Fig. 6
KCa3.1 involved in microglia SOCE and ER stress. a, b Representative images of GRP78, p-PERK, and p-eIF2α in KCa3.1−/− microglia, responses to 500 μM MPP+ (a) or 1 μM Tg (b) vs. WT cells. Mean values of GRP78, p-PERK, and p-eIF2α relative to β-actin. Data are presented as the mean ± SEM (n = 3). Western blot was repeated three times and showed similar results. *p < 0.05, **p < 0.01, unpaired, two-tailed Student’s t test. c Primary cultured microglia were treated with 500 μM MPP+ for 12 h with or without pretreatment of 1 μM senicapoc or 10 μM 2-APB. Fluorescence intensities of [Ca2+]i are shown. Fluorescence intensity was measured in the presence of 1 μM Tg with or without 2 mM Ca2+. Data are presented as the mean ± SEM (n = 10). ###p < 0.001 vs. control, ***p < 0.001 vs. MPP+-treated cells. One-way ANOVA followed by Dunnett’s multiple comparison test. d Western blot analysis of KCa3.1 and Orai1 expression after 500 μM MPP+-treatment for 3, 6, 12 h. Data represent the mean ± SEM of KCa3.1 and Orai1 density normalized to β-actin values for n = 3 cultures. *p < 0.05, **p < 0.01, ***p < 0.001, one-way ANOVA followed by Dunnett’s multiple comparison test compared with control. e–g Levels of the dendritic marker MAP2 were compared between neurons cocultured with KCa3.1−/− (e, f) or control microglia (g). e Neuron dendrites were immunostained with MAP2, and nuclei were stained with DAPI (blue). e, f Neuron cocultured with WT or KCa3.1−/− microglia treated with 500 μM MPP+ for 12 h. g Neuron cocultured with/without microglia treated with 500 μM MPP+ for 12 h with pretreatment of 1 μM senicapoc for 30 min. Cell body area, neurite length, and branch point counts were analyzed by extended neurite outgrowth bioapplication software. Data represent mean ± SEM (n = 3). *p < 0.05, **p < 0.01, ***p < 0.001. One-way ANOVA followed by Dunnett’s multiple comparison test. Tg, thapsigargin; Se, senicapoc