# Single-Particle Cryo-EM Characterization of AMPA Receptor Functional States

> **NIH NIH R01** · COLUMBIA UNIVERSITY HEALTH SCIENCES · 2024 · $398,780

## Abstract

PROJECT SUMMARY
AMPA-subtype ionotropic glutamate receptors mediate fast signaling between neurons and contribute to high
cognitive processes. Since AMPA receptors are also implicated in numerous neurological disorders, including
Alzheimer’s disease, amyotrophic lateral sclerosis, epilepsy, and ischemia, the ability to regulate them
represents an important clinical goal. However, there is an unmet need for drugs to regulate AMPA receptor
activity in pathological conditions, which highlights a profound gap in our knowledge of AMPA receptor
structure and function. Our long-term goal is to understand how AMPA receptor molecular machinery operates
at the atomic level. We plan to study AMPA receptor structure and function using single-particle cryo-electron
microscopy (cryo-EM). Advances in cryo-EM that were signified by the “resolution revolution” brought us a
number of AMPA receptor structures in different gating conformations and in complex with several small
molecules and regulatory proteins. Nevertheless, only a little fraction of AMPA receptor conformational states
and binding partners have been characterized structurally and the resolution of the reported structures remains
relatively low, limiting our ability to see the atomic details. We, therefore, plan to use cryo-EM advances to fill
up this knowledge gap and to focus on the following Specific Aims: (1) unravel structural principles of AMPA
receptor regulation by auxiliary subunits, (2) reveal molecular determinants of AMPA receptor activation and
conductance, and (3) determine molecular mechanisms of AMPA receptor desensitization. To reach our
research goals, we will use cryo-EM to obtain structures of AMPA receptors alone or in complex with different
auxiliary proteins, in the presence of agonists, competitive antagonists, positive or negative allosteric
modulators, and in conditions favoring various conformational states. We will use Fluorescence-detection Size
Exclusion Chromatography (FSEC) and thermostability assays to assess protein expression, assembly,
homogeneity, and stability. We will also employ site-directed mutagenesis combined with single-channel and
whole-cell patch-clamp electrophysiological recordings to study functional mechanisms and to critically test our
structural models. Reaching our research goals will have a significant impact on understanding the
mechanisms of excitatory neurotransmission and will provide molecular-level knowledge essential to facilitate
the design of new drugs for the treatment of neurological disorders.

## Key facts

- **NIH application ID:** 10839881
- **Project number:** 5R01NS107253-07
- **Recipient organization:** COLUMBIA UNIVERSITY HEALTH SCIENCES
- **Principal Investigator:** Alexander Sobolevsky
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2024
- **Award amount:** $398,780
- **Award type:** 5
- **Project period:** 2018-08-01 → 2028-05-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10839881

## Citation

> US National Institutes of Health, RePORTER application 10839881, Single-Particle Cryo-EM Characterization of AMPA Receptor Functional States (5R01NS107253-07). Retrieved via AI Analytics 2026-05-28 from https://api.ai-analytics.org/grant/nih/10839881. Licensed CC0.

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