Patch-Clamp Single-Cell Proteomics in Acute Brain Slices: A Framework for Recording, Retrieval, and Interpretation.

Area of Science:

• Neuroscience
• Proteomics
• Electrophysiology

Background:

• Single-cell proteomics (SCP) offers deep molecular insights into individual neurons but faces challenges in acute brain slices.
• Patch-clamp electrophysiology records neuronal function but requires physical neuron retrieval, potentially affecting proteomic data quality.

Purpose of the Study:

• To develop a framework for contextualizing single-cell proteomics outcomes from patch-clamped neurons.
• To assess the impact of neuron retrieval quality on proteomic measurements.

Main Methods:

• Utilized an indiscriminate shotgun proteomics strategy on patch-clamped rat medial prefrontal cortex pyramidal neurons.
• Correlated electrophysiological parameters (e.g., capacitance) and retrieval characteristics with proteomic data (protein identifications, synaptic enrichment).

Main Results:

• Neuron capacitance during retrieval correlated with proteome yield, suggesting soma size influences protein identification.
• Preservation of neuronal spiking during retrieval was linked to greater synaptic enrichment and recovery of transmembrane proteins.
• Torn or aspirated neurons yielded smaller proteomes with poor synaptic representation.

Conclusions:

• The study demonstrates that patch-SCP can assess soma retrieval quality.
• Provides a framework for interpreting single-neuron proteomic data by considering electrophysiological context and retrieval fidelity in semi-intact circuits.