D. Salyer

Retinal oximetry using intravitreal illumination.

Purpose: To demonstrate spectroscopic retinal oximetry measurements on arteries and veins in swine using intravitreal illumination. Retinal arterial and venous saturations are measured for a range of inspired O2 levels after pars plana vitrectomy. Methods: Pars plana vitrectomy and intravitreal manipulations were performed on two female American Yorkshire domestic swine. Light from a scanning monochromator was coupled into a fiberoptic intraocular illuminator inserted into the vitreous. The retinal vessels were illuminated obliquely, minimizing vessel glints. Multispectral images of the retinal vasculature were obtained as the swines arterial blood oxygen saturation was decreased from 100% to 67% in decrements of approximately 10%. Retinal vessel spectra were used to calculate oxygen saturation in selected arteries and veins. Arterial oxygen saturations were calibrated using blood gas analysis on blood drawn from a Swan-Ganz catheter placed in the femoral artery. Results: Oblique illumination of retinal vessels using an intravitreal fiberoptic illuminator provided a substantial reduction in the central vessel glint usually seen in fundus images, thus simplifying the analysis of spectral data. The vessel shadows were displaced from the vessel image simplifying the light paths in the eye. Using a full spectral analysis simplified by the light path reductions, we calculated retinal vessel saturations. The reduction of glint allowed for increased accuracy in measuring retinal vessel spectral optical density. Abnormally low retinal venous oxygen saturations were observed shortly after pars plana vitrectomy. Conclusions: Retinal oximetry using intravitreal illumination has been demonstrated. As a research tool, intravitreal illumination addresses several difficulties encountered when performing retinal oximetry with transcorneal illumination.

Blue-green spectral minimum correlates with oxyhemoglobin saturation in vivo

An imaging multi-spectral retinal oximeter with intravitrial illumination is used to perform the first in vivo test of the blue-green minima shift oximetry method (BGO) in swine eyes [K. R. Dennighoff, R. A. Chipman, and L. W. Hillman, Opt. Lett. 31, 924-926 (2006); J. Biomed. Opt. 12, 034020 (2007).] A fiber optic intravitreal illuminator inserted through the pars plana was coupled to a monochromator and used to illuminate the retina from an angle. A camera viewing through the cornea recorded a series of images at each wavelength. This intravitreal light source moves the specular vessel glint away from the center of the vessel and directly illuminates the fundus behind most blood vessels. These two conditions combine to provide accurate measurements of vessel and perivascular reflectance. Equations describing these different light paths are solved, and BGO is used to evaluate large retinal vessels. In order to test BGO calibration in vivo, data were acquired from swine with varied retinal arterial oxyhemoglobin saturations (60-100% saturation.). The arterial saturations determined using BGO to analyze the multispectral image sets showed excellent correlation with co-oximeter data (r2=0.98, and residual error +/-3.4% saturation) and are similar to results when hemoglobin and blood were analyzed using this technique.

Diffuse spectral fundus reflectance measured using subretinally placed spectralon

The diffuse fundus reflectance and the spectral transmittance of the swine sensory retina was measured in vivo using intravitreal illumination. Pars plana vitrectomy and intravitreal manipulations were performed on a female American Yorkshire domestic swine. Light from a scanning monochromator was coupled into a fiber optic intraocular illuminator inserted into the vitreous. A 1.93-mm(2) region of the illuminated fundus was imaged from an oblique illumination angle. Multispectral retinal images were acquired for four experimental conditions: the eye (1) prior to vitrectomy, (2) after vitrectomy, (3) after insertion of a Spectralon disk super-retinally, and (4) after subretinal insertion of the disk. The absorption of melanin and hemoglobin in the red wavelengths was used to convert relative spectral reflectance to absolute reflectance. The flux scattered from the super-retinal Spectralon was used to correct for scattering in the globe. The transmittance of the sensory retina was measured in vivo using the scatter corrected subretinal Spectralon disk reflectance. The hemoglobin and melanin components of the spectrum due to scattered light were removed from the retinal transmission spectrum. The in vivo spectral transmittance of the sensory retina in this swine was essentially flat across the visible spectrum, with an average transmittance >90%.

Wavelength dependence of the apparent diameter of retinal blood vessels

Imaging of retinal blood vessels may assist in the diagnosis and monitoring of diseases such as glaucoma, diabetic retinopathy, and hypertension. However, close examination reveals that the contrast and apparent diameter of vessels are dependent on the wavelength of the illuminating light. In this study multispectral images of large arteries and veins within enucleated swine eyes are obtained with a modified fundus camera by use of intravitreal illumination. The diameters of selected vessels are measured as a function of wavelength by cross-sectional analysis. A fixed scale with spectrally independent dimension is placed above the retina to isolate the chromatic effects of the imaging system and eye. Significant apparent differences between arterial and venous diameters are found, with larger diameters observed at shorter wavelengths. These differences are due primarily to spectral absorption in the cylindrical blood column.