Immunohistochemical localization of STIM1&2 in cultured hippocampal neurons with and without extracellular calcium. (A,B) Sample dendrites were taken from 10- to 20-day-old cultures and stained for STIM1(red) and STIM2 (blue) in cells transfected with EGFP (green) to visualize morphology in the presence and absence of extracellular calcium. It is apparent that the 10-day-old culture contains more STIM1 than STIM2 puncta, and the opposite is seen in the 20-day-old neuron. Under the calcium-free condition, STIM2 flows into protrusions in young and, especially, old culture. (C) Bar graphs quantification of the results illustrated on the left. The difference between STIM1 and 2 in 10 DIV in both conditions is highly significant (control conditions: n = 10 dendrites from five cells for each group, ANOVA p < 0.0001; in calcium-free medium: n = 10 dendrites from five cells for each group, ANOVA p < 0.0001). (D) Bar graphs quantification of the results illustrated on the right. The difference between STIM1 and 2 in 20 DIV in both conditions is highly significant (control conditions: n = 10 dendrites from five cells for each group, ANOVA p < 0.0002; in calcium-free medium: n = 10 dendrites from five cells for each group, ANOVA p < 0.0001). *Significant, 0.05 > p > 0.01; **very significant, 0.01 > p > 0.001; ***highly significant, p < 0.001.