April 2010
Volume 51, Issue 13
Free
ARVO Annual Meeting Abstract  |   April 2010
The Photoreceptor Ribbon Synapse in the Absence of Piccolo: An RNAi Approach
H. Regus-Leidig; J. Atorf; A. Feigenspan; S. Boye; V. Chiodo; G. M. Seigel; W. W. Hauswirth; J. Kremers; J. H. Brandstätter
Author Affiliations & Notes
  • H. Regus-Leidig
    Dept. of Biology, Animal Physiology, University of Erlangen-Nuremberg, Erlangen, Germany
  • J. Atorf
    Dept. of Ophthalmology, University Hospital Erlangen, Erlangen, Germany
  • A. Feigenspan
    Dept. of Biology, Animal Physiology, University of Erlangen-Nuremberg, Erlangen, Germany
  • S. Boye
    Dept. of Ophthalmology, University of Florida, Gainesville, Florida
  • V. Chiodo
    Dept. of Ophthalmology, University of Florida, Gainesville, Florida
  • G. M. Seigel
    The Center for Hearing and Deafness, SUNY at Buffalo, Buffalo, New York
  • W. W. Hauswirth
    Dept. of Ophthalmology, University of Florida, Gainesville, Florida
  • J. Kremers
    Dept. of Ophthalmology, University Hospital Erlangen, Erlangen, Germany
  • J. H. Brandstätter
    Dept. of Biology, Animal Physiology, University of Erlangen-Nuremberg, Erlangen, Germany
  • Footnotes
    Commercial Relationships  H. Regus-Leidig, None; J. Atorf, None; A. Feigenspan, None; S. Boye, None; V. Chiodo, None; G.M. Seigel, None; W.W. Hauswirth, None; J. Kremers, None; J.H. Brandstätter, None.
  • Footnotes
    Support  DFG BR 1643/4-1
Investigative Ophthalmology & Visual Science April 2010, Vol.51, 4120. doi:
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      H. Regus-Leidig, J. Atorf, A. Feigenspan, S. Boye, V. Chiodo, G. M. Seigel, W. W. Hauswirth, J. Kremers, J. H. Brandstätter; The Photoreceptor Ribbon Synapse in the Absence of Piccolo: An RNAi Approach. Invest. Ophthalmol. Vis. Sci. 2010;51(13):4120.

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Abstract

Purpose: : The mammalian photoreceptor ribbon complex is specialized for the tonic release of neurotransmitter. It separates into two spatially and functionally distinct molecular compartments: the ribbon compartment and the arciform density/plasma membrane compartment. The huge cytomatrix protein Piccolo is located at the ribbon compartment, where it colocalizes and interacts with RIBEYE, the major constituent of the ribbon. As slowly the role of Piccolo is being unravelled at conventional chemical synapses, nothing is known about its function at photoreceptor ribbon synapses. In this study we use RNA interference to specifically knock down Piccolo in photoreceptors already during synaptogenesis.

Methods: : Targeted delivery of Piccolo specific shRNA is achieved via subretinal injection of shRNA carrying AAV5 in postnatal day 5 old C57/Bl6 mice. For detection of transduced photoreceptors, hGFP is expressed under the control of an opsin promoter.

Results: : Knockdown efficiency of Piccolo mRNA was verified in HEK293 cells, R28 cells and isolated transduced photoreceptors. The transduction rate of individual retinae varies between 10% and 100% depending on the distance to the injection site. So far, immunocytochemical analysis indicates that lack of Piccolo results in a drastic reduction in ribbon size, while expression and distribution of other presynaptic molecules, e.g. SNARE and synaptic vesicle proteins, remain unchanged. This indicates a scaffolding role of Piccolo at the photoreceptor ribbon.

Conclusions: : As an alternative to the generation of knockout animals, our tissue specific knockdown approach is a very promising tool to study the contribution of presynaptic proteins to photoreceptor ribbon structure and function. Apart from immunocytochemical examinations, in the future our analyses will include electrophysiological and multifocal ERG recordings, as well as immunoelectron microscopy.

Keywords: photoreceptors • synapse 
© 2010, The Association for Research in Vision and Ophthalmology, Inc., all rights reserved. Permission to republish any abstract or part of an abstract in any form must be obtained in writing from the ARVO Office prior to publication.
Copyright © 2025 Association for Research in Vision and Ophthalmology.
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