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G. Baerveldt, D.S. Minckler, N. Morishige, J.V. Jester, D.J. Brown; Dynamic Non–Invasive Visualization of the Human Lamina Cribrosa With Femosecond Laser Energy . Invest. Ophthalmol. Vis. Sci. 2006;47(13):3376.
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This study investigated the feasibility of non–invasively examining the three–dimensional structure of the human lamina cribrosa using second harmonic generated (SHG) signals from collagen fibers. The ability to visualize the lamina cribrosa with this technique may help to clarify variations in topographic vulnerability of the laminar structures during glaucomatous injury and damage to optic nerve axons.
Post mortem human eyes were dissected and a trephine employed to cut a 6 mm diameter button containing the optic nerve head. Laminar tissues were analyzed by SHG using a chameleon femtosecond laser tuned to 800 nm. Backscattered and forward scattered SHG signals were collected using a 350–450 nm band pass filter. Stacks of images were collected and reconstructed using Zeiss 510 LSM software.
The method allows for visualization of the "channels" (openings or pores) between collagen beams of the lamina cribrosa through which nerve fiber bundles pass into the orbital optic nerve. At the anterior aspect of the lamina, the laminar pores appeared narrower with thinner walls than more posterior regions next to the optic nerve. At its most posterior aspect, the laminar pores are thicker and better defined with this technique. Three–dimensional reconstruction clearly illustrates that the bundle pores often branch into smaller openings or merge into larger channels as they course posteriorly.
Imaging of SHG signals from collagen allows rapid, non–invasive, real– time visualization of the lamina cribrosa without artifacts common to standard histologic methods requiring embedding and sectioning in paraffin or plastic media. This technique may be a valuable addition to other methodologies to study experimental animal and human optic nerve tissue responses to elevated intraocular pressure.
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