Abstract
Stoichiometric labeling of endogenous synaptic proteins for high-contrast live-cell imaging in brain tissue remains challenging. Here, we describe a conditional mouse genetic strategy termed endogenous labeling via exon duplication (ENABLED), which can be used to fluorescently label endogenous proteins with near ideal properties in all neurons, a sparse subset of neurons, or specific neuronal subtypes. We used this method to label the postsynaptic density protein PSD-95 with mVenus without overexpression side effects. We demonstrated that mVenus-tagged PSD-95 is functionally equivalent to wild-type PSD-95 and that PSD-95 is present in nearly all dendritic spines in CA1 neurons. Within spines, while PSD-95 exhibited low mobility under basal conditions, its levels could be regulated by chronic changes in neuronal activity. Notably, labeled PSD-95 also allowed us to visualize and unambiguously examine otherwiseunidentifiable excitatory shaft synapses in aspiny neurons, such as parvalbumin-positive interneurons and dopaminergic neurons. Our results demonstrate that the ENABLED strategy provides a valuable new approach to study the dynamics of endogenous synaptic proteins in vivo.
Original language | English (US) |
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Pages (from-to) | 16698-16712 |
Number of pages | 15 |
Journal | Journal of Neuroscience |
Volume | 34 |
Issue number | 50 |
DOIs | |
State | Published - Dec 10 2014 |
Keywords
- Cell-type-specific labeling
- Conditional knock-in
- Live imaging
- Protein labeling
- Psd-95
- Sparse labeling
ASJC Scopus subject areas
- General Neuroscience