Fig. 1.

Thapsigargin causes a decrease in CFTR-mediated conductance. a Representative fast whole cell current traces obtained by holding the membrane potential at 0 mV and applying voltage steps between ± 100 mV in 20 mV increments under (i) basal conditions and after (ii) 8 min, (iii) 18 min and (iv) 28 min perfusion with forskolin (fsk; 5 µM) and (v) fsk and CFTR_inh-172 (10 µM). b Representative fast whole cell current traces obtained as in a under (i) basal conditions (ii) after 8 min perfusion with forskolin (fsk; 5 µM) (iii) 10 min perfusion with fsk and thapsigargin (TG; 200 nM) (iv) 10 min perfusion with fsk alone and (v) 8 min perfusion with fsk and CFTR_inh-172. Dashed line indicates the zero current level. c Mean changes in conductance (G) plotted relative to the maximum current reached at + 100 mV when cells were perfused with fsk or after TG exposure as indicated. ^ǂp < 0.05 when compared to equivalent timepoint in cells exposed to forskolin only. d Changes in conductance under the conditions indicated. Conductance was normalised to cell size. Data was taken from timepoints indicated in a and b. Data are mean ± SEM (n = 4–10 cells). Open bars represent cells exposed to thapsigargin, closed bars represent cells exposed to forskolin only. *p < 0.05 when compared to baseline in fsk only treated cells. ^†p < 0.05 when compared to maximal forskolin stimulated conductance at 8 min in fsk only treated cells. ^#p < 0.05 when compared to baseline in fsk + TG treated cells ^•p < 0.05 when compared to maximal forskolin stimulated conductance at 8 min in cells treated with TG. e Representative images showing the effect of vehicle and thapsigargin on CFTR at timepoints indicated. f Changes in intracellular and membrane fluorescence. A single exponential was fitted to the data to give a half-life of 11.9 min and a τ of 17.2. Data are mean ± SEM (n = 63–86 cells from 6 coverslips). Scale bar represents 25 µm