Figure 2

Blocking SK3 channels in microglia reduces their neurotoxic behavior. Microglia on Transwell™ inserts were incubated with lipopolysaccharide (LPS; 100 ng/ml, 24 h), with or without 100 nM apamin, 5 nM tamapin or 250 pM tamapin. The inserts were then washed to remove the drugs; thus, target neurons were never exposed to LPS or channel blockers. Each microglia-bearing insert was placed in a Transwell™ chamber above healthy neurons, incubated for 24 h (for caspase 3 activation) or 48 h (for TUNEL), and then the target neuron cultures were removed and assessed. Results are presented as mean±SEM for the number of separate cultures indicated on the bars. A. TUNEL-positive neuronal nuclei were counted and expressed as a percentage of all DAPI-stained nuclei. LPS-stimulated microglia killed more neurons than untreated microglia (^††p < 0.01), and killing was significantly reduced by treating the microglia with 100 nM apamin (*p < 0.05) or 5 nM tamapin (*p < 0.05), but not with 250 pM tamapin. B. Average caspase 3 activity in each neuron-containing well was measured with a fluorogenic substrate (Ac-DEVD-AMC) and the fluorescence plate reader. Treating microglia with either 100 nM apamin or 5 nM tamapin reduced caspase-3 activation in the target neurons (*p < 0.05); whereas, 250 pM tamapin treatment had no effect.