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At least 379 records · Page 21Linked to original sources

All-optical image processing with cavity type II second-harmonic generation.

The possibilities offered by type II intracavity second-harmonic generation for all-optical parallel processing of images are investigated. By injecting an image in a linearly polarized pump beam and a homogeneous field with orthogonal polarization, we obtain, according to the value of the latter, either frequency and polarization transfer or contrast enhancement and contour recognition. Noise-filtering effects are also predicted.

Journal Article↗

Topographic, electrochemical, and optical images captured using standing approach mode scanning electrochemical/optical microscopy.

We developed a high-resolution scanning electrochemical microscope (SECM) for the characterization of various biological materials. Electrode probes were fabricated by Ti/Pt sputtering followed by parylene C-vapor deposition polymerization on the pulled optical fiber or glass capillary. The effective electrode radius estimated from the cyclic voltammogram of ferrocyanide was found to be 35 nm. The optical aperture size was less than 170 nm, which was confirmed from the cross section of the near-field scanning optical microscope (NSOM) image of the quantum dot (QD) particles with diameters in the range of 10-15 nm. The feedback mechanism controlling the probe-sample distance was improved by vertically moving the probe by 0.1-3 microm to reduce the damage to the samples. This feedback mode, defined as "standing approach (STA) mode" (Yamada, H.; Fukumoto, H.; Yokoyama, T.; Koike, T. Anal. Chem. 2005, 77, 1785-1790), has allowed the simultaneous electrochemical and topographic imaging of the axons and cell body of a single PC12 cell under physiological conditions for the first time. STA-mode feedback imaging functions better than tip-sample regulation by the conventionally available AFM. For example, polystyrene beads (diameter approximately 6 microm) was imaged using the STA-mode SECM, whereas imaging was not possible using a conventional AFM instrument.

Animals↗

Optical imaging of dynamic horizontal spread of excitation in rat auditory cortex slices.

Optical recordings using a voltage-sensitive dye (RH482) were conducted in brain slice preparations to investigate spatiotemporal patterns of excitation in the rat auditory cortex. Electrical stimulation of the border between the white matter and layer VI evoked vertical as well as horizontal spreading responses. While velocities of vertical and horizontal propagation of excitation were similar to those reported in non-disinhibited preparations, the horizontal propagation was widespread and strong especially in layers II/III in auditory cortex slices. This horizontal spread was blocked by 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) but not by D-2-amino-5-phosphonopentanoic acid (D-AP5). These results suggest that the horizontal responses, especially in layers II/III, are prominent and are mediated primarily by non-N-methyl-D-aspartic acid (NMDA) receptors in the auditory cortex.

2-Amino-5-phosphonovalerate↗

Modular activation and suppression of neocortical activity in the monkey revealed by optical imaging.

To examine functional local connections in the primate neocortex, we examined changes in optical signal by intracortical microstimulation (ICMS), using a voltage-sensitive dye (RH795) in frontal and parietal cortical areas of the anesthetized monkey. ICMS increased the optical signal at the stimulated site, followed by increases and decreases in a modular fashion at regions that were from hundreds of micrometres to a few millimetres from the stimulated site. These changes lasted for 100-250 ms after the offset of ICMS. Further, this increase in the optical signal was accompanied by an increase in electrical activity in the neurones. These findings suggest the presence of excitatory and inhibitory modular connections within the neocortical areas. It is likely that some functional 'modules' are activated concurrently by excitatory loops, which suppress the activity of other modules.

Animals↗

Near-infrared fluorescent deoxyglucose analogue for tumor optical imaging in cell culture and living mice.

2-deoxy-2-[18F]fluoro-D-glucose ([18F]FDG) has extensively been used for clinical diagnosis, staging, and therapy monitoring of cancer and other diseases. Nonradioactive glucose analogues enabling the screening of the glucose metabolic rate of tumors are of particular interest for anticancer drug development. A nonradioactive fluorescent deoxyglucose analogue may have many applications for both imaging of tumors and monitoring therapeutic efficacy of drugs in living animals and may eventually translate to clinical applications. We found that a fluorescent 2-deoxyglucose analogue, 2-[N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino]-2-deoxy-D-glucose (2-NBDG), can be delivered in several tumor cells via the glucose transporters (GLUTs). We therefore conjugated D-glucosamine with a near-infrared (NIR) fluorphor Cy5.5 and tested the feasibility of the Cy5.5-D-glucosamine (Cy5.5-2DG) conjugate for NIR fluorescence imaging of tumors in a preclinical xenograft animal model. Cy5.5-2DG was prepared by conjugating Cy5.5 monofunctional N-hydroxysuccinimide ester (Cy5.5-NHS) and D-glucosamine followed by high-performance liquid chromatography purification. The accumulation of Cy5.5-2DG and Cy5.5-NHS in different tumor cell lines at 37 and 4 degrees C were imaged using a fluorescence microscope. Tumor targeting and retention of Cy5.5-2DG and Cy5.5-NHS in a subcutaneous U87MG glioma and A375M melanoma tumor model were evaluated and quantified by a Xenogen IVIS 200 optical cooled charged-coupled device system. Fluorescence microscopy imaging shows that Cy5.5-2DG and Cy5.5-NHS are taken up and trapped by a variety of tumor cell lines at 37 degrees C incubation, while they exhibit marginal uptake at 4 degrees C. The tumor cell uptake of Cy5.5-2DG cannot be blocked by the 50 mM D-glucose, suggesting that Cy5.5-2DG may not be delivered in tumor cells by GLUTs. U87MG and A375M tumor localization was clearly visualized in living mice with both NIR fluorescent probes. Tumor/muscle contrast was clearly visible as early as 30 min postinjection (pi), and the highest U87MG tumor/muscle ratios of 2.81 +/- 0.10 and 3.34 +/- 0.23 were achieved 24 h pi for Cy5.5-2DG and Cy5.5-NHS, respectively. While as a comparison, the micropositron emission tomography imaging study shows that [18F]FDG preferentially localizes to the U87MG tumor, with resulting tumor/muscle ratios ranging from 3.89 to 4.08 after 30 min to 2 h postadministration of the probe. In conclusion, the NIR fluorescent glucose analogues, Cy5.5-2DG and Cy5.5-NHS, both demonstrate tumor-targeting abilities in cell culture and living mice. More studies are warranted to further explore their application for optical tumor imaging. To develop NIR glucose analogues with the ability to target GLUTs/hexokinase, it is highly important to select NIR dyes with a reasonable molecular size.

Animals↗

Time-resolved white light transillumination for optical imaging.

PURPOSE: To describe a new breast-imaging method with the potential of multi-spectral optical transillumination based on a time-resolved technique. MATERIAL AND METHODS: A breast phantom was irradiated with ultra-short laser pulses of white light generated by self-phase modulation of an incident high-power laser pulse in water. Time-resolved detection of the transmitted light was performed. Contrast resolution was studied using different absorbers located inside the breast phantom. RESULTS AND CONCLUSION: The results showed that simultaneous, multi-spectral transillumination is possible. The technique can also be used for measurements of optical properties in tissue.

Breast↗

High-resolution near-field optical imaging of single nuclear pore complexes under physiological conditions.

Scanning near-field optical microscopy (SNOM) circumvents the diffraction limit of conventional light microscopy and is able to achieve optical resolutions substantially below 100 nm. However, in the field of cell biology SNOM has been rarely applied, probably because previous techniques for sample-distance control are less sensitive in liquid than in air. Recently we developed a distance control based on a tuning fork in tapping mode, which is also well-suited for imaging in solution. Here we show that this approach can be used to visualize single membrane protein complexes kept in physiological media throughout. Nuclear envelopes were isolated from Xenopus laevis oocytes at conditions shown recently to conserve the transport functions of the nuclear pore complex (NPC). Isolated nuclear envelopes were fluorescently labeled by antibodies against specific proteins of the NPC (NUP153 and p62) and imaged at a resolution of approximately 60 nm. The lateral distribution of epitopes within the supramolecular NPC could be inferred from an analysis of the intensity distribution of the fluorescence spots. The different number densities of p62- and NUP153-labeled NPCs are determined and discussed. Thus we show that SNOM opens up new possibilities for directly visualizing the transport of single particles through single NPCs and other transporters.

Animals↗

Fluorescence-enhanced optical imaging of large phantoms using single and simultaneous dual point illumination geometries.

Fluorescence-enhanced optical tomography is typically performed using single point illumination and multiple point collection measurement geometry. Single point illumination is often insufficient to illuminate greater volumes of large phantoms and results in an inadequate fluorescent signal to noise ratio (SNR) for the majority of measurements. In this work, the use of simultaneous multiple point illumination geometry is proposed for acquiring a large number of fluorescent measurements with a sufficiently high SNR. As a feasibility study, dual point excitation sources, which are in-phase, were used in order to acquire surface measurements and perform three-dimensional reconstructions on phantoms of large volume and/or significant penetration depth. Measurements were acquired in the frequency-domain using a modulated intensified CCD imaging system under different experimental conditions of target depth (1.4-2.8 cm deep) with a perfect uptake optical contrast. Three-dimensional reconstructions of the fluorescence absorption from the dual point illumination geometry compare well with the reconstructions from the single point illumination geometry. Targets located up to 2 cm deep were located successfully, establishing the feasibility of reconstructions from simultaneous multiple point excitation sources. With improved excitation light rejection, multiple point illumination geometry may prove useful in reconstructing more challenging domains containing deeply embedded targets. Image quality assessment tools are required to determine the optimal measurement geometry for the largest set off imaging tasks.

Breast↗

Optical imaging of calcium transients in neurons and pharyngeal muscle of C. elegans.

Electrophysiology and optical indicators have been used in vertebrate systems to investigate excitable cell firing and calcium transients, but both techniques have been difficult to apply in organisms with powerful reverse genetics. To overcome this limitation, we expressed cameleon proteins, genetically encoded calcium indicators, in the pharyngeal muscle of the nematode worm Caenorhabditis elegans. In intact transgenic animals expressing cameleons, fluorescence ratio changes accompanied muscular contraction, verifying detection of calcium transients. By comparing the magnitude and duration of calcium influx in wild-type and mutant animals, we were able to determine the effects of calcium channel proteins on pharyngeal calcium transients. We also successfully used cameleons to detect electrically evoked calcium transients in individual C. elegans neurons. This technique therefore should have broad applications in analyzing the regulation of excitable cell activity in genetically tractable organisms.

Animals↗

Wide-field optical imaging of surface nanostructures.

We present a new technique that increases the sensitivity of incoherent light optical microscopy to a point where it becomes possible to directly visualize ultrathin films (approximately nanometers) and isolated nano-objects. The technique is based on the use of nonreflecting substrate surfaces for cross-polarized reflected light microscopy. These surfaces generate a contrast enhancement of about 2 orders of magnitude, extending the application field of wide-field optical microscopy toward the nanoworld. The efficiency of the method is proven experimentally on well-characterized samples. Wide-field imaging of a nonlabeled lambda-DNA molecule is also presented.

DNA↗

Odour-evoked responses to queen pheromone components and to plant odours using optical imaging in the antennal lobe of the honey bee drone Apis mellifera L.

The primordial functional role of honey bee males (drones) is to mate with virgin queens, a behaviour relying heavily on the olfactory detection of queen pheromone. In the present work I studied olfactory processing in the drone antennal lobe (AL), the primary olfactory centre of the insect brain. In drones, the AL consists of about 103 ordinary glomeruli and four enlarged glomeruli, the macroglomeruli (MG). Two macroglomeruli (MG1 and MG2) and approximately 20 ordinary glomeruli occupy the anterior surface of the antennal lobe and are thus accessible to optical recordings. Calcium imaging was used to measure odour-evoked responses to queen pheromonal components and plant odours. MG2 responded specifically to the main component of the queen mandibular pheromone, 9-ODA. The secondary components HOB and HVA each triggered activity in one, but not the same, ordinary glomerulus. MG1 did not respond to any of the tested stimuli. Plant odours induced signals only in ordinary glomeruli in a combinatorial manner, as in workers. This study thus shows that the major queen pheromonal component is processed in the most voluminous macroglomerulus of the drone antennal lobe, and that plant odours, as well as some queen pheromonal components, are processed in ordinary glomeruli.

Animal Communication↗

Method for the three-dimensional localization of intramyocardial excitation centers using optical imaging.

This study explores the possibility of localizing the excitation centers of electrical waves inside the heart wall using voltage-sensitive dyes (fluorescent or absorptive). In the present study, we propose a method for the 3-D localization of excitation centers from pairs of 2-D images obtained in two modes of observation: reflection and transillumination. Such images can be obtained using high-speed charge-coupled device (CCD) cameras and photodiode arrays with time resolution up to 0.5 ms. To test the method, we simulate optical signals produced by point sources and propagating ellipsoidal waves in 1-cm-thick slabs of myocardial tissue. Solutions of the optical diffusion equation are constructed by employing the method of images with Robin boundary conditions. The coordinates of point sources as well as of the centers of expanding waves can be accurately determined using the proposed algorithm. The method can be extended to depth estimations of the outer boundaries of the expanding wave. The depth estimates are based on ratios of spatially integrated images. The method shows high tolerance to noise and can give accurate results even at relatively low signal-to-noise ratios. In conclusion, we propose a novel and efficient algorithm for the localization of excitation centers in 3-D cardiac tissue.

Action Potentials↗

Imaging optical reflectance in rodent barrel and forelimb sensory cortex.

Novel neuroimaging techniques are extending the scope for studying dynamic brain function. We have developed a system which enables the repeatable imaging of rapid function in rodent primary somatosensory cortex (S-I), based on activity-related changes in its optical reflectance (intrinsic signals). The S-I cortices of anesthetized male Sprague-Dawley rats were exposed. Images were acquired with a slow-scan, cooled, charge-coupled device camera (CCD) through filters at 550, 610, and 850 nm before, during, and after contralateral stimulation (vibrissal deflection or forepaw stimulation). Images were divided by prestimulus controls and then averaged across 9-27 trials to produce maps of stimulus-related reflectance change. Optical activity had magnitude 10(-3) of baseline reflectance and consistently comprised two distinct spatiotemporal components over cortex, depending on paradigm. The diffuse signal at 610 nm begins 0.5-1 s after stimulus onset and has a duration of 4-5 s. The second signal is macrovenous and is delayed by 1 s. Similar response patterns were observed at 550 and 850 nm. Evoked potentials, recorded at sites inside and outside the zone of optical activity, confirmed the functional nature of these signals. Using a CCD we have imaged functional reflectance changes over rodent S-I which commence, peak, and extinguish over a time scale of seconds. This optical activity is consistent with the etiologies of microvascular recruitment and chromophore redox change.

Afferent Pathways↗

Event-related optical imaging reveals the temporal dynamics of right temporal and frontal cortex activation in pre-attentive change detection.

The mismatch negativity (MMN) is a pre-attentive brain response to auditory environmental change. Temporal and frontal cortex generators of pre-attentive change detection have been proposed based on source localization of event-related potentials (ERP), positron emission tomography (PET), and functional magnetic resonance imaging (fMRI) studies. The temporal cortex generators are believed to underlie change detection, whereas the frontal cortex generators are thought to subserve reorientation of attention in response to change. The present study used the event-related optical signal (EROS), an imaging technique that is sensitive to activity related changes in the light scattering properties of neurons, to investigate the pre-attentive brain response to stimulus omissions. The stimulus train comprised 10 ms tone pips presented with a stimulus onset asynchrony (SOA) of 84 ms. Occasional tone omissions elicited a significant increase in right superior temporal gyrus (STG) activity 140 ms after the omitted stimulus, followed 60 ms later by right inferior frontal gyrus (IFG) activity. This result provides support for a temporal-frontal cortical network that underlies pre-attentive change detection.

Acoustic Stimulation↗