Visualizing excitation waves inside cardiac muscle using transillumination

Voltage-sensitive fluorescent dyes have become powerful tools for the visualization of excitation propagation in the heart. However, until recently they were used exclusively for surface recordings. Here we demonstrate the possibility of visualizing the electrical activity from inside cardiac muscle via fluorescence measurements in the transillumination mode (in which the light source and photodetector are on opposite sides of the preparation). This mode enables the detection of light escaping from layers deep within the tissue. Experiments were conducted in perfused (8 mm thick) slabs of sheep right ventricular wall stained with the voltage-sensitive dye di-4-ANEPPS. Although the amplitude and signal-to-noise ratio recorded in the transillumination mode were significantly smaller than those recorded in the epi-illumination mode, they were sufficient to reliably determine the activation sequence. Penetration depths (spatial decay constants) derived from measurements of light attenuation in cardiac muscle were 0.8 mm for excitation (520 +/- 30 nm) and 1.3 mm for emission wavelengths (640 +/- 50 nm). Estimates of emitted fluorescence based on these attenuation values in 8-mm-thick tissue suggest that 90% of the transillumination signal originates from a 4-mm-thick layer near the illuminated surface. A 69% fraction of the recorded signal originates from > or =1 mm below the surface. Transillumination recordings may be combined with endocardial and epicardial surface recordings to obtain information about three-dimensional propagation in the thickness of the myocardial wall. We show an example in which transillumination reveals an intramural reentry, undetectable in surface recordings.