Abstract

Astrocytes are responsible for a wide variety of essential functions throughout the central nervous system. The protein markers glial fibrillary acidic protein (GFAP), glutamate aspartate transporter (GLAST), glutamate transporter-1 (GLT-1), glutamine synthetase (GS), 10-formyltetrahydrofolate dehydrogenase (ALDH1L1), and the transcription factor SOX9 are routinely used to label astrocytes in primary rodent cultures. However, GLAST, GLT-1, GS, and SOX9 are also produced by microglia and oligodendrocytes and GFAP, GLAST, GLT-1, and GS production levels are affected by astrocyte phenotypic changes associated with reactive astrogliosis. No group has performed a comprehensive immunocytochemical evaluation to quantify the percentage of cells labeled by these markers in vitro, nor compared changes in staining between cortex- and spinal cord-derived cells in naïve and stimulated cultures. Here, we quantified the percentage of cells positively stained for these six markers in astrocyte, microglia, and oligodendrocyte cultures isolated from neonatal rat cortices and spinal cords. Additionally, we incubated the astrocytes with transforming growth factor (TGF)-β1 or TGF-β3 to determine if the labeling of these markers is altered by these stimuli. We found that only SOX9 in cortical cultures and ALDH1L1 in spinal cord cultures labeled more than 75% of the cells in naïve and stimulated astrocyte cultures and stained less than 5% of the cells in microglia and oligodendrocyte cultures. Furthermore, significantly more cortical than spinal cord astrocytes stained for GFAP, GLAST, and ALDH1L1 in naïve cultures, whereas significantly more spinal cord than cortical astrocytes stained for GLAST and GS in TGF-β1-treated cultures. These findings are important as variability in marker staining may lead to misinterpretation of the astrocyte response in cocultures, migration assays, or engineered disease models.

Department(s)

Chemical and Biochemical Engineering

Publication Status

Full Access

Comments

National Institute of Neurological Disorders and Stroke, Grant R01NS092754

Keywords and Phrases

astrocyte marker; cortical astrocytes; primary astrocyte culture; RRID:AB_10013382; RRID:AB_10712968; RRID:AB_141607; RRID:AB_177576; RRID:AB_2534074; RRID:AB_2534095; RRID:AB_259853; RRID:AB_2620155; RRID:AB_2715497; RRID:AB_304334; RRID:AB_490574; RRID:AB_941782; RRID:AB_94975; spinal cord astrocytes; TGF-β1; TGF-β3

International Standard Serial Number (ISSN)

1097-4547; 0360-4012

Document Type

Article - Journal

Document Version

Citation

File Type

text

Language(s)

English

Rights

© 2025 Wiley, All rights reserved.

Publication Date

01 Mar 2021

PubMed ID

33295039

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