Jul 30 2019
A new microscope has exceeded a long-standing speed limit by recording footage of brain activity 15 times faster than researchers once thought could be achievable.
It collects data rapid enough to record voltage spikes of neurons and discharge of chemical messengers across wide regions, monitoring hundreds of synapses at the same time—a significant step for the powerful imaging method known as two-photon microscopy.
The knack is not in bending the laws of physics, but in using insight about a sample to constrict the same information into fewer measurements.
Researchers at the Howard Hughes Medical Institute’s Janelia Research Campus reported on July 29th, 2019, in Nature Methods that they have employed the new microscope to observe patterns of neurotransmitter discharge onto mouse neurons. To date, it has been impossible to capture these millisecond-timescale patterns in the brains of living animals.
Researchers employ two-photon imaging to view inside opaque samples—like living brains—that cannot be penetrated with traditional light microscopy. These microscopes employ a laser to excite fluorescent molecules and subsequently measure the light emitted. In typical two-photon microscopy, each measurement takes a few nanoseconds; in order to create a video, it requires making measurements for every pixel in the image in every frame.
Theoretically, that restricts how fast one can take a picture, states study lead author Kaspar Podgorski, a fellow at Janelia.
You’d think that'd be a fundamental limit—the number of pixels multiplied by the minimum time per pixel. But we've broken this limit by compressing the measurements.
Kaspar Podgorski, Study Lead Author and Fellow, Janelia Research Campus, Howard Hughes Medical Institute
Earlier, that sort of speed could be attained only over small areas.
The new tool—Scanned Line Angular Projection microscopy, or SLAP—renders the time-consuming data-collection stage more efficient in some methods. It packs multiple pixels into one measurement and scans only pixels in areas of interest, with the help of a device that can control which portions of the image are illuminated.
A high-resolution image of the sample acquired prior to the two-photon imaging starts, directs the scope, and enables researchers to decompress the data to make comprehensive videos.
Quite similar to a CT scanner that constructs an image by scanning a patient from different viewpoints, SLAP sweeps a beam of light across a sample along four different planes.
Rather than recording each pixel in the beam’s path as an individual data point, the scope crams the points in that line together into one number. After that, computer programs sort out the lines of pixels to obtain data for every point in the sample—quite similar to solving a big Sudoku puzzle.
By the time SLAP scans the entire sample, a conventional scope going pixel-by-pixel would scan just a tiny part of an image. This speed enabled Podgorski’s group to comprehensively observe how glutamate—a vital neurotransmitter—is discharged onto various parts of mouse neurons.
For example, in the mouse visual cortex, they discovered areas on neurons’ dendrites where several synapses appear to be active at the same time. And they monitored neural activity patterns transferring across the mouse’s cortex as an object moved across its visual field.
Podgorski’s eventual aim is to image every signal entering a single neuron, to gain insight into how neurons convert incoming signals into outgoing signals. This existing scope is “only a step along the way—but we’re already building a second generation. Once we have that, we won’t be limited by the microscope anymore,” he states.
His group is advancing the scope’s scanners to increase its speed. They are also looking for ways to monitor other neurotransmitters so they can completely understand the symphony of neural communication.
Source: https://www.hhmi.org/