Purchase this article with an account.
B.A. Hirons, J.F. Reynaud, A.J. Bellezza, P. Zhou, J.C. Downs, C.F. Burgoyne; Visualization of Digital 3-Dimensional Reconstructions of the Connective Tissues of the Optic Nerve Head . Invest. Ophthalmol. Vis. Sci. 2003;44(13):3341.
Download citation file:
© ARVO (1962-2015); The Authors (2016-present)
Purpose: To describe our current method for reconstruction of the ONH connective tissues with emphasis on visualization and analysis software. Methods: From a fixed eye, the ONH and surrounding peripapillary sclera is trephined (6 mm), pierced with four to seven alignment sutures (10-0 Prolene), embedded in paraffin, and mounted on a serial imaging device (Leica RM2165 microtome fitted with a Diagnostic Instruments SpotRT 1.6 mega-pixel CCD camera on a computer-controlled X translation table with additional manual Y and Z control) capable of generating images at a resolution of 2.5 µm2 per pixel. As each 3.0 µm section is cut away, Ponceau S/Basic Fuchsin stain is applied to the block surface staining only the exposed connective tissues, and an image is acquired. Each embedded ONH yields 250 to 600 serial section images, beginning at the vitreous/retinal interface and ending well into the orbital optic nerve. Each image is then aligned by registering the cut ends of the embedded prolene sutures, which are visible within each section. The scleral canal wall, Bruch’s membrane and the Border Tissue of Elschnig, sclera, lamina cribrosa, optic nerve septa, pial sheath, and vasculature are delineated as discrete objects using manual and semi-automated selection tools in custom and commercial software. The aligned, delineated images are then compiled into a binary volume of 2.5x2.5x3.0 µm voxels, which range in size from 300 to 500 megabytes and can be queried by voxel to generate properties for mathematical modeling or by planar or cubic section for visualization. The volumes can be rendered in whole and in part using the open source OpenDX data visualization software or custom visualization software built using the Visualization Toolkit (VTK) library. Specialized software has also been constructed in Visual Basic and C++ to simultaneously render contralateral views along transverse, vertical sagittal and horizontal coronal axes for each individual subject to aid in direct qualitative comparative analysis. Results: To date, high-resolution, digital 3D geometries of the optic nerve head connective tissues have been constructed for the normal, early-glaucoma and endothelin-treated eyes of five monkeys. Conclusions: Within these geometries, hypothesis testing regarding peripapillary scleral thickness, the anterior and posterior laminar insertions, the intrascleral and intralaminar branches of the posterior ciliary arteries and regional differences in laminar pore size, tortuosity and connective tissue density will be possible.
This PDF is available to Subscribers Only