Many types of questions in neuroscience require the detection and mapping of synapses in the complex mammalian brain. A tool, mammalian GFP reconstitution across synaptic partners (mGRASP), offers a relatively easy, quick and economical approach to this technically challenging task. Here we describe in step-by-step detail the protocols for virus production, gene delivery, brain specimen preparation, fluorescence imaging and image analysis, calibrated substantially and specifically to make mGRASP-assisted circuit mapping (mGRASPing) practical in the mouse brain. The protocol includes troubleshooting suggestions and solutions to common problems. The mGRASP method is suitable for mapping mammalian synaptic connectivity at multiple scales: microscale for synapse-by-synapse or neuron-by-neuron analysis, and mesoscale for revealing local and long-range circuits. The entire protocol takes 5–6 weeks, including time for incubation and virus expression.
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We thank M. Son for technical support. This work was supported by the KIST Institutional Program (project no. 2E24210) and by the World Class Institute (WCI) Program of the Nation Research Foundation (NRF) of Korea (NRF grant no. WCI 2009-003).
The authors declare no competing financial interests.
AAV production and titer determination. (PDF 391 kb)
The zipped mGRASP detection package named 'puncta'. (ZIP 160 kb)
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Feng, L., Kwon, O., Lee, B. et al. Using mammalian GFP reconstitution across synaptic partners (mGRASP) to map synaptic connectivity in the mouse brain. Nat Protoc 9, 2425–2437 (2014). https://doi.org/10.1038/nprot.2014.166
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