Figure 3

Migration and chemotaxis are affected by the microglial activation state. Cells were untreated (control) or exposed to 10 ng/ml lipopolysaccharide (LPS) or 20 ng/ml IL4. A) Migration into a scratch wound in a monolayer of microglia. Cells were fixed and stained after 24 hr with the microglial marker, FITC-conjugated tomato lectin (green), and the nuclear marker, DAPI (blue). Scale bar, 100 μm. A′) For each slide, confocal images of five random fields were taken along the border of the scratch (delineated by dashed red lines), and all lectin-positive cells within the scratch region were counted. B) Transmigration of microglia in Transwell™ chambers. After each 24-hr treatment, cells that had migrated to the underside of each filter were counted in five random fields. C) Chemotactic response to 300 μM ATP added to the lower Transwell™ chamber (striped bars) compared with unstimulated transmigration (solid bars). Results were analyzed as in panel B, and then normalized to control transmigration without ATP. Results are reported as mean ± SEM, with the number of individual cultures indicated on each bar. Statistical differences were determined using either one-way ANOVA with Tukey’s post-hoc test (A, B) or two-way ANOVA with Bonferroni post-hoc test (C). # indicates an effect of ATP; * (A, B) or † (C) indicates treatment differences (control versus LPS or IL4). One symbol, P <0.05; two symbols, P <0.01; three symbols, P <0.001.