| 1. |
- Immerstrand, Charlotte, 1974-, et al.
(author)
-
Altered impedance during pigment aggregation in Xenopus laevis melanophores
- 2003
-
In: Medical and Biological Engineering and Computing. - 0140-0118 .- 1741-0444. ; 41:3, s. 357-364
-
Journal article (peer-reviewed)abstract
- Melanophores are dark-brown pigment cells located in the skin of amphibia, fish and many invertebrates. The skin colour of these organisms is regulated by the translocation of pigment organelles, and the pigment distribution can be altered by external stimuli. The ability to change colour in response to stimuli makes these cells of interest for biosensing applications. It was investigated whether pigment aggregation in Xenopus laevis melanophores can be detected by impedance measurements performed in transparent microvials. The results show that cell attachment, cell spreading and pigment aggregation all resulted in impedance changes, seen particularly at the highest frequency tested (10 kHz). The mechanisms behind the impedance changes were investigated by the addition of latrunculin or melatonin, both of which cause pigment aggregation. The latrunculin-induced aggregation was associated with cell area decrease and filamentous actin (F-actin) breakdown, processes that can influence the impedance. Lack of F-actin breakdown and an increase in cell area during melatonin-induced aggregation suggest that some other intracellular process also contributes to the impedance decrease seen for melatonin. It was shown that impedance measurements reflect not only cell attachment and cell spreading, but also intracellular events.
|
|
| 2. |
- Immerstrand, Charlotte, 1974-
(author)
-
Biophysical studies of pigment transport in frog melanophores : impedance measurements and advanced microscopy analyses
- 2003
-
Doctoral thesis (other academic/artistic)abstract
- Small proteins, other molecules and large organelles are frequently transported from one location to another within cells. These transports employ cytoskeletal networks and enable cells to maintain regions with different functions and attain an asymmetrical shape.The aim of this work was to explore biophysical methods for monitoring intracellular transport processes and associated structural changes. For these studies we have used pigment cells, melanophores, from the African clawed frog Xenopus laevis. In response to external stimuli, these cells can change colour by redistributing pigment granules in the cytoplasm.Transparent "cell clinics" equipped with gold electrodes were developed for impedance studies. The results show that impedance measurements at different frequencies not only can be used to monitor cell attachment and spreading, but also events like pigment aggregation. Significant F-actin breakdown and a cell area decrease may explain the impedance decrease seen during latrunculin-induced aggregation. In aggregation induced by melatonin there was, however, a small increase of the cell area, no F-actin breakdown and still lowered impedance, indicating that some other, likely intracellular mechanism is involved. In addition, confocal laser scanning microscopy (CLSM) studies showed that aggregation was associated with an increase in the cell height, more prominent for latrunculin than for melatonin. This height increase did not seem to involve influx of water through aquaporin channels at the cell membrane, or newly formed or remodelled microtubules in the cells.Besides impedance measurements, Image Correlation Spectroscopy (ICS) was applied to analyse pigment aggregation. The study shows for the first time that ICS can be used to analyse aggregation of non-fluorescent particles and suggests that the method may provide new information on the state of aggregation of granules in pigment cells.
|
|
| 3. |
- Immerstrand, Charlotte, 1974-, et al.
(author)
-
Height changes associated with pigment aggregation in Xenopus laevis melanophores
- 2004
-
In: Bioscience Reports. - : Portland Press Ltd.. - 0144-8463 .- 1573-4935. ; 24:3, s. 203-214
-
Journal article (peer-reviewed)abstract
- Melanophores are pigment cells found in the skin of lower vertebrates. The brownish-black pigment melanin is stored in organelles called melanosomes. In response to different stimuli, the cells can redistribute the melanosomes, and thereby change colour. During melanosome aggregation, a height increase has been observed in fish and frog melanophores across the cell centre. The mechanism by which the cell increases its height is unknown. Changes in cell shape can alter the electrical properties of the cell, and thereby be detected in impedance measurements. We have in earlier studies of Xenopus laevis melanophores shown that pigment aggregation can be revealed as impedance changes, and therefore we were interested in investigating the height changes associated with pigment aggregation further. Accordingly, we quantified the changes in cell height by performing vertical sectioning with confocal microscopy. In analogy with theories explaining the leading edge of migrating cells, we investigated the possibility that the elevation of plasma membrane is caused by local swelling due to influx of water through HgC12-sensitive aquaporins. We also measured the height of the microtubule structures to assess whether they are involved in the height increase. Our results show that pigment aggregation in X. laevis melanophores resulted in a significant height increase, which was substantially larger when aggregation was induced by latrunculin than with melatonin. Moreover, the elevation of the plasma membrane did not correlate with influx of water through aquaporins or formation of new microtubules, Rather, the accumulation of granules seemed to drive the change in cell height.
|
|
| 4. |
- Immerstrand, Charlotte, 1974-, et al.
(author)
-
Organelle transport in melanophores analyzed by white light image correlation spectroscopy
- 2007
-
In: Journal of Microscopy. - : Wiley. - 0022-2720 .- 1365-2818. ; 225:3, s. 275-282
-
Journal article (peer-reviewed)abstract
- Intracellular transport of organelles, vesicles and proteins is crucial in all eukaryotic cells, and is accomplished by motor proteins that move along cytoskeletal filaments. A widely used model of intracellular transport is Xenopus laevis melanophores. These cells help the frog to change color by redistributing melanin-containing organelles in the cytoplasm. The high contrast of the pigment organelles permits changes in distribution to be observed by ordinary light microscopy; other intracellular transport systems often require fluorescence labeling. Here we have developed white light Image Correlation Spectroscopy (ICS) to monitor aggregation and dispersion of pigment. Hitherto in ICS, images of fluorescent particles from Confocal Laser Scanning Microscopy (CLSM) have been used to calculate autocorrelation functions from which the density can be obtained. In the present study we show that ICS can be modified to enable analysis of light-microscopy images; it can be used to monitor pigment aggregation and dispersion, and distinguish between different stimuli. This new approach makes ICS applicable not only to fluorescent but also to black-and-white images from light or electron microscopy, and is thus very versatile in different studies of movement of particles on the membrane or in the cytoplasm of cells without potentially harmful fluorescence labeling and activation.
|
|
| 5. |
- Jager, Edwin W.H., et al.
(author)
-
The cell clinic : closable microvials for single cell studies
- 2002
-
In: Biomedical microdevices (Print). - 1387-2176 .- 1572-8781. ; 4:3, s. 177-187
-
Journal article (peer-reviewed)abstract
- We present the development of a cell clinic. This is a micromachined cavity, or microvial, that can be closed with a lid. The lid is activated by two polypyrrole/Au microactuators. Inside the microvials two Au electrodes have been placed in order to perform impedance studies on single or a small number of cells. We report on impedance measurements on Xenopus leavis melanophores. We could measure a change in the impedance upon cell spreading and identify intracellular events such as the aggregation of pigment granules. The electrical data is correlated to optical microscopy.
|
|
