Sharper, Deeper, Faster: Interdisciplinary Team Develops Advanced Live-Imaging Approach

PASADENA, Calif.— For modern biologists, the ability to capture high-quality, three-dimensional (3D) images of living tissues or organisms over time is necessary to answer problems in areas ranging from genomics to neurobiology and developmental biology. The better the image, the more detailed the information that can be drawn from it. Looking to improve upon current methods of imaging, researchers from the California Institute of Technology (Caltech) have developed a novel approach that could redefine optical imaging of live biological samples by simultaneously achieving high resolution, high penetration depth (for seeing deep inside 3D samples), and high imaging speed.

The imaging technique is explained in a paper in the advance online publication of the journal Nature Methods, released on July 14. It will also appear in an upcoming print version of the journal.  

"Before our work, the state-of-the-art imaging techniques typically excelled in only one of three key parameters: resolution, depth, or speed. With our technique, it's possible to do well in all three and, critically, without killing, damaging, or adversely affecting the live biological samples," says biologist Scott Fraser, director of the Biological Imaging Center at Caltech's Beckman Institute and senior author of the study. 

The research team achieved this imaging hat trick by first employing an unconventional imaging method called light-sheet microscopy, where a thin, flat sheet of light is used to illuminate a biological sample from the side, creating a single illuminated optical section through the sample. The light given off by the sample is then captured with a camera oriented perpendicularly to the light sheet, harvesting data from the entire illuminated plane at once. This allows millions of image pixels to be captured simultaneously, reducing the light intensity that needs to be used for each pixel. This not only enables fast imaging speed but also decreases the light-induced damage to the living samples, which the teams demonstrated using the embryos of fruit fly and zebrafish.

To achieve sharper image resolution with light-sheet microscopy deep inside the biological samples, the team used a process called two-photon excitation for the illumination. This process has been used previously to allow deeper imaging of biological samples; however, it usually is used to collect the image one pixel at a time by focusing the exciting light to a single small spot.

"The conceptual leap for us was to realize that two-photon excitation could also be carried out in sheet-illumination mode," says Thai Truong, a postdoctoral fellow in Fraser's laboratory and first author of the paper. This novel side-illumination with a two-photon illumination is the topic of a pending patent.

"With this approach, we believe that we can make a contribution to advancing biological imaging in a meaningful way," continues Truong, who did his PhD training in physics. "We did not want to develop a fanciful optical imaging technique that excels only in one niche area, or that places constraints on the sample so severe that the applications will be limited. With a balanced high performance in resolution, depth, and speed, all achieved without photo-damage, two-photon light-sheet microscopy should be applicable to a wide variety of in vivo imaging applications." He credits this emphasis on wide applicability to the interdisciplinary nature of the team—which includes two biologists, two physicists, and one electrical engineer.

"We believe the performance of this imaging technique will open up many applications in life sciences and biomedical research—wherever it is useful to observe, non-invasively, dynamic biological process in 3D and with cellular or subcellular resolution," says Willy Supatto, co–author of the paper and a former postdoctoral fellow in Fraser's laboratory (now at the Centre National de la Recherche Scientifique, in France).

One example of such an application would be to construct 3D movies of the entire embryonic development of an organism, covering the entire embryo in space and time. These movies could capture what individual cells are doing, as well as important genes' spatiotemporal expression patterns—elucidating the activation of those genes within specific tissues at specific times during development.

"The goal is to create 'digital embryos,' providing insights into how embryos are built, which is critical not only for basic understanding of how biology works but also for future medical applications such as robotic surgery, tissue engineering, or stem-cell therapy," says Fraser. The team's first attempt at this can be seen in the accompanying movie, in which the cell divisions and movements that built the entire fruit fly embryo were captured without perturbing its development. 

The Nature Methods paper is titled "Deep and fast live imaging with two-photon scanned light-sheet microscopy." David Koos, senior research scientist at Caltech's Beckman Institute, and John Choi, a former postdoctoral fellow in Fraser's laboratory, also contributed to the study.

The research was supported by the Beckman Institute and the U.S. National Institutes of Health.


Katie Neith

Neutralizing HIV

Each time a virus invades a healthy individual, antibodies created by the body fight to fend off the intruders. For some viruses, like HIV, the antibodies are very specific and are generated too slowly to combat the rapidly changing virus. However, in the past few years, scientists have found that some HIV-positive people develop highly potent antibodies that can neutralize different subtypes of the HIV virus.

Now, a study involving researchers at Caltech points to the possibility of using these neutralizing antibodies in the development of a vaccine. The paper, published in the July 14 issue of Science Express, describes a group of novel antibodies that were isolated from HIV-infected individuals using a new cloning approach.

These antibodies are the most potent anti-HIV antibodies targeting the CD4 binding site— a functional site on the surface of HIV needed for cell entry and infection—that have ever been identified, says Ron Diskin, a post-doctoral scholar at Caltech who worked on the paper. David Ho (BS '74), scientific director of the Aaron Diamond AIDS Research Center in New York, also contributed to the study, which was led by researchers at Rockefeller University.

At Caltech, the researchers conducted structural studies and were able to show, based on similarity to a previously known antibody (VRC01), that the new antibodies indeed target the CD4 receptor binding site. CD4 positive cells are the point of HIV infection and where the virus multiplies.  

This study is important for several different reasons, according to Diskin. "First, it provides extremely useful reagents that can be used for passive immunization to treat infected individuals," he says. "Second, it demonstrates that a comparable and highly effective anti-HIV immune response was elicited in different individuals, which strongly supports the idea that an effective vaccine will be feasible to develop."

Next, researchers at Caltech will address the structural mechanisms that make those antibodies so potent. In fact, they are currently investigating those structural aspects of the neutralization mechanisms.

"We're very excited to have the opportunity to use structural biology to learn what makes these new antibodies so potent against HIV," says Pamela Bjorkman, Caltech's Delbruck Professor of Biology and a co-author of the study. "We hope that visualizing how these antibodies interact with HIV proteins will allow the design of even more potent anti-HIV reagents and provide critical information for vaccine design."

Katie Neith
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Tracking Bacteriophages

Viruses are parasites; they need to live inside another organism to survive. Figuring out just who a virus's host is can be difficult, however. Especially when you're talking about bacteriophages—a particularly pervasive group of bacteria-infecting viruses.

The problem lies in identifying which bacteriophages are infecting which bacteria, without having to culture either the viruses or their hosts in the lab, which can often be a difficult if not impossible task.

To address that issue, a team led by Caltech biophysicist Rob Phillips has created a genetic-analysis technique—called microfluidic digital polymerase chain reaction (PCR)—that can "physically link single bacterial cells harvested from a natural environment with a viral marker genes," the scientists report in the July 1 issue of the journal Science.

The team looked at the bacterial community found in the hindgut of a termite—specifically, a termite collected in 2005 in Costa Rica. The typical termite hindgut contains over 250 different species of bacteria, "making it ideally suited to explore many potential, diverse phage-host interactions," the scientists say.

What they found was that microfluidic digital PCR allowed them to see interactions between the viruses and their bacterial hosts in uncultivated cells harvested directly from the hindgut. They noted, for instance, that variants of a viral packaging gene had made their way into hosts across an entire genus of bacteria; in another gene-tracking experiment, however, they found little evidence of lateral gene transfer or switching of the gene from one host to another, despite plenty of opportunity.

"Our approach does not require culturing hosts or viruses," the researchers write, "and provides a method for examining virus-bacteria interactions in many environments."

For more on the new technology and its potential applications, check out the paper's abstract or this press release from the National Science Foundation, one of the funders of the research.

Lori Oliwenstein

$6 Million Gift To Spur Innovative Research Collaborations Between Caltech, City of Hope

PASADENA, Calif.—As part of a program to foster innovative biomedical research projects, an anonymous donor has pledged $3 million each to the California Institute of Technology (Caltech) and City of Hope to strengthen scientific collaborations between the two leading research institutions.

The gifts to Caltech and City of Hope will build upon support for research projects featuring partnerships between Caltech and City of Hope investigators. They will carry forward a program of collaborations between the two institutions that began in 2008 and has since helped start several promising projects in cancer, AIDS, and diabetes research.

"By combining the intellect, creativity, and passion of researchers from both of our institutions, we hope to accelerate the speed at which potentially game-changing discoveries in medical science are moved from the laboratory to patient care," says Caltech president Jean-Lou Chameau. "Private support is absolutely critical to achieving this goal, so we greatly appreciate this generous gift."

The biomedical initiative is meant to support those original, interdisciplinary, early-stage translational medical research projects that do not yet qualify for traditional sources of funding. Translational medicine is also known as bench-to-bedside research, in which basic science discoveries are developed into new therapies or treatments. Through the Caltech–City of Hope program, investigators will be able to leverage their unique strengths by working together. Eventually, these projects could lead to a clinical-trial phase with patients, which could then lead to new pharmaceuticals, medical devices, or other treatments.

"When we effectively apply the strengths of both City of Hope and Caltech to scientific research, we can more easily leverage groundbreaking discovery into the development of improved therapies for people facing serious illness such as cancer, HIV or diabetes," says Michael A. Friedman, president and chief executive officer of City of Hope. "We are very grateful for the generosity of donors who support our combined efforts to help patients everywhere."

Some of the Caltech–City of Hope projects that have already received seed funding include the investigation of a chemical compound that has been unexpectedly effective in reducing breast cancer tumors, the study of a gene that plays a role in tumor suppression, and a novel method of selectively killing HIV-infected cells using synthetic RNA molecules.

The gifts to the two institutions also provide support for an annual public event. The most recent event was held May 17 at Caltech. It featured presentations by Nobel Laureate David Baltimore, president emeritus and Robert Andrews Millikan Professor of Biology at Caltech; and John A. Zaia, the Aaron D. and Edith Miller Chair in Gene Therapy; Chair and Professor of Virology; and Deputy Director for Clinical Research at the Comprehensive Cancer Center at City of Hope.

As part of the pledge, Caltech and City of Hope have been challenged to raise an additional $3 million each toward their collaboration. The Caltech challenge is eligible for the Gordon and Betty Moore Matching Program, which, for a limited time, will contribute $1 for every $2 raised by Caltech.

Caltech has an extensive history of discovery in the biological sciences, and its faculty and alumni have won nine Nobel Prizes in medically related fields. City of Hope is a leading research, treatment, and education center for cancer, diabetes, and other life-threatening diseases.

Mike Rogers
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Caltech Scientist Awarded $5 Million Grant for Plant Research

PASADENA, Calif.—Elliot Meyerowitz, a plant genetics and developmental biology expert at the California Institute of Technology (Caltech), has been awarded one of 15 five-year, $5 million grants for fundamental plant science research.

The awards were made by the Howard Hughes Medical Institute (HHMI) and the Gordon and Betty Moore Foundation (GBMF).

Meyerowitz, the George W. Beadle Professor of Biology at Caltech, is an expert in the study of Arabidopsis thaliana, a small flowering plant in the mustard family and a model organism for plant genetics and molecular biology studies. During three decades of study, his work revealed the mechanism by which plants create their characteristic patterns of leaves and flowers; uncovered the first plant hormone receptor; and led to the sequencing of the Arabidopsis genome.

Traditionally, fundamental plant research often has been overlooked and underfunded. Indeed, despite the importance of plant science to food production, human health, environmental protection, and renewable-energy science, basic plant research represents only 2 percent of the funding for life sciences from the federal government.

The $75 million in awards represents an unprecedented influx of cash.

"I think this sort of funding is overdue, but not surprising," Meyerowitz says. "Medical researchers and policy makers are becoming increasingly aware that health and agriculture are critically related. The World Health Organization estimates that 10 percent of the entire world disease burden is due to undernutrition. The United Nations Food and Agriculture Organization estimates 925 million people are undernourished, and most estimates indicate that more people die of starvation every year, worldwide, than of cancer. Plants are also at the heart of many solutions to our energy problems and the relation of energy use to climate change," he adds, "because plants absorb carbon dioxide and emit oxygen."

"We think the creation of our joint program underscores the importance of investing in fundamental plant science, and we hope it will encourage others in the United States to make analogous commitments," said HHMI President Robert Tjian, in an announcement about the awards.

Meyerowitz is currently on leave from Caltech and serving as the inaugural director of the Sainsbury Laboratory at the University of Cambridge, which concentrates on understanding plant development and plant diversity through experimental and computational approaches. Meyerowitz will use the funding from the HHMI and the GBMF to develop a new interface between plant developmental biology and computational modeling, based on a method known as computational morphodynamics—the study of the three-way interaction of physical, informational, and geometrical processes that influences the changing form, shape, and structure of living cells, tissues, and organisms.

Kathy Svitil
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From Pre-Gut Cells to Glory

Caltech researchers discover a genomic control system that regulates gut formation in sea-urchin embryos

PASADENA, Calif.—For all animals, development begins with the embryo. It is here that uniform cells divide and diversify, and blueprints are laid for future structures, like skeletal and digestive systems. Although biologists have known for some time that signaling processes—messages that tell a cell to express certain genes so as to become certain parts of these structures—exist at this stage, there has not been a clear framework explanation of how it all comes together.

Now, a research team at the California Institute of Technology (Caltech) has outlined exactly how specific sets of cells in sea-urchin embryos differentiate to become the endoderm, the early domain of the embryo that eventually forms the gut. Their findings were reported in a paper entitled "A gene regulatory network controlling the embryonic specification of endoderm," published by the journal Nature online on May 29, in advance of the print version.

"If you only look at the genetic information of cells in an embryo, they all have the same genome and they all start from the single-cell zygote," says Isabelle S. Peter, a senior postdoctoral scholar at Caltech and coauthor of the study. "But then cells start to divide and, at some point, these cells are no longer identical in the genes that they express. We wanted to know how this process is achieved—how differences are established in cells in the right place and at the right time."

In order for undifferentiated cells to change their state and become a specific part of the body, the right genes need to be expressed, and the wrong ones repressed. The most important genes are regulatory genes, which control the expression of other genes, and form a gene regulatory network (GRN) that doles out differentiation instructions by turning genes on or off at specific times during embryonic development.

In the work described in the Nature paper, Peter and Eric H. Davidson, the Norman Chandler Professor of Cell Biology at Caltech and the other coauthor on the study, were able to analyze systematically the specification process controlled by the GRN and map out a master plan that, for the first time, shows the relationships between all the regulatory genes in specific parts of the embryo.

They studied sea-urchin embryos over a 24-hour period, beginning at hour eight of the embryo's existence. During this period, different physical domains exist in the embryo, each of which represents a future structure in the body, like the gut. All the regulatory genes known to exist in the sea-urchin genome and to be expressed in the embryo have been studied, and it was found that certain regulatory genes are expressed in the cells of each domain. Some of the domains will express certain regulatory genes in common, but the combination of genes found in each domain is unique. In addition, they found that this process is dynamic—where the genes are expressed changes over time. For example, two genes that are coexpressed in one domain at an early stage of the process may then be expressed in different domains at a later stage.

"It's like you are building a complicated edifice," explains Davidson. "And before anything is actually there, the building instructions have already been handed to all the workers. They all know what they are going to have to do once the bulldozer comes in and starts moving earth around."

The team focused on pinning down the precise regulatory genes in the progression of pre-gut cells (which eventually form the gut), following them from their initial stage as undifferentiated cells, to the point of gastrulation. During gastrulation the endoderm cells reorganize from a single layer into an internal tube with three regions that serve as the foundation for the future foregut, midgut, and hindgut structures. The researchers were able to pinpoint which regulatory genes were expressed at which specific times in the 24-hour period, and how those genes interacted over time to turn each other on and off.

"The instructions for development have to be in the genome somewhere, but you would be surprised how fragmentary the information about how that works was until we did this system-level analysis," says Davidson. "You can never understand it by looking at one gene. You can never understand it by looking at a third of the genes. You really have to get the whole system mapped out—and that's what we did."

In 2008, Davidson—who has been studying the biological processes of sea urchins for many years—led a research team that sketched a rough outline, for the first time, of how the GRN works to produce the sea urchin's skeletal system. "We are light-years beyond that with this new study," he says. "That was about solving network subcircuits, but now we have a framework that causally explains the far more complex process of development required for gut formation in terms of the genome's regulatory instruction code." This advance opens a much larger range of developmental scenarios to causal network analysis.

Sea urchins' gene regulatory systems, Davidson points out, are the closest—among the thoroughly studied invertebrate systems—to those of mammals, in terms of evolutionary relationships. This means the mechanisms the team uncovered in their work are likely to illuminate our own developmental regulatory systems. This could have implications for human health.

"If you believe that medicine consists of putting Band-Aids on things, then we have no relevance to that," says Davidson. "But if you believe that we should understand how life works before trying to find a cure when something goes wrong, then understanding biological processes from their initial stages comes first."

The team would next like to take their framework analysis and apply it to later stages in development—to when the gut is actually present. "We would like to understand how the different compartments in the gut are established, which would also make the work more directly informative to the development of the human gut," says Peter.

They would also like to extend the analysis to as much of the sea-urchin embryo as they can, says Davidson, as well as to formalize their findings to make an abstract computer model of how the gene regulatory system works. This will allow them to validate this particular network and to eventually do experiments that involve manipulating the cells to produce different results.

"Basically everything that happens in us, or in any animal during development, is encoded in genomic regulatory instructions," says Davidson. "Now we have an explanation as to how that works, which is very exciting. We can only move forward from here."

The work was funded by the Swiss National Science Foundation and the National Institutes of Health.

Katie Neith

Learning to Tolerate Our Microbial Self

PASADENA, Calif.—The human gut is filled with 100 trillion symbiotic bacteria—ten times more microbial cells than our own cells—representing close to one thousand different species. "And yet, if you were to eat a piece of chicken with just a few Salmonella, your immune system would mount a potent inflammatory response," says Sarkis K. Mazmanian, assistant professor of biology at the California Institute of Technology (Caltech).

Salmonella and its pathogenic bacterial kin don't look that much different from the legion of bacteria in our gut that we blissfully ignore, which raises the question: What decides whether we react or don't? Researchers have pondered this paradox for decades.

In the case of a common "friendly" gut bacterium, Bacteroides fragilis, Mazmanian and his colleagues have figured out the surprising answer: "The decision is not made by us," he says. "It's made by the bacteria. Since we are their home, they hold the key to our immune system."

What's more, the bacteria enforce their "decision" by hijacking cells of the immune system, say Mazmanian and his colleagues, who have figured out the mechanism by which the bacteria accomplish this feat—and revealed an explanation for how the immune system distinguishes between beneficial and pathogenic organisms.

In addition, the work, described in the April 21 issue of Science Express, "suggests that it's time to reconsider how we define self versus non-self," Mazmanian says.

Like other commensal gut bacteria—those that provide nutrients and other benefits to their hosts, without causing harm—B. fragilis was thought to live within the interior of the gut (the lumen), and thus far away from the immune system. "The dogma is that the immune system doesn't respond to symbiotic bacteria because of immunological ignorance," Mazmanian explains. "If we can't see them, we won't react to them."

But using a technique called whole-mount confocal microscopy to study the intestines of mice, he and his colleagues found that the bacteria actually live in a unique ecological niche, deep within the crypts of the colon, "and thus in intimate contact with the gut mucosal immune system," he says.

"The closeness of this association highlights that an active communication is occurring between the bacteria and their host," says Caltech postdoctoral scholar June L. Round.

From that vantage point, the bacteria are able to orchestrate control over the immune system—and, specifically, over the behavior of immune cells known as regulatory T cells, or Treg cells. The normal function of Treg cells is to prevent the immune system from reacting against our own tissues, by shutting down certain immune responses; they therefore prevent autoimmune reactions (which, when uncontrolled, can lead to diseases such as multiple sclerosis, type 1 diabetes, lupus, psoriasis, and Crohn's disease).

Bacteroides fragilis has evolved to produce a molecule that tricks the immune system into activating Treg cells in the gut, but in this case, Mazmanian says, "the purpose is to keep the cells from attacking the bugs. Beautiful, right?"

In their Science paper, Mazmanian and colleagues describe the entire molecular pathway that produces this effect. It starts with the bacteria producing a complex sugar molecule called polysaccharide A (PSA). PSA is sensed by particular receptors, known as Toll-like receptors, on the surfaces of Treg cells, thus activating those cells specifically. In response, Treg cells suppress yet another type of cell, the T helper 17 (Th17) cells. Normally, Th17 cells induce pro-inflammatory responses—those that would result, for example, in the elimination of foreign bacteria or other pathogens from the body. By shutting those cells down, B. fragilis gets a free pass to colonize the gut. "Up until now, we have thought that triggering of Toll-like receptors resulted solely in the induction of pathways that eliminate bacteria," says Round. "However, our studies suggest that multiple yet undiscovered host pathways allow us to coexist with our microbial partners."

When Mazmanian and his colleagues blocked this mechanism—by removing the PSA molecule, by removing the Toll-like receptor for PSA, or by eliminating the Treg cells themselves—the bacteria were attacked by the immune system and expelled. "They can no longer co-opt the immune system into inducing an anti-inflammatory response, so the formerly benign bacterium now looks like a pathogen," he says, "although the bug itself is exactly the same."

"Our immune system arose in the face of commensal colonization and thus likely evolved specialized molecules to recognize good bacteria," says Round. Mazmanian suspects that genetic mutations in these pathways could be responsible for certain types of immune disorders, including inflammatory bowel disease: "The question is, do patients get sick because they are rejecting bacteria they shouldn't reject?"

On a more philosophical level, Mazmanian says, the findings suggest that our concept of "self" should be broadened to include our many trillions of microbial residents. "These bacteria live inside us for our entire lives, and they've evolved to look and act like us, as part of us," he says. "As far as our immune system is concerned, the molecules made by gut bacteria should be tolerated similarly to our own molecules. Except in this case, the bacteria 'teaches' us to tolerate them, for both our benefit and theirs."

The other coauthors on the paper, "The Toll-like receptor 2 pathway establishes colonization by a commensal of the human microbiota," are S. Melanie Lee, Jennifer Li, and Gloria Tran of Caltech; Bana Jabri of the University of Chicago; and Talal A. Chatila of the David Geffen School of Medicine at UCLA. June L. Round was supported by a Jane Coffin Childs Memorial Fund postdoctoral fellowship. The work was supported by the National Institutes of Health, the Damon Runyon Cancer Research Foundation, and the Crohn's and Colitis Foundation of America.

Kathy Svitil

Caltech Biologist Recognized for Cellular Noise Research

Nearly ten years ago, Michael Elowitz, Caltech Bren Scholar and professor of biology, bioengineering, and applied physics, first amplified the idea that stochasticity—or noise—plays an important role in the process of gene expression. Prior to his work, such cellular noise was treated as a mysterious property.

For his pioneering work on gene expression noise, Elowitz has been named the winner of the 2011 Human Frontier Science Program (HFSP) Nakasone Award. The HFSP is a program that funds frontier research in the life sciences and the award is for breakthrough contributions at the frontier of the life sciences, either conceptual or methodological, that have had a major impact on basic biological research.

Elowitz's work introduced conceptual and experimental tools to detect gene expression noise, to quantify its level, and to evaluate its effect on cellular function. Genetic noise is now considered a core aspect of biology–one that functions actively in diverse cellular functions, including differentiation, regulation, and evolution.

Because of Elowitz's findings, noise has gone from being considered a curiosity of cellular life to being recognized as a key process whose effects must be considered in almost any analysis of biological systems. Stochastic processes are thought to enable stem cell differentiation and reprogramming, and developmental cell fates are controlled by noise. Thanks to Elowitz's work, noise is now recognized as an essential and functional element that distinguishes and enables the core cellular behaviors of life. 

Elowitz will give the HFSP Nakasone Lecture at the annual meeting of HFSP awardees to be held in Montreal, Canada in June

Katie Neith
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Pamela Bjorkman Named Among Most Powerful Moms

When Working Mother magazine recently compiled its list of the Most Powerful Moms in STEM (Science, Technology, Engineering, and Math), it included Caltech's Pamela Bjorkman—a pioneer in the study of cell-surface recognition in the immune system, and a mother of two—among its 10 honorees.

"I'm honored to be included among the very accomplished other women in this list," says Bjorkman, who is Caltech's Max Delbruck Professor of Biology and a Howard Hughes Medical Institute investigator.

"I think it's great to publicize that women can combine a career in science with having children. The more we can communicate this message to young women, the more likely we will be to keep women in the pipeline for STEM careers."

Working Mother says that the women on its list—which also includes Xerox Chairman and CEO Ursula Burns and Padmasree Warrior, Cisco's Chief Technology Officer—are all "shattering the illusion that women can't succeed in STEM fields."

"For all of the bunk that women aren't interested in careers in math or science," writes Leah Bourne in the magazine's introduction to the profiles of its selectees, "the numbers of women entering STEM careers has been quietly growing."

In its profile of Bjorkman, the magazine points to her "many awards," including election to the National Academy of Sciences in 2001, the L'Oreal-UNESCO Women in Science North American Laureate for her discovery of how the immune system recognizes targets in 2006 and a National Institute of Health Director's Pioneer Award in 2010. 

Despite her feeling of pride at being recognized by the magazine—"I can't wait to tell my kids that I'm one of the most powerful moms," she laughed when told of her inclusion on the list—Bjorkman says there is still a way to go in terms of women's parity in such traditionally male-dominated fields.

Working Mother agrees, noting that US Bureau of Labor Statistics show women hold only 14 percent of engineering positions, and a quarter of mathematics positions.

"Women are not so much deliberately excluded as they are not thought about," Bjorkman told the magazine. "It's human nature for people to have friends like themselves, and when a question comes up of who to invite to a meeting or who should give a talk, you tend to think of your friends. If all your friends are white males, then you'll tend to invite a white male. It's the same thing for minorities in science. It's this vicious circle. It's almost impossible to create a normal atmosphere for women in science when they are in such low numbers."

Lori Oliwenstein
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Caltech-Led Team Pinpoints Aggression Neurons in the Brain

Finding could lead to new treatments for impulsive violence

PASADENA, Calif.—Where does violence live in the brain? And where, precisely, does it lay down its biological roots? With the help of a new genetic tool that uses light to turn nerve cells on and off, a team led by researchers at the California Institute of Technology (Caltech) has tracked down the specific location of the neurons that elicit attack behaviors in mice, and defined the relationship of those cells to the brain circuits that play a key role in mating behaviors.

The researchers’ hope is that these insights might lead to treatments that can specifically address impulsive violence, a category of behavior that has been historically difficult to grapple with from a medical or psychological perspective.

In a study published in this week's issue of the journal Nature, the researchers were not only able to localize the neuronal circuits mediating attack behavior in mice, but were able to determine that these circuits are "intimately associated, deeply intertwined," with another basic social-behavioral drive—mating—according to David J. Anderson, the Benzer Professor of Biology at Caltech and a Howard Hughes Medical Institute investigator.

Indeed, the neurons for violence and mating live so close together, in a brain region known as the ventrolateral subdivision of the ventromedial hypothalamus (VMH), that "they are like a salt-and-pepper mixture," says Anderson.

And if you think of the brain as the world and the hypothalamus as a country, he adds, then the ventromedial hypothalamus is like a state and the ventrolateral subdivision is like a city within that state. "We've found that these 'mating' and 'fighting' neurons are not only located in the same city, but potentially in the same neighborhood," he says.

To determine whether aggression and mating involve the same—or, rather, distinct but intermingled—neurons, Dayu Lin, a former postdoctoral fellow in Anderson's lab and now an assistant professor at New York University, carried out a series of challenging electrophysiological recording experiments in the VMH. Because the VMH is located deep within the brain, it is exceedingly difficult to target accurately. But by inserting a bundle of 16 wire electrodes into this region, Lin was able to get recordings from multiple neurons during repeated episodes of fighting or mating. "It's the first time in which it's been possible to record electrical activity from deep-brain hypothalamic structures in animals while they are engaging in aggressive and mating behaviors," says Anderson.

Using software developed by Allen E. Puckett Professor of Electrical Engineering Pietro Perona and senior postdoctoral scholar Piotr Dollar—and with the help of several Caltech undergraduates—the researchers annotated behavioral changes on a frame-by-frame basis from video taken at the same time as the electrical recordings were performed. This annotation allowed them to make correlations between neuronal activity and behavior with a temporal resolution of approximately 30 milliseconds.

These experiments indicated that while there is some overlap between "mating" neurons and "fighting" neurons, the majority of these cells are distinct, despite their close proximity. Perhaps most surprising, Anderson notes, is the way that the neurons responsible for aggression and mating communicate—or, rather, how they shut each other up. Sex and violence, it seems, are actually at odds: a neuron that is turned on during aggressive behavior will turn off during mating, and vice versa. "We found that they talk to each other in an inhibitory way," he says.

But a correlation between neuronal activity and fighting behavior doesn't indicate whether the activity causes the behavior or the behavior causes the activity. And so Lin, Anderson, and colleagues carried out experiments to activate or inhibit VMH neurons, to distinguish between those alternatives and to determine the effect of such manipulations on behavior.

In order to activate neurons in the VMH, they used a technique known as optogenetic stimulation. Using a disabled virus as a kind of "disposable molecular syringe," Lin injected VMH neurons with DNA that carries the code for channelrhodopsin-2, a protein from blue-green algae that increases neuronal activity in response to blue light. The sensitized neurons could then be turned on or off with the literal flip of a light switch, allowing the scientists to watch what happens to the behavior of an individual mouse.

Remarkably, says Anderson, for mice in which the injection was targeted to the correct location, blue light induced an attack—even toward an inanimate object such as an inflated latex glove. Conversely, using a technique developed by Caltech’s Bren Professor of Biology Henry A. Lester that allowed the scientists to genetically inhibit neuronal activity, Lin and colleagues were able to show that neuronal activity in the VMH was necessary for normal aggressive behavior, as well.

"This answers an important, long-standing question in the field," says Anderson. "Are regions of the brain that can evoke aggression when artificially stimulated actually necessary for normal aggressive behavior? In this case, the answer is clearly 'yes.'"

The researchers also found that stimulating a male to be aggressive toward a female became more difficult as a mating encounter progressed to its consummatory phase. This result was consistent with the observation that neurons activated during fighting appear to become inhibited in the presence of a female. "The question," says Anderson, "is how that inhibition is achieved."

The answer may lead to new areas of research—and, perhaps, to new treatments for impulsive, violent behaviors. Specifically, notes Anderson, scientists can begin thinking about treatments that target violence-begetting neurons while sparing those involved in normal sexual behavior.

"For the last 500 years, we've really had no viable treatments for pathological violence other than execution or imprisonment," says Anderson. "And part of the reason is that we haven't understood enough about the basic neurobiology of aggression. The new studies are an important step in that direction."

In addition, he says, "mapping out the brain circuitry of aggression will provide a framework for understanding where and how in the brain genetic and environmental influences—nature vs. nurture—exert their influences on aggressive behavior."

The other authors on the Nature paper, "Functional identification of an aggression locus in the mouse hypothalamus," in addition to Anderson, Lin, Dollar, and Perona, are Caltech postdoctoral scholar Hyosang Lee and Maureen Boyle and Ed Lein from the Allen Institute for Brain Science in Seattle.

Their work was funded by the Weston-Havens Foundation, the Jane Coffin Childs Foundation, and the Howard Hughes Medical Institute.


Lori Oliwenstein


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