September 2016
Volume 57, Issue 12
Open Access
ARVO Annual Meeting Abstract  |   September 2016
Loss of cone function without degeneration in a novel Gnat2 knock-out mouse line
Author Affiliations & Notes
  • Kaitryn Ronning
    University of California Davis, Davis, California, United States
  • Gabriel Peinado
    University of California Davis, Davis, California, United States
  • Mayank Goswami
    University of California Davis, Davis, California, United States
  • Robert J Zawadzki
    University of California Davis, Davis, California, United States
  • Edward N Pugh
    University of California Davis, Davis, California, United States
  • Rolf Herrmann
    University of California Davis, Davis, California, United States
  • Marie E Burns
    University of California Davis, Davis, California, United States
  • Footnotes
    Commercial Relationships   Kaitryn Ronning, None; Gabriel Peinado, None; Mayank Goswami, None; Robert Zawadzki, None; Edward Pugh, None; Rolf Herrmann, None; Marie Burns, None
  • Footnotes
    Support  National Eye Institute R01EY14047 and the NEI UC Davis Vision Core Grant P-30 EY012576
Investigative Ophthalmology & Visual Science September 2016, Vol.57, 639. doi:
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    • Get Citation

      Kaitryn Ronning, Gabriel Peinado, Mayank Goswami, Robert J Zawadzki, Edward N Pugh, Rolf Herrmann, Marie E Burns; Loss of cone function without degeneration in a novel Gnat2 knock-out mouse line. Invest. Ophthalmol. Vis. Sci. 2016;57(12):639.

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

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Abstract

Purpose : Pure rod function has been difficult to study in vivo because of the paucity of mutants that abolish cone signaling without causing photoreceptor degeneration. For example, a loss-of-function mutation in the G-protein alpha subunit of cones, the Gnat2cpfl3 mutant mouse line, exhibits retinal degeneration by 27 weeks (Chang et al., 2006 IOVS). Here we characterize a novel Gnat2 knock-out mouse line (Gnat2-/-) for its potential utility for dissecting rod and cone function.

Methods : Gnat2 mRNA expression levels were examined using quantitative real-time PCR, and protein expression was quantified using Western blot analysis. Protein localization and photoreceptor health were examined using immunohistochemistry and in vivo optical coherence tomography (OCT). Corneal electroretinograms (ERGs) were used to characterize retinal responses to calibrated flashes delivered in darkness or in the presence of background light. ERG a- and b-wave amplitudes were recorded from dark-adapted mice in response to increasingly intense flashes of both green and UV light. In order to isolate cone responses, a bright 1 ms UV flash (2.75x106 photons mm-2) was delivered on top of an intense green background light that saturated the rods.

Results : Immunohistochemical staining with PNA revealed normal numbers and morphologies of cones in the knockout, although there was no detectable immunoreactivity to Gnat2 antibody. DAPI staining revealed a normal outer nuclear layer thickness in retinal cross-sections and OCT B-scans revealed normal appearance of retinal layers in the Gnat2-/- mice at 5 months of age in vivo. Gnat2-/- mice exhibited nearly normal dark-adapted responses to both dim and bright flashes, presumably due to the predominance of rod responses, which were normal. When cone responses were isolated with a rod-saturating background, non-mutant mice displayed normal cone-driven ERGs. However, in the presence of the same rod-saturating background, there were no detectable electrical responses from the Gnat2-/- retinas.

Conclusions : Loss of the Gnat2 gene does not cause retinal degeneration in this knock out line. Despite healthy-looking morphology, Gnat2-/- cones are rendered completely unresponsive to light by the absence of the heteromeric G protein alpha subunit 2. Thus, this mouse line will be a very useful tool for studying the interactions of rod and cone signaling in the intact, non-degenerating retina.

This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.

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