Peripapillary Rat Sclera Investigated Using Polarization Sensitive Optical Coherence Tomography Imaging and Histological Analysis

Abstract

Purpose: To demonstrate polarization sensitive (PS) optical coherence tomography (OCT) for quantitative imaging of the birefringent properties of the peripapillary sclera in in vivo rat eyes. To correlate the findings in PS-OCT images to histological analysis of the same tissues.

Methods: A prototype high-speed PS-OCT system was modified for imaging the rat eye. Healthy male Sprague Dawley rats were anaesthetized and densely sampled PS-OCT raster scans covering an area of ~1.5 mm x 1.5 mm centered at the papilla were acquired. In addition to conventional OCT images based on reflectivity, cross-sectional and en-face PS-OCT images displaying the birefringent tissue properties were computed from the 3D data of the retina and sclera. The eyes were enucleated post mortem for histological analysis. After fixation, cryosectioning and staining, confocal microscopy imaging of ocular structures in the posterior globe was performed.

Results: High-speed PS-OCT enables 3D imaging of the retinal and scleral polarization properties with dense sampling and reduced motion artifacts. In reflectivity images, the sclera appears as a hyperreflective layer between retina and orbital connective tissue. Strong birefringence can be observed in the sclera as a dramatic increase of phase retardation values. En-face phase retardation images reveal the strongest birefringence at ~0.5 mm distance from the papilla where a donut shaped ring of severely increasing retardation values can be observed. The birefringent axis orientation was investigated in en-face images. Around the optic nerve, axis orientation varies considerably dependent on the location and exhibits a radial dependence in the posterior sclera. The histological analysis revealed the scleral microstructure. As in the PS-OCT images, varying collagen fiber orientation was observed in the peripapillary region.

Conclusions: PS-OCT enables quantitative imaging of polarizing tissue properties in addition to conventional OCT imaging based on reflectivity. In the sclera, PS-OCT provides access to birefringence and birefringent axis orientation, which is related to the arrangement of collagen fiber compounds. Since scleral birefringence has been related to its biomechanical properties, PS-OCT might be a powerful tool for scleral imaging and quantification in longitudinal studies using rodent models of glaucoma.

Keywords: 552 imaging methods (CT, FA, ICG, MRI, OCT, RTA, SLO, ultrasound) • 708 sclera • 630 optical properties