FIGURE 5.

Cholesterol attenuated the thapsigargin-activated capacitative calcium entry (CCE) similarly to SOC inhibition. Prior to experiment, cells were incubated with PBS with Ca⁺² for 30 min (solid line), 3.5 mM water-soluble cholesterol for 30 min (dashed line) or 50 μM of SOC inhibitor SKF-96365 for 10 min (dotted line). (a) Shows representative traces of the calcium response to 1 μM Tg was added to cells in Ca⁺² free PBS. After the initial response, the solution is replaced with 500 μL of PBS with Ca⁺². (b) Bar graph representing average responses for each condition at initial response peak and after re-addition Ca⁺², which represents capacitative calcium entry (CCE). Cholesterol enriched cells showed an increase in the ER calcium response and an attenuated CCE response, which was significantly different from unenriched cells. In addition, the CCE response in cholesterol-treated cells was not statistically different from cells treated with the SOC inhibitor SKF. (c) Representative images of the fluorescence signal at baseline and peak stimulation for unenriched and cholesterol treated coverslips. Average responses represent multiple coverslips (cs) and total cell count among coverslips (n) Unenriched: cs = 4, n = 225, Cholesterol: cs = 3, n = 169, SKF: cs = 3, n = 135. Mean and SEM were plotted Two-tailed t test ***p <0.0001.