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Related Experiment Video

Updated: Jun 6, 2025

Subcellular Fractionation for the Isolation of Synaptic Components from the Murine Brain

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Published on: September 14, 2022

Functional specificity of liquid-liquid phase separation at the synapse.

Natalie J Guzikowski^1,2, Ege T Kavalali^3,4

¹Department of Pharmacology, Vanderbilt University, Nashville, TN, USA.

Nature Communications

|November 21, 2024

Summary

Liquid-liquid phase separation (LLPS) is crucial for precise synaptic signaling. Disrupting neuronal LLPS impairs action potential-dependent neurotransmission but not spontaneous release, highlighting LLPS

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Area of Science:

Neuroscience
Cell Biology
Biophysics

Background:

Synaptic function relies on precise molecular organization at the nanoscale.
Liquid-liquid phase separation (LLPS) is a key mechanism for subcellular organization.
The role of synaptic LLPS in neurotransmission remains largely undefined.

Purpose of the Study:

To investigate the impact of disrupting neuronal liquid-liquid phase separation (LLPS) on synaptic function.
To determine if LLPS is essential for action potential-dependent and spontaneous neurotransmission.
To elucidate the role of LLPS in organizing synaptic nanostructures.

Main Methods:

Utilized rat primary hippocampal cultures.
Disrupted neuronal LLPS using aliphatic alcohols.
Assessed action potential-dependent and spontaneous neurotransmission.
Examined synaptic vesicle pool clustering and recycling.
Analyzed the organization of active zone RIM1/2 and Munc13 nanoclusters.

Main Results:

Disruption of neuronal LLPS severely dysregulated action potential-dependent neurotransmission.
Spontaneous neurotransmission persisted despite LLPS disruption.
Synaptic LLPS was found to maintain synaptic vesicle pool clustering and recycling.
LLPS is critical for the precise organization of active zone nanoclusters (RIM1/2, Munc13).
LLPS supports fast calcium-dependent neurotransmitter release.

Conclusions:

Liquid-liquid phase separation (LLPS) is essential for the nano-organization of the synapse.
Synaptic LLPS specifically regulates action potential-evoked neurotransmitter release.
Disruption of synaptic LLPS spares spontaneous neurotransmission, indicating functional specificity.
In vitro observations of LLPS in larger condensates translate to functional roles at the nanoscale within synapses.