Resolving Native GABAA Receptor Structures from the Human Brain: A Breakthrough Study

Academic Background

GABAA receptors (γ-aminobutyric acid type A receptors) are among the most important inhibitory neurotransmitter receptors in the brain, responsible for regulating fast inhibitory signaling in neurons. These receptors are not only key drug targets for treating epilepsy, anxiety, depression, and insomnia but are also widely studied for their role in the mechanisms of anesthetic drugs. GABAA receptors are composed of 19 different subunits, forming pentameric ligand-gated ion channels. Although previous studies have revealed the structure and function of some GABAA receptors through recombinant expression and mouse models, the subunit composition and three-dimensional structure of native GABAA receptors in the human brain remain unclear. In particular, the subunit combinations of GABAA receptors in the human brain are more complex than those in rodents, and their interactions with auxiliary proteins have not been fully explored.

To address this issue, researchers isolated GABAA receptors containing the α1 subunit from brain tissue resected from epilepsy patients and resolved the three-dimensional structures of 12 native GABAA receptors using cryo-electron microscopy (cryo-EM). This study not only revealed the diversity of GABAA receptors in the human brain but also uncovered unexpected binding sites for antiepileptic drugs on the receptors, providing a crucial structural foundation for understanding GABAA receptor signaling and targeted pharmacology.

Source of the Paper

This paper was co-authored by Jia Zhou, Colleen M. Noviello, Jinfeng Teng, Haley Moore, Bradley Lega, and Ryan E. Hibbs, with research teams from UC San Diego, UT Southwestern Medical Center, and other institutions. The paper was published in Nature in 2024 under the title Resolving native GABAA receptor structures from the human brain.

Research Process and Results

1. Sample Collection and Processing

The research team obtained samples from brain tissue resected from 81 epilepsy patients, primarily from the temporal and frontal lobes. The samples were divided into two groups: the first group included tissue from 45 patients, and the second group included tissue from 36 patients. All samples were flash-frozen immediately after resection to ensure the integrity of protein structures.

2. Purification of GABAA Receptors

The researchers used a high-affinity antibody fragment (Fab 1F4) targeting the α1 subunit to purify GABAA receptors containing the α1 subunit from brain tissue. During purification, a detergent (lauryl maltose neopentyl glycol, LMNG) was used to maintain interactions between the receptors and synaptic binding partners. The purified receptors were then reconstituted into lipid nanodiscs for cryo-EM analysis.

3. Cryo-EM Data Collection and Processing

The researchers used cryo-EM to image the purified GABAA receptors at high resolution. Through two-dimensional and three-dimensional classification, they successfully resolved the three-dimensional structures of 12 different GABAA receptor subunit assemblies. The overall resolution of these structures ranged from 2.5 to 3.3 Å, sufficient to distinguish details of different subunits.

4. Diversity of Subunit Assemblies

The study identified multiple GABAA receptor subunit assemblies, with the most common being β2–α1–β2–α1–γ2. Additionally, several assemblies containing α2, α3, β1, β3, and γ2 subunits were discovered. These combinations revealed the high diversity of GABAA receptors in the human brain, particularly the involvement of the β3 subunit, which had not been extensively reported in previous studies.

5. Discovery of Drug-Binding Sites

During the structural analysis, the researchers observed drug-like densities at multiple subunit interfaces. Further experiments showed that the antiepileptic drugs lamotrigine and levetiracetam could bind to GABAA receptors and modulate their function. Notably, the binding site of lamotrigine overlapped with that of benzodiazepines, providing new insights into its antiepileptic mechanism.

6. Interactions with Auxiliary Proteins

Through mass spectrometry and cryo-EM data, the researchers also identified interactions between GABAA receptors and auxiliary proteins, such as neuroligin 2 and garlh4. These auxiliary proteins play important roles in regulating the localization and function of GABAA receptors, further revealing the complex regulatory mechanisms of receptors at synapses.

Research Conclusions and Significance

This study is the first to systematically resolve the three-dimensional structures of native GABAA receptors in the human brain, revealing the diversity and complexity of their subunit assemblies. The research not only provides a structural foundation for understanding GABAA receptor signaling mechanisms but also uncovers new binding sites for antiepileptic drugs, offering important insights for developing more precise drug-targeting strategies.

Additionally, the study revealed interactions between GABAA receptors and auxiliary proteins, providing new perspectives on the functional regulation of receptors at synapses. These findings have significant scientific value and open new directions for future drug development and the treatment of neurological disorders.

Research Highlights

1. First Resolution of Native GABAA Receptor Structures in the Human Brain: The study revealed 12 different subunit assemblies, providing critical data on the diversity of GABAA receptors.
2. Discovery of New Drug-Binding Sites: The binding sites of lamotrigine and levetiracetam revealed potential mechanisms of action for these drugs.
3. Revealing Interactions with Auxiliary Proteins: The study identified the important roles of auxiliary proteins such as neuroligin 2 and garlh4 in regulating receptor function.
4. Application of High-Resolution Cryo-EM: The research demonstrated the powerful capabilities of cryo-EM in resolving complex protein structures.

Additional Valuable Information

The study also found that subunit assemblies of GABAA receptors varied among patients, potentially related to factors such as sex, age, epilepsy duration, and medication history. Future research could further explore the impact of these factors on the structure and function of GABAA receptors, providing a basis for personalized treatment.

This study represents a significant breakthrough in understanding the structure and function of GABAA receptors and opens new research directions for the treatment of neurological disorders.