Protocol for in vitro analysis of pro-inflammatory and metabolic functions of cultured primary murine astrocytes

• Published in: STAR protocols Vol. 3; no. 1; p. 101033
• Main Authors: Gutiérrez-Vázquez, Cristina, Quintana, Francisco J.
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 18.03.2022; Elsevier
• Subjects: Animals; Astrocytes - metabolism; Brain - metabolism; Cell culture; Cell isolation; Immunology; Metabolism; Mice; Neuroscience; Oxygen Consumption; Protocol; Cell culture; Neuroscience; Immunology; Metabolism; Cell isolation
• ISSN: 2666-1667; 2666-1667
• DOI: 10.1016/j.xpro.2021.101033

Robust protocols are required to investigate in vitro the molecular mechanisms that control astrocyte metabolism and pro-inflammatory activities. In the present protocol, we describe step by step the isolation and culture of primary murine astrocytes from neonatal brains, followed by their genetic manipulation with siRNA. We further describe cytokine activation of the cultured astrocytes for the analysis of their pro-inflammatory responses, and the oxygen consumption analysis to assess their metabolic function. For complete details on the use and execution of this protocol, please refer to Chao et al. (2019), Clark et al. (2021), and Rothhammer et al. (2018). [Display omitted] •Primary murine astrocyte culture establishment•Effective knockdown studies on primary astrocytes in vitro•Stimulation with cytokines for downstream assays•Oxygen consumption analysis to assess metabolic function Robust protocols are required to investigate in vitro the molecular mechanisms that control astrocyte metabolism and pro-inflammatory activities. In the present protocol, we describe step by step the isolation and culture of primary murine astrocytes from neonatal brains, followed by their genetic manipulation with siRNA. We further describe cytokine activation of the cultured astrocytes for the analysis of their pro-inflammatory responses, and the oxygen consumption analysis to assess their metabolic function.