FRET response of T306-Rv-Mermaid^D129E/Y235R and T306-Rv-Mermaid during Ca²⁺ release in the absence of intracellular EGTA. (A) FRET response to 0.5-s-long voltage-clamp depolarizing pulses to values ranging between −30 and +10 mV from a muscle fiber expressing T306-Rv-Mermaid^D129E/Y235R, equilibrated in the presence of intracellular BAPTA. The inset shows the mean response from four fibers to large depolarizing pulses in the same condition (see text for details). Successive single depolarizing pulses were separated by a time interval of 30 s. (B) Mean rhod-2 cytosolic Ca²⁺ transient elicited by a depolarizing pulse from −80 to +40 mV in seven fibers equilibrated with intracellular EGTA (black trace) and four fibers equilibrated with intracellular BAPTA (red trace). Gray shading corresponds to SEM. (C) FRET response to 0.5-s-long depolarizing voltage-clamp pulses to −35, −30, and −20 mV from a muscle fiber expressing T306-Rv-Mermaid^D129E/Y235R in the presence of extracellular BTS. Successive single depolarizing pulses were separated by 30 s. (D) FRET response of a muscle fiber expressing T306-Rv-Mermaid^D129E/Y235R, equilibrated with intracellular EGTA, to a train of action potentials generated at 100 Hz. (E) FRET response of a muscle fiber expressing T306-Rv-Mermaid^D129E/Y235R, equilibrated without intracellular EGTA and in the presence of extracellular BTS, to a train of action potentials generated at 20 Hz. (F) FRET response of a fiber expressing T306-Rv-Mermaid, equilibrated without intracellular EGTA and in the presence of extracellular BTS, to a train of action potentials at 100 Hz.