Jeffrey Perkel has been a scientific writer and editor since 2000. He holds a PhD in Cell and Molecular Biology from the University of Pennsylvania, and did postdoctoral work at the University of Pennsylvania and at Harvard Medical School.
On October 1, the U.S. National Institutes of Health (NIH) announced some $38 million in new research funding awarded as part of President Obama’s Brain Research through Advancing Innovative Neurotechnologies (BRAIN) Initiative.
In a press release announcing the funding, the National Institute of Neurological Disorders and Stroke (NINDS), one the NIH institutes overseeing the BRAIN Initiative, emphasized the awards’ focus on new tools and technologies.
The neuroscience field has in fact been inundated with new technologies over the past decade or so, from Brainbow and fMRI to optogenetics and connectomics. The goal of the BRAIN Initiative is to use such tools to figure out how the brain works not at a cellular or even circuit level, but holistically.
Neuroscientists, explains Edmund Talley, program director for extramural research at NINDS, know a great deal about how neurons work at the cellular level. “What we don’t know is how neurons function together to code information across the whole brain.” The BRAIN Initiative, he says, should help close that gap. (Synopses of all funded projects are available here: http://www.braininitiative.nih.gov/nih-brain-awards.htm.)
According to Michelle Freund, a program officer at the NIH’s National Institute of Mental Health (NIMH), funding will be channeled into technology development during the first five years in an effort to achieve the nine major goals set forth by an Advisory Committee to the NIH Director. One goal to be addressed, a “census of cell types,” is intended to catalog what Freund calls the brain’s “parts list”—that is, a comprehensive index of all cell types in the brain.
“The key question is, how do you call a cell unique?” Freund explains. Is it defined by cell shape? Connectivity? Gene expression? The NIH is funding 10 research projects that take different approaches to this problem, based on such methods as RNA-Seq and histology, she says.
Another program, “tools for cells and circuits,” funds development of novel methods “to dissect the circuitry of the brain,” Freund says. One project, directed by Bryan Roth at the University of North Carolina, Chapel Hill, “will provide an enhanced chemogenetic toolbox that allows non-invasive, multiplexed spatiotemporal control of neuronal activity in domains ranging from single synapses to ensembles of neurons,” according to the funding announcement. Specifically, the researchers plan an updated and enhanced version of a technology called “Designer Receptors Exclusively Activated by Designer Drugs” (DREADDs), which are mutant cell-surface receptors activated by “otherwise inert drug-like small molecules .”
DREADDs isn’t a new technology, Freund notes; the award is funding work to refine it and increase its specificity. “A lot of the applications do exactly that,” she says, “take existing technology and make them better.”
Another project in this area, directed by Ed Boyden at Massachusetts Institute of Technology, proposes what Boyden calls “a high-risk, high-payoff, and as far as we know entirely novel agenda: to develop tools capable of resolving the molecular proteomic composition of synapse types, testing them in cultured neurons and intact brain tissues.” The team proposes using a super-resolution imaging method called DNA-PAINT (Points Accumulation for Imaging in Nanoscale Topography) to probe the protein complement (proteome) of neuronal synapses.
According to Michael Hasselmo, director of the Center for Systems Neuroscience at Boston University, which includes several BRAIN Initiative-funded researchers, one particularly exciting facet of the BRAIN Initiative involves optogenetics, which encompasses tools for controlling neuronal activity with light as well as large-scale recording of neuronal activity.
This latter goal is typically accomplished using genetically encoded calcium indicators (GECIs), such as GCaMP6. In one recent study, Jim Heys, a post-doc at Northwestern University (and Hasselmo’s former graduate student), used GCaMP6 and two-photon microscopy to simultaneously record the activity of several hundred neurons in the medial entorhinal cortex of a mouse, as it ran on a treadmill [1]. “It’s a head-fixed mouse, but it’s running on a ball,” Hasselmo explains. “As it runs, its feet move the ball and that moves the virtual environment that it’s seeing on these computer screens in front of it.”
According to Hasselmo, that technology represents an order-of-magnitude improvement in throughput compared with the technology he uses in his own lab, tetrode recording. Using that approach, in which microelectrodes are implanted directly into the brain, researchers can record 20 to 100 neurons simultaneously in a behaving animal. But with optical approaches, experiments can scale to 10,000 cells, or even a million cells at once—a fact he discussed in a recent essay entitled (with a wink and a nod to the Barenaked Ladies) “If I Had a Million Neurons” [2].
Genetically encoded voltage indicators also are being developed. Vincent Pieribone of the John B. Pierce Laboratory in New Haven, Conn., for instance, won BRAIN Initiative funding for a proposal “to discover protein-based fluorescent voltage probes with signal to noise characteristics that allow routine optical recording of action potentials from single cortical neurons in vivo.”
Novel microscope technologies also are in development. According to Jochen Tham, senior director for global marketing at Carl Zeiss Microscopy, popular neuroscience microscopy techniques include two-photon or multiphoton microscopy and light-sheet microscopy. Neither can image an entire mouse brain, Tham notes—multiphoton microscopy can penetrate dense neural tissue to about 0.5 mm, and light-sheet microscopy can produce a complete volumetric dataset in a single scan of a sample about the size of a Drosophila embryo.
"To image the whole mouse brain, you would have to image in small blocks," Tham says. "That’s the challenge."
Alipasha Vaziri at Rockefeller University and Elizabeth Hillman at Columbia University have received funding to develop tools to accelerate volumetric imaging. The problem, Talley explains, is that although today’s microscopes are fast, they’re not fast enough to capture large tissue volumes on the time scale of neuronal action potentials, which fire over milliseconds. “We’re funding a number of different strategies … to make it possible to image whole volumes at the subsecond scale, and potentially down to the tens of millisecond scales,” he says.
Hillman, for instance, proposes refining a technique developed in her lab, called SCAPE (Swept, Confocally-Aligned Planar Excitation) microscopy—“a hybrid between light-sheet microscopy and laser scanning confocal which overcomes the major speed barriers of both techniques”—into a tool for routine use in both Drosophila and mice. Vaziri is developing a strategy called Multiplexed Scanned Temporal Focusing (MuST), for “unprecedentedly large” and rapid volumetric imaging.
At Boston College, electrical engineer Siddharth Ramachandran received BRAIN Initiative funding for development of a new multiphoton-microscopy imaging system, which can penetrate tissue to a depth of 2 mm, Hasselmo says.
Other researchers are using scanning electron microscopy (SEM) and automated sample preparation and analysis methods to map neural connections at the nanoscale level, a monumental technical challenge. In a recent paper in Cell, Jeff Lichtman, at Harvard University, and colleagues described the effort required to map the synaptic connectivity in a tiny piece of mouse neocortex. Though it measured just 1,500 ?m3, that piece of tissue contained some 1,407 axons and 193 dendrites, and 1,700 synapses overall—which the authors painstakingly reconstructed in silico from 2,250 tissue slices, each 29 nm thick [3]. “Surprisingly, analysis of the connectomic data turned out to be even more challenging than creating the image data or annotating it,” they wrote.
Several applications funded in 2015 support technologies to bypass the difficult (and nontranslatable) genetic engineering required to achieve cell-type restricted transgene expression. For example, Daniel Schmidt and colleagues at the University of Minnesota are developing “a novel viral delivery method able to deliver transgenes selectively to neurons that express, on the cell surface, a targeted set of ion channels and receptors.” Seth Blackshaw and co-workers at Johns Hopkins University are developing an alternative to a method called CRE-DOG (Cre Recombinase Dependent on GFP)—basically, a form of Cre recombinase that functions only in the presence of GFP—“to induce cell-specific activation of expression of reporter and effector constructs delivered by electroporation or viral vector.”
Some BRAIN Initiative research also is being funded by the National Science Foundation (NSF). Among the researchers that the NSF supports are Steven Chase and Byrun Yu at Carnegie Mellon University in Pittsburgh. According to Alison Barth, interim director of the university’s BrainHub Initiative, Chase and Yu are studying how neural activity in the motor cortex encodes movement. Among other things, she says, such information could be used to drive brain-computer interfaces. “If we could forget the brain-to-muscle connection and use those signals to drive a robotic arm or a cursor, we could go a long way to restore quality of life for paralyzed individuals,” she says.
And that’s not all; the BRAIN Initiative also supports novel imaging modalities, minimally invasive brain-recording strategies—and especially data coordination. The BRAIN Initiative, says Freund, will be generating reams of data of diverse types, on a range of size scales from the smallest synapse to brain-scale circuits. The challenge is to integrate those into a unified and navigable dataset. “How do we put all of these diverse datasets and data types together to learn from them?” Stay tuned to find out.
References
[1] Heys, JG, et al., “The functional micro-organization of grid cells revealed by cellular-resolution imaging,” Neuron, 84:1079-90, 2014. [PMID: 25467986]
[2] Hasselmo, ME, “If I had a million neurons: Potential tests of cortico-hippocampal theories,” Progress in Brain Research, 219:1-19, 2015. [PMID: 26072231]
[3] Kasthuri, N, et al., “Saturated reconstruction of a volume of neocortex,” Cell, 162:648-61, 2015. [PMID: 26232230]
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