Concomitant Isolation of Primary Astrocytes and Microglia for Protozoa Parasite Infection

• Published in: Journal of visualized experiments no. 157
• Main Authors: Pacheco, Aline de Oliveira Lima, Amaral, Marcelo Pires, de Farias, Ingrid Sancho, Bottino, Luiza Zainotti Miguel Fahur, Bortoluci, Karina Ramalho
• Format: Journal Article
• Language: English
• Published: United States 18.03.2020
• ISSN: 1940-087X; 1940-087X
• DOI: 10.3791/60680

Astrocytes and microglia are the most abundant glial cells. They are responsible for physiological support and homeostasis maintenance in the central nervous system (CNS). The increasing evidences of their involvement in the control of infectious diseases justify the emerging interest in the improvement of methodologies to isolate primary astrocytes and microglia in order to evaluate their responses to infections that affect the CNS. Considering the impact of Trypanosoma cruzi (T. cruzi) and Toxoplasma gondii (T. gondii) infection in the CNS, here we provide a method to extract, maintain, dissociate and infect murine astrocytes and microglia cells with protozoa parasites. Extracted cells from newborn cortices are maintained in vitro for 14 days with periodic differential media replacement. Astrocytes and microglia are obtained from the same extraction protocol by mechanical dissociation. After phenotyping by flow cytometry, cells are infected with protozoa parasites. The infection rate is determined by fluorescence microscopy at different time points, thus enabling the evaluation of differential ability of glial cells to control protozoan invasion and replication. These techniques represent simple, cheap and efficient methods to study the responses of astrocytes and microglia to infections, opening the field for further neuroimmunology analysis.