Figure 4. Changes in cytosolic and ER calcium levels upon PAC-1 and S-PAC-1 treatment.

Graphs of single representative experiments show a) an increase in the FRET ratio in four individual HeLa cells expressing genetically-encoded cytosolic calcium sensor D3cpV upon treatment with 100 µM PAC-1 but not S-PAC-1. Following treatment with PAC-1 or S-PAC-1, cells were washed and treated with ionomycin/Ca²⁺ to demonstrate the maximum FRET ratio and confirm sensor functionality in each cell. b) A greater decrease in the FRET ratio of genetically-encoded ER calcium sensor D1ER was observed upon treatment of 100 µM PAC-1 compared to S-PAC-1 (average of at least 9 cells or n=3 experiments) and c) the absolute change in FRET value is shown to demonstrate that 100 µM PAC-1 elicits a significantly different ER calcium response than S-PAC-1. Error bars represent standard deviation, p < 0.0001.