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Adjusting the neuron to astrocyte ratio with cytostatics in hippocampal cell cultures from postnatal rats: A comparison of cytarabino furanoside (AraC) and 5-fluoro-2’-deoxyuridine (FUdR)

Cell culture studies offer the unique possibility to investigate the influence of pharmacological treatments with quantified dosages applied for defined time durations on survival, morphological maturation, protein expression and function as well as the mutual interaction of various cell types. Cult...

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Autores principales: Lesslich, Heiko M., Klapal, Lars, Wilke, Justus, Haak, Annika, Dietzel, Irmgard D.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Public Library of Science 2022
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8906639/
https://www.ncbi.nlm.nih.gov/pubmed/35263366
http://dx.doi.org/10.1371/journal.pone.0265084
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author Lesslich, Heiko M.
Klapal, Lars
Wilke, Justus
Haak, Annika
Dietzel, Irmgard D.
author_facet Lesslich, Heiko M.
Klapal, Lars
Wilke, Justus
Haak, Annika
Dietzel, Irmgard D.
author_sort Lesslich, Heiko M.
collection PubMed
description Cell culture studies offer the unique possibility to investigate the influence of pharmacological treatments with quantified dosages applied for defined time durations on survival, morphological maturation, protein expression and function as well as the mutual interaction of various cell types. Cultures obtained from postnatal rat brain contain a substantial number of glial cells that further proliferate with time in culture leading to an overgrowth of neurons with glia, especially astrocytes and microglia. A well-established method to decrease glial proliferation in vitro is to apply low concentrations of cytosine arabinoside (AraC). While AraC primarily effects dividing cells, it has been reported repeatedly that it is also neurotoxic, which is the reason why most protocols limit its application to concentrations of up to 5 μM for a duration of 24 h. Here, we investigated 5-fluoro-2’-deoxyuridine (FUdR) as a possible substitute for AraC. We applied concentrations of both cytostatics ranging from 4 μM to 75 μM and compared cell composition and cell viability in cultures prepared from 0-2- and 3-4-day old rat pups. Using FUdR as proliferation inhibitor, higher ratios of neurons to glia cells were obtained with a maximal neuron to astrocyte ratio of up to 10:1, which could not be obtained using AraC in postnatal cultures. Patch-clamp recordings revealed no difference in the amplitudes of voltage-gated Na(+) currents in neurons treated with FUdR compared with untreated control cells suggesting replacement of AraC by FUdR as glia proliferation inhibitor if highly neuron-enriched postnatal cultures are desired.
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spelling pubmed-89066392022-03-10 Adjusting the neuron to astrocyte ratio with cytostatics in hippocampal cell cultures from postnatal rats: A comparison of cytarabino furanoside (AraC) and 5-fluoro-2’-deoxyuridine (FUdR) Lesslich, Heiko M. Klapal, Lars Wilke, Justus Haak, Annika Dietzel, Irmgard D. PLoS One Research Article Cell culture studies offer the unique possibility to investigate the influence of pharmacological treatments with quantified dosages applied for defined time durations on survival, morphological maturation, protein expression and function as well as the mutual interaction of various cell types. Cultures obtained from postnatal rat brain contain a substantial number of glial cells that further proliferate with time in culture leading to an overgrowth of neurons with glia, especially astrocytes and microglia. A well-established method to decrease glial proliferation in vitro is to apply low concentrations of cytosine arabinoside (AraC). While AraC primarily effects dividing cells, it has been reported repeatedly that it is also neurotoxic, which is the reason why most protocols limit its application to concentrations of up to 5 μM for a duration of 24 h. Here, we investigated 5-fluoro-2’-deoxyuridine (FUdR) as a possible substitute for AraC. We applied concentrations of both cytostatics ranging from 4 μM to 75 μM and compared cell composition and cell viability in cultures prepared from 0-2- and 3-4-day old rat pups. Using FUdR as proliferation inhibitor, higher ratios of neurons to glia cells were obtained with a maximal neuron to astrocyte ratio of up to 10:1, which could not be obtained using AraC in postnatal cultures. Patch-clamp recordings revealed no difference in the amplitudes of voltage-gated Na(+) currents in neurons treated with FUdR compared with untreated control cells suggesting replacement of AraC by FUdR as glia proliferation inhibitor if highly neuron-enriched postnatal cultures are desired. Public Library of Science 2022-03-09 /pmc/articles/PMC8906639/ /pubmed/35263366 http://dx.doi.org/10.1371/journal.pone.0265084 Text en © 2022 Lesslich et al https://creativecommons.org/licenses/by/4.0/This is an open access article distributed under the terms of the Creative Commons Attribution License (https://creativecommons.org/licenses/by/4.0/) , which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
spellingShingle Research Article
Lesslich, Heiko M.
Klapal, Lars
Wilke, Justus
Haak, Annika
Dietzel, Irmgard D.
Adjusting the neuron to astrocyte ratio with cytostatics in hippocampal cell cultures from postnatal rats: A comparison of cytarabino furanoside (AraC) and 5-fluoro-2’-deoxyuridine (FUdR)
title Adjusting the neuron to astrocyte ratio with cytostatics in hippocampal cell cultures from postnatal rats: A comparison of cytarabino furanoside (AraC) and 5-fluoro-2’-deoxyuridine (FUdR)
title_full Adjusting the neuron to astrocyte ratio with cytostatics in hippocampal cell cultures from postnatal rats: A comparison of cytarabino furanoside (AraC) and 5-fluoro-2’-deoxyuridine (FUdR)
title_fullStr Adjusting the neuron to astrocyte ratio with cytostatics in hippocampal cell cultures from postnatal rats: A comparison of cytarabino furanoside (AraC) and 5-fluoro-2’-deoxyuridine (FUdR)
title_full_unstemmed Adjusting the neuron to astrocyte ratio with cytostatics in hippocampal cell cultures from postnatal rats: A comparison of cytarabino furanoside (AraC) and 5-fluoro-2’-deoxyuridine (FUdR)
title_short Adjusting the neuron to astrocyte ratio with cytostatics in hippocampal cell cultures from postnatal rats: A comparison of cytarabino furanoside (AraC) and 5-fluoro-2’-deoxyuridine (FUdR)
title_sort adjusting the neuron to astrocyte ratio with cytostatics in hippocampal cell cultures from postnatal rats: a comparison of cytarabino furanoside (arac) and 5-fluoro-2’-deoxyuridine (fudr)
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8906639/
https://www.ncbi.nlm.nih.gov/pubmed/35263366
http://dx.doi.org/10.1371/journal.pone.0265084
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