TetraCultures of Neurons, Astrocytes, Oligodendrocytes, and Microglia: A Murine In Vitro Model to Dissect Region-Specific CNS Immune Dynamics
Abstract
Neuroinflammation and glial scar formation in the central nervous system (CNS) emerge from coordinated signaling between neurons, astrocytes, oligodendrocytes and microglia. Conventional embryonic neuron-astrocyte-oligodendrocyte (NAO) TriCultures lack microglia and cannot model innate immune responses. Here, we introduce postnatal neuron-astrocyte-oligodendrocyte-microglia (NAOM) TetraCultures, in which resident microglia are retained from the original brain tissue and co-develop with other CNS cell populations from the onset of culture. Hippocampal and cortical cultures were maintained for 14 days in vitro and challenged with LPS, poly I or scratch injury. Only NAOM TetraCultures mounted robust, region- and stimulus-specific inflammatory responses. LPS induced marked TNF-αand IL-1βsecretion and increased IBA1 in hippocampal microglia, whereas poly I elicited IFN-βrelease without increasing IBA1. CD68 regulation differed between hippocampus and cortex, and microglial morphology showed pronounced amoeboid transitions after LPS. Astrocytes in NAOM TetraCultures expanded their territorial coverage after LPS, an effect absent in NAO TriCultures. Following scratch injury, IL-1βincreased, microglia infiltrated lesion zones and astrocytes formed lesion borders. Thus, NAOM TetraCultures recapitulate key features of CNS immunity, glial scar formation and region-specific glial responses while providing a physiologically relevant microglia-containing model.
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BibTeX
@misc{gabelekutsche2026tetracultures,
title = {{TetraCultures of Neurons, Astrocytes, Oligodendrocytes, and Microglia: A Murine In Vitro Model to Dissect Region-Specific CNS Immune Dynamics}},
author = {Lea Gabele-Kutsche and Yaning Zhao and Tabea Spengler and Kay Michael Gottlieb and Christian Deppe and Martin Korte and Kristin Michaelsen-Preusse},
howpublished = {arXiv},
year = {2026},
month = oct,
doi = {10.48550/arxiv.2610.16279},
eprint = {2610.16279},
archivePrefix = {arXiv},
url = {https://arxiv.org/abs/2610.16279},
}
RIS
TY - UNPB
TI - TetraCultures of Neurons, Astrocytes, Oligodendrocytes, and Microglia: A Murine In Vitro Model to Dissect Region-Specific CNS Immune Dynamics
AU - Lea Gabele-Kutsche
AU - Yaning Zhao
AU - Tabea Spengler
AU - Kay Michael Gottlieb
AU - Christian Deppe
AU - Martin Korte
AU - Kristin Michaelsen-Preusse
PY - 2026
DA - 2026/10/12
PB - arXiv
DO - 10.48550/arxiv.2610.16279
UR - https://arxiv.org/abs/2610.16279
AB - Neuroinflammation and glial scar formation in the central nervous system (CNS) emerge from coordinated signaling between neurons, astrocytes, oligodendrocytes and microglia. Conventional embryonic neuron-astrocyte-oligodendrocyte (NAO) TriCultures lack microglia and cannot model innate immune responses. Here, we introduce postnatal neuron-astrocyte-oligodendrocyte-microglia (NAOM) TetraCultures, in which resident microglia are retained from the original brain tissue and co-develop with other CNS cell populations from the onset of culture. Hippocampal and cortical cultures were maintained for 14 days in vitro and challenged with LPS, poly I or scratch injury. Only NAOM TetraCultures mounted robust, region- and stimulus-specific inflammatory responses. LPS induced marked TNF-αand IL-1βsecretion and increased IBA1 in hippocampal microglia, whereas poly I elicited IFN-βrelease without increasing IBA1. CD68 regulation differed between hippocampus and cortex, and microglial morphology showed pronounced amoeboid transitions after LPS. Astrocytes in NAOM TetraCultures expanded their territorial coverage after LPS, an effect absent in NAO TriCultures. Following scratch injury, IL-1βincreased, microglia infiltrated lesion zones and astrocytes formed lesion borders. Thus, NAOM TetraCultures recapitulate key features of CNS immunity, glial scar formation and region-specific glial responses while providing a physiologically relevant microglia-containing model.
ER -