makes statistical comparisons possible. The tessellation also provides information concerning spatial distribution; the areas of the Voronoi polygons do not vary much when the cells are regularly distributed. On the contrary, small and large polygons are found when cellular clusters are present. The coefficient of variation of the polygon areas is an objective measurement of their variability and helps to define 'regular', 'clustered' and 'random' distributions. When cells are clustered, small polygons are contiguous and may be objectively identified by simple algorithms. Voronoi tessellations are easily performed in 2-D. On an average the area of a polygon times the thickness of the section equals the volume of the corresponding polyhedron. 3-D tessellations that are theoretically possible and for which algorithms have been published remain to be adapted to histological works.

Algorithms↗
PubMed1984

Spatial distribution of DNA loop attachment and replicational sites in the nuclear matrix.

Biochemical fractionation was combined with high resolution electron microscopic autoradiography to study the localization in rat liver nuclear matrix of attached DNA fragments, in vivo replicated DNA, and in vitro synthesized DNA. In particular, we determined the distribution of these DNA components with the peripheral nuclear lamina versus more internally localized structural elements of isolated nuclear matrix. Autoradiography demonstrated that the bulk of in vivo newly replicated DNA associated with the nuclear matrix (71%) was found within internal matrix regions. A similar interior localization was observed in isolated nuclei and in situ in whole liver tissue. Likewise, isolated nuclear lamina contained only a small amount (12%) of the total matrix-bound, newly replicated DNA. The structural localization of matrix-bound DNA fragments was examined following long-term in vivo labeling of the DNA. The radioactive DNA fragments were found predominantly within interior regions of the matrix structure (77%), and isolated nuclear lamina contained less than 15% of the total nuclear matrix-associated DNA. Most of the endogenous DNA template sites for the replicative enzyme DNA polymerase alpha (approximately 70%) were also sequestered within interior regions of the matrix. In contrast, a majority of the endogenous DNA template sites for DNA polymerase beta (a presumptive repair enzyme) were closely associated with the peripheral nuclear lamina. A similar spatial distribution for both polymerase activities was measured in isolated nuclei before matrix fractionation. Furthermore, isolated nuclear lamina contained only a small proportion of total matrix-bound DNA polymerase alpha endogenous and exogenous template activities (3-12%), but a considerable amount of the corresponding beta polymerase activities (47-52%). Our results support the hypothesis that DNA loops are both anchored and replicated at nuclear matrix-bound sites that are predominantly but not exclusively associated with interior components of the matrix structure. Our results also suggest that the sites of nuclear DNA polymerase beta-driven DNA synthesis are uniquely sequestered within the characteristic peripheral heterochromatin shell and associated nuclear envelope structure, where they may potentially participate in DNA repair and/or replicative functions.

Animals↗
PubMed2004

SPET assessment of auditory cortex stimulation and tonotopic spatial distribution in auditory brainstem implant.

Activation of the auditory cortex by multifrequency acoustic stimuli has been evaluated using Single Photon Emission Tomography in a case of auditory brainstem implant after activation of 6 and 11 electrodes. Before implantation, no activation of the auditory cortex has been observed after acoustic stimulation. Following auditory brainstem implant, the stimulation of 11 electrodes showed an activation value, in terms of blood flow increase, of the contralateral temporal cortex similar to that obtained with 6 electrodes (47.70 vs. 43.76%), but a significantly stronger activation was present in the contralateral parietal region (29.59 vs. 14.73%), in the homolateral temporal area (22.02 vs. 10.46%) and, especially, in the homolateral parietal zone (16.6 vs. 4.33%). The strongest activation in the contralateral temporal cortex was detected in the sagittal tomogram at 26.25 mm from the midline, that is in the areas where high frequencies are projected, both with 6 and 11 active electrodes. The medio-lateral auditory cortex, where the middle and lower frequencies are projected, showed an overall lower activation which was however significantly lower with 6-electrode stimulation. Stimulation of the surface of cochlear nuclei determines mainly an activation of the high frequency domain, independently of the electrodes number. This finding may explain the better results of cochlear implants in comparison with auditory brainstem implant and could justify the use of needle electrodes in auditory brainstem implant. In conclusion, Single Photon Emission Tomography can be considered useful in evaluating auditory brainstem implant placement and function. It is also able to define the effectiveness of acoustic stimulation, the degree and tonotopic spatial distribution of auditory cortex activation.

Acoustic Stimulation↗
PubMed2002

Selectivity and spatial distribution of signals from the receptive field surround in macaque V1 neurons.

The responsiveness of neurons in V1 is modulated by stimuli placed outside their classical receptive fields. This nonclassical surround provides input from a larger portion of the visual scene than originally thought, permitting integration of information at early levels in the visual processing stream. Signals from the surround have been reported variously to be suppressive and facilitatory, selective and unselective. We tested the specificity of influences from the surround by studying the interactions between drifting sinusoidal gratings carefully confined to conservatively defined center and surround regions. We found that the surround influence was always suppressive when the surround grating was at the neuron's preferred orientation. Suppression tended to be stronger when the surround grating also moved in the neuron's preferred direction, rather than its opposite. When the orientation in the surround was 90 degrees from the preferred orientation (orthogonal), suppression was weaker, and facilitation was sometimes evident. The tuning of surround signals therefore tended to match the tuning of the center, though the tuning of the surround was somewhat broader. The tuning of suppression also depended on the contrast of the center grating-when the center grating was reduced in contrast, orthogonal surround stimuli became relatively more suppressive. We also found evidence for the tuning of the surround being dependent to some degree on the stimulus used in the center-suppression was often stronger for a given center stimulus when the parameters of the surround grating matched the parameters of the center grating even when the center grating was not itself of the optimal direction or orientation. We also explored the spatial distribution of surround influence and found an orderly relationship between the orientation of grating patches presented to regions of the surround and the position of greatest suppression. When surround gratings were oriented parallel to the preferred orientation of the receptive field, suppression was strongest at the receptive field ends. When surround gratings were orthogonal, suppression was strongest on the flanks. We conclude that the surround has complex effects on responses from the classical receptive field. We suggest that the underlying mechanism of this complexity may involve interactions between relatively simple center and surround mechanisms.

Algorithms↗
PubMed1991

Specific temporal and spatial distribution of JUN, FOS, and KROX-24 proteins in spinal neurons following noxious transsynaptic stimulation.

We present the first comparative investigation of the basal and transsynaptically induced expression of c-JUN, JUN B, JUN D, c-FOS, FOS B, and KROX-24 proteins in the spinal cord, using immunocytochemistry with specific antibodies. We demonstrate that electrical stimulation of the sciatic nerve at A delta/C-fiber (not A alpha/beta-fiber) intensity strongly induces the expression of these immediate-early gene-encoded proteins. Basal immunoreactivity was found for c-JUN in motoneurons, for JUN D in almost every cell of the gray matter, and for KROX-24 in the superficial dorsal horn. One hour after electrical stimulation of the sciatic nerve at A delta/C-fiber intensity, expression of all proteins except JUN D reached its maximum. Initially immunoreactivity was restricted to the ipsilateral dorsal horn, but after 4 hours appeared contralaterally. Expression of JUN D was increased only after 4 hours. Within the dorsal horn, the expression of c-JUN, JUN B, FOS B, and KROX-24 was mainly restricted to the superficial layers. Immunoreactivity decreased to basal levels between 8 and 16 hours. c-FOS and JUN D were expressed in both the superficial and deep dorsal horn; in the latter, c-FOS and JUN D persisted longer. Induced JUN D was present the longest and was still visible after 32 hours. In motoneurons of the ipsilateral ventral horn, c-JUN, JUN D, and c-FOS appeared after 8 hours. Surgical exposure of the sciatic nerve evoked a strikingly prolonged expression of all proteins compared to that following electrical stimulation of the sciatic nerve. Our results demonstrate that stimulation of nociceptive A delta- and C-fibers induces early and late expression of proteins encoded by immediate-early genes with a specific temporal and spatial distribution of the expression of each protein. Furthermore, the extent of protein expression reflects the intensity of noxious stimulation.

Animals↗
PubMed1997

Immunocytochemical detection and spatial distribution of myosin light-chain kinase in preimplantation mouse embryos.

As a follow-up to our previous study on the role of myosin light-chain kinase (MLCK), a Ca2+/calmodulin-dependent enzyme, in the development of preimplantation mouse embryos, we examined the presence and pattern of distribution of MLCK during preimplantation development of the mouse by whole-mount, indirect immunocytochemistry and by Western blotting, using a monoclonal antibody against MLCK. At all stages of preimplantation development, the nucleus was brightly stained with an unstained region around the nucleus, and regions near the cell membrane were also brightly stained. Using the optical sectioning capability of the confocal laser scanning microscope, we found that, up to the eight-cell stage, the regions of cell contact were mostly unstained, but along with the process of compaction, cell contact regions showed a clear staining pattern along with clearing of the cytoplasm. During formation of the blastocyst, a ring of immunofluorescence was found at the margin of the blastocoel. In the blastocyst, cells of the inner cell mass were less immunofluorescent than trophectoderm cells. These staining results appear to be due to specific immunoreaction between MLCK and the antibody, because the staining patterns were abolished when the antibody was preabsorbed by MLCK purified from chicken gizzard smooth muscle. In Western blotting of blastocysts, we found a band at 130 kD. We also show by immunoblotting and immunohistochemistry of various mouse tissues that the antibody used in this study has cross-reactivity to MLCK of various muscle and non-muscle tissues of the mouse. The presence and spatial distribution of MLCK at various stages of preimplantation development of the mouse suggest that it could play a crucial role in the regulation of the contractile events involved in the initial differentiation that occurs during formation of the mouse blastocyst.

Animals↗
PubMed2005

Spatial distribution of suppressive signals outside the classical receptive field in lateral geniculate nucleus.

A suppressive surround modulates the responsiveness of cells in the lateral geniculate nucleus (LGN), but we know nothing of its spatial structure or the way in which it combines signals arising from different locations. It is generally assumed that suppressive signals are either uniformly distributed or balanced in opposing regions outside the receptive field. Here, we examine the spatial distribution and summation of suppressive signals outside the receptive field in extracellular recordings from 46 LGN cells in anesthetized marmosets. The receptive field of each cell was stimulated with a drifting sinusoidal grating of the preferred size and spatial and temporal frequency; we probed different positions in the suppressive surround with either a large half-annular grating or a small circular grating patch of the preferred spatial and temporal frequency. In many of the cells with a strong suppressive surround (29/46), the spatial distribution of suppression showed clear deviation from circular symmetry. In the majority of these of cells, suppressive signals were spatially asymmetrical or balanced in opposing areas outside the receptive field. A suppressive area was larger than the classical receptive field itself and spatial summation within and between these areas was nonlinear. There was no bias for suppression to arise from foveal or nasal retina where cone density is higher and no other sign of a systematic spatial organization to the suppressive surround. We conclude that nonclassical suppressive signals in LGN deviate from circular symmetry and are nonlinearly combined.

Action Potentials↗