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The Geisler Method: Tracing Activity-Dependent cGMP Plasticity Changes upon Double Detection of mRNA and Protein on Brain Slices

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Guanylate Cyclase and Cyclic GMP

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1020))

Abstract

We recently demonstrated that an increase of guanosine 3′,5′-cyclic monophosphate (cGMP) signaling could protect the inner ear from noise-induced hair cell damage. Noise exposure not only damages hair cells but also alters the central responsiveness to sound leading to plasticity changes. cGMP signaling has long been known to play a crucial role for plasticity changes and long-term potentiation (LTP). To get a first insight into the role of cGMP for noise-induced plasticity changes we aimed to co-trace the mRNA and protein of plasticity-related genes as, e.g., the immediate early gene Arc (activity-regulated cytoskeletal protein) with markers for the cGMP pathway. We developed a method that permits the simultaneous monitoring of mRNA and protein through light microscopy to visualize gene expression in neurons and synapses of its processes. Accordingly, different from previous fluorescence-based assays that detect, e.g., fluorochrome-labeled Arc antibodies and Arc mRNA, we describe here a methodology that allows the detection of mRNA and protein of synaptic genes using nonfluorescent stable tracers for high-resolution observation of activity-dependent plasticity changes using light microscopy even after weeks or months.

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Authors and Affiliations

  1. Department of Otolaryngology, Hearing Research Centre Tübingen (THRC), Molecular Physiology of Hearing, Universität Tübingen, Tübingen, Germany

    Wibke Singer, Hyun-Soon Geisler & Marlies Knipper

Authors
  1. Wibke Singer
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  2. Hyun-Soon Geisler
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  3. Marlies Knipper
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Editor information

Editors and Affiliations

  1. Addenbrooke’s Hospital, Department of Medicine, University of Cambridge, Hills Road, Cambridge, CB2 2QQ, United Kingdom

    Thomas Krieg

  2. Toxicology and Clinical Pharmacy, Department of Pharmacology, Universität Tübingen, Auf der Morgenstelle 8, Tübingen, 72076, Germany

    Robert Lukowski

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Singer, W., Geisler, HS., Knipper, M. (2013). The Geisler Method: Tracing Activity-Dependent cGMP Plasticity Changes upon Double Detection of mRNA and Protein on Brain Slices. In: Krieg, T., Lukowski, R. (eds) Guanylate Cyclase and Cyclic GMP. Methods in Molecular Biology, vol 1020. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-62703-459-3_15

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  • DOI: https://doi.org/10.1007/978-1-62703-459-3_15

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  • Publisher Name: Humana Press, Totowa, NJ

  • Print ISBN: 978-1-62703-458-6

  • Online ISBN: 978-1-62703-459-3

  • eBook Packages: Springer Protocols

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