Assessing protein distribution and dendritic spine morphology relationships using structured illumination microscopy in cultured neurons

Summary

This research provides a detailed step-by-step guide for scientists to visualize and measure how proteins are organized inside tiny structures called dendritic spines, which are the connection points between nerve cells in the brain. Using advanced microscopy techniques, researchers can see how different proteins cluster together and how this organization relates to the shape and size of these synaptic connections. Understanding protein arrangement in dendritic spines is important because it helps explain how brain cells communicate and adapt, which has implications for learning, memory, and neurological disorders.

Background

Dendritic spines are protrusions on dendrites that form the postsynaptic aspect of excitatory connections in the brain. Spine morphology is associated with synaptic functional strength, and the spatial regulation of protein nanodomains within dendritic spines is an important determinant of spine structure and function.

Objective

To present a protocol for resolving the nanoscale localization of proteins within dendritic spines using structured illumination microscopy. The protocol enables quantitative assessment of protein distribution as a function of spine morphology and spatial overlap between distinct proteins.

Results

The workflow produces comprehensive dendritic spine morphology data including spine area, head area, neck area, and neck length for spine type determination. Additionally, the protocol provides protein nanodomain information including area, intensity, location within spine sub-compartments, and colocalization patterns with other proteins.

Conclusion

This protocol provides a systematic workflow for analyzing the sub-compartmentalization of protein nanodomains in dendritic spines as a function of spine morphology. The method enables quantitative assessment of whether protein distribution is uniform across spine types or influenced by morphology, with implications for understanding synaptic function and structure.
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