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Open Access Highly Accessed Research article

Nanoscopic spine localization of Norbin, an mGluR5 accessory protein

Linda Westin1, Matthias Reuss2, Maria Lindskog3, Anita Aperia1 and Hjalmar Brismar12*

Author Affiliations

1 Department of Women’s and Children’s Health, Science for Life Laboratory, Karolinska Institutet, 17165 Solna, Sweden

2 Department of Applied Physics, Science for Life Laboratory, Royal Institute of Technology, 10691 Stockholm, Sweden

3 Department of Neuroscience, Karolinska Institutet, 17177 Stockholm, Sweden

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BMC Neuroscience 2014, 15:45  doi:10.1186/1471-2202-15-45

Published: 26 March 2014

Abstract

Background

Norbin is a neuron-specific, cytosolic protein that interacts with the metabotropic glutamate receptor 5 (mGluR5) and has a profound impact on mGluR5 signaling. Yet, little is known about its synaptic distribution.

Results

Here we have analyzed the spatial relationship between Norbin, postsynaptic density protein 95 (PSD-95), actin and mGluR5 in spines using super-resolution microscopy. Norbin was found to have a high degree of colocalization with actin and a lower degree of colocalization with PSD-95. Co-immunoprecipitation studies confirmed that interaction occurs between Norbin and actin, but not between Norbin and PSD-95. Norbin was also found to have a high degree of colocalization with the perisynaptically located mGluR5. Findings based on structured illumination microscopy (3D-SIM) of exogenous expressed Norbin-GFP were confirmed by stimulated emission depletion microscopy (STED) of immunolabeled endogenous Norbin.

Conclusions

Norbin associates with actin rather than with PSD-95 in dendritic spines. Results regarding protein localization and colocalization performed with conventional confocal microscopy must be interpreted with great caution. The now available super-resolution microscopy techniques provide more accurate information about sub-cellular protein localization than previously was possible.

Keywords:
Super-resolution microscopy; Colocalization; Norbin; Dendritic spines