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Getting the skinny on glutamate
GOTHENBURG, Sweden—Glutamate, like serotonin and dopamine, is a neurotransmitter. Despite being the brain’s major excitatory neurotransmitter, specifics about its involvement in various neurological functions is limited due to a lack of technology that can rapidly measure its use within the brain.
That technological hurdle has been largely overcome by a team of scientists from Chalmers University of Technology and Gothenburg University, who recently published a pair of papers detailing a method for counting the molecules of glutamate released as signals are transferred between brain cells. The most recent paper was published in the Journal of the American Chemical Society under the title “Counting the number of glutamate molecules in single synaptic vesicles.”
“Here, we introduce a new enzyme-based amperometric sensor technique for the counting of glutamate molecules stored inside single synaptic vesicles,” the authors explained. “In this method, an ultra-fast enzyme-based glutamate sensor is placed into a solution of isolated synaptic vesicles, which stochastically rupture at the sensor surface in a potential-dependent manner at a constant negative potential. The continuous amperometric signals are sampled at high speed (10 kHz) to record sub-millisecond spikes, which represent glutamate release from single vesicles that burst open. Glutamate quantification is achieved by a calibration curve that is based on measurements of glutamate release from vesicles pre-filled with various glutamate concentrations. Our measurements show that an isolated single synaptic vesicle encapsulates about 8,000 glutamate molecules and is comparable to the measured exocytotic quantal glutamate release in amperometric glutamate sensing in the nucleus accumbens of mouse brain tissue.”
The enzyme is only a molecule thick and is placed on a nano-structured sensor surface, making it roughly a thousand times faster than previous sensing techniques. And that speed is necessary, because glutamate is released from synaptic vesicles in less than a thousandth of a second.
“When we started, everybody said ‘this will never work’. But we didn’t give in. Now we have a beautiful example of how multi-disciplinary basic science can yield major breakthroughs, and deliver real benefit,” commented Ann-Sofie Cans, an associate professor of chemistry at Chalmers and leader of the research group.
This research has overturned more than a few previous suppositions about glutamate. One discovery is that the amount of glutamate contained in a given synaptic vesicle is higher than previously thought, and is in fact essentially quantitatively equal to serotonin and dopamine. According to Cans, “it seems that transport and storage of glutamate in synaptic vesicles is not as different as we thought, when compared with other neurotransmitters like serotonin and dopamine.” In addition, nerve cells can adjust the strength of their chemical signals by throttling glutamate release from single synaptic vesicles.
“The level of measurement offered by this ultra-fast glutamate sensor opens up countless possibilities to truly understand the function of glutamate in health and disease. Our knowledge of the brain function, and dysfunction, is limited by the experimental tools we have, and this new ultra-fast tool will allow us to examine neuronal communication at a level we did not have access to before,” said Karolina Patrycja Skibicka, an associate professor of neuroscience and physiology at Gothenburg University. “The new finding, that glutamate-based communication is regulated by the quantity of glutamate released from synaptic vesicles, begs the question of what happens to this regulation in brain diseases thought to be linked to glutamate—for example, epilepsy.”
The ability to better measure glutamate usage in the brain could lead to more answers on a variety of neurological conditions. As a neurotransmitter, glutamate plays a role in functions such as cognition, memory and learning, but an overabundance of this molecule can lead to neurotoxicity and is implicated in a number of conditions. Excessive amounts of glutamate can lead to conditions such as stroke, multiple sclerosis, Parkinson’s disease, amyotrophic lateral sclerosis and Alzheimer’s disease.
As noted in the book Excitatory Amino Acid Transmission in Health and Disease, “Glutamate can cause neuronal cell death by a mechanism referred to as excitotoxicity, a process of cell death of neurons and sonic glial cells resulting from excessive or prolonged activation of excitatory amino acid receptors. Excitotoxicity is a convergence point in the neurodegenerative cascade of many acute and chronic neurodegenerative disorders. In fact, a role for excitotoxicity has been implicated in the etiology of many neurodegenerative diseases, including ischemic stroke, Alzheimer’s disease, Parkinson’s disease, amyotrophic lateral sclerosis, and epilepsy. Because Glu can cause neuronal degeneration, there are two major therapeutic approaches for neurological diseases: prevent the degeneration of neurons by controlling excitotoxicity, and modulate glutamatergic synaptic transmission and function in the surviving neurons by positive or negative regulation of glutamate receptors.”
With this new technology enabling researchers to precisely measure how much glutamate is being released, such a therapeutic approach may one day become an option.
Funding for this work came from the Swedish Research Council, the Swedish Brain Foundation, Ragnar Söderberg Foundation, the Novo Nordisk Foundation, the Wallenberg Center for Molecular and Translational Medicine at the University of Gothenburg, Ernst and Fru Rådman Colliander Stiftelse, Wilhelm and Martina Lundgren Stiftelse and Magnus Bergvall Stiftelse.