July 2019
Volume 60, Issue 9
Open Access
ARVO Annual Meeting Abstract  |   July 2019
Serial Block-Face Scanning EM (SBEM) Provides Evidence for Retrolaminar Demyelination of Structurally Intact Axons in Non-Human Primate (NHP) Early Experimental Glaucoma (EG)
Author Affiliations & Notes
  • Cheri Stowell
    Discoveries in Sight, Devers Eye Institute, Beaverton, Oregon, United States
    Legacy Research Institute, Portland, Oregon, United States
  • Eric Bushong
    National Center for Microscopy & Imaging Research, La Jolla, California, United States
  • Howard Lockwood
    Discoveries in Sight, Devers Eye Institute, Beaverton, Oregon, United States
    Legacy Research Institute, Portland, Oregon, United States
  • Imee Williams
    Discoveries in Sight, Devers Eye Institute, Beaverton, Oregon, United States
    Legacy Research Institute, Portland, Oregon, United States
  • Juan Reynaud
    Discoveries in Sight, Devers Eye Institute, Beaverton, Oregon, United States
    Legacy Research Institute, Portland, Oregon, United States
  • Stuart Keith Gardiner
    Discoveries in Sight, Devers Eye Institute, Beaverton, Oregon, United States
    Legacy Research Institute, Portland, Oregon, United States
  • Nicholas Marsh-Armstrong
    Department of Ophthalmology, University of California, Davis, California, United States
  • Mark Ellisman
    National Center for Microscopy & Imaging Research, La Jolla, California, United States
  • Claude F Burgoyne
    Discoveries in Sight, Devers Eye Institute, Beaverton, Oregon, United States
    Legacy Research Institute, Portland, Oregon, United States
  • Footnotes
    Commercial Relationships   Cheri Stowell, None; Eric Bushong, None; Howard Lockwood, None; Imee Williams, None; Juan Reynaud, None; Stuart Gardiner, None; Nicholas Marsh-Armstrong, None; Mark Ellisman, None; Claude Burgoyne, None
  • Footnotes
    Support  NIH Grants R01-EY011610 (CFB), Legacy Good Samaritan Foundation, AGS Mid-Career Award (CFB), Alcon Research Institute (CFB), Sears Medical Trust (CFB)
Investigative Ophthalmology & Visual Science July 2019, Vol.60, 5658. doi:https://doi.org/
  • Views
  • Share
  • Tools
    • Alerts
      ×
      This feature is available to authenticated users only.
      Sign In or Create an Account ×
    • Get Citation

      Cheri Stowell, Eric Bushong, Howard Lockwood, Imee Williams, Juan Reynaud, Stuart Keith Gardiner, Nicholas Marsh-Armstrong, Mark Ellisman, Claude F Burgoyne; Serial Block-Face Scanning EM (SBEM) Provides Evidence for Retrolaminar Demyelination of Structurally Intact Axons in Non-Human Primate (NHP) Early Experimental Glaucoma (EG). Invest. Ophthalmol. Vis. Sci. 2019;60(9):5658. doi: https://doi.org/.

      Download citation file:


      © ARVO (1962-2015); The Authors (2016-present)

      ×
  • Supplements
Abstract

Purpose : To test the hypothesis that structurally intact retrolaminar optic nerve (ON) axons are demyelinated in NHP early EG.

Methods : Unilateral early EG1 NHPs (n=8) were perfusion fixed, EG and Control (C) eyes were enucleated, and ONs were axon counted2. For each eye, the optic nerve head (ONH) was trephined, and serial 100µm vibratome sectioned parallel to the OCT fovea to Bruch’s membrane opening (FoBMO) axis (Fig.1). Each section was imaged and each image colocalized to a fundus photograph allowing sectoral post-mortem ON axon loss to be estimated.2 To guide locations of 3D reconstruction data by SBEM, x-ray microscopy and tomographic reconstructions (μCT) were obtained from the same resin-embedded samples, and used to precisely select the x-y-z location of the 200 x 200 x 50 μm region of interest (ROI). The posterior-most laminar beams were segmented and best fit with a posterior laminar reference surface (PLRS). All axons were manually identified within a digital section-image parallel to the vitreous block surface. Axons were randomly selected and 3D traced, until they ended (non-intact) or left the block (intact). For each axon, myelin onset was marked, if present, and myelin onset distance (MOD) was measured along the axon from the PLRS (Fig. 2). Intact axons without myelin were labeled unmyelinated and the full retrolaminar distance was used. Axons were traced until 50 axons with a MOD were identified. Cox proportional hazards models were used to compare MOD between axons in each EG ROI and axons in a pooled C of 4 ROIs.

Results : To date, 6 ROIs have been studied: 2 ROIs from 2 sectors of a single early EG eye (NHP1-EG1 with 8% estimated axon loss, and NHP1-EG2 with 15% loss), 2 ROIs from the matched contralateral C eye sectors (NHP1-C1 and NHP1-C2) (Fig. 2) and 2 ROIs from a second NHP C eye (NHP2-C1 and NHP2-C2). Axons in both EG eye ROIs were unmyelinated further from the posterior lamina than in the pooled C eye ROIs (p<0.0001, Fig. 2). The median MODs from the survival analysis were 49.6μm for the pooled C eye ROIs; 165.6μm for NHP-EG1; and 125.5μm for NHP1-EG2.

Conclusions : We describe a novel SBEM colocalization technique and report preliminary findings that suggest structurally intact axons are demyelinated in the immediate retrolaminar ON in NHP early EG1.

1He et al. IOVS. 2014
2Reynaud, et al. IOVS. 2012

This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.

 

 

×
×

This PDF is available to Subscribers Only

Sign in or purchase a subscription to access this content. ×

You must be signed into an individual account to use this feature.

×