Figure 5.

Addition of saponin to the depolarized t-system has little effect on the fluorescence signal. Saponin (10 µg/ml) causes small perforations (pores) in the t-system membrane and rapid equilibration of the pH in the t-system lumen with that in the cytosolic environment. In this experiment, the t-system was loaded with an additional fluorescent probe that is not sensitive to pH (SRB, see Materials and methods) to correct for the loss of HTPS signal associated with the diffusion of HTPS from the t-system lumen through the pores made by saponin. Accordingly, the fluorescence signal is shown as the ratio between the HTPS and SRB fluorescence. Solution exchanges are indicated by the vertical pale bars, and the composition of the internal solution is indicated by the horizontal lines above the trace.