Within the brain, neurons\u00a0compute\u00a0by\u00a0generating\u00a0electrical impulses. These signals travel throughout\u00a0neurons,\u00a0which\u00a0are\u00a0in turn\u00a0connected in vast\u00a0networks\u00a0that\u00a0control brain functions such as sensory perception, memory formation, and control of movement.\u00a0<\/p>\n
In an advance that could help neuroscientists map those neural networks,\u00a0leading to a better understanding of how neural activity underlies behavior and other brain functions,\u00a0MIT engineers have invented a new\u00a0microscope\u00a0that can image electrical activity\u00a0in neurons distributed\u00a0across the brain of an entire organism, the experimental model\u00a0Danio rerio\u00a0(zebrafish).\u00a0<\/p>\n
Using a microscope that they adapted for fast, high-volumetric rate\u00a0imaging, the researchers were able to track electrical activity across the brain on the scale of milliseconds. This method revealed patterns of\u00a0neural activity from neurons\u00a0throughout the brain that were activated in response to\u00a0ultraviolet\u00a0light.\u00a0<\/p>\n
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All of the parts of the brain are connected together,\u00a0so if you want to\u00a0truly\u00a0understand\u00a0the\u00a0brain, you have to understand\u00a0how\u00a0all the\u00a0neurons work together as an emergent whole.”<\/p>\n
\n <\/p>\nEd Boyden,\u00a0the Y. Eva Tan Professor in Neurotechnology at MIT<\/p>\n
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Ed Boyden is a professor of biological engineering, media arts and sciences, and brain and cognitive sciences; and a member of MIT’s McGovern Institute for Brain Research, Yang Tan Collective,\u00a0and\u00a0the\u00a0Koch Institute for Integrative Cancer Research.\u00a0<\/p>\n
Boyden is the senior author of the study, which appears today in\u00a0Nature Methods.\u00a0Former J. Douglas Tan Postdoctoral Fellow\u00a0Zeguan Wang\u00a0PhD\u00a0’24\u00a0and\u00a0former MIT postdoc\u00a0Jie Zhang are the lead authors of the paper.\u00a0Other authors include former MIT postdoc Panagiotis\u00a0Symvoulidis,\u00a0Picower\u00a0Institute research scientist Wei Guo,\u00a0graduate students\u00a0Davy Deng\u00a0and Lige Zhang, Koch Institute research scientist Adam Amsterdam,\u00a0Picower\u00a0Institute research scientist Takato Honda, Boston College undergraduate Steven Roche,\u00a0and Matthew Wilson, the\u00a0Sherman Fairchild Professor of Neuroscience at MIT and a member of the\u00a0Picower\u00a0Institute.\u00a0<\/p>\n
High-speed imaging\u00a0<\/p>\n
One technique often used to measure neuron activity in the brain is calcium imaging. Calcium flows into neurons\u00a0after they fire an electrical impulse, so measuring calcium\u00a0levels in the cells can\u00a0serve as a proxy for neural activity. However, this type of imaging\u00a0isn’t\u00a0fast enough to capture single spikes of activity.\u00a0<\/p>\n
“Calcium imaging inherently is\u00a0very slow, so\u00a0you’re\u00a0talking about imaging activity\u00a0on\u00a0the order of seconds or even minutes.\u00a0Typically\u00a0that is too slow for us to be able to\u00a0see\u00a0a lot of these high-speed neural activities,” Zhang says. “Neurons compute using electrical activity, so with\u00a0voltage imaging, you\u00a0can\u00a0get direct observation of that.”\u00a0<\/p>\n
To enable\u00a0direct imaging of\u00a0voltage, researchers have developed proteins called genetically encoded voltage indicators – fluorescent proteins that can be genetically\u00a0expressed\u00a0in neurons. When a neuron fires an impulse, the protein\u00a0fluoresces, which\u00a0can be detected with a fluorescence microscope.\u00a0<\/p>\n
In\u00a0previous\u00a0work, researchers\u00a0have used these proteins to image small populations of neurons, usually focusing on one\u00a0localized\u00a0part of the brain. Until now, there\u00a0hasn’t\u00a0been a way to\u00a0image\u00a0a large volume, such as the entire brain, with the millisecond-scale resolution needed to see electrical impulses from individual neurons.\u00a0<\/p>\n
To achieve that, the MIT team decided to\u00a0modify\u00a0a commonly used microscope known as a light sheet microscope. This type of microscope uses a sheet of laser light to illuminate a thin slice of a sample. By\u00a0imaging\u00a0many layers in sequence, this technique can generate 3D images of a large volume. However,\u00a0with\u00a0previous\u00a0microscopes,\u00a0the scanning of\u00a0an\u00a0entire volume\u00a0would take too long to be able to capture neuronal impulses\u00a0across the volume\u00a0at single cell resolution.\u00a0<\/p>\n
“Different groups of neurons\u00a0that are\u00a0distributed across the brain\u00a0coordinate together at millisecond timescales\u00a0to generate a lot of behaviors and brain computations,” Wang says. “To understand the principles, we need the technology to observe their activity at the same time,\u00a0across the whole brain,\u00a0so we are not missing any important participant neurons.”\u00a0<\/p>\n
To make the imaging process fast enough to image millisecond-scale activity, the researchers increased the image acquisition speed of the microscope’s camera, and they also boosted the scanning speed of the microscope using a technique called\u00a0remote refocusing.\u00a0<\/p>\n
Using this approach, the researchers showed that they could scan the entire zebrafish brain 200 times per second, or once every five milliseconds.\u00a0<\/p>\n
Mapping brain activity\u00a0<\/p>\n
To test the new microscope, the researchers\u00a0engineered\u00a0neurons in larval zebrafish to express a voltage indicator\u00a0called Positron2-Kv.\u00a0Although they had hoped that the indicator would end up in every neuron, it produced signals in\u00a0neurons distributed throughout the brain, with about\u00a0one\u00a0quarter\u00a0of the neurons\u00a0exhibiting\u00a0acceptable signals. This was enough, however, to\u00a0observe\u00a0patterns of activity\u00a0across the brain. The researchers\u00a0imaged\u00a0the brain as the fish were\u00a0resting, and\u00a0they\u00a0were able to\u00a0observe\u00a0single voltage spikes from neurons, as well as\u00a0rapid\u00a0bursts of spikes.\u00a0<\/p>\n
Additionally, this technique revealed patterns in how the brain is activated following a stimulus such as ultraviolet light. Immediately following the stimulus, activity was seen in the optic\u00a0tectum, which receives and processes visual input from the retina. This activity propagated\u00a0from one side of a part of the brain called the\u00a0tectum\u00a0to the other. Stimulus-independent activity also occurred in sequences across sets of neurons in\u00a0the cerebellum and hindbrain.\u00a0<\/p>\n
The researchers now hope to increase the percentage of neurons that they can\u00a0image across the brain, as well as the microscope’s speed and resolution. They are also working on expanding the use of this technique to other experimental models, including mice.\u00a0<\/p>\n
This approach, they say, could offer neuroscientists a new way to generate hypotheses about what happens in the brain when it engages in specific behaviors, or about how brain activity is linked to states of mind such as daydreaming.\u00a0<\/p>\n
“A\u00a0big question is simply to understand how neurons work together as a network. And this might be the first time that you could do that,\u00a0because you\u00a0can\u00a0image\u00a0the voltage of neurons distributed throughout\u00a0the network,” Boyden says.<\/p>\n
The research was funded by the National Institutes of Health,\u00a0the BRAIN Initiative,\u00a0the\u00a0Picower\u00a0Institute Innovation Fund, K. Lisa Yang, Ashar Aziz, the K. Lisa Yang and Hock E. Tan Center for Molecular Therapeutics\u00a0in Neuroscience\u00a0at MIT, the Hock E. Tan and K. Lisa Yang Center for Autism Research, the Alana Down Syndrome Center, John Doerr, Jed McCaleb, James Fickel, and the Howard Hughes Medical Institute.\u00a0<\/p>\n
Source:<\/p>\n
Massachusetts Institute of Technology<\/a><\/p>\n
Journal reference:<\/p>\n