Confocal Laser Microscopy of Physiologically Characterized Fluorescently Labeled Central Nervous System Neurons

1988 ◽  
Vol 46 ◽  
pp. 274-275
Author(s):  
J.N. Turner ◽  
J. Swann ◽  
K. Smith ◽  
M. Siemens ◽  
D. Szarowski ◽  
...  
Keyword(s):  
Central Nervous System ◽  
Nervous System ◽  
Laser Scanning ◽  
Tissue Sample ◽  
Lucifer Yellow ◽  
Single Cells ◽  
Brain Slices ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Efficient Detection

Confocal laser scanning microscopy (CLSM) is capable of three-dimensional imaging of fluorescently labeled single cells. Efficient detection via a photomultiplier and optical sectioning with high rejection of light from other specimen levels make it possible to image cells surrounded by either labeled or unlabeled tissue. It is no longer necessary to restrict high resolution light microscopy to cultured cells or those near the surface of a tissue sample. Cells can be observed üin situ” in a physiologically characterized environment. Central nervous system neurons can be electrophysiologically characterized and then injected with a fluorescent dye such as lucifer yellow. The CLSM can excite the dye and image the fluorescent emission in thick tissue preparations (hundreds of micrometers) making possible a new approach to the correlation of physiology and anatomy.Brain slices 350 μm thick were obtained from hippocampus and inferior colliculus of immature rats and incubated in oxygenated artificial cerebrospinal fluid. Cells were penetrated with micropipets, characterized electrophysiologically and ionophoretically injected with 5% lucifer yellow in LiAc.

scholarly journals Heparin Inhibits Leukocyte Rolling in Pial Vessels and Attenuates Inflammatory Changes in a Rat Model of Experimental Bacterial Meningitis

1997 ◽  
Vol 17 (11) ◽  
pp. 1221-1229 ◽  
Author(s):  
Joerg R. Weber ◽  
Klemens Angstwurm ◽  
Thomas Rosenkranz ◽  
Ute Lindauer ◽  
Dorette Freyer ◽  
...  
Keyword(s):  
Central Nervous System ◽  
Nervous System ◽  
Bacterial Meningitis ◽  
Laser Scanning ◽  
Laser Scanning Microscopy ◽  
Confocal Laser ◽  
Leukocyte Rolling ◽  
Central Nervous System Inflammation ◽  
Inflammatory Processes

Heparin is a natural proteoglycan that was first described in 1916. In addition to its well characterized effect on blood coagulation, it is becoming clear that heparin also modulates inflammatory processes on several levels, including the interference with leukocyte–endothelium interaction. Anecdotal observations suggest a better clinical outcome of heparin-treated patients with bacterial meningitis. The authors demonstrate that heparin, a glycosaminoglycan, inhibits significantly in the early phase of experimental pneumococcal meningitis the increase of 1) regional cerebral blood flow (125 ± 18 versus 247 ± 42%), 2) intracranial pressure (4.5 ± 2.0 versus 12.1 ± 2.2 mm Hg), 3) brain edema (brain water content: 78.23 ± 0.33 versus 79.49 ± 0.46%), and 4) influx of leukocytes (571 ± 397 versus 2400 ± 875 cells/μL) to the cerebrospinal fluid compared with untreated rats. To elucidate the possible mechanism of this observation, the authors investigated for the first time leukocyte rolling in an inflammatory model in brain venules by confocal laser scanning microscopy in vivo. Heparin significantly attenuates leukocyte rolling at 2, 3, and 4 hours (2.8 ± 1.3 versus 7.9 ± 3.2/100 μm/min), as well as leukocyte sticking at 4 hours (2.1 ± 0.4 versus 3.5 ± 1.0/100 μm/min) after meningitis induction compared with untreated animals. The authors conclude that heparin can modulate acute central nervous system inflammation and, in particular, leukocyte–endothelium interaction, a key process in the cascade of injury in bacterial meningitis. They propose to evaluate further the potential of heparin in central nervous system inflammation in basic and clinical studies.

scholarly journals Cell Adherence and Drug Delivery from Particle Based Mesoporous Silica Films

2019 ◽  
Author(s):  
Emma Björk ◽  
Bernhard Baumann ◽  
Florian Hausladen ◽  
Rainer Wittig ◽  
mika lindén
Keyword(s):  
Drug Delivery ◽  
Laser Scanning ◽  
Single Cells ◽  
Silicon Wafers ◽  
Laser Scanning Microscopy ◽  
Drug Uptake ◽  
C2c12 Cells ◽  
Confocal Laser ◽  
Particle Sizes

Spatially and temporally controlled drug delivery is important for implant and tissue engineering applications, as the efficacy and bioavailability of the drug can be enhanced, and can also allow for drugging stem cells at different stages of development. Long-term drug delivery over weeks to months is however difficult to achieve, and coating of 3D surfaces or creating patterned surfaces is a challenge using coating techniques like spin- and dip-coating. In this study, mesoporous films consisting of SBA-15 particles grown onto silicon wafers using wet processing were evaluated as a scaffold for drug delivery. Films with various particle sizes (100 – 900 nm) and hence thicknesses were grown onto OTS-functionalized silicon wafers using a direct growth method. Precise patterning of the areas for film growth could be obtained by local removal of the OTS functionalization through laser ablation. The films were incubated with the model drug DiO, and murine myoblast cells (C2C12 cells) were seeded onto films with different particle sizes. Confocal laser scanning microscopy (CLSM) was used to study the cell growth, and a vinculin-mediated adherence of C2C12 cells on all films was verified. The successful loading of DiO into the films was confirmed by UV-vis and CLSM. It was observed that the drugs did not desorb from the particles during 24 hours in cell culture. During adherent growth on the films for 4 h, small amounts of DiO and separate particles were observed inside single cells. After 24 h, a larger number of particles and a strong DiO signal were recorded in the cells, indicating a particle mediated drug uptake. A substantial amount of DiO loaded particles were however attached on the substrate after 24 making the films attractive as a long-term reservoir for drugs on e.g. medical implants.<br>

scholarly journals Localization and quantification of epidermal growth factor receptors on single cells by confocal laser scanning microscopy.

1992 ◽  
Vol 40 (9) ◽  
pp. 1353-1361 ◽  
Author(s):  
M J Good ◽  
W J Hage ◽  
C L Mummery ◽  
S W De Laat ◽  
J Boonstra
Keyword(s):  
Growth Factor ◽  
Epidermal Growth Factor ◽  
Laser Scanning ◽  
Single Cells ◽  
Laser Scanning Microscopy ◽  
Confocal Laser Scanning ◽  
Confocal Laser ◽  
Scatchard Analysis ◽  
Epidermal Growth

We have established a method for quantifying binding of fluorescence-labeled growth factors to their receptors on single cells in situ with the confocal laser scanning microscope (CLSM). Biotinylated epidermal growth factor (EGF) coupled to phycoerythrin-labeled anti-biotin was used to compare the levels of fluorescence on three different cell types for which the number of EGF factors was known from Scatchard analysis of [125I]-EGF binding. The results showed that as few as 10,000 receptors/cell were detectable above back-ground. This method will provide a rapid and quantifiable alternative to autoradiography for ligand binding to single cells in situ.

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