Major discovery about mammalian brain surprises researchers

Major discovery about mammalian brain surprises researchers

Overview: V-ATPase, a vital enzyme that enables neurotransmission, can be switched on and off randomly, even with hours-long breaks.

Source: University of Copenhagen

In a new breakthrough to understand more about the mammalian brain, researchers at the University of Copenhagen have made an incredible discovery. Namely, a vital enzyme that enables brain signals turns on and off randomly, even taking hour-long “breaks from work.”

These findings could have a major impact on our understanding of the brain and drug development.

Today the discovery is on the cover of Nature.

Millions of neurons are constantly messaging each other to shape thoughts and memories and allow us to move our bodies at will. When two neurons meet to exchange a message, neurotransmitters are transported from one neuron to the other using a unique enzyme.

This process is crucial for neuronal communication and the survival of all complex organisms. Until now, researchers worldwide thought that these enzymes were active at all times to continuously transmit essential signals. But this is far from the case.

Using an innovative method, researchers from the Department of Chemistry at the University of Copenhagen have closely studied the enzyme and found that its activity switches on and off at random intervals, contradicting our previous understanding.

“This is the first time anyone has studied these mammalian brain enzymes molecule by molecule, and we are impressed with the result. Contrary to popular belief, and unlike many other proteins, these enzymes can take minutes to hours to stop working, yet the brains of humans and other mammals are miraculously able to function,” says Professor Dimitrios Stamou, who led the study from the Center for Geometrically Constructed Cellular Systems at the Department of Chemistry at the University of Copenhagen.

Until now, such studies have been carried out with highly stable enzymes from bacteria. Using the new method, the researchers examined mammalian enzymes isolated from rat brains for the first time.

The study was published today in Nature.

Enzyme switching can have far-reaching consequences for neuronal communication

Neurons communicate using neurotransmitters. To transfer messages between two neurons, neurotransmitters are first pumped into small membrane vesicles (synaptic vesicles). The bladders act as containers that store the neurotransmitters and only release them between the two neurons when it’s time to transmit a message.

The central enzyme of this study, known as V-ATPase, is responsible for providing the energy for the neurotransmitter pumps in these containers. Without it, neurotransmitters would not be pumped into the containers and the containers would not be able to send messages between neurons.

But the study shows that there is only one enzyme in each container; when this enzyme shuts off, there would be no more energy to drive the loading of neurotransmitters into the containers. This is a completely new and unexpected discovery.

“It is almost incomprehensible that the extremely critical process of loading neurotransmitters into containers is delegated to only one molecule per container. Especially when we discover that these molecules are switched off 40% of the time,” says Professor Dimitrios Stamou.

This shows a v-atpases on a synaptic vesicle
The cover illustration shows vacuolar-type adenosine triphosphatases (V-ATPases, large blue structures) on a synaptic vesicle of a nerve cell in the mammalian brain. Image: C. Kutzner, H. Grubmüller and R. Jahn/Max Planck Institute for Multidisciplinary Sciences. Credits: C. Kutzner, H. Grubmüller and R. Jahn/Max Planck Institute for Multidisciplinary Sciences.

These findings raise many intriguing questions:

“Does cutting off the energy source of the containers mean that many of them are indeed depleted of neurotransmitters? Would a high proportion of empty containers significantly affect communication between neurons? If so, could that be a “problem” that neurons have evolved to get around, or could it possibly be an entirely new way of encoding important information in the brain? Only time will tell,” he says.

A revolutionary method to screen drugs for the V-ATPase

The V-ATPase enzyme is an important drug target because it plays a critical role in cancer, cancer metastases and several other life-threatening diseases. Thus, the V-ATPase is a lucrative target for anticancer drug development.

Existing assays to screen drugs for V-ATPase are based on simultaneous averaging of the signal from billions of enzymes. Knowing the average effect of a drug is sufficient as long as an enzyme works constantly over time or when enzymes work together in large numbers.

“However, we now know that neither is necessarily true for the V-ATPase. As a result, it has suddenly become critical to have methods that measure the behavior of individual V-ATPases in order to understand and optimize the desired effect of a drug,” says first author of the paper Dr. Elefterios Kosmidis, Department of Chemistry, University of Copenhagen, who conducted experiments in the laboratory.

The method developed here is the first ever to measure the effects of drugs on the proton pumping of individual V-ATPase molecules. It can detect currents over a million times smaller than the gold standard patch clamp method.

Facts About The V-ATPase Enzyme:

Also see

This shows a brain
  • V-ATPases are enzymes that break down ATP molecules to pump protons across cell membranes.
  • They are found in all cells and are essential for controlling pH/acidity levels inside and/or outside cells.
  • In neuronal cells, the proton gradient established by V-ATPases provides energy for loading neurochemical messengers called neurotransmitters into synaptic vesicles for subsequent release at synaptic junctions.

About this neuroscience research news

Author: Press Office
Source: University of Copenhagen
Contact: Press Service – University of Copenhagen
Image: The image is in the public domain

Original research: Closed access.
Regulation of the mammalian brain V-ATPase by ultraslow mode switchingby Dimitrios Stamou et al. Nature


Abstract

Regulation of the mammalian brain V-ATPase by ultraslow mode switching

Vacuolar-type adenosine triphosphatases (V-ATPases) are electrogenic rotary mechanoenzymes structurally related to F-type ATP synthases. They hydrolyze ATP to establish electrochemical proton gradients for a plethora of cellular processes.

In neurons, the loading of all neurotransmitters into synaptic vesicles is powered by approximately one V-ATPase molecule per synaptic vesicle. To shed light on this bona fide single-molecule biological process, we investigated electrogenic proton pumps by single mammalian brain V-ATPases in single synaptic vesicles.

Here we show that V-ATPases do not pump continuously over time as suggested by observing the rotation of bacterial homologs and assuming strict ATP-proton coupling.

Instead, they stochastically switch between three ultra-long-lived modes: proton-pumping, inactive, and proton-leakage. In particular, direct observation of pumping revealed that physiologically relevant ATP concentrations do not regulate intrinsic pumping rate.

ATP regulates V-ATPase activity through proton pump mode switching probability. In contrast, electrochemical proton gradients control pumping speed and switching between pumping and idle mode.

A direct consequence of mode switching are all-or-nothing stochastic fluctuations in the electrochemical gradient of synaptic vesicles that are expected to introduce stochasticity to proton-driven secondary active load of neurotransmitters and thus may have important implications for neurotransmission.

This work reveals and highlights the mechanistic and biological importance of ultraslow mode switching.



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