Figure 4.

Temperature dependence of Ca²⁺ store filling and basal [Ca²⁺]_i. (A) Solid circles show mean whole-cell fluorescence signals recorded from SH-SY5Y cells after bath addition of 1 μM ionomycin (indicated by arrows) in solution containing no added Ca²⁺ and 1 mM EGTA at 12°C, 22°C, and 36°C (n = 66, 65, and 80 cells, respectively). The mean responses to the same treatment in cells that were incubated with 1 μM thapsigargin for 1 h before recording are shown by open circles (n = 26, 45, and 20 cells, respectively). (B) Bars indicate the mean thapsigargin-sensitive response to ionomycin in the cells in panel A. A Student’s t-test indicated no significant difference between responses at 36°C and either 12°C or 22°C (C) Measurements of mean basal [Ca²⁺]_i derived from ratiometric fura-2 recordings in SH-SY5Y cells incubated at 12°C, 22°C, and 36°C (n = 5 replicate experiments, with ≥25 cells per experiment) for 15 min (black bars). The raw F₃₄₀/F₃₈₀ ratios used to derive basal [Ca²⁺]_i are superimposed in light gray. Corresponding estimates of basal [Ca²⁺]_i were determined by comparison with in vitro calibration of fura-2 ratios at 22°C after correcting for temperature-dependent changes in K_d (325, 275, and 224 nM at 12°C, 22°C, and 36°C, respectively (12)). A Student’s t-test indicated no significant differences between responses at any of the temperatures examined.