Background: Traditional methods for identifying ischemia in ex-vivo models often rely on permanent dyes or late-stage electrophysiological markers. We propose a dynamic method using Indocyanine Green (ICG) stabilized with Bovine Serum Albumin (BSA) to quantify perfusion kinetics and precisely delineate ischemic boundaries in real-time.
Methods: Langendorff-perfused rabbit hearts (n=5) were subjected to regional ischemia via Left Anterior Descending (LAD) coronary ligation. A bolus of ICG, stabilized with 1% BSA to prevent non-specific tissue binding, was injected into the perfusate. High-speed Near-Infrared (NIR) imaging captured the dye transit. We developed an intensity-slope analysis algorithm to calculate wash-in and wash-out rates. These kinetic parameters were used to generate a perfusion-defect map, which was subsequently validated against high-resolution voltage mapping using RH-237 voltage sensitive dye.
Results: BSA-stabilized ICG provided a contrast between well-perfused and ischemic regions. In deoxygenated ischemic zones, we observed significantly higher dye retention and increased absorption, likely due to reduced localized clearance and altered tissue binding affinity in the absence of adequate flow. Ischemic zones were identified by a marked reduction in the peak intensity slope compared to baseline. The boundaries defined by these high-retention regions showed a high spatial correlation with areas of significant Action Potential (AP) triangulation and APD shortening. Intensity slope analysis successfully distinguished between total flow obstruction and partial "low-flow" penumbras.
Conclusion: Dynamic ICG intensity slope analysis provides a repeatable, non-destructive, and quantitative framework for mapping cardiac ischemia. The proportional relationship between dye retention and deoxygenated regions allows for the precise correlation of perfusion deficits with subsequent electrophysiological instability and AP triangulation in rabbit models.