By chilling a molecule on the surface of brain cells to around minus 180 degrees Celsius — nearly double the cold of the coldest spots in Antarctica — researchers have discovered how a commonly used epilepsy drug reduces the excitability of brain cells and helps to manage, but not cure, seizures.
By super cooling a molecule on the surface of brain cells down to about minus 180 degrees Celsius — nearly twice as cold as the coldest places in Antarctica — scientists at Johns Hopkins Medicine say they have determined how a widely-used epilepsy drug works to dampen the excitability
Brain cells are affected by seizures, but they cannot be cured by drugs. The research, which was published in Nature Structural & Molecular Biology, links the activity of the epilepsy drug perampanel to the movements of the AMPA receptor on brain cells. The researchers believe that this discovery could lead to the development of new drugs for treating other neurological conditions such as Alzheimer’s disease, schizophrenia, learning disabilities, glioblastoma brain cancers, and chronic pain.
The glutamate, a major neurotransmitter in the brain, plays a crucial role in activating brain cells through its connection with the AMPA protein. This interaction is similar to a Pac-man game, where AMPA receptors engulf the glutamate molecules. A single AMPA receptor can bind up to four glutamate molecules, creating a connection that allows positively-charged ions to flood into the neuron, thereby exciting and activating it.
According to Edward Twomey, Ph.D., an assistant professor of biophysics, AMPA receptors and glutamate are essential for various life processes, such as learning, memory, and encoding experiences.biophysical chemistry at the Johns Hopkins University School of Medicine. “Most neurological diseases can be linked in some way to AMPA receptors and glutamate.”
Twomey was approached by neuroscientist Richard Huganir, Ph.D., who has been studying AMPA receptors for 40 years, to work together on research aimed at gaining a better understanding of the receptors’ structure and their process of binding with glutamate.
Overactivation (excitation) of AMPA receptors is known to lead to epilepsy. Perampanel, which focuses on the AMPA receptor, is currently the only medication approved by the U.S. Food and Drug Administration to target AMPA receptors, but numerous pharmaceutical companies are also interested in developing medications that target this receptor.The researchers are working on compounds that are similar to this drug, according to Twomey. The drug was first discovered in the 1980s, and its exact mechanism has been a long-standing mystery. Huganir, a Bloomberg Distinguished Professor of Neuroscience and Psychological and Brain Sciences and director of the Solomon H. Snyder Department of Neuroscience, explains that they know the drug does not block or get stuck in the receptor’s ion channels. Other scientists have found where perampanel binds to AMPA receptors in pockets around the ion channel, but they have not discovered how this connection disrupts ion flow.To understand the process, the scientists used cryo-electron microscopy (cryoEM), a technique that has become a powerful tool for studying structures a million times smaller than the width of a human hair over the past two decades.
W. Dylan Hale, Ph.D., a postdoctoral fellow at Johns Hopkins, conducted most of the experiments and analysis in the Beckman Center for CryoEM, where biological molecules are super-cooled and imaged at different time points. Hale worked in the Twomey and Huganir labs.
For the research, the team examined millions of images of AMPA receptors in brain cells from mice and rat models, as well as their interactions.Using the original version of the perampanel drug, GYKI-52466, researchers studied the interactions at a very small scale, about 1 to 2 angstroms, according to Twomey. An angstrom is 1 10 billionth of a meter. The researchers examined the binding of the GYKI-52466 drug with and without glutamate. They also conducted electrical recordings of the ion flow and physiological studies in mice to complement the cryoEM images. Additionally, the scientists utilized artificial intelligence and machine learning tools to average and combine the cryoEM images into a 3D reconstruction of the receptor. When glutamate binds to the AMPA recepThe researchers discovered that when the AMPA receptor is in one of its four positions, a part of the receptor pulls down and opens the ion channel, allowing ions to flow through. This is similar to how a pull chain releases water from a shower head. The study found that two of the four positions where glutamate binds are crucial for the drug GYKI-52466 to block the ion flow. According to Twomey, the drug binds to the AMPA receptor and stops the ion channel from opening by preventing glutamate from pulling on the strand that opens the ion channel. This action seems to separate the glutamate binding regions from each other and puts the receptor into a desensitized state.
Huganir also intends to collaborate with Twomey to utilize cryoEM for investigating the effects of mutations on the AMPA receptor.
“We aim to identify the structural abnormalities in the receptor that lead to its dysfunction,” Huganir explains. “In principle, we could develop medications to enhance the receptor’s activity in order to treat conditions associated with changes in its structure.”
Along with Hale, Twomey, and Huganir, the study also involved contributions from researchers Alejandra Montaño Romero and Albert Lau at Johns Hopkins, as well as Cuauhtemoc Gonzalez and Vasanthi Jayaraman at the University of Texas Health Science Center at Houston.
The research was funded by the Searle Scholars Program, the Diana Helis Henry Medical Research Foundation, and the National Institutes of Health (R37 NS036715, R01 GM094495, R35 GM122528, F99NS130928, K99 MH132811).
