Novel cell separation method for molecular analysis of neuron-astrocyte cocultures
Over the last decade, the importance of astrocyte-neuron communication in neuronal development and synaptic plasticity has become increasingly clear. Since neuron-astrocyte interactions represent highly dynamic and reciprocal processes, we hypothesized that many astrocyte genes may be regulated as a...
| Main Authors: | , , , , , |
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| Format: | Article |
| Language: | English |
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Frontiers Media S.A.
2014-01-01
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| Series: | Frontiers in Cellular Neuroscience |
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| Online Access: | http://journal.frontiersin.org/Journal/10.3389/fncel.2014.00012/full |
| _version_ | 1811337135415885824 |
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| author | Andrea eGoudriaan Nutabi eCamargo Karen eCarney Karen eCarney Karen eCarney Stéphane H.R. Oliet Stéphane H.R. Oliet August B. Smit Mark H.G. Verheijen |
| author_facet | Andrea eGoudriaan Nutabi eCamargo Karen eCarney Karen eCarney Karen eCarney Stéphane H.R. Oliet Stéphane H.R. Oliet August B. Smit Mark H.G. Verheijen |
| author_sort | Andrea eGoudriaan |
| collection | DOAJ |
| description | Over the last decade, the importance of astrocyte-neuron communication in neuronal development and synaptic plasticity has become increasingly clear. Since neuron-astrocyte interactions represent highly dynamic and reciprocal processes, we hypothesized that many astrocyte genes may be regulated as a consequence of their interactions with maturing neurons. In order to identify such neuron-responsive astrocyte genes in vitro, we sought to establish an expedite technique for separation of neurons from co-cultured astrocytes. Our newly established method makes use of cold jet, which exploits different adhesion characteristics of subpopulations of cells (Jirsova et al., 1997), and is rapid, performed under ice-cold conditions and avoids protease-mediated isolation of astrocytes or time-consuming centrifugation, yielding intact astrocyte mRNA with approximately 90% of neuronal RNA removed. Using this purification method, we executed genome-wide profiling in which RNA derived from astrocyte-only cultures was compared with astrocyte RNA derived from differentiating neuron-astrocyte co-cultures. Data analysis determined that many astrocytic mRNAs and biological processes are regulated by neuronal interaction. Our results validate the cold jet as an efficient method to separate astrocytes from neurons in co-culture, and reveals that neurons induce robust gene-expression changes in co-cultured astrocytes. |
| first_indexed | 2024-04-13T17:49:35Z |
| format | Article |
| id | doaj.art-553fee5688db40589e4ae50293e2c3a3 |
| institution | Directory Open Access Journal |
| issn | 1662-5102 |
| language | English |
| last_indexed | 2024-04-13T17:49:35Z |
| publishDate | 2014-01-01 |
| publisher | Frontiers Media S.A. |
| record_format | Article |
| series | Frontiers in Cellular Neuroscience |
| spelling | doaj.art-553fee5688db40589e4ae50293e2c3a32022-12-22T02:36:45ZengFrontiers Media S.A.Frontiers in Cellular Neuroscience1662-51022014-01-01810.3389/fncel.2014.0001268855Novel cell separation method for molecular analysis of neuron-astrocyte coculturesAndrea eGoudriaan0Nutabi eCamargo1Karen eCarney2Karen eCarney3Karen eCarney4Stéphane H.R. Oliet5Stéphane H.R. Oliet6August B. Smit7Mark H.G. Verheijen8CNCR, Neuroscience Campus Amsterdam, VU UniversityCNCR, Neuroscience Campus Amsterdam, VU UniversityCNCR, Neuroscience Campus Amsterdam, VU UniversityINSERM U862, Neurocentre MagendieUniversité de BordeauxINSERM U862, Neurocentre MagendieUniversité de BordeauxCNCR, Neuroscience Campus Amsterdam, VU UniversityCNCR, Neuroscience Campus Amsterdam, VU UniversityOver the last decade, the importance of astrocyte-neuron communication in neuronal development and synaptic plasticity has become increasingly clear. Since neuron-astrocyte interactions represent highly dynamic and reciprocal processes, we hypothesized that many astrocyte genes may be regulated as a consequence of their interactions with maturing neurons. In order to identify such neuron-responsive astrocyte genes in vitro, we sought to establish an expedite technique for separation of neurons from co-cultured astrocytes. Our newly established method makes use of cold jet, which exploits different adhesion characteristics of subpopulations of cells (Jirsova et al., 1997), and is rapid, performed under ice-cold conditions and avoids protease-mediated isolation of astrocytes or time-consuming centrifugation, yielding intact astrocyte mRNA with approximately 90% of neuronal RNA removed. Using this purification method, we executed genome-wide profiling in which RNA derived from astrocyte-only cultures was compared with astrocyte RNA derived from differentiating neuron-astrocyte co-cultures. Data analysis determined that many astrocytic mRNAs and biological processes are regulated by neuronal interaction. Our results validate the cold jet as an efficient method to separate astrocytes from neurons in co-culture, and reveals that neurons induce robust gene-expression changes in co-cultured astrocytes.http://journal.frontiersin.org/Journal/10.3389/fncel.2014.00012/fullAstrocytesTranscriptional ActivationMethodneuron-glia interactionisolation approachesco-culture |
| spellingShingle | Andrea eGoudriaan Nutabi eCamargo Karen eCarney Karen eCarney Karen eCarney Stéphane H.R. Oliet Stéphane H.R. Oliet August B. Smit Mark H.G. Verheijen Novel cell separation method for molecular analysis of neuron-astrocyte cocultures Frontiers in Cellular Neuroscience Astrocytes Transcriptional Activation Method neuron-glia interaction isolation approaches co-culture |
| title | Novel cell separation method for molecular analysis of neuron-astrocyte cocultures |
| title_full | Novel cell separation method for molecular analysis of neuron-astrocyte cocultures |
| title_fullStr | Novel cell separation method for molecular analysis of neuron-astrocyte cocultures |
| title_full_unstemmed | Novel cell separation method for molecular analysis of neuron-astrocyte cocultures |
| title_short | Novel cell separation method for molecular analysis of neuron-astrocyte cocultures |
| title_sort | novel cell separation method for molecular analysis of neuron astrocyte cocultures |
| topic | Astrocytes Transcriptional Activation Method neuron-glia interaction isolation approaches co-culture |
| url | http://journal.frontiersin.org/Journal/10.3389/fncel.2014.00012/full |
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