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Capturing the intricate brain activity requires resolution, scale and speed-when millions of neurons actively send out signals from remote corners of the cerebral cortex in less than a second, we can distinguish with extreme clarity Rate to see them
.
Now, researchers have developed a microscope technology that allows scientists to accomplish this feat, capturing detailed images of a large number of cell activities at different depths of the brain with unprecedented speed and clarity
.
The study published in the journal Nature Methods demonstrates the innovative power of this kind of light bead microscope.
"To understand the nature of the brain's tightly connected network, it is necessary to develop new imaging techniques that can capture the activity of neurons in a wide range of brain regions at high speed and single-cell resolution," said Rockefeller's Alipasha Vaziri
.
"Light beads microscopy will enable us to study biological problems in a way that was not possible before
Focusing microscope
Whether it is looking for a dangerous beard by swinging back and forth, or helping humans to hit a baseball's hand-eye coordination, animals rely on the brain's sense, movement, and visual areas of the call and response
.
Cells from the distant part of the cerebral cortex coordinate this feat through a network of neural activity that weaves the distant areas of the brain into an interconnected symphony
With the help of cutting-edge microscope technology, scientists are just now beginning to unravel this net
.
The combination of two-photon scanning microscopy and fluorescent labels is the gold standard for imaging neuronal activity in opaque brain tissue, which easily scatters light
But the two-photon microscope has a basic limitation
.
Neurobiologists need to record the simultaneous interaction between sensory, motor, and visual areas of the brain, but it is difficult to capture activity in such a wide area of the brain without sacrificing resolution and speed
Designing an ideal microscope to visualize the interactions between the various areas of the brain feels like blocking a hole in a shipwreck
.
In order to obtain high resolution, scientists often have to sacrifice scale-or shrink the image to obtain a larger structure, at the expense of resolution
Vaziri said: "We need to capture many neurons in the remote part of the brain at the same time with high resolution
.
These parameters are almost mutually exclusive
An innovative solution
The light bead microscope provides a creative solution and pushes the limit of imaging speed to the maximum achievable-only limited by the physical properties of fluorescence itself
.
This is done by eliminating the "dead time" between consecutive laser pulses, when no neural activity is recorded and scanning is required at the same time
This technique involves breaking a strong pulse into 30 smaller sub-pulses—each of which has a different intensity—these sub-pulses sneak into the brains of scattered mice at 30 different depths, but induce the same number at each depth.
Fluorescence
.
This is achieved through a mirror cavity, which ignites each pulse staggeredly and ensures that they can reach the target depth through a microscope focusing lens
.
With this method, the only limit to the sample recording rate is the time for the fluorescent label to glow
.
This means that a large number of brain regions can be recorded at the same time, while a traditional two-photon microscope can only capture a small number of brain cells
.
Vaziri and colleagues then integrated the light bead microscope into a microscope platform for testing, which allowed optical access to large brain volumes, thus recording the activity of more than 1 million neurons throughout the mouse cerebral cortex for the first time
.
Since Vaziri's method is an innovation based on two-photon microscopy, many laboratories already have or can commercially obtain the technology necessary to implement light bead microscopy, as described in the paper
.
Laboratories less familiar with these technologies can benefit from the more widely used simplified, stand-alone modules that Vaziri is currently developing
.
"We have no good reason not to do this five years ago," he said
.
"This was possible because microscopy and laser technology already existed
.
No one thought of this
.
"
"Ultimately, our goal is to complement, not replace, existing technologies
.
For some neurobiological problems, standard two-photon microscopes are sufficient," Vaziri said
.
"But the light bead microscope allows us to solve problems that cannot be solved by existing methods
.
"
"High-speed, cortex-wide volumetric recording of neuroactivity at cellular resolution using light beads microscopy" by Jeffrey Demas, Jason Manley, Frank Tejera, Kevin Barber, Hyewon Kim, Francisca Martínez Traub, Brandon Chen and Alipasha Vaziri, 30 August 2021, Nature Methods.
DOI: 10.
1038/s41592-021-01239-8
