Cytoplasmic streaming in Chara corallina studied by laser light scattering.
(9/3679)
An apparatus is described by means of which the power versus frequency spectrum of the photomultiplier current can be obtained for laser light scattered by streaming cytoplasm in the algal cell Chara corallina. A Doppler peak is noted in the spectrum which is abolished when cytoplasmic streaming is arrested by electrical stimulation. For 5 cells of Chara, this simple laser-Doppler velocimeter gave streaming velocities (46-7 mum s-1, S.D. +/- 4-8 at 20 degrees C) similar to those obtained for the same cells using the light microscope (44-3 mum s-1, S.D. +/- 5-3 at 20 degrees C). A narrow distribution of streaming velocities is indicated. The technique described provides a rapid, quantitative assay of the in vivo rheological properties of cytoplasm. (+info)
Electrophysiological recording in primary visual cortex (VI) was performed both prior to and in the hours immediately following the creation of a discrete retinal lesion in one eye with an argon laser. Lesion projection zones (LPZs; 21-64 mm2) were defined in the visual cortex by mapping the extent of the lesion onto the topographic representation in cortex. There was no effect on neuronal responses to the unlesioned eye or on its topographic representation. However, within hours of producing the retinal lesion, receptive fields obtained from stimulation of the lesioned eye were displaced onto areas surrounding the scotoma and were enlarged compared with the corresponding field obtained through the normal eye. The proportion of such responsive recording sites increased during the experiment such that 8-11 hours post-lesion, 56% of recording sites displayed neurons responsive to the lesioned eye. This is an equivalent proportion to that previously reported with long-term recovery (three weeks to three months). Responsive neurons were evident as far as 2.5 mm inside the border of the LPZ. The reorganization of the lesioned eye representation produced binocular disparities as great as 15 degrees, suggesting interactions between sites in VI up to 5.5 mm apart. (+info)
Evanescent-wave microscopy: a new tool to gain insight into the control of transmitter release.
(11/3679)
Evanescent-wave excitation was used to visualize individual fluorescently labelled vesicles in an optical slice near the plasma membrane of bovine adrenal chromaffin cells. A standard upright microscope was modified to accommodate the optics used for directing a laser beam under a supracritical angle on to the glass-water interface on top of which the cells are grown. Whereas epi-illumination images appeared blurred and structureless, evanescent-wave excitation highlighted acridine orange-labelled vesicles as individual pinpoints. Three-dimensional (3D) trajectories of individual vesicles were obtained from time-resolved image stacks and used to characterize vesicles in terms of their average fluorescence F and mobility, expressed here as the 3D diffusion coefficient D(3). Based on the single-vesicle analysis, two groups of vesicles were identified. Transitions between these states were studied before and after stimulation of exocytosis by repetitive or maintained membrane depolarizations by elevated extracellular [K+]. Findings were interpreted as sequential transitions between the previously characterized pools of vesicles preceding the fusion step. The observed approach of vesicles to their docking sites was not explained in terms of free diffusion: most vesicles moved unidirectionally as if directed to their binding sites at the plasma membrane. Vesicle mobility at the membrane was low, such that the sites of docking and fusion were in close vicinity. Both the rim region and confined areas in the centre of the footprint region were the site of intense vesicle trafficking. (+info)
Estimation of the retinal nerve fibre layer thickness in the papillomacular area of long standing stage IV macular holes.
(12/3679)
AIM: To compare the thickness of the retinal nerve fibre layer (RNFL) in the papillomacular area of patients with long standing stage IV macular holes with age matched controls, using a scanning laser polarimeter. METHODS: The nerve fibre analyser (NFA) was used to measure the mean thickness of the RNFL around the optic nerve head, the thickness values of temporal and nasal 45 degrees sectors and the integral values in 10 patients with macular holes and in 10 age matched controls. RESULTS: The mean RNFL thickness around the optic nerve head was 79.71 (SD 15.06) microm in the macular hole group and 75.1 (10.8) microm in the control group (p = 0.44). The mean thickness in the temporal sector was 63.69 (12.08) microm in the macular hole group and 58.65 (8.9) microm in the control group (p = 0.3). The mean ratio between the temporal and nasal sector thickness values was 0.8441 in the macular hole group and 0.7819 in the controls (p = 0.42). CONCLUSIONS: There was no significant difference in the thickness of the RNFL in the papillomacular area in the two groups. This suggests that there may be no changes in the thickness of the RNFL in patients with long standing macular holes. (+info)
Sensitive detection of Escherichia coli O157:H7 in food and water by immunomagnetic separation and solid-phase laser cytometry.
(13/3679)
Rapid, direct methods are needed to assess active bacterial populations in water and foods. Our objective was to determine the efficiency of bacterial detection by immunomagnetic separation (IMS) and the compatibility of IMS with cyanoditolyl tetrazolium chloride (CTC) incubation to determine respiratory activity, using the pathogen Escherichia coli O157:H7. Counterstaining with a specific fluorescein-conjugated anti-O157 antibody (FAb) following CTC incubation was used to allow confirmation and visualization of bacteria by epifluorescence microscopy. Broth-grown E. coli O157:H7 was used to inoculate fresh ground beef (<17% fat), sterile 0.1% peptone, or water. Inoculated meat was diluted and homogenized in a stomacher and then incubated with paramagnetic beads coated with anti-O157 specific antibody. After IMS, cells with magnetic beads attached were stained with CTC and then an anti-O157 antibody-fluorescein isothiocyanate conjugate and filtered for microscopic enumeration or solid-phase laser cytometry. Enumeration by laser scanning permitted detection of ca. 10 CFU/g of ground beef or <10 CFU/ml of liquid sample. With inoculated meat, the regression results for log-transformed respiring FAb-positive counts of cells recovered on beads versus sorbitol-negative plate counts in the inoculum were as follows: intercept = 1.06, slope = 0.89, and r2 = 0. 95 (n = 13). The corresponding results for inoculated peptone were as follows: intercept = 0.67, slope = 0.88, and r2 = 0.98 (n = 24). Recovery of target bacteria on beads by the IMS-CTC-FAb method, compared with recovery by sorbitol MacConkey agar plating, yielded greater numbers (beef, 6.0 times; peptone, 3.0 times; water, 2.4 times). Thus, within 5 to 7 h, the IMS-CTC-FAb method detected greater numbers of E. coli O157 cells than were detected by plating. The results show that the IMS-CTC-FAb technique with enumeration by either fluorescence microscopy or solid-phase laser scanning cytometry gave results that compared favorably with plating following IMS. (+info)
Video-rate scanning two-photon excitation fluorescence microscopy and ratio imaging with cameleons.
(14/3679)
A video-rate (30 frames/s) scanning two-photon excitation microscope has been successfully tested. The microscope, based on a Nikon RCM 8000, incorporates a femtosecond pulsed laser with wavelength tunable from 690 to 1050 nm, prechirper optics for laser pulse-width compression, resonant galvanometer for video-rate point scanning, and a pair of nonconfocal detectors for fast emission ratioing. An increase in fluorescent emission of 1.75-fold is consistently obtained with the use of the prechirper optics. The nonconfocal detectors provide another 2.25-fold increase in detection efficiency. Ratio imaging and optical sectioning can therefore be performed more efficiently without confocal optics. Faster frame rates, at 60, 120, and 240 frames/s, can be achieved with proportionally reduced scan lines per frame. Useful two-photon images can be acquired at video rate with a laser power as low as 2.7 mW at specimen with the genetically modified green fluorescent proteins. Preliminary results obtained using this system confirm that the yellow "cameleons" exhibit similar optical properties as under one-photon excitation conditions. Dynamic two-photon images of cardiac myocytes and ratio images of yellow cameleon-2.1, -3.1, and -3.1nu are also presented. (+info)
Radixin is involved in lamellipodial stability during nerve growth cone motility.
(15/3679)
Immunocytochemistry and in vitro studies have suggested that the ERM (ezrin-radixin-moesin) protein, radixin, may have a role in nerve growth cone motility. We tested the in situ role of radixin in chick dorsal root ganglion growth cones by observing the effects of its localized and acute inactivation. Microscale chromophore-assisted laser inactivation (micro-CALI) of radixin in growth cones causes a 30% reduction of lamellipodial area within the irradiated region whereas all control treatments did not affect lamellipodia. Micro-CALI of radixin targeted to the middle of the leading edge often split growth cones to form two smaller growth cones during continued forward movement (>80%). These findings suggest a critical role for radixin in growth cone lamellipodia that is similar to ezrin function in pseudopodia of transformed fibroblasts. They are consistent with radixin linking actin filaments to each other or to the membrane during motility. (+info)
Radiation-induced leukocyte entrapment in the rat retinal microcirculation.
(16/3679)
PURPOSE: To evaluate the effects of irradiation on leukocyte dynamics in the rat retinal microcirculation. METHODS: Thirty-five Brown-Norway rats received a dose of 10 Gy irradiation in one fraction. Leukocyte dynamics were studied with acridine orange digital fluorography, in which a nuclear fluorescent dye of acridine orange is used and examined by scanning laser ophthalmoscope. This technique allowed visualization of fluorescent leukocytes in vivo. The leukocyte dynamics were evaluated at 0, 4, 7, 14, 30, and 60 days after the irradiation. RESULTS: Mean leukocyte velocity in the retinal capillaries decreased immediately. It partially recovered on day 4 but then gradually decreased up to the 2-month mark. Low-dose irradiation induced entrapment of leukocytes in the retinal microcirculation. The number of entrapped leukocytes gradually increased with time. The major retinal vessels significantly constricted immediately after irradiation. The diameter was reduced by 76% in arteries and 75% in veins, 2 months after irradiation. CONCLUSIONS: Entrapped leukocytes may be activated and exacerbate vascular injury and microinfarction and thus may participate in the pathogenesis of radiation retinopathy. (+info)