Microglia were purified from the brains of newborn (day 0) Lewis rats. Brains were stripped of their meninges and minced with scissors under a microscope (Stemi DV4; Zeiss) in medium (Leibovitz-15; Biological Industries). After trypsinization (0.5% trypsin, 10 minutes, 37°C, 5% CO2), the tissue was triturated. The cell suspension was washed in culture medium for glial cells (DMEM supplemented with 10% FCS, l-glutamine [1 mM], sodium pyruvate [1 mM], penicillin [100 U/mL], and streptomycin [100 mg/mL]) and was cultured at 37°C, 5% CO2 in 75-cm2 tissue-culture flasks (Falcon; BD Biosciences, San Diego, CA) coated with poly-d-lysine (PDL; 10 mg/mL; Sigma-Aldrich). Half the medium was changed after 6 hours in culture and every second day thereafter starting on day 2, for a total culture time of 10 to 14 days. Microglia were shaken off the primary mixed brain glial cell cultures (150 rpm, 37°C, 4 hours) with maximum yields between days 10 and 14. They were then seeded on a PDL-coated culture flask (1 hour, 37°C, 5% CO2), and nonadherent cells were rinsed off and grown in culture medium for microglia (RPMI-1640 medium [Sigma-Aldrich] supplemented with 10% FCS, l-glutamine [1 mM], sodium pyruvate [1 mM], β-mercaptoethanol [50 mM], penicillin [100 U/mL], and streptomycin [100 mg/mL]).