Do Nanoparticles Affect Neuronal Function? - Establishment of a Microelectrode-Based Assay
(2020) KBK820 20151Pure and Applied Biochemistry
- Abstract
- Nanomaterials are used extensively in industry and daily life but comparatively little is known about the possible health effects. Silver nanoparticles (AgNPs) and gold nanoparticles (AuNPs) are two metal nanoparticles that have been intensively studied. AgNPs due to their antibacterial effect and AuNPs due to good intrinsic properties and the fact that they are highly biocompatible. Extracellular recordings of spontaneous electrical activity from in vitro cultured neurons using microelectrode arrays (MEAs) can be used in order to study if AgNPs and AuNPs affect neuronal function. Neuronal cell cultures were exposed to AgNPs and AuNPs with a diameter of either 20 nm or 80 nm in low concentrations (800 particles/cell). To this date,... (More)
- Nanomaterials are used extensively in industry and daily life but comparatively little is known about the possible health effects. Silver nanoparticles (AgNPs) and gold nanoparticles (AuNPs) are two metal nanoparticles that have been intensively studied. AgNPs due to their antibacterial effect and AuNPs due to good intrinsic properties and the fact that they are highly biocompatible. Extracellular recordings of spontaneous electrical activity from in vitro cultured neurons using microelectrode arrays (MEAs) can be used in order to study if AgNPs and AuNPs affect neuronal function. Neuronal cell cultures were exposed to AgNPs and AuNPs with a diameter of either 20 nm or 80 nm in low concentrations (800 particles/cell). To this date, neurotoxicity assessment still relies on in vivo animal testing. However, human induced pluripotent stem cell (hiPSC)-derived neurons (iCell® Neurons commercially) were used in this thesis since animal experiments have several drawbacks and do not always mimic the human physiology. Immunocytochemical analysis of the iCell® Neurons as well as images taken with a scanning electron microscope concludes that the cells develop into functional neuronal networks and attach to electrodes on the MEAs. This provides a good basis for electrophysiological recordings. Significant multiunit extracellular activity associated with spontaneous action potential waveforms was recorded from the iCell® Neurons with the setup presented in this thesis. However, no quantitative analysis of how AgNPs and AuNPs affect the neuronal function was carried out due to a lack of experimental data. Only 10% of the electrodes on the MEAs recorded neuronal activity and further studies are needed in order to develop a fully functioning assay. Nevertheless, hiPSC-derived neurons cultured on MEAs may still enable animal free neurotoxicity testing in the future and could be a valuable tool when studying possible health effects of nanoparticles and other compounds. (Less)
- Popular Abstract
- Despite the advances in nanotechnologies and the abundance of nanoparticles in commercial applications comparatively little is known about possible health effects. If nanoparticles of gold and silver affect the function of neurons is still a question that remains to be answered.
Nanoparticles are objects that are in the range of 1-100 nm in at least one dimension. Nanoparticles are so small that they can enter the human body through a variety of different ways and they are also able to cross the blood-brain barrier and reach the central nervous system (CNS). This could introduce new unwanted toxic effects. Gold and silver nanoparticles are two very common and intensively studied nanoparticles. Silver nanoparticles, due to their... (More) - Despite the advances in nanotechnologies and the abundance of nanoparticles in commercial applications comparatively little is known about possible health effects. If nanoparticles of gold and silver affect the function of neurons is still a question that remains to be answered.
Nanoparticles are objects that are in the range of 1-100 nm in at least one dimension. Nanoparticles are so small that they can enter the human body through a variety of different ways and they are also able to cross the blood-brain barrier and reach the central nervous system (CNS). This could introduce new unwanted toxic effects. Gold and silver nanoparticles are two very common and intensively studied nanoparticles. Silver nanoparticles, due to their antibacterial effect, are used in e.g. cosmetics, textiles and household goods whereas gold nanoparticles are used to create new drug and gene delivery applications.
The CNS consists of the brain and spinal cord where neurons and glial cells are the basic cell types. Glial cells provide support and protection for neurons. The neurons in the central nervous system, however, control all major functions of the human body by processing and transmitting information with a specific type of electric signal, an action potential.
Action potential waveforms can be recorded and studied by culturing neuronal cells directly on top of microelectrodes. When electrical activity occurs, ions flow across the cell membrane. The moving ions generate changes in the electric field which in turn can be recorded by the metal microelectrodes. Arranging several microelectrodes in an array creates a microelectrode array (MEA). MEAs enable simultaneous recording of extracellular activity from a large number of neurons. The MEAs used in this thesis to record neuronal activity before and after exposure to nanoparticles have 60 microelectrodes with a diameter of 10 µm.
To this date, neurotoxicity assessment still relies on animal experiments. However, human cells were used in this thesis since animal tests are expensive, time consuming and ethically debatable. Human neural progenitor cells (hNPCs) was the first cell type that was tested. hNPCs are neural stem cells taken from a human embryo and these cells can differentiate into cell types that populate the CNS. No significant neuronal activity was recorded from the hNPCs with the MEA plates. Therefore, human induced pluripotent stem cell (hiPSC)-derived neurons (iCell® Neurons commercially) were used instead. hiPSCs are generated from adult human cells. The cells have been reprogrammed into stem cells that in turn can develop into any cell type in the human body, in this case neurons. Significant electrical activity was successfully recorded from the iCell® Neurons. Action
potential waveforms from several different neurons were able to be recorded simultaneously with the MEAs. For instance, one microelectrode recorded activity from four different neurons at the same time. However, only 10% of the microelectrodes on the MEAs showed signs of neuronal activity. This means that no quantitative analysis of how gold and silver nanoparticles affect neuronal function could be carried out. Further studies are needed in order to improve the experimental setup. In conclusion, MEA recordings of hiPSC-derived neurons do show a lot of promise and may enable a new approach to efficient animal free
neurotoxicity testing in the future and might very well turn out to be a valuable tool when investigating nanoparticles and other compounds. (Less)
Please use this url to cite or link to this publication:
http://lup.lub.lu.se/student-papers/record/9009035
- author
- Nylander, Markus LU
- supervisor
- organization
- course
- KBK820 20151
- year
- 2020
- type
- H2 - Master's Degree (Two Years)
- subject
- keywords
- Microelectrode array, gold nanoparticles, silver nanoparticles, immunocytochemistry, scanning electron microscopy, human neural progenitor cells, human induced pluripotent stem cells, applied biochemistry, tillämpad biokemi
- language
- English
- id
- 9009035
- date added to LUP
- 2020-05-26 07:23:51
- date last changed
- 2020-05-26 07:23:51
@misc{9009035,
abstract = {{Nanomaterials are used extensively in industry and daily life but comparatively little is known about the possible health effects. Silver nanoparticles (AgNPs) and gold nanoparticles (AuNPs) are two metal nanoparticles that have been intensively studied. AgNPs due to their antibacterial effect and AuNPs due to good intrinsic properties and the fact that they are highly biocompatible. Extracellular recordings of spontaneous electrical activity from in vitro cultured neurons using microelectrode arrays (MEAs) can be used in order to study if AgNPs and AuNPs affect neuronal function. Neuronal cell cultures were exposed to AgNPs and AuNPs with a diameter of either 20 nm or 80 nm in low concentrations (800 particles/cell). To this date, neurotoxicity assessment still relies on in vivo animal testing. However, human induced pluripotent stem cell (hiPSC)-derived neurons (iCell® Neurons commercially) were used in this thesis since animal experiments have several drawbacks and do not always mimic the human physiology. Immunocytochemical analysis of the iCell® Neurons as well as images taken with a scanning electron microscope concludes that the cells develop into functional neuronal networks and attach to electrodes on the MEAs. This provides a good basis for electrophysiological recordings. Significant multiunit extracellular activity associated with spontaneous action potential waveforms was recorded from the iCell® Neurons with the setup presented in this thesis. However, no quantitative analysis of how AgNPs and AuNPs affect the neuronal function was carried out due to a lack of experimental data. Only 10% of the electrodes on the MEAs recorded neuronal activity and further studies are needed in order to develop a fully functioning assay. Nevertheless, hiPSC-derived neurons cultured on MEAs may still enable animal free neurotoxicity testing in the future and could be a valuable tool when studying possible health effects of nanoparticles and other compounds.}},
author = {{Nylander, Markus}},
language = {{eng}},
note = {{Student Paper}},
title = {{Do Nanoparticles Affect Neuronal Function? - Establishment of a Microelectrode-Based Assay}},
year = {{2020}},
}