June 2013
Volume 54, Issue 15
Free
ARVO Annual Meeting Abstract  |   June 2013
Rpe65 gene therapy restores precise visual responses in vitro to many retinal ganglion cells in Rpe65-/- mice
Author Affiliations & Notes
  • Steven Stasheff
    Pediatrics,Ophthalmology,Neurosci & BME, Univ of Iowa-Children's Hospital, Iowa City, IA
  • Frederick Blodi
    Pediatrics, University of Iowa, Iowa City, IA
  • Malini Shankar
    Pediatrics, University of Iowa, Iowa City, IA
  • Jeannette Bennicelli
    Ophthalmology, Univ of Penna-Children's Hosp of Philadelphia, Philadelphia, PA
  • Jean Bennett
    Ophthalmology, Univ of Penna-Children's Hosp of Philadelphia, Philadelphia, PA
  • Sajag Bhattarai
    Ophthalmology & Visual Sciences, University of Iowa, Iowa City, IA
  • Arlene Drack
    Ophthalmology & Visual Sciences, University of Iowa, Iowa City, IA
  • Stewart Thompson
    Ophthalmology & Visual Sciences, University of Iowa, Iowa City, IA
  • Footnotes
    Commercial Relationships Steven Stasheff, None; Frederick Blodi, None; Malini Shankar, None; Jeannette Bennicelli, None; Jean Bennett, Gensight Biologics (S); Sajag Bhattarai, None; Arlene Drack, None; Stewart Thompson, None
  • Footnotes
    Support None
Investigative Ophthalmology & Visual Science June 2013, Vol.54, 720. doi:
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      Steven Stasheff, Frederick Blodi, Malini Shankar, Jeannette Bennicelli, Jean Bennett, Sajag Bhattarai, Arlene Drack, Stewart Thompson; Rpe65 gene therapy restores precise visual responses in vitro to many retinal ganglion cells in Rpe65-/- mice. Invest. Ophthalmol. Vis. Sci. 2013;54(15):720.

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

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Abstract
 
Purpose
 

To help further improve Rpe65 gene therapy for Leber's congenital amaurosis (LCA), we sought to better understand its effectiveness at the resolution of single cells and retinal circuits. Here we report substantial improvements in retinal ganglion cell (RGC) responsiveness to light in vitro, but also persistent background hyperactivity and other abnormalities in visual signaling. Further understanding of mechanisms underlying these abnormalities may help explain the better response of children than adults and modest improvements of other visual function seen in early clinical trials.

 
Methods
 

We used in vitro multielectrode recordings to measure RGC activity in rd12 (Rpe65-/-) mouse retinas after subretinal injection with a viral vector to infect retinal pigment epithelium (RPE) cells with normal RPE65 (AAV2/1-hRPE65). We compared in vitro spontaneous and light-evoked activity in both treated and untreated eyes at several time points between 7-90 days following injection. Light stimuli were full field flashes of graded intensities, or pseudorandom checkerboard sequences to map receptive fields.

 
Results
 

Ganglion cell responses to full field flashes were robust in treated eyes, including multiple recognizable response types (e.g., ON/OFF, brisk/sluggish, transient/sustained) and reliable receptive field maps in many cells. However, others had weak or imprecise responses. Retinotopic regions receiving gene therapy corresponded to regions with light responsiveness. No light-evoked responses were detected in any untreated eyes. Spontaneous hyperactivity persisted after gene therapy at a level similar to that in untreated eyes.

 
Conclusions
 

In vitro multielectrode recording provides a powerful system for evaluating the effectiveness of gene therapy at high resolution not possible in human patients. It can provide more detailed understanding of mechanisms that underlie imperfect responsiveness to gene therapy, and guide further improvements in treatment. The results of our current study suggest that spontaneous hyperactivity corrupts the neural code of RGCs, decreasing the precision of some RGC responses. This is expected to degrade the quality of vision and lead to disruption in downstream visual pathways in the brain. We are now investigating whether earlier treatment may prevent RGC hyperactivity.

 
 
A-Treated (GFP) retina. B-Responses recover, hyperactivity persists. C-Receptive fields
 
A-Treated (GFP) retina. B-Responses recover, hyperactivity persists. C-Receptive fields
 
Keywords: 508 electrophysiology: non-clinical • 696 retinal degenerations: hereditary • 538 gene transfer/gene therapy  
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