Summary: Researchers have uncovered the molecular structure of three major complexes of glutamate receptors in the hippocampus. The findings shed new light on the mechanisms behind memory and learning in the brain.
Source: Oregon Health and Sciences University
Scientists have for the first time revealed the structure surrounding important receptors in the brain’s hippocampus, the seat of memory and learning.
The study, carried out at Oregon Health & Science University, published today in the journal Nature.
The new study focuses on the organization and function of glutamate receptors, a type of neurotransmitter receptor involved in sensing signals between nerve cells in the hippocampus region of the brain. The study reveals the molecular structure of three major complexes of glutamate receptors in the hippocampus.
The findings may be immediately useful in drug development for conditions such as epilepsy, said senior author Eric Gouaux, Ph.D., senior scientist in the OHSU Vollum Institute, Jennifer and Bernard Lacroute Endowed Chair in Neuroscience Research and an Investigator with the Howard Hughes Medical Institute.
“Epilepsy or seizure disorders can have many causes,” he said. “If one knows the underlying cause for a particular person’s seizure activity, then you may be able to develop small molecules to modulate that activity.”
Working with a mouse model, the OHSU researchers made the breakthrough by developing a chemical reagent based on monoclonal antibodies to isolate the receptor and the complex of subunits surrounding it. They then imaged the assemblage using state-of-the-art cryo-electron microscopy at the Pacific Northwest Cryo-EM Center, housed in OHSU’s South Waterfront campus in Portland.
Gouaux anticipates the technique will transform structural biology.
“It really opens the door to specifically target the molecules that need to be targeted in order to treat a particular condition,” he said. “A great deal of drug development is structure-based, where you see what the lock looks like and then you develop a key. If you don’t know what the lock looks like, then it’s much harder to develop a key.”
Previously, scientists had to rely on mimicking the actual receptors by artificially engineering receptors by combining DNA segments in tissue culture. However, that technique has obvious shortcomings.
“It doesn’t work perfectly because the real receptors are surrounded by a constellation of additional, sometimes previously unknown, subunits,” Gouaux said.
The new monoclonal antibody reagents, also developed at OHSU, enabled scientists to isolate actual glutamate receptors from the brain tissue of mice. They then were able to image those samples in near-atomic detail using cryo-EM, which allowed them to capture the entire assemblage of three types of glutamate receptors along with their auxiliary subunits.
“Previously, it’s been impossible to do this because we had no good way to isolate molecules and no way to see what they looked like,” Gouaux said. “So this is a super exciting development.”
Co-authors on the study included Jie Yu, Ph.D., a postdoctoral fellow in the Gouaux lab at OHSU; Prashant Rao, a graduate student in the Gouaux lab; Sarah Clark, Ph.D., a postdoctoral fellow in the Gouaux lab; Taekjip Ha, Ph.D., professor of biophysics and biomedical engineering at Johns Hopkins University; and Jaba Mitra, a graduate student in the Ha lab at Johns Hopkins.
Funding: Funding for the research was supported, in part, by the National Institute of Neurological Disorders and Stroke award number RO1NS038631.
Hippocampal AMPA receptor assemblies and mechanism of allosteric inhibition
AMPA-selective glutamate receptors mediate the transduction of signals between the neuronal circuits of the hippocampus. The trafficking, localization, kinetics and pharmacology of AMPA receptors are tuned by an ensemble of auxiliary protein subunits, which are integral membrane proteins that associate with the receptor to yield bona fide receptor signalling complexes.
Thus far, extensive studies of recombinant AMPA receptor–auxiliary subunit complexes using engineered protein constructs have not been able to faithfully elucidate the molecular architecture of hippocampal AMPA receptor complexes.
Here we obtain mouse hippocampal, calcium-impermeable AMPA receptor complexes using immunoaffinity purification and use single-molecule fluorescence and cryo-electron microscopy experiments to elucidate three major AMPA receptor–auxiliary subunit complexes.
The GluA1–GluA2, GluA1–GluA2–GluA3 and GluA2–GluA3 receptors are the predominant assemblies, with the auxiliary subunits TARP-γ8 and CNIH2–SynDIG4 non-stochastically positioned at the B′/D′ and A′/C′ positions, respectively.
We further demonstrate how the receptor–TARP-γ8 stoichiometry explains the mechanism of and submaximal inhibition by a clinically relevant, brain-region-specific allosteric inhibitor.