Identification of Structures Labeled by Indocyanine Green in the Rat Choroid and Retina Can Guide Interpretation of Indocyanine Green Angiography.

Purpose: Indocyanine green (ICG) is an albumin and lipoprotein binding dye absorbing in the far red used in angiography to visualize choroidal vessels (ICG angiography [ICGA]). To guide interpretation, ICG transport in the choroid, RPE, and retina of rats was studied. Methods: Two conditions were used: RPE/choroid organoculture, incubated for 45 minutes in DMEM medium, 1% fetal bovine serum containing 0.25 mg/mL ICG and RPE/choroid and neural retina flat-mounts at 1 and 6 hours after intravenous ICG injection. Early and late sequences of ICGA were recorded until 6 hours. Ultra-deep red confocal microscope was used to localize ICG in flat-mounts and immunohistochemistry was performed for caveolin-1, tryptase (mast cell marker), and tubulin β3 (a nerve marker). Results: In the organoculture, ICG penetrated homogeneously in the cytoplasm and stained the membranes of the RPE. At 1 hour after intravenous injection, ICG appeared in fine granules in RPE, partly labeled with caveolin-1 and decreasing at 6 hours. At 1 hour and 6 hours, ICG was found in the retinal vessels, faintly in the inner retina, and in the photoreceptor outer segments at 6 hours. In the choroid, ICG colocalized with mast cells, immunostained with tryptase, and accumulated along the large tubulin β3-labeled nerve bundles. The hypothesis was raised on the interpretation of late ICGA infrared photography in case of transthyretin amyloidosis with neuropathy. Conclusions: Beside being a vascular dye, ICG is transported from the vessels to the RPE toward the outer retina. It stains mast cells and large choroidal nerves. These observations could help the analysis of ICGA images.