April 2014
Volume 55, Issue 13
ARVO Annual Meeting Abstract  |   April 2014
Optimizing rAAV vectors to target ON bipolar cells
Author Affiliations & Notes
  • Miranda L White
    University of Florida, Gainesville, FL
  • Frank M Dyka
    University of Florida, Gainesville, FL
  • Charles N de Leeuw
    University of British Colombia, Vancouver, BC, Canada
  • Seok-Hong Min
    University of Florida, Gainesville, FL
  • Qing Ruan
    University of Florida, Gainesville, FL
  • Sanford L Boye
    University of Florida, Gainesville, FL
  • Neal S Peachey
    Cole Eye Institute, Cleveland Clinic Foundation, Cleveland, OH
    Cleveland VA Medical Center, Cleveland, OH
  • Elizabeth M Simpson
    University of British Colombia, Vancouver, BC, Canada
  • William W Hauswirth
    University of Florida, Gainesville, FL
  • Shannon Boye
    University of Florida, Gainesville, FL
  • Footnotes
    Commercial Relationships Miranda White, None; Frank Dyka, None; Charles de Leeuw, None; Seok-Hong Min, None; Qing Ruan, None; Sanford Boye, None; Neal Peachey, None; Elizabeth Simpson, None; William Hauswirth, AGTC (C); Shannon Boye, None
  • Footnotes
    Support None
Investigative Ophthalmology & Visual Science April 2014, Vol.55, 3336. doi:
  • Views
  • Share
  • Tools
    • Alerts
      This feature is available to authenticated users only.
      Sign In or Create an Account ×
    • Get Citation

      Miranda L White, Frank M Dyka, Charles N de Leeuw, Seok-Hong Min, Qing Ruan, Sanford L Boye, Neal S Peachey, Elizabeth M Simpson, William W Hauswirth, Shannon Boye; Optimizing rAAV vectors to target ON bipolar cells. Invest. Ophthalmol. Vis. Sci. 2014;55(13):3336.

      Download citation file:

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

  • Supplements

Inaccessible by conventional vectors following subretinal or intravitreal delivery, bipolar cells in the middle retina remain a largely unexploited target for gene therapy. Mutations in bipolar specific genes are associated with Congenital Stationary Night Blindness (CSNB). X-linked CSNB (XCSNB1) is caused by mutations in NYX, which encodes nyctalopin, a protein involved in glutamate signaling in ON bipolar cells (BCs). In addition to night blindness, XCSNB1 patients have reduced acuity, high myopia, nystagmus, and strabismus. Despite having preserved retinal structure, ERG analysis reveals a lack of a b-wave. The Nyxnob mouse carries a frameshift mutation in Nyx. Like patients, it lacks a b-wave but its retina does not degenerate. The purpose of this study is to develop a novel Adeno-associated virus (AAV) vector that will drive expression of therapeutic Nyx exclusively in BCs of Nyxnob mice thereby restoring normal ERG phenotype.


eYFP- tagged Nyx was delivered by a rationally designed, AAV2-based capsid mutant- AAV2(quadY-F+T-V). “Ple155” (based on purkinje cell promoter PCP2) was chosen to drive transgene expression. AAV2(quadY-F+T-V)-Ple155-eYFP-NYX or control vector containing GFP (~2.0E13 vg/ml) were injected intravitreally or subretinally into Nyxnob or C57BL/6J mice. Four weeks post-injection, Nyxnob mice were analyzed by ERG and retinas from all mice were stained with antibodies raised against GFP and either PCP2 or PKCα to label BCs.


AAV2(quadY-F+T-V)-Ple155- mediated GFP and eYFP-NYX expression was restricted to ON BCs. No improvements in retinal function were detected in vector treated Nyxnob mice evaluated by full field ERG.


We identified an AAV serotype/cellular promoter combination that is capable of robust and highly selective transduction of murine ON BCs. This is achievable with either subretinal or intravitreal injection. Functional rescue may not have been observed because 1) an insufficient number of BCs expressed Nyx or 2) our ERG protocol lacked sufficient sensitivity. Work is underway to modify our vectors to increase transgene expression and to increase ERG sensitivity. We conclude that specific targeting of ON BCs is possible, a result which has implications for the treatment of CSNB and also for optogenetic strategies aimed to impart light sensitivity specifically to this cell type.

IHC following IV injection of GFP control vector (left) and eYFP-NYX vector (right) in wt mice
IHC following IV injection of GFP control vector (left) and eYFP-NYX vector (right) in wt mice
Keywords: 538 gene transfer/gene therapy • 435 bipolar cells  

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.