Cholesterol depletion and augmentation altered both phagocytosis and the release of Il1β, TNFα, and BDNF from microglia. Isolated rat microglial cultures were treated with 4 mM mβcd for 30 min, washed and allowed to rest 30 min, then treated with 1 mM hpβcd-cholesterol for 30 min, washed and allowed to rest 30 min. Cultures were activated with 1 μg/ml LPS overnight (18 h) and media was collected and assayed for release of Il1β (A), TNFα (B), and BDNF (C). All analyses were normalized to the total protein of the corresponding cell lysate to control for cell viability. Phagocytosis was assayed after cholesterol depletion (D-E) as in Figure 3. Dagger represents P <0.05 by Mann–Whitney U-test while asterisks represent P <0.05, 0.01, and 0.001 by 2-way ANOVA with Bonferroni’s post-hoc from N = 3 independent preparations.