Complex response of Rv-Mermaid^D129E/Y235R to t-tubule membrane depolarization from −80 mV. (A) Schematic structure of Rv-Mermaid^D129E/Y235R and the changes in mUKG and mKOκ fluorescence in response to the indicated voltage-clamp pulse protocol. Insets show the corresponding raw records before correction for voltage-independent changes in fluorescence; the fit used for correction is shown in white. (B) FRET ratio calculated from the fluorescent traces shown in A. (C) Voltage dependence of the end-pulse FRET ratio measured in response to the pulse protocol shown in A and B. Datasets are from seven muscle fibers, six of which were the ones used to generate mean values shown in Fig. 2 D. (D) Asymmetrical response to a depolarization from −80 mV compared with a hyperpolarization from 0 mV. Left: The FRET ratio (bottom trace) was measured in response to the pulse protocol shown at top. The pink portions of traces were synchronized and summed to produce the pink trace on the right.