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Redox-Activated Probes Enable High-Contrast Live Imaging of Native Postsynaptic Scaffolds.

Christiane Huhn1,2, Clémence Mille3, Sheng-Yang Ho4

  • 1Rudolf Virchow Center for Integrative and Translational Bioimaging, University of Würzburg, Würzburg, Germany.

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Summary
This summary is machine-generated.

Researchers developed Sylives, synthetic fluorescent peptides for high-contrast visualization of postsynaptic scaffolds in living neurons. This method bypasses genetic engineering, offering a new tool for studying synaptic plasticity.

Keywords:
PSDfluorescent probegephyrinlive‐cell imagingneuronpeptidesynapse

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Area of Science:

  • Neuroscience
  • Cell Biology
  • Biochemistry

Background:

  • Direct visualization of postsynaptic scaffolds is crucial for understanding synaptic dynamics and plasticity.
  • Current live imaging methods are limited by genetic engineering or incompatible reagents, leading to poor contrast and delivery issues.

Purpose of the Study:

  • To introduce Sylives, compact synthetic fluorescent peptides for high-contrast live imaging of inhibitory and excitatory postsynaptic scaffolds in native neurons.
  • To overcome limitations of existing probes and delivery strategies for accurate visualization of synaptic structures.

Main Methods:

  • Development of Sylives, redox-cleavable CPP-probe conjugates designed for efficient cytosolic delivery and intracellular reduction-restored scaffold binding.
  • Quantitative evaluation of Sylive uptake and off-target binding to define parameters for effective intracellular delivery.
  • Validation of Sylive specificity and near-traceless uptake through comparison with transiently expressed proteins and immunolabeling.

Main Results:

  • Sylives enable high-contrast live imaging of gephyrin and PSD-95 postsynaptic scaffolds in native neurons without genetic modification.
  • Pre-purification of redox-cleavable conjugates prevents side-product formation, ensuring reliable delivery and restored binding.
  • Defined transferrable parameters for effective intracellular delivery, achieving nanomolar probe levels versus micromolar CPP.

Conclusions:

  • Sylives offer a modular platform for targeting intracellular proteins in living neurons, enabling specificity-restored labeling of endogenous postsynaptic sites.
  • This approach overcomes major constraints of existing live synapse visualization techniques.
  • Sylives facilitate high-contrast imaging without genetic manipulation, advancing the study of synaptic plasticity.