Anterior Segment Enhanced-Depth Imaging Optical Coherence Tomography for Imaging the Lamina Cribrosa Ex Vivo

Abstract

 

Purpose

 

To investigate the use of anterior segment enhanced-depth imaging optical coherence tomography (EDI OCT) in imaging the lamina cribrosa (LC) of ex vivo eyes.

 

Methods

 

After removing the anterior segment and the vitreous from fresh enucleated pig eyes, posterior segment tissue containing the optic nerve head and peripapillary sclera (Fig A) was placed on a custom-designed eye holder (Fig 1B). To stabilize the eye holder on the OCT device, an eye holder frame was used (Fig 1C). Serial horizontal (15°x10°) and vertical (10°x15°) EDI OCT B-scans (distance between scans: ~32 µm) of the optic nerve head were obtained from the prepared tissue using the anterior segment module of spectral-domain OCT (Spectralis; Heidelberg Engineering GmbH, Heidelberg, Germany). Different conditions were tested to obtain better-quality OCT images of the LC. After EDI OCT, serial horizontal or vertical histological sections were obtained (distance between sections: ~5 µm), stained (Periodic Acid-Schiff) and photographed. Structures identified in the histological sections were compared with structures in the matched EDI OCT B-scans. 3-dimensional images of the LC were reconstructed using serial EDI OCT B-scans after manually delineating the LC beams.

 

Results

 

Optic nerve heads of 3 enucleated pig eyes were imaged using EDI OCT and then examined histologically. The LC was more clearly visualized when the retina and part of prelaminar tissue were removed (Fig 1D, 1E) and when the tissue surface was kept moist during EDI OCT (Fig 1F, 1G). The LC image quality was similar between pre-fixation and post-fixation tissues (Fig 1H, 1I). The LC was also visualized successfully using the OCT’s posterior segment module when a +20-diopter lens was placed between the tissue and OCT using a custom-designed lens holder, to substitute for the refractive power of the anterior portion of the eyeball (Fig 1J, 1K). EDI OCT B-scans accurately matched serial histological sections and identified the LC beams and its continuity with the retrolaminar glial columns (Fig 2A-2F). 3-dimensional images of the LC were successfully reconstructed using EDI OCT B-scans (Fig 2G, 2H).

 

Conclusions

 

High-resolution cross-sectional images can be obtained using anterior segment EDI OCT when the retina and part of prelaminar tissue are removed. Anterior segment EDI OCT may be useful in evaluating the LC in ex vivo eyes for glaucoma research.

 

 

 

Keywords: 577 lamina cribrosa • 551 imaging/image analysis: non-clinical • 637 pathology: experimental