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Melanophores: cell biophysics and sensor applications
Linköping University, Department of Biomedical Engineering. Linköping University, The Institute of Technology.
2001 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

This thesis deals with studies of melanophores and melanosomes by means of a physical approach. Melanophores are pigment cells that give the black colour to many vertebrates, e.g. fishes, frogs, and reptiles. Covering large areas of the skin, these cells are approximately 0.1 mm in diameter, and they contain black pigment granules called melanosomes.

The geometry and the electric charge of isolated melanosomes were two physical properties that were studied. The electric charge was measured by electrophoresis and was found to be (-1.7 ± 0.2)·10-16 Coulomb in average. The geometry of melanosomes was measured using Scanning Force Microscopy, SFM, and resulted in an ellipsoidal shape with an average major diameter of 840 ± 20 nm.

Under nervous and hormonal control, melanophores rearrange the intracellular melanosomes from a scattered distribution, called dispersed, to a state where all melanosomes are accumulated in the cell centre, called the aggregated state. In this way, melanophores change from black towards transparent. This gives an animal the ability to change not only between being pale or dark, but also between different colours by using melanophores to cover and uncover the colours of different types of pigment cells from underlying layers.

The volume of melanosomes was measured with SFM. This study resulted in a difference of 18% when individual melanosomes from aggregated and dispersed melanophores were measured separately.

Magnetic field exposure of melanophores has been reported to affect the aggregation. However, contradicting results are presented in the literature. To clarify the possible effect of magnetic fields on melanophores, experiments by aggregating fish melanophores under exposure to strong (8 and 14 Tesla), homogenous, static magnetic fields were carried out. Both the magnetic field-induced Lorentz force on the charged melanosomes and the reorientation of the cytoskeleton were considered as possible explanations of any effects. Whenfield experiments were compared to control experiments with zero field, no difference in aggregation levels were found. However, a more irregular speed of aggregation was seen in the 8 Tesla field than in the control experiments.

A theoretical model was developed to explain switch-like responses in biological systems. A switch-like response to a graded stimulation was sometimes seen in the case of melanophores but was shown not to have a very large so-called Hill coefficient. The model is simple in its approach. It may be applied to general phenomena and is based on the assumption of a simultaneous desorption of an activator (agonist; substrate molecule; ... ) and an inactivator (antagonist, inhibitor; ... ) caused by a collision or interaction between two effector molecules (e.g. receptors or enzymes).

Melanosomes are also found in the human body and have a remarkably capacity to bind other molecules. A well-established forensic application of this is to detect (illegal) drugs that have bound to melanosomes in hair shafts. So far this application is only qualitative. This thesis includes a characterisation of the binding of flunitrazepam to melanin. Flunitrazepam is the active substance of Rohypnol, which is a sedative that is illegal in several countries and sometimes called the "date-rape-drug".

Melanophores are excellent model systems for studies of cellular phenomena. Moreover, melanophores are also interesting in sensing aspects. The change from black to transparent is a highly visible response to substances in their surroundings and has previously been the measurand in melanophore-based biosensors. The physical approach of these studies of melanophores also had the objective of evaluating new biosensor solutions.

Place, publisher, year, edition, pages
Linköping: Linköpings universitet , 2001. , 58 p.
Series
Linköping Studies in Science and Technology. Dissertations, ISSN 0345-7524 ; 687
National Category
Medical and Health Sciences
Identifiers
URN: urn:nbn:se:liu:diva-29443Local ID: 14789ISBN: 91-7373-015-7 (print)OAI: oai:DiVA.org:liu-29443DiVA: diva2:250258
Public defence
2001-05-11, Föreläsningssal Elsa Brändström, Universitetssjukhuset, Linköping, 09:15 (English)
Opponent
Supervisors
Available from: 2009-10-09 Created: 2009-10-09 Last updated: 2013-02-22
List of papers
1. The electric charge of pigment granules in pigment cells
Open this publication in new window or tab >>The electric charge of pigment granules in pigment cells
2001 (English)In: Biosensors & bioelectronics, ISSN 0956-5663, E-ISSN 1873-4235, Vol. 16, no 1-2, 31-36 p.Article in journal (Refereed) Published
Abstract [en]

Black pigment cells called melanophores change colour in response to environmental changes and have lately been studied as promising biosensors. To further elucidate the intracellular processes involved in the colour changes of these cells, and to find optimal biosensing principles, the electric charge of intracellular pigment granules, melanosomes, has been determined in vitro by electrophoresis. Melanosomes from the two extreme states in the cell colour change (aggregated and dispersed melanosomes) were measured. The charge was found to be −1.5·10−16 and −1.7·10−16 C, aggregated and dispersed melanosomes, respectively, without significant difference between the two conditions. This charge is of the same order of magnitude as the one of 1000 electrons. The origin of the melanosome charge, and the use of these findings in new biosensor principles, is discussed.

National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:liu:diva-32560 (URN)10.1016/S0956-5663(00)00130-5 (DOI)18473 (Local ID)18473 (Archive number)18473 (OAI)
Available from: 2009-10-09 Created: 2009-10-09 Last updated: 2013-02-22
2. Melanophore aggregation in strong static magnetic fields
Open this publication in new window or tab >>Melanophore aggregation in strong static magnetic fields
2002 (English)In: Bioelectromagnetics, ISSN 0197-8462, E-ISSN 1521-186X, Vol. 23, no 6, 444-449 p.Article in journal (Refereed) Published
Abstract [en]

Contradicting results can be found in the literature on effects from magnetic exposure to pigment cells. We have studied the influence of strong, static, homogenous magnetic fields of 8 and 14 T on melanophore aggregation during exposure to the field. Melanophores, black pigment cells, in fish are large flat cells having intracellular black pigment granules. Due to large size, high optical contrast, and quick response to drugs, melanophores are attractive as biosensors as well as for model studies of intracellular processes. This is especially true for modeling nerve cells, since melanophores share stem cells with axons. Twenty experiments on black tetra fish fins were carried out in the two magnetic flux densities. The same number of control experiments were carried out in the magnet with the magnetic field turned off. Several factors, such as the degree of maximal aggregation, speed of aggregation, and irregularity of the speed, were examined. The statistical analysis showed no significant field effects on the aggregation, with one exception: the irregularity in aggregation speed in the 8 T field, compared to control. The believed reorientation of the cytoskeleton (microtubules) in the field or the induced Lorentz force on moving pigment granules, did not affect the aggregation.

Keyword
Pigment Cell, DC field, Lorentz force, Melanosome, Nerve stem cell, Pigment cell
National Category
Engineering and Technology
Identifiers
urn:nbn:se:liu:diva-46794 (URN)10.1002/bem.10040 (DOI)
Available from: 2009-10-11 Created: 2009-10-11 Last updated: 2013-02-22
3. Volume changes of individual melanosomes measured by scanning force microscopy
Open this publication in new window or tab >>Volume changes of individual melanosomes measured by scanning force microscopy
2001 (English)In: Pigment Cell Research, ISSN 0893-5785, E-ISSN 1600-0749, Vol. 14, no 6, 445-449 p.Article in journal (Refereed) Published
Abstract [en]

Black pigment cells, melanophores, e.g. located in the epidermis and dermis of frogs, are large flat cells having intracellular black pigment granules, called melanosomes. Due to a large size, high optical contrast, and quick response to drugs, melanophores are attractive as biosensors as well as for model studies of intracellular processes; e.g. organelle transport and G-protein coupled receptors. The geometry of melanosomes from African clawed toad, Xenopus laevis, has been measured using scanning force microscopy (SFM). Three-dimensional images from SFM were used to measure height, width, and length of the melanosomes (100 from aggregated cells and 100 from dispersed cells). The volumes of melanosomes isolated from aggregated and dispersed melanophores were significantly different (P<0.05, n=200). The average ellipsoidal volume was 0.14±0.01 (aggregated) and 0.17±0.01 μm3 (dispersed), a difference of 18%. The average major diameter was 810±20 and 880±20 nm for aggregated and dispersed melanosomes, respectively. To our knowledge, this is the first time SFM has been used to study melanosomes. This may provide an alternative non-destructive technique that may be particularly suitable for studying morphological aspects of various melanin granules.

National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:liu:diva-26717 (URN)10.1034/j.1600-0749.2001.140604.x (DOI)11311 (Local ID)11311 (Archive number)11311 (OAI)
Available from: 2009-10-08 Created: 2009-10-08 Last updated: 2013-02-22
4. Characterization of [3H]flunitrazepam binding to melanin
Open this publication in new window or tab >>Characterization of [3H]flunitrazepam binding to melanin
Show others...
2001 (English)In: Analytical Biochemistry, ISSN 0003-2697, E-ISSN 1096-0309, Vol. 298, no 2, 259-264 p.Article in journal (Refereed) Published
Abstract [en]

In both clinical and forensic toxicology, the analysis of hair for drugs is an important tool to determine drug use in the past or to verify abstinence from illegal drugs during extended periods. Melanin is proposed as one of the factors that influences drug incorporation to hair and we have characterized the binding of the drug flunitrazepam to melanin in vitro. The drug was 3H labeled and melanin granules from cuttlefish, Sepia officinalis, were used according to the suggested standard for melanin studies. We observed a rapid Langmuir-like binding followed by a slower diffusion-limited binding that may be interpreted as an initial surface binding followed by deeper bulk binding. From three concentrations of melanin, with a 60-min incubation time, a mean saturation value of 180 ± 20 pmol/mg was calculated. The binding of a group of benzodiazepines and tranquilizers was compared to the binding of [3H]flunitrazepam by means of displacement experiments. These drugs showed binding characteristics similar to [3H]flunitrazepam except phenobarbital, which had a lower affinity to melanin. The method presented in this study allowed measurements with low melanin and drug concentrations and it has the strength of directly measuring the amount of drug bound to melanin, in contrast to previous indirect methods.

Keyword
Affinity, Benzodiazepines, Binding, Hair, Melanin
National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:liu:diva-47156 (URN)10.1006/abio.2001.5364 (DOI)
Available from: 2009-10-11 Created: 2009-10-11 Last updated: 2013-02-22Bibliographically approved
5. A model for switch-like phenomena in biological systems
Open this publication in new window or tab >>A model for switch-like phenomena in biological systems
Show others...
2001 (English)In: Biophysical Chemistry, ISSN 0301-4622, E-ISSN 1873-4200, Vol. 94, no 1-2, 1-9 p.Article in journal (Refereed) Published
Abstract [en]

We present a model for the activity of protein clusters based on a simultaneous desorption of an activator (agonist, substrate molecule, etc.) and an inactivator (antagonist, inhibitor, etc.) caused by the collision or interaction between two effector molecules (e.g. receptors, enzymes). This model gives rise to switch-like dose–response curves, which are difficult to explain by ordinary co-operativity. It fits with recent experimental results obtained on single cells. Some other interesting aspects of the model are also pointed out. The model is similar to the model used to explain steep ‘dose–response curves’ in heterogeneous catalysis, caused by the reaction between two different molecules or atoms on the surface of the catalyst.

National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:liu:diva-26718 (URN)10.1016/S0301-4622(01)00231-9 (DOI)11312 (Local ID)11312 (Archive number)11312 (OAI)
Available from: 2009-10-08 Created: 2009-10-08 Last updated: 2015-09-18

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