Discovery may lead to relief for victims of a range of neurological disorders.
Researchers at the University of Toronto have identified a protein complex that helps control the balance between excitation and inhibition in the brain — a balance whose disruption is linked to disorders such as epilepsy, schizophrenia, autism spectrum disorder and neuropathic pain. This finding marks an important advance toward therapies that could better restore normal brain function.
“Neurons communicate at synapses, and those signals can either excite or inhibit other neurons,” said Professor Melanie Woodin of the Department of Cell and Systems Biology at the University of Toronto, lead investigator of the study published in Cell Reports. “When excitation overwhelms inhibition, or vice versa, normal circuit function breaks down and pathological events such as seizures can follow. We have identified a key protein complex that directly regulates this excitation–inhibition balance at the cellular level.”
The newly characterized complex links three proteins — the potassium-chloride cotransporter KCC2, the auxiliary subunit Neto2, and the kainate receptor subunit GluK2. KCC2 maintains low intracellular chloride and is essential for inhibitory synaptic transmission. GluK2 is part of the kainate-type glutamate receptors that mediate excitation, and Neto2 is a modulatory protein that interacts with both KCC2 and GluK2. Until now, KCC2 and kainate receptors were thought to occupy separate cellular compartments and operate independently; the discovery that they coexist in a functional complex reveals a direct molecular mechanism for coordinating inhibitory and excitatory signaling.
“Demonstrating that these proteins directly interact and can co-regulate each other provides the first clear example of a molecular system capable of tuning excitation–inhibition balance between neurons themselves,” said Vivek Mahadevan, a PhD candidate in Woodin’s laboratory and the study’s lead author.
The team used a combination of biochemical methods, fluorescence imaging and electrophysiology on mouse brain tissue. A central technique was Blue Native PAGE, an advanced gel electrophoresis method that preserves native protein complexes and reveals macromolecular assemblies that standard denaturing gels would disrupt. By maintaining the biochemical environment in which complexes exist, Blue Native PAGE allowed the researchers to identify the intact KCC2–kainate receptor complex in neuronal tissue.
Functional experiments showed that kainate receptor subunits are important for KCC2 oligomerization and surface expression. Both acute and chronic genetic deletion of kainate receptor components reduced KCC2 function and weakened inhibitory synaptic transmission in hippocampal neurons. These results identify kainate receptors as direct regulators of KCC2 and, by extension, as contributors to chloride homeostasis and inhibitory strength in the mature central nervous system.
“Understanding the molecular machinery that sets the excitation–inhibition balance opens clear avenues for drug discovery,” said Woodin. “Current epilepsy treatments largely suppress symptoms like seizures but do not cure the underlying imbalance. Targeting the proteins that modulate KCC2 and kainate receptor interactions could lead to therapies that prevent seizures and other dysfunctions from arising in the first place.”
Mahadevan added, “Pinpointing the cellular mechanisms that govern inhibitory strength was essential. With KCC2 now identified as a key mediator modulated by excitatory receptor partners, future research can explore how its regulation sometimes fails in disease and how those failures might be corrected.”
The research team included collaborators from the Hospital for Sick Children Research Institute, Vanderbilt University School of Medicine, Jewish General Hospital at McGill University, and the Institute of Biomedicine, Anatomy at the University of Helsinki. The findings are reported in the article titled “Kainate Receptors Coexist in a Functional Complex with KCC2 and Regulate Chloride Homeostasis in Hippocampal Neurons,” published online in Cell Reports. Funding for the study came from the Canadian Institutes of Health Research, the U.S. National Institutes of Health, and the Academy of Finland.
Submitted to NeuroscienceNews.com by Melanie Woodin
Contact: Melanie Woodin – University of Toronto
Source: University of Toronto press release
Image Source: Image credited to geralt and is in the public domain
Original Research: Full open-access article in Cell Reports: “Kainate Receptors Coexist in a Functional Complex with KCC2 and Regulate Chloride Homeostasis in Hippocampal Neurons” by Vivek Mahadevan et al.
Kainate Receptors Coexist in a Functional Complex with KCC2 and Regulate Chloride Homeostasis in Hippocampal Neurons
KCC2 is a neuron-specific K+-Cl− cotransporter required to maintain low intracellular chloride concentrations, which are essential for fast inhibitory synaptic transmission in the mature central nervous system. Despite KCC2’s critical role, cellular mechanisms controlling its expression and function have been poorly understood. Using blue native-polyacrylamide gel electrophoresis (BN-PAGE) to analyze native protein assemblies in vivo, the researchers found KCC2 in a macromolecular complex with kainate-type glutamate receptors (KARs). KAR subunits were necessary for KCC2 oligomerization and for maintaining KCC2 at the cell surface. Genetic deletion of KARs reduced KCC2 function and impaired synaptic inhibition in hippocampal neurons, revealing KARs as regulators of KCC2 and highlighting a tight molecular interplay between excitation and inhibition.