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From synapse to behavior: rapid modulation of defined neuronal types with engineered GABAA receptors

Nature Neuroscience volume 10, pages 923–929 (2007)Cite this article

Abstract

In mammals, identifying the contribution of specific neurons or networks to behavior is a key challenge. Here we describe an approach that facilitates this process by enabling the rapid modulation of synaptic inhibition in defined cell populations. Binding of zolpidem, a systemically active allosteric modulator that enhances the function of the GABAA receptor, requires a phenylalanine residue (Phe77) in the γ2 subunit. Mice in which this residue is changed to isoleucine are insensitive to zolpidem. By Cre recombinase–induced swapping of the γ2 subunit (that is, exchanging Ile77 for Phe77), zolpidem sensitivity can be restored to GABAA receptors in chosen cell types. We demonstrate the power of this method in the cerebellum, where zolpidem rapidly induces significant motor deficits when Purkinje cells are made uniquely sensitive to its action. This combined molecular and pharmacological technique has demonstrable advantages over targeted cell ablation and will be invaluable for investigating many neuronal circuits.

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Figure 1: Strategy to restrict zolpidem sensitivity to selected neural types, using Cre recombinase to drive a γ2I77 to γ2F77 subunit swap.
Figure 2: Summary of mouse genotypes.
Figure 3: Synaptic expression of the γ2F77GFP subunit in Purkinje cells of adult PC-γ2–swap mice.
Figure 4: Potentiation of GABAA receptor–mediated mIPSCs by zolpidem in Purkinje cells of PC-γ2–swap mice.
Figure 5: Motor performance on a rotarod and horizontal beam after systemic administration of zolpidem.

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Acknowledgements

We thank E. Sigel for pointing out the γ2I77 mutation; J. Oberdick for the L7 expression cassette; S.J. Moss for the γ2F77GFP plasmid; M. Meyer for the L7Cre line; R. Tomioka and E. Mugnaini for antibodies to EGFP and GAD, respectively; H. Monyer for discussion and support; F. Zimmermann for the transgene and stem cell injections; I. Preugschat-Gumprecht for help with mouse genotyping; D. Andersson, T. Karayannis and M. Capogna for contributing to initial electrophysiological recordings; and S. Brickley, M. Capogna, S.G. Cull-Candy, C. De Zeeuw, N. Franks, T. Klausberger and Z. Nusser for comments on the manuscript. This work was funded by the VolkswagenStiftung (grant I/78 554 to W.W., E.R.K., W.S. and P.S.), the Deutsche Forschungsgemeinschaft (grant WI 1951/2 to W.W. and P.W.), the UK Medical Research Council (grant G0501584 to W.W.), the J. Ernest Tait Estate (to W.W. and T.G.), a Heidelberg Young Investigator Award (to P.W.), the Academy of Finland (to E.R.K. and A.-M.L.), the Sigrid Juselius Foundation (to E.R.K. and E.L.), the Institute Pasteur-Fondazione Cenci Bolognetti (to M.R.), the Austrian Federal Government (W.S.), the Medical University Vienna (W.S.) and a Wellcome Trust Programme Grant (to M.F.).

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Author notes

  1. Peer Wulff, Thomas Goetz & William Wisden

    Present address: Present address: Institute of Medical Sciences, University of Aberdeen, Foresterhill, Aberdeen AB25 2ZD, UK.,

  2. Peer Wulff and Thomas Goetz: These authors contributed equally to this work.

Authors and Affiliations

  1. Department of Clinical Neurobiology, University of Heidelberg, Im Neuenheimer Feld 364, Heidelberg, 69120, Germany

    Peer Wulff, Thomas Goetz & William Wisden

  2. Institute of Biomedicine, Pharmacology, Biomedicum Helsinki, POB 63 (Haartmaninkatu 8), University of Helsinki, Helsinki, FI-00014, Finland

    Elli Leppä, Anni-Maija Linden, Olga Y Vekovischeva & Esa R Korpi

  3. Department of Pharmacology, University College London, Gower Street, London, WC1E 6BT, UK

    Massimiliano Renzi & Mark Farrant

  4. MRC Anatomical Neuropharmacology Unit, Oxford University, Mansfield Road, Oxford, OX1 3TH, UK

    Jerome D Swinny & Peter Somogyi

  5. Center for Brain Research, Section of Biochemistry and Molecular Biology of the Nervous System, Medical University of Vienna, Spitalgasse 4, Vienna, A-1090, Austria

    Werner Sieghart

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Contributions

The original idea was conceived by W.S. and developed with P.S., W.W. and E.R.K. Experiments were designed by M.F., E.R.K., P.S., P.W., W.S. and W.W. Experiments were performed by M.F., T.G., A.-M.L., E.L., M.R., J.D.S., P.S., O.Y.V., P.W. and W.W. Behavioral data were analyzed by E.L., A.-M.L. and E.R.K. Electrophysiological data were analyzed by M.F. The manuscript was written by M.F., P.W. and W.W. All authors commented on and helped to revise the text.

Corresponding author

Correspondence to Peer Wulff.

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The authors declare no competing financial interests.

Supplementary information

Supplementary Fig. 1

Generation of γ2I77lox and L7γ2F77GFP mice. (PDF 3384 kb)

Supplementary Fig. 2

Confirmation of Purkinje cell–specific Cre activity in the L7Cre line using ROSA26 indicator mice. (PDF 8956 kb)

Supplementary Fig. 3

Expression of the γ2F77GFP subunit in cerebellar Purkinje cells of PC-γ2–swap mice. (PDF 1678 kb)

Supplementary Fig. 4

In slices from PC-γ2–swap mice, potentiation of GABAA receptor–mediated mIPSCs by zolpidem is restricted to Purkinje cells. (PDF 152 kb)

Supplementary Methods (PDF 189 kb)

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Wulff, P., Goetz, T., Leppä, E. et al. From synapse to behavior: rapid modulation of defined neuronal types with engineered GABAA receptors. Nat Neurosci 10, 923–929 (2007). https://doi.org/10.1038/nn1927

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