Simultaneous imaging of local [Ca²⁺]_i and SL upon electrical stimulation at 5 Hz. (A) Time course of changes in F_yellow/F_cyan (red) and SL (blue) in a cardiomyocyte. Electrical stimulation was performed for 10 s (from 5 to 15 s on the x axis). Bar, period for electrical stimulation. See Video 3. (B) Enlarged view of the graph showing time-dependent changes in F_yellow/F_cyan and SL in A from 10 to 12 s. Note that SL changes in a reciprocal manner in response to a change in F_yellow/F_cyan (as previously demonstrated by us using Fluo-4: see Shintani et al., 2014). (C, top) FFT analysis for the change in SL. (bottom) Same as in the top, for the change in F_yellow/F_cyan. Note that only one peak is present at 5 Hz for both SL and F_yellow/F_cyan, indicating that the EC coupling (i.e., an increase in local [Ca²⁺]_i and the ensuing actomyosin interaction) occurs locally at the single sarcomere level under the present condition. Data were averaged from four individual sarcomeres for both F_yellow/F_cyan and SL.