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J Vis Exp. 2015 May 6;(99):e52588. doi: 10.3791/52588.

Imaging Ca2+ dynamics in cone photoreceptor axon terminals of the mouse retina.

Author information

1
Institute for Ophthalmic Research, University of Tübingen; Graduate School of Cellular & Molecular Neuroscience, University of Tübingen.
2
Institute for Ophthalmic Research, University of Tübingen; Bernstein Centre for Computational Neuroscience, University of Tübingen.
3
Institute for Ophthalmic Research, University of Tübingen; Graduate School of Cellular & Molecular Neuroscience, University of Tübingen; Bernstein Centre for Computational Neuroscience, University of Tübingen.
4
Molecular Genetics Laboratory, University of Tübingen; Centre for Ophthalmology, University of Tübingen.
5
Institute for Ophthalmic Research, University of Tübingen; Graduate School of Cellular & Molecular Neuroscience, University of Tübingen; Bernstein Centre for Computational Neuroscience, University of Tübingen; thomas.euler@cin.uni-tuebingen.de.
6
Institute for Ophthalmic Research, University of Tübingen; francois.paquet-durand@klinikum.uni-tuebingen.de.

Abstract

Retinal cone photoreceptors (cones) serve daylight vision and are the basis of color discrimination. They are subject to degeneration, often leading to blindness in many retinal diseases. Calcium (Ca(2+)), a key second messenger in photoreceptor signaling and metabolism, has been proposed to be indirectly linked with photoreceptor degeneration in various animal models. Systematically studying these aspects of cone physiology and pathophysiology has been hampered by the difficulties of electrically recording from these small cells, in particular in the mouse where the retina is dominated by rod photoreceptors. To circumvent this issue, we established a two-photon Ca(2+) imaging protocol using a transgenic mouse line that expresses the genetically encoded Ca(2+) biosensor TN-XL exclusively in cones and can be crossbred with mouse models for photoreceptor degeneration. The protocol described here involves preparing vertical sections ("slices") of retinas from mice and optical imaging of light stimulus-evoked changes in cone Ca(2+) level. The protocol also allows "in-slice measurement" of absolute Ca(2+) concentrations; as the recordings can be followed by calibration. This protocol enables studies into functional cone properties and is expected to contribute to the understanding of cone Ca(2+) signaling as well as the potential involvement of Ca(2+) in photoreceptor death and retinal degeneration.

PMID:
25993489
PMCID:
PMC4542458
DOI:
10.3791/52588
[Indexed for MEDLINE]
Free PMC Article

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