Fig. 1

[Ca²⁺]_i either low and rising or high and falling is permissive for filopodia. Cells were treated with 5 µM CPA or 1.5 µM EGTA in Ca²⁺-free HBSS to deplete the ER of Ca²⁺. Results in panels A-C and E-F represent 3–7 experiments. (A, B) ER depletion followed by CPA washout and readdition of extracellular Ca²⁺. At various times, samples were collected and the fraction of cells with filopodia and percent coverage of the perimeter determined. a Significance by ANOVA, P = 0.0017. *Treatments differ at P = 0.0015. b Significance by ANOVA, P = 0.0030. **Treatments differ at P = 0.0028. c Fluorescence intensity of groups of cells preloaded with calcium orange and exposed to Ca²⁺-free HBSS with CPA. Areas with 1–3 cells and colonies of 4–9 cells are analyzed separately, and emission is shown as the ratio of the intensity over initial intensity (F₁/F₀). Significance of experiment by ANOVA repeated measures, P < 0.0001. d Fluorescence intensity of calcium orange during ER depletion and Ca²⁺ readdition. Depletion is represented by negative numbers, and Ca²⁺ readdition is at time = 0 (arrow). The means, tested by ANOVA repeated measures, differed significantly at P = 0.0042. e Filopodia extension and retraction. Typical extension (blue) and retraction (black) rates are 1.7 µm/min and 1.4 µm/min, respectively. The mean length was 5.7 µm (S.D. 1.0 µm, filopodia in 8 images). f Increase in F₁/F₀ after CPA washout followed by Ca²⁺ with 25 mM KCl, supplied at time = 0 (arrow). Values after time = 0 differed significantly by the ANOVA repeated measures test, P < 0.0001. Error bars represent ± one standard error of the mean (S.E.M.)