Video-based Imaging System Research Articles

Morphometry of Dorsal Raphe Nucleus Serotonergic Neurons in Alcoholism

Reduced serotonergic function is hypothesized in alcohol abuse and dependence. Serotonergic innervation of the cortex arises predominantly from the dorsal raphe nucleus (DRN). We sought to determine the number and morphometric characteristics of DRN serotonergic neurons postmortem in alcoholic individuals (n=9; age: 16-66 years; 8M:1F) compared with psychiatrically normal, nonalcoholic controls (n=6; age: 17-74 years; 4M:2F). Brainstems were collected at autopsy, fixed and cryoprotected. Alcohol dependence or abuse was determined by psychological autopsy (DSM-IV), the presence of liver fatty changes or cirrhosis and/or high blood alcohol level. Tissue was sectioned at 50 microm (-25 degrees C). A series of 1:10 sections was immunoreacted with antiserum to tryptophan hydroxylase (TPH), the rate-limiting enzyme in the biosynthesis of serotonin. The total number of TPH-immunoreactive (IR) DRN neurons was determined by stereology. Neuron morphometry indices were determined using a video-based imaging system attached to a microscope. We identified TPH-IR neurons every 1,000 microm in each brainstem and measured neuron area, total cross sectional neuron area, and the total area and density of immunolabeled processes. Dorsal raphe nucleus neuron number (controls: 80,386+/-10,238; alcoholic individuals: 85,884+/-12,478) was not different between groups but TPH-IR was greater in alcoholic individuals throughout the rostrocaudal extent of the DRN. The volume of the DRN was 66+/-9 mm3 in controls and 55+/-5 mm3 in alcoholic individuals (p>0.05). The average size of DRN neurons did not differ between groups (353+/-12 microm2 for controls vs 360+/-15 microm2 for alcoholic subjects). However, the area occupied by neuron processes (area of processes/DRN area) was 2.2-fold greater in alcoholic individuals compared with controls (p<0.05). The increased area occupied by neuron processes in alcoholic individuals may represent sprouting and, together with greater TPH-IR, be a compensatory response to impaired serotonergic transmission or cumulative effects of alcohol on the serotonin system.

Noise aliasing in interline-video-based fluoroscopy systems.

Video-based imaging systems for continuous (nonpulsed) x-ray fluoroscopy use a variety of video formats. Conventional video-camera systems may operate in either interlaced or progressive-scan modes, and CCD systems may operate in interline- or frame-transfer modes. A theoretical model of the image noise power spectrum corresponding to these formats is described. It is shown that with respect to frame-transfer or progressive-readout modes, interline or interlaced cameras operating in a frame-integration mode will result in a spectral shift of 25% of the total image noise power from low spatial frequencies to high. In a field-integration mode, noise power is doubled with most of the increase occurring at high spatial frequencies. The differences are due primarily to the effect of noise aliasing. In interline or interlaced formats, alternate lines are obtained with each video field resulting in a vertical sampling frequency for noise that is one half of the physical sampling frequency. The extent of noise aliasing is modified by differences in the statistical correlations between video fields in the different modes. The theoretical model is validated with experiments using an x-ray image intensifier and CCD-camera system. It is shown that different video modes affect the shape of the noise-power spectrum and therefore the detective quantum efficiency. While the effect on observer performance is not addressed, it is concluded that in order to minimize image noise at the critical mid-to-high spatial frequencies for a specified x-ray exposure, fluoroscopic systems should use only frame-transfer (CCD camera) or progressive-scan (conventional video) formats.

Clinical evaluation of replacement of class V resin based composite restorations

Objectives: The purpose of this clinical study was to evaluate any increase in size of Class V resin based composite restorations that required replacement. Emphasis was placed on measurements of the loss of tooth structure in areas distant from the site of failure. Methods: 14 Class V restorations scheduled for replacement were recorded with pre-operative impressions and intra-oral photographs. Restorations were then removed and cavity preparations were completed. A second impression and photograph were taken and stone casts were prepared for each impression. The perimeter and area of each restoration and cavity preparation were measured with a video-based imaging system (Dage 725 CCD camera) equipped with a macro lens, at a fixed magnification of ×10–15. The initial measurements of the restoration image were compared to the measurements of the new cavity preparation, in both instances excluding the site of failure. Intra-class correlation coefficient (ICR) and paired t-test were used for statistical analysis. Results: A significant difference between the pre and post-operative measurements for both area ( P=0.0125) and perimeter ( P=0.003) was found. Conclusion: The study showed that replacement of Class V resin based composite restorations resulted in increased size of the cavity preparation in areas distant from the site of failure.

In vitro evaluation of margins of replaced resin-based composite restorations.

To evaluate in vitro any increase in size of Class V resin-based composite (RBC) restorations as a result of replacement. For group 1, 20 Class V cavity preparations were prepared in extracted incisor and canine teeth. The cavosurface margin was prepared as a butt joint 90-degree angle toward the occlusal wall. Impressions were made of all preparations with a polyvinyl siloxane material. The cavities then received a 1.5-mm 45-degree bevel at the cavosurface margin of the occlusal wall. Impressions were again made of the preparations. The cavities were then restored with a conventional multistep bonding system (Scotchbond Multi-Purpose, 3M Dental Products, St. Paul, Minnesota) and a microfilled RBC (Silux Plus, 3M Dental Products). As a second part of the study (group 2), another 20 Class V cavity preparations were prepared. Half of the cavities received a bevel at the cavosurface margin of the occlusal wall, and half were finished with a butt joint. Impressions were made from all 20 specimens followed by resin-based composite restoration in the same manner as previously described. All the restorations from both group 1 and group 2 were then removed with carbide burs. New impressions were made, and 100 stone models were fabricated. The perimeter and area of the cavity preparations, including the beveled area, were recorded using a video-based imaging system equipped with macro lens, at 10 to 15 times magnification. Both perimeter and area of the postoperative cavity preparation were compared to the initial cavity size for both butt joint and bevel finishing. Intraclass correlation coefficient and paired t-test showed no significant difference between the preoperative bevel type of cavity preparation and its postoperative counterparts for both group 1 and group 2 (p > .05). However, a significant increase in the size of the cavity preparation was found preoperatively when a bevel was placed at the cavosurface margin. Postoperatively, a significant increase in size of the cavities was found for butt joint type finishing for both area (p = .0001) and perimeter (p = .0001), compared to initial cavity size.

Observation study of electronic portal images for off-line verification

Purpose: The goals of this study were to evaluate the use of electronic portal imaging device (EPID) paper images as off-line verification tools and to assess the feasibility of replacing portal films by EPID printed images. Materials and methods: Electronic portal images were acquired using a video-based imaging system. After contrast enhancement, these images were printed and compared to portal films when prescribed, and judged about their usefulness for off-line verification. A total of 2025 images were acquired from 322 fields on 137 patients. The images were shown to eight radiation oncologists and two senior residents in radiation oncology, each one of them judging fields relevant to his (her) daily practice. The questions asked were related to the choice of important anatomical structures and the visibility of such structures, the usefulness of the printed images, the comparison with portal films and the possible replacement of such films by paper images. Results: Answers to the different questions were treated as quantitative scores. For the visibility question, means and standard deviations were calculated for each individual structure, then a global score was obtained for a given treatment site. Means and standard deviations were also computed for the comparison question. Proportions and confidence intervals were used for the other questions. The results show that EPID paper images are useful for some treatment sites such as breast, thorax, prostate, abdomen, pelvis (other than rectum) and axilla. The image quality remains insufficient for some other sites such as head and neck and spine. Conclusion: Although global anatomical landmarks scores are good, the usefulness score is not always as high because some essential anatomical structures scores must be taken into account. There is also a strong habit factor related to acceptance of EPID printed images as verification tools. As long as they see more and more images, radiation oncologists can more easily visualize anatomical structures and are less stringent when evaluating the efficiency of EPID paper images as off-line verification tools.

Light source-induced error in computer-assisted image analysis with a video-based system

Objectives We tested the hypothesis that two different light sources, an alternating current fluorescent viewbox and a direct current halogen viewbox, do not differ with respect to their ability to illuminate reproducibly a radiograph during image capture. Study design Two radiographs were taken: one with four hydroxyapatite chips mounted against a dry mandible and onewithout the chips. They were digitally subtracted with a video-based imaging system. The procedure was repeated at different times. Results A statistically significant difference among optical density measurements was found when the alternating currentfluorescent viewbox ( p<0.001) was used and was related to light intensity variation. Such effect was not observed with the direct current halogen viewbox ( p=0.873). Conclusions Study design efficiency was increased by 212% with the use of the direct current halogen viewbox so that todetect a specified treatment effect with a given level of statistical confidence, the sample size has to be 2.12 times greater if the alternating current fluorescent viewbox is used.

Measurements of coal particle shape, mass, and temperature histories: Impact of particle irregularity on temperature predictions and measurements

Individual coal and carbon particles were levitated in an electrodynamic balance (EDB) and characterized using high-speed diode array and video-based imaging systems to determine particle surface area, volume, drag, mass, and density. These same particles were then heated bidirectionally using a long-pulsed Nd:YAG laser to simulate combustion-level heating fluxes (heating rates on order of 10 4 –10 5 K/s). Measurements of particle surface temperature, size, and laser temporal power variation were made and recorded during each heating experiment. Measured temperature histories were compared with a heat transfer analysis that accounted for variations in particle shape, mass, density, and laser heating power. Results of this study indicate that with well-characterized materials of known properties, agreement, between measurement and model of within 20 K is typical throughout an entire heating and cooling profile. Large particle-to-particle variations are observed in coal particle temperature histories during rapid heating. These variations can be explained inlarge part by accounting for particle-to-particle property (shape, mass, and density) variations. Even when accounting for particle-to-particle shape and density variation, however, model predictions greatly underestimate observed temperature histories. It is concluded that these discrepancies are largely due to uncertainties in the thermal properties (heat capacity and thermal conductivity) typically used to model coal combustion behavior.

Schwann Cells Exhibit P2Y Purinergic Receptors that Regulate Intracellular Calcium and Are Up‐Regulated by Cyclic AMP Analogues

Schwann cells establish close contact with axons during development, and this is maintained throughout life. Signaling by neurotransmitters may play an important role in Schwann cell-axon interaction. Schwann cells were examined for the presence of neuroligland receptors that are linked to increases in levels of cytoplasmic calcium. Schwann cell cultures were prepared from neonatal rat sciatic nerve and, after 0.25, 1, 4, 7, and 14 days in vitro (DIV), loaded with the calcium indicator dye fura 2-AM. The influence of neuroligands on the cytosolic free calcium concentration ([Ca2+]i) was then examined at each time point using a video-based imaging system. Approximately 80-95% of all freshly isolated Schwann cells responded to 10 microM ATP with a three-fold rise in [Ca2+]i. Bradykinin, glutamate, and histamine had no or only partial and inconsistent responses. The ATP-induced calcium response disappeared within 4 DIV. Culturing cells in the presence of cyclic AMP (cAMP) analogues (which induce proliferation and differentiation in vitro) restored the ability of Schwann cells to respond to ATP with increased [Ca2+]i. In the presence of cAMP analogues the extent of recovery of ATP responsiveness was dependent on serum concentration. Fifty to ninety percent of cells regained calcium responsiveness to ATP when grown in medium containing cAMP analogues and 1% serum. These cells also exhibited immunoreactivity to P0 antibody, characteristic of the myelinating lineage. In contrast, only 15-30% of the Schwann cells regained calcium responsiveness when grown in medium containing cAMP analogues and 10% serum.(ABSTRACT TRUNCATED AT 250 WORDS)

Regulation of astroglial responsiveness to neuroligands in primary culture

Previous studies from this laboratory indicate that type-1 astroglia in primary culture are pharmacologically heterogeneous. Two competing hypotheses were proposed to explain the development of glial heterogeneity. First, that the heterogeneity may reflect stable subclasses of astroglia that express a set of receptor-signalling systems. Second, that astroglia can undergo qualitative changes in their expression of receptor-signalling systems with time in vitro. To distinguish between these two hypotheses, experiments were designed to examine neuroligand-evoked calcium responses within clones of type-1 astroglia. If stable and distinct subsets of astroglia were present, a clone derived from a single cell would exhibit uniform responses to a given set of neuroligands. Alternatively, if the pharmacological properties of astroglia underwent qualitative changes, astroglial clones should contain pharmacologically distinct cells. A video-based imaging system and the Ca 2+ indicator dye Fura-2 were used to monitor receptor-mediated increases in Ca 2+ i upon receptor activation. Interestingly, only a fraction of the cells within a given clone responded to carbachol or histamine with an increase in Ca 2+ i, whereas treatment with a P 2Y purinergic receptor agonist generally increased Ca 2+ i in 100% of the cells within the clone. To examine the stability of the receptor signalling over time, individual astroglia within a number of clones were tested on different days for their ability to respond to neuroligands. The results of these experiments indicated that individual astroglial cells tended to lose their responsiveness to certain ligands such as carbachol and histamine as they developed responsiveness to others such as norepinephrine. Our data indicate that during development neurotransmitter receptors on astroglial cells are regulated by both internal and external mechanisms. Glial proliferation produces a variety of pharmacologically distinct astroglial cells. Exposure to neurotransmitters can qualitatively turn off some, but not all, astroglial receptor systems.

Oligodendroglial lineage cells express neuroligand receptors.

This study was designed to determine whether cells of the oligodendroglial lineage express neuroligand receptors linked to Ca2+ mobilization. Intracellular Ca2+ levels were monitored with a video-based imaging system and cells were characterized with immunocytochemical markers. O-2A progenitor cells (A2B5+/GFAP-) and mature oligodendroglia (GC+/MBP+) responded to norepinephrine, glutamate, ATP, and histamine with increased intracellular Ca2+ levels. As O-2A progenitor cells differentiated into mature oligodendroglia, there was an increase in the percentage of cells that responded to ATP and histamine with an increase in intracellular Ca2+ levels. Both O-2A progenitor cells and mature oligodendroglia were pharmacologically heterogeneous with respect to their ability to respond to neuroligands with an increase in intracellular Ca2+. Treatment with bradykinin, carbachol, and substance P also increased intracellular Ca2+ levels in O-2A progenitor cells and mature oligodendroglia. Whereas the percentage of cells that responded to bradykinin and substance P increased with differentiation of O-2A progenitor cells into mature oligodendroglia, the trend was reversed with respect to the percentage of cells responding to carbachol. These results suggest that cells of the oligodendroglial lineage exhibit neuroligand receptors linked to Ca2+ mobilization and that the ability of these cells to respond to neuroligands is developmentally regulated.

Pharmacologically-distinct subsets of astroglia can be identified by their calcium response to neuroligands

Type 1 astroglia have been reported to exhibit in excess of 20 different neuroligand receptors in vitro. The aim of this study was to determine if astroglia, like neurons, are heterogeneous with respect to their ability to respond to different neuroligand receptor agonists. Type 1 astroglia were loaded with the calcium indicator dye fura-2 and the influence of six different neuroligands on their intracellular calcium levels was examined using a video-based imaging system. Each of the six different neuroligands tested increased calcium levels in a subpopulation of glial fibrillary acidic protein immunopositive cells (astroglia). The percentage of cerebral cortical type 1 astroglia responding to a given neuroligand varied with the agonist and generally followed the order: 2-methylthio-ATP > phenylephrine > carbachol = serotonin > glutamate = histamine. The results also indicate that a single astroglial cell can respond to one agonist with a sustained rise in calcium levels and to an alternate agonist with oscillations in calcium levels. The pharmacological heterogeneity evident among astroglia does not appear to depend on cell proliferation or association with neurons. The results of our studies indicate that cultures of cerebral cortical type 1 astroglia are composed of distinct subsets of cells that can be distinguished by qualitative differences in their ability to respond to specific neuroligands with a rise in intracellular calcium.