Understanding how glutamate receptors are organised in the human brain, and how that organisation changes in disease, has long been limited by the lack of tools capable of characterising receptor complexes directly from small amounts of human tissue. A central gap has been whether metabotropic glutamate receptors (mGluRs), known to form both homodimers and heterodimers, co-exist as both assemblies in living human brain, how their composition varies across brain regions, and whether these properties shift in psychiatric conditions like depression.

El-Baba et al. address these gaps by developing a nanobody-based immunoprecipitation platform that captures multiple native synaptic receptor complexes from single, anatomically dissected human and mouse brain specimens in under 24 hours. This platform feeds into an integrated mass spectrometry (MS) workflow combining native MS, native top-down MS, glycoproteomics, lipidomics, and chemical crosslinking MS, enabling detailed molecular dissection of intact endogenous receptor complexes, including stoichiometry, glycosylation, lipid interactions, and protein binding partners.

For the crosslinking MS component, peptides were separated on an IonOpticks Aurora® Ultimate™ 25×75 CSI C18 UHPLC column coupled to a nanoElute 2 and timsTOF Ultra 2 – a configuration chosen to deliver the sensitivity and peak capacity needed to confidently identify low-abundance crosslinked peptide pairs from complex brain immunoprecipitates.

Leveraging this workflow, El-Baba et al. demonstrate for the first time that mGluR2/3 heterodimers are abundant in human prefrontal cortex (~70% in OFC, ~50% in sgACC), with region-specific binding partners including CRMP2 directly interacting with the mGluR3 subunit in the sgACC. mGluR2/3 heterodimer abundance is selectively elevated in the OFC of individuals with severe depression, without any change in total receptor expression levels.

These findings suggest that receptor assembly state, rather than expression level, is a molecular correlate of depression, potentially reframing how glutamatergic drug targets are conceptualised and how precision medicine approaches to psychiatric disorders might be developed.

Publication
bioRxiv

Authors

Tarick J. El-Baba, Corinne A. Lutomski, Jack L. Bennett, Sophie A. S. Lawrence, Sean A. Burnap, Frances I. Butroid, Olivia B. Ramsay, Titas Radzevičius, Di Wu, Haigang Song, Kenny L. Chan, Lyonna F. Parise, Eric M. Parise, Weston B. Struwe, James W. Murrough, Scott J. Russo, & Carol V. Robinson;

Title

Molecular dissection of protein complexes isolated from sections of human brain