Posted on by Dr. Francis Collins
All of us make many decisions every day. For most things, such as which jacket to wear or where to grab a cup of coffee, there’s usually no right answer, so we often decide using values rooted in our past experiences. Now, neuroscientists have identified the part of the mammalian brain that stores information essential to such value-based decision making.
Researchers zeroed in on this particular brain region, known as the retrosplenial cortex (RSC), by analyzing movies—including the clip shown about 32 seconds into this video—that captured in real time what goes on in the brains of mice as they make decisions. Each white circle is a neuron, and the flickers of light reflect their activity: the brighter the light, the more active the neuron at that point in time.
All told, the NIH-funded team, led by Ryoma Hattori and Takaki Komiyama, University of California at San Diego, La Jolla, made recordings of more than 45,000 neurons across six regions of the mouse brain . Neural activity isn’t usually visible. But, in this case, researchers used mice that had been genetically engineered so that their neurons, when activated, expressed a protein that glowed.
Their system was also set up to encourage the mice to make value-based decisions, including choosing between two drinking tubes, each with a different probability of delivering water. During this decision-making process, the RSC proved to be the region of the brain where neurons persistently lit up, reflecting how the mouse evaluated one option over the other.
The new discovery, described in the journal Cell, comes as something of a surprise to neuroscientists because the RSC hadn’t previously been implicated in value-based decisions. To gather additional evidence, the researchers turned to optogenetics, a technique that enabled them to use light to inactivate neurons in the RSC’s of living animals. These studies confirmed that, with the RSC turned off, the mice couldn’t retrieve value information based on past experience.
The researchers note that the RSC is heavily interconnected with other key brain regions, including those involved in learning, memory, and controlling movement. This indicates that the RSC may be well situated to serve as a hub for storing value information, allowing it to be accessed and acted upon when it is needed.
The findings are yet another amazing example of how advances coming out of the NIH-led Brain Research through Advancing Innovative Neurotechnologies® (BRAIN) Initiative are revolutionizing our understanding of the brain. In the future, the team hopes to learn more about how the RSC stores this information and sends it to other parts of the brain. They note that it will also be important to explore how activity in this brain area may be altered in schizophrenia, dementia, substance abuse, and other conditions that may affect decision-making abilities. It will also be interesting to see how this develops during childhood and adolescence.
 Area-Specificity and Plasticity of History-Dependent Value Coding During Learning. Hattori R, Danskin B, Babic Z, Mlynaryk N, Komiyama T. Cell. 2019 Jun 13;177(7):1858-1872.e15.
Komiyama Lab (UCSD, La Jolla)
NIH Support: National Institute of Neurological Disorders and Stroke; National Eye Institute; National Institute on Deafness and Other Communication Disorders
Posted on by Dr. Francis Collins
It’s been 25 years since researchers coaxed a bacterium to synthesize an unusual jellyfish protein that fluoresced bright green when irradiated with blue light. Within months, another group had also fused this small green fluorescent protein (GFP) to larger proteins to make their whereabouts inside the cell come to light—like never before.
To mark the anniversary of this Nobel Prize-winning work and show off the rainbow of color that is now being used to illuminate the inner workings of the cell, the American Society for Cell Biology (ASCB) recently held its Green Fluorescent Protein Image and Video Contest. Over the next few months, my blog will feature some of the most eye-catching entries—starting with this video that will remind those who grew up in the 1980s of those plasma balls that, when touched, light up with a simulated bolt of colorful lightning.
This video, which took third place in the ASCB contest, shows the cytoskeleton of a frequently studied human breast cancer cell line. The cytoskeleton is made from protein structures called microtubules, made visible by fluorescently tagging a protein called doublecortin (orange). Filaments of another protein called actin (purple) are seen here as the fine meshwork in the cell periphery.
The cytoskeleton plays an important role in giving cells shape and structure. But it also allows a cell to move and divide. Indeed, the motion in this video shows that the complex network of cytoskeletal components is constantly being organized and reorganized in ways that researchers are still working hard to understand.
Jeffrey van Haren, Erasmus University Medical Center, Rotterdam, the Netherlands, shot this video using the tools of fluorescence microscopy when he was a postdoctoral researcher in the NIH-funded lab of Torsten Wittman, University of California, San Francisco.
All good movies have unusual plot twists, and that’s truly the case here. Though the researchers are using a breast cancer cell line, their primary interest is in the doublecortin protein, which is normally found in association with microtubules in the developing brain. In fact, in people with mutations in the gene that encodes this protein, neurons fail to migrate properly during development. The resulting condition, called lissencephaly, leads to epilepsy, cognitive disability, and other neurological problems.
Cancer cells don’t usually express doublecortin. But, in some of their initial studies, the Wittman team thought it would be much easier to visualize and study doublecortin in the cancer cells. And so, the researchers tagged doublecortin with an orange fluorescent protein, engineered its expression in the breast cancer cells, and van Haren started taking pictures.
This movie and others helped lead to the intriguing discovery that doublecortin binds to microtubules in some places and not others . It appears to do so based on the ability to recognize and bind to certain microtubule geometries. The researchers have since moved on to studies in cultured neurons.
This video is certainly a good example of the illuminating power of fluorescent proteins: enabling us to see cells and their cytoskeletons as incredibly dynamic, constantly moving entities. And, if you’d like to see much more where this came from, consider visiting van Haren’s Twitter gallery of microtubule videos here:
 Doublecortin is excluded from growing microtubule ends and recognizes the GDP-microtubule lattice. Ettinger A, van Haren J, Ribeiro SA, Wittmann T. Curr Biol. 2016 Jun 20;26(12):1549-1555.
Lissencephaly Information Page (National Institute of Neurological Disorders and Stroke/NIH)
Wittman Lab (University of California, San Francisco)
Green Fluorescent Protein Image and Video Contest (American Society for Cell Biology, Bethesda, MD)
NIH Support: National Institute of General Medical Sciences
Posted on by Dr. Francis Collins
Can you identify a familiar pattern in this image’s square grid? Yes, it’s the outline of the periodic table! But instead of organizing chemical elements, this periodic table sorts 46 different types of neurons present in the visual cortex of a mouse brain.
Scientists, led by Hongkui Zeng at the Allen Institute for Brain Science, Seattle, constructed this periodic table by assigning colors to their neuronal discoveries based upon their main cell functions . Cells in pinks, violets, reds, and oranges have inhibitory electrical activity, while those in greens and blues have excitatory electrical activity.
For any given cell, the darker colors indicate dendrites, which receive signals from other neurons. The lighter colors indicate axons, which transmit signals. Examples of electrical properties—the number and intensity of their “spikes”—appear along the edges of the table near the bottom.
To create this visually arresting image, Zeng’s NIH-supported team injected dye-containing probes into neurons. The probes are engineered to carry genes that make certain types of neurons glow bright colors under the microscope.
This allowed the researchers to examine a tiny slice of brain tissue and view each colored neuron’s shape, as well as measure its electrical response. They followed up with computational tools to combine these two characteristics and classify cell types based on their shape and electrical activity. Zeng’s team could then sort the cells into clusters using a computer algorithm to avoid potential human bias from visually interpreting the data.
Why compile such a detailed atlas of neuronal subtypes? Although scientists have been surveying cells since the invention of the microscope centuries ago, there is still no consensus on what a “cell type” is. Large, rich datasets like this atlas contain massive amounts of information to characterize individual cells well beyond their appearance under a microscope, helping to explain factors that make cells similar or dissimilar. Those differences may not be apparent to the naked eye.
Just last year, Allen Institute researchers conducted similar work by categorizing nearly 24,000 cells from the brain’s visual and motor cortex into different types based upon their gene activity . The latest research lines up well with the cell subclasses and types categorized in the previous gene-activity work. As a result, the scientists have more evidence that each of the 46 cell types is actually distinct from the others and likely drives a particular function within the visual cortex.
Publicly available resources, like this database of cell types, fuel much more discovery. Scientists all over the world can look at this table (and soon, more atlases from other parts of the brain) to see where a cell type fits into a region of interest and how it might behave in a range of brain conditions.
 Classification of electrophysiological and morphological neuron types in the mouse visual cortex. N Gouwens NW, et al. Neurosci. 2019 Jul;22(7):1182-1195.
 Shared and distinct transcriptomic cell types across neocortical areas. Tasic B, et al. Nature. 2018 Nov;563(7729):72-78.
Brain Basics: The Life and Death of a Neuron (National Institute of Neurological Disorders and Stroke/NIH)
Cell Types: Overview of the Data (Allen Brain Atlas/Allen Institute for Brain Science, Seattle)
Hongkui Zeng (Allen Institute)
NIH Support: National Institute of Mental Health; Eunice Kennedy Shriver National Institute of Child Health & Human Development
Posted on by Dr. Francis Collins
The NIH-led Brain Research through Advancing Innovative Neurotechnologies® (BRAIN) Initiative is revolutionizing our understanding of how the brain works through its creation of new imaging tools. One of the latest advances—used to produce this rainbow of images—makes it possible to view dozens of proteins in rapid succession in a single tissue sample containing thousands of neural connections, or synapses.
Apart from their colors, most of these images look nearly identical at first glance. But, upon closer inspection, you’ll see some subtle differences among them in both intensity and pattern. That’s because the images capture different proteins within the complex network of synapses—and those proteins may be present in that network in different amounts and locations. Such findings may shed light on key differences among synapses, as well as provide new clues into the roles that synaptic proteins may play in schizophrenia and various other neurological disorders.
Synapses contain hundreds of proteins that regulate the release of chemicals called neurotransmitters, which allow neurons to communicate. Each synaptic protein has its own specific job in the process. But there have been longstanding technical difficulties in observing synaptic proteins at work. Conventional fluorescence microscopy can visualize at most four proteins in a synapse.
As described in Nature Communications , researchers led by Mark Bathe, Massachusetts Institute of Technology (MIT), Cambridge, and Jeffrey Cottrell, Broad Institute of MIT and Harvard, Cambridge, have just upped this number considerably while delivering high quality images. They did it by adapting an existing imaging method called DNA PAINT . The researchers call their adapted method PRISM. It is short for: Probe-based Imaging for Sequential Multiplexing.
Here’s how it works: First, researchers label proteins or other molecules of interest using antibodies that recognize those proteins. Those antibodies include a unique DNA probe that helps with the next important step: making the proteins visible under a microscope.
To do it, they deliver short snippets of complementary fluorescent DNA, which bind the DNA-antibody probes. While each protein of interest is imaged separately, researchers can easily wash the probes from a sample to allow a series of images to be generated, each capturing a different protein of interest.
In the original DNA PAINT, the DNA strands bind and unbind periodically to create a blinking fluorescence that can be captured using super-resolution microscopy. But that makes the process slow, requiring about half an hour for each protein.
To speed things up with PRISM, Bathe and his colleagues altered the fluorescent DNA probes. They used synthetic DNA that’s specially designed to bind more tightly or “lock” to the DNA-antibody. This gives a much brighter signal without the blinking effect. As a result, the imaging can be done faster, though at slightly lower resolution.
Though the team now captures images of 12 proteins within a sample in about an hour, this is just a start. As more DNA-antibody probes are developed for synaptic proteins, the team can readily ramp up this number to 30 protein targets.
Thanks to the BRAIN Initiative, researchers now possess a powerful new tool to study neurons. PRISM will help them learn more mechanistically about the inner workings of synapses and how they contribute to a range of neurological conditions.
 Multiplexed and high-throughput neuronal fluorescence imaging with diffusible probes. Guo SM, Veneziano R, Gordonov S, Li L, Danielson E, Perez de Arce K, Park D, Kulesa AB, Wamhoff EC, Blainey PC, Boyden ES, Cottrell JR, Bathe M. Nat Commun. 2019 Sep 26;10(1):4377.
 Super-resolution microscopy with DNA-PAINT. Schnitzbauer J, Strauss MT, Schlichthaerle T, Schueder F, Jungmann R. Nat Protoc. 2017 Jun;12(6):1198-1228.
Schizophrenia (National Institute of Mental Health)
Mark Bathe (Massachusetts Institute of Technology, Cambridge)
Jeffrey Cottrell (Broad Institute of MIT and Harvard, Cambridge)
NIH Support: National Institute of Mental Health; National Human Genome Research Institute; National Institute of Neurological Disorders and Stroke; National Institute of Environmental Health Sciences
Posted on by Dr. Francis Collins
A major aim of the NIH-led Brain Research through Advancing Innovative Neurotechnologies® (BRAIN) Initiative is to develop new technologies that allow us to look at the brain in many different ways on many different scales. So, I’m especially pleased to highlight this winner of the initiative’s recent “Show Us Your Brain!” contest.
Here you get a close-up look at pyramidal neurons located in the hippocampus, a region of the mammalian brain involved in memory. While this tiny sample of mouse brain is densely packed with many pyramidal neurons, researchers used new ExLLSM technology to zero in on just three. This super-resolution, 3D view reveals the intricacies of each cell’s structure and branching patterns.
The group that created this award-winning visual includes the labs of X. William Yang at the University of California, Los Angeles, and Kwanghun Chung at the Massachusetts Institute of Technology, Cambridge. Chung’s team also produced another quite different “Show Us Your Brain!” winner, a colorful video featuring hundreds of neural cells and connections in a part of the brain essential to movement.
Pyramidal neurons in the hippocampus come in many different varieties. Some important differences in their functional roles may be related to differences in their physical shapes, in ways that aren’t yet well understood. So, BRAIN-supported researchers are now applying a variety of new tools and approaches in a more detailed effort to identify and characterize these neurons and their subtypes.
The video featured here took advantage of Chung’s new method for preserving brain tissue samples . Another secret to its powerful imagery was a novel suite of mouse models developed in the Yang lab. With some sophisticated genetics, these models make it possible to label, at random, just 1 to 5 percent of a given neuronal cell type, illuminating their full morphology in the brain . The result was this unprecedented view of three pyramidal neurons in exquisite 3D detail.
Ultimately, the goal of these and other BRAIN Initiative researchers is to produce a dynamic picture of the brain that, for the first time, shows how individual cells and complex neural circuits interact in both time and space. I look forward to their continued progress, which promises to revolutionize our understanding of how the human brain functions in both health and disease.
 Protection of tissue physicochemical properties using polyfunctional crosslinkers. Park YG, Sohn CH, Chen R, McCue M, Yun DH, Drummond GT, Ku T, Evans NB, Oak HC, Trieu W, Choi H, Jin X, Lilascharoen V, Wang J, Truttmann MC, Qi HW, Ploegh HL, Golub TR, Chen SC, Frosch MP, Kulik HJ, Lim BK, Chung K. Nat Biotechnol. 2018 Dec 17.
 Genetically-directed Sparse Neuronal Labeling in BAC Transgenic Mice through Mononucleotide Repeat Frameshift. Lu XH, Yang XW. Sci Rep. 2017 Mar 8;7:43915.
Chung Lab (Massachusetts Institute of Technology, Cambridge)
Yang Lab (University of California, Los Angeles)
Show Us Your Brain! (BRAIN Initiative/NIH)
NIH Support: National Institute of Mental Health; National Institute of Neurological Disorders and Stroke; National Institute of Biomedical Imaging and Bioengineering