Single-neuron labeling with inducible Cre-mediated knockout in transgenic mice

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Abstract

To facilitate a functional analysis of neuronal connectivity in a mammalian nervous system that is tightly packed with billions of cells, we developed a new technique that uses inducible genetic manipulations in fluorescently labeled single neurons in mice. Our technique, single-neuron labeling with inducible Cre-mediated knockout (SLICK), is achieved by coexpressing a drug-inducible form of Cre recombinase and a fluorescent protein in a small subsets of neurons, thus combining the powerful Cre recombinase system for conditional genetic manipulation with fluorescent labeling of single neurons for imaging. Here, we demonstrate efficient inducible genetic manipulation in several types of neurons using SLICK. Furthermore, we applied SLICK to eliminate synaptic transmission in a small subset of neuromuscular junctions. Our results provide evidence for the long-term stability of inactive neuromuscular synapses in adult animals and demonstrate a Cre-loxP compatible system for dissecting gene functions in single identifiable neurons.

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Figure 1: Strategy used for coexpression of YFP and Cre in SLICK transgenic mice.
Figure 2: YFP labeling of distinct neuronal populations in SLICK transgenic mice.
Figure 3: Genetic manipulation in SLICK transgenic mice.
Figure 4: Inhibition of neurotransmission in subsets of motor neurons through Chat knockout in SLICK-A mice.

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Acknowledgements

We thank J. Sanes for providing the Chat conditional-knockout mice and the antibody to β-galactosidase. We thank A. Mizrahi for performing two-photon imaging of SLICK-3 mice and R. Kotloski for advice regarding tamoxifen administration. We also thank the Duke Neurotransgenic Core Facility for generating the SLICK mice. We thank J. McNamara, M.D. Ehlers and M. McCaffrey for access to confocal microscopes. We are grateful to B. Arenkiel, M.D. Ehlers, A. West, K. Dean and members of the Feng laboratory for critically reading and discussing the manuscript. This work was supported by the Ruth K. Broad Biomedical Research Foundation, the Science Foundation Ireland and the US National Institutes of Health.

Author information

P.Y. and G.F. conceived the SLICK method, designed the experiments and wrote the manuscript. P.Y. and J.G. generated the SLICK transgenic mice. P.Y., L.Q., S.Z. and D.W. characterized the SLICK mice. P.Y. and S.Z. carried out the quantification and data analysis.

Correspondence to Paul Young or Guoping Feng.

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Supplementary Figures 1–6, Methods and Discussion (PDF 6860 kb)

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Young, P., Qiu, L., Wang, D. et al. Single-neuron labeling with inducible Cre-mediated knockout in transgenic mice. Nat Neurosci 11, 721–728 (2008) doi:10.1038/nn.2118

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