Fig. 2.

STIM1 expression during differentiation of SH-SY5Y cells. a Top: SH-SY5Y cells were differentiated with RA + BDNF, and bright-field microscopy images of cells were recorded (left panel, non-differentiated; right panel, differentiated after 9 DIV). Bottom: Neurite length was measured in undifferentiated cells (n = 52), and differentiated cells (n = 45), from two independent cultures. Scale bar = 100 μm. b Top: Expression of STIM1 and TUBB3 was assessed by immunoblot from undifferentiated cells and cells differentiated after 9–10 DIV with RA + BDNF. Level of GAPDH was assessed as a loading control of the immunoblot. Bottom: The expression of STIM1 and TUBB3 was quantified by immunoblotting with lysates from three independent assays. c Store-operated Ca²⁺ entry was evaluated in undifferentiated (black line) and differentiated cells (red line). Fura-2-loaded cells were incubated in a Ca²⁺-free HBSS (assay medium), and 1 μM thapsigargin (Tg) was added to the cells for 6 min. Ca²⁺ (2 mM CaCl₂) was finally added to the cells to evaluate the extension of Ca²⁺-entry. The experiment was performed at controlled temperature (36–37 °C). Data are presented as the mean ± s.d. of three independent experiments (n > 60 cells for each condition)