Multiparametric optical mapping of rat right ventricle (original traces, unfiltered, obtained from tissue within 4 × 4 pixel square in top panel). Em1–4 correspond to emitted fluorescence during exposure to excitation sources Ex1–4, respectively. A: In the presence of the Vm-sensitive di-4-ANBDQPQ only, no significant fluorescence signal is detected when illuminated by excitation sources 3 and 4 (excitation sources for fura-2). B: Simultaneous ratiometric Vm and ratiometric [Ca2+]i optical mapping after additional loading of the same heart with fura-2. EM1/EM2 and EM3/EM4 show normalized Vm and CaT signals after calculation of corresponding source signal ratios (EM3/EM4 signals in A are normalized with respect to corresponding signal-amplitudes in B). Scale bar = 5 mm. Note two to 20-fold difference in Y-axis scaling for Em3 and Em4, compared to EM1 and EM2.