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Turrigiano lab uncovers sources of neuronal heterogeneity

December 18, 2020 By

High activity neurons have greater instrinsic excitability and response to local inputs, but no difference in total input type or amount

Mammalian cortex has long been one of the most widely studied systems in neuroscience, dating back to the pioneering work of Santiago Ramon y Cajal in the late 19th century. The cortex is much larger in primates than other mammals, and is thought to be responsible for the advanced cognitive abilities of humans. Today, models of cortical connections and computations form the basis for some of the most powerful deep learning paradigms. However, despite this success, there is still much that is unknown about how cortex functions. One feature of cortex that has recently been discovered is that neurons that appear to be similar to each other can have very different baseline activity levels: some neurons are 100x more active than their neighbors. We don’t know how neurons that are otherwise highly similar in shape and genetic makeup can maintain such different activity levels, or if the neurons with high and low activity levels have different functions in the brain. These neurons are otherwise so similar to each other that it is difficult to tell them apart without recording their activity directly, and current techniques for recording the activity of many neurons simultaneously in live animals do not allow us to later re-identify them for further study.

In a paper recently published in Neuron, the Turrigiano lab, led by postdoctoral researcher Nick Trojanowski, reported a new approach for permanently labeling high and low activity neurons in live animals, and then determining what makes them different. To do this they used a fluorescent protein called CaMPARI2 that changes from green to red as activity increases, but only when exposed to UV light. By shining UV light into the brain, they caused neurons with high activity to turn red, while neurons with low activity remained green. This procedure allowed them to run a series of tests on high and low activity neurons to identify differences between them. They found that high activity neurons would intrinsically generate more activity than low activity neurons when presented with the same stimulus. These high activity neurons also receive more excitatory input specifically from nearby neurons of the same type. Surprisingly, however, they found that the total amount of excitatory and inhibitory input that high and low activity neurons received from other neurons was not a major factor in determining their activity levels. Together, these results tell us that the differences in activity between neurons are due to intrinsic differences, as well as their pattern of connectivity to their nearby partners. This has deep implications for how the networks that underlie cortical computations are built and maintained.

With these tools in hand, it is now possible to further explore the differences between high and low activity neurons. Do these neurons serve different functions? Are the baseline activity levels specified from birth? How do these activity levels affect the mechanisms of plasticity that are responsible for learning and memory? The recently published results represent just the tip of the iceberg of information that can be learned with this new technique, in the mammalian cortex as well as other brain regions.

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Andrea Guerrero and Gina Turrigiano Receive 2020 Gilliam Fellowship

July 31, 2020 By
Gina Turrigiano & Andrea Guerrero

Andrea Guerrero (left) and Gina Turrigiano (right)

Andrea Guerrero, a Neuroscience PhD student working in the Turrigiano lab, a 2020 Gilliam Fellowships for Advanced Study recipient. The Gilliam Fellowship is awarded to the student and dissertation adviser, therefore Gina Turrigiano will also participate in this fellowship. Turrigiano said, “I am really pleased that Andrea was awarded this fellowship, which recognizes her potential to become a scientific leader.  I am also really excited at the opportunity to improve my mentoring skills that this terrific program provides to me as her PhD advisor.”

The purpose of the Gilliam Fellowship is to increase diversity among scientists who are preparing for leadership roles, particularly as college and university faculty members.  Fellows receive up to three years of support for dissertation research, typically in years three, four, and five of their PhD study. The Gilliam Fellowship is part of HHMI.

In response to receiving the award Guerrero said, “I am honored and excited to be selected as a 2020 HHMI Gilliam Fellowship recipient as it will aid my own advancement in an academic-track career and will importantly promote diversity and inclusion within the Brandeis science community.”

Andrea describes her research as follows:

“The human Shank3 gene is strongly associated with Autism Spectrum Disorder (ASD). Shank3 protein functions as a scaffold that plays a crucial role in synapse formation and maintenance. Prior work in our lab supports the idea that differential Shank3 phosphorylation alters its activity. Phosphomimetic and phosphodeficient mutants show dysfunction in the mechanisms that normally maintain brain circuitry homeostasis. In order to understand how Shank3 is able to do this, I will investigate how the phosphorylation state of Shank3 changes its synaptic localization, protein binding interactions, and cellular signaling pathways in vitro. Additionally, I will assess the effects of overexpression of Shank3 phosphorylation mutants on synaptic plasticity within the rodent primary visual cortex. My research project has the potential to uncover novel cellular pathways that can be targeted for ASD therapeutic development.”

 

 

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Autism-linked Gene Keeps Brains in Balance

April 3, 2020 By

Mutations in the human Shank3 gene – so called “Shankopathies” – are strongly associated with Autism-spectrum disorders and intellectual disability, and appear to increase risk for a number of other disorders such as bipolar disorder and epilepsy. How it is that loss of function of this single gene generates pervasive disfunction within the neural circuits that underlie cognition and behavior is not understood. Now a recent report from the Turrigiano lab at Brandeis (Autism-Associated Shank3 Is Essential for Homeostatic Compensation in Rodent V1. Neuron. 2020 Mar 10. ) sheds light into this process, by showing how Shank3 loss disables mechanisms that normally act to keep brain circuitry in balance. Much as your body maintains a constant temperature through the use of internal thermostats and negative feedback mechanisms, brain circuits maintain balanced activity – neither too low and unresponsive, nor too high and hyperactive – by using a set of so-called “homeostatic” plasticity mechanisms to keep circuit excitability within an ideal range. This process is especially important during childhood and adolescence, because developing circuits can easily get out of balance as brain circuitry changes as a result of normal developmental processes.

Using mouse and rat models of human Shankopathies, the team, led by Research Associate Vedakumar Tatavarty, found that loss of Shank3 disables these homeostatic plasticity mechanisms and prevents brain circuits from compensating for changes to sensory drive. These defects in homeostatic plasticity are due to acute loss of Shank3 within individual neurons, meaning they are not an indirect effect of messed-up circuit wiring caused by loss of the gene throughout development. This finding suggests that Shank3 is a fundamental part of the cellular machinery that normally mediates homeostatic plasticity. The team went on to show that homeostatic plasticity could be restored after Shank3 loss by treatment with Lithium – a drug with a long history of use to treat neuropsychiatric disorders such as bipolar disorder – and that Lithium was also able to reduce a repetitive grooming behavior in mice that lack Shank3. These mice normally groom to excess, even to the point of self-injury, but a week of lithium treatment was able to reduce grooming to normal levels.

So do these findings suggest that Lithium might be useful in treating human Shankopathies? While Lithium remains the frontline treatment for some human disorders such as bipolar disorder, it is not well-tolerated, says Turrigiano, “and of course we cannot extrapolate from findings in mice directly to humans. Instead, we hope to use Lithium as a tool to reveal the pathways that can restore homeostatic plasticity in Shankopathies, which in the long term may allow us to design better, more specific interventions”. Defects in homeostatic plasticity have been implicated in a wide range of human brain disorders ranging from Autism spectrum disorders to Alzheimer’s disease, so these studies are likely to have important implications for overall brain health.

Autism-Associated Shank3 Is Essential for Homeostatic Compensation in Rodent V1. Tatavarty V, Torrado Pacheco A, Groves Kuhnle C, Lin H, Koundinya P, Miska NJ, Hengen KB, Wagner FF, Van Hooser SD, Turrigiano GG. Neuron. 2020 Mar 10. pii: S0896-6273(20)30184-7. doi: 10.1016/j.neuron.2020.02.033.

Filed Under: Biology, labs Tagged With: , , ,

Raul Ramos Pays It Forward in His Home State of Texas

May 3, 2018 By


photo credit: Simon Goodacre

Helen Wong | Graduate School of Arts and Sciences

Raul Ramos, a fourth-year Ph.D. candidate in Neuroscience, spent the five-hour flight from Boston to Austin, Texas trying to think of what to say to a classroom full of adolescents who had been sentenced to juvenile detention, like he had been once when he was a teenager.

“I was trying to get into the mindset of it all,” he says of those nerve-wracking hours before arriving in Austin. “I was trying to remember how I felt when I had been in their shoes.” He had put together a talk and a script, but the moment he entered the first classroom at the Austin Alternative Learning Center, all of it went out the window. “Instead of giving a lecture, I had an actual conversation with the kids,” says Ramos. “They could relate to me. I was someone who looked like them, talked like them, moved like them. So they listened when I told them about my story and how, despite what they were facing now, their outcomes could be different too.”

Ramos first started working with high school students after he moved to Waltham. Anique Olivier-Mason PhD’12, Director of Education, Outreach and Diversity at the Materials Research Science and Engineering Center had arranged “Pizza Talks,” a program where graduate students in the sciences visit classrooms at Waltham High School and discuss their decisions to pursue careers in science, their experiences as investigators and their research. The program has been a great success and now serves as the model for similar talks taking place nationally, sponsored by the American Association for the Advancement of Science (AAAS). Ramos volunteered to give a talk when he first heard about the program.

“Waltham High has a large Hispanic student population,” says Ramos. “These groups underrepresented in science. I really liked going to speak to them and talking about my own journey and its relation to my identity.” AAAS became aware of this community outreach and contacted the university to learn more. Ramos has always been open about the troubles in his own past, so when AAAS were looking for scientists to speak to students in alternative learning centers in Austin, they asked him if he would like to go. “I said yes, of course,” says Ramos. “I’m from Texas originally, so I agreed to fly down and talk to the kids.”

What began as originally just one or two schools became six upon his arrival in Austin as word got around of his visit. During the trip, Ramos gave sixteen talks and spoke to around two hundred students. “I went to juvie centers, alternative learning centers, drug rehabilitation facilities,” he says. “The level of engagement was amazing. For every kid that didn’t want to engage, there were a few more who wanted to talk to me and learn about how I’d gotten to where I am. One of the most frequent questions they asked me was, ‘Sir, what do I do when I get out of here?’ and I would tell them the truth. I told them that once they got out, they would have to actively avoid situations and people that would get them in trouble. I said that if that meant having to hole up in their room to study and get away from it all, then doing that would absolutely be worth it in the long run. Their environment matters.”

But even after telling them his advice, Ramos knew that advice alone wasn’t going to be enough for many of the kids he was speaking to. “You need a support network,” he says. “A lot of these kids don’t have that. Some of them are safer in detention than at home. So many of them are angry–why wouldn’t they be? They’re supposed to become upstanding members of society, but the way the system goes about that is to lock them up and isolate them. That’s not how rehabilitation should work.”

At some of the facilities he visited, Ramos saw kids as young as eleven or twelve being escorted by armed guards from classroom to classroom despite some of them being barely half his size. For Ramos, the sight was jarring. “It looks like overkill,” says Ramos. “I know they’re here because they did something wrong, but at the end of the day, they’re just kids.”

It also struck Ramos, as he made the rounds in each facility, that the kids incarcerated at these centers were all people of color despite Austin being in a majority white part of Texas. “Brandeis is all about recruiting underrepresented minorities into its science programs,” he says. However, the challenges of recruiting students of color for doctoral programs in science are significant, and Ramos realized during his trip to Texas that “part of the reason for the absence of black and brown individuals in science was that so many of them, who could potentially be scientists someday, are stuck in juvie–stuck in environments that deprive them of opportunities and healthy role models.

“And people like me that manage to get an education, we make it out and we leave. We come over here to go to college, we leave Laredo [Ramos’ hometown], and these kids don’t get to have good role models. They make mistakes fueled by a terrible home environment and get stuck in the juvie-to-prison pipeline. They repent and feel bad in juvie, but once they get out, if they don’t have a support network, it starts all over again. The system tries them as full adults at seventeen, when they’re not even old enough to vote. Things have to change. I want to help make that happen and to show them that right now, there are still opportunities open to them.”

Despite all of the system’s shortcomings, the alternative learning centers and similar institutions are making a tangible difference. “The system’s not perfect,” says Ramos. “It’s deeply flawed. But things are already better now than when I was in. Back then, I was put in what would conventionally be considered a prison cell. At least most of these kids get an education, space to walk, and are surrounded by people who care about them. Everyone working at the Austin Alternative Learning Center was so motivated and clearly cared about the kids.”

Upon his return to Brandeis, Ramos decided that he would dedicate more time to community outreach and consider the possibility of working in science policy after earning his doctorate. He wants to do work that not only has value in the scientific world, but that also actively helps bolster diversity and inclusion in the field, helping fight back against larger societal and institutional structures that disadvantage people of color.

“We need representation to show kids that the journey is possible,” says Ramos. “The cards feel like they’re stacked almost the entire way through. I’m going to do whatever I have to do to get the message out there to those kids who are hardest to reach and who need to hear from us the most.”

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Recycling is good for your brain

August 25, 2016 By

If you were able to remember where you put your keys on your way out the door this morning, it’s because – somehow – synapses in your brain changed their properties to encode this information and store it until you needed it. This process, known as “synaptic plasticity”, is essential for the continuity of our memory and sense of self, and yet we are only beginning to grasp the molecular mechanisms that enable this amazing feat of constant information storage and retrieval. Now a collaborative paper from the Turrigiano and Nelson labs just published in Cell Reports sheds important new light into how experience interacts with the genome to allow synapses to change their strength to store information.

Synapses are the connections between neurons, and it has long been appreciated that information is stored in large part through changes in the strength of these connections. Changes in strength at many synapses are in turn determined by the number of neurotransmitter receptors that are clustered at synaptic sites – the more receptors synapses have, the easier it is for neurons to excite each other to transmit information. Synapses are highly complicated molecular machines that utilize at least 300 different proteins that interact to traffic these receptors to synapses and sequester them there, and exactly how a change in experience alters the function of this nano-machine to enhance the number of synaptic receptors is still a matter of puzzlement.

In this study the Brandeis team devised a way to screen for candidate proteins that are critical for a particular form of synaptic plasticity: “synaptic scaling”, thought to be especially important for maintaining brain stability during learning and development. They were able to induce synaptic scaling within specific labelled neurons in the intact mouse brain (layer 4 star pyramidal neurons), and then sort out those labelled neurons from the rest of the brain and probe for changes in gene expression that were correlated with (and potentially causally involved in) the induction of plasticity.  This approach produced a small number of candidate genes that were up- or down-regulated during plasticity, to produce more or less of a given protein.  The team then went on to show that – when upregulated – one of these candidates (known as µ3A) acts to prevent neurotransmitter receptors from going into the cellular garbage bin (the lysosomes, where proteins are degraded) and instead recycles them to the synapse. Thus increased µ3A flips a switch within cells to enhance receptor recycling, and this in turn increases synaptic strength.

µ3A plays a critical role in the recycling of AMPA-type neurotransmitter receptors

A screen for genes with altered expression during synaptic plasiticity in specific neurons revealed that µ3A plays a critical role in the recycling of AMPA-type neurotransmitter receptors at the synapse. When this protein is upregulated, it prevents receptors from being trafficked into lysosomes, and instead allows them to be recycled back to synapses, increasing synapse number and enhancing synaptic strength.

It turns out that many other forms of synaptic plasticity use the same receptor recycling machinery as synaptic scaling, so it is likely that this mechanism represents  an important and general way for neurons to alter synaptic strength. This study also raises the possibility that defects in this pathway might contribute to the genesis of neurological disorders in which the stability of brain circuits is disrupted, such as epilepsy and autism. So next time you complain about having to sort your garbage, consider that your neurons do it all the time –  and what’s good for the planet turns out to be good for your brain as well.

Steinmetz CC, Tatavarty V, Sugino K, Shima Y, Joseph A, Lin H, Rutlin M, Lambo M, Hempel CM, Okaty BW, Paradis S, Nelson SB, Turrigiano G. Upregulation of μ3A Drives Homeostatic Plasticity by Rerouting AMPAR into the Recycling Endosomal Pathway. Cell reports. 2016.

Filed Under: Biology, Neuroscience Tagged With: , , , ,

Sleep suppresses brain rebalancing

March 18, 2016 By

Why humans and other animals sleep is one of the remaining deep mysteries of physiology. One prominent theory in neuroscience is that sleep is when the brain replays memories “offline” to better encode them (“memory consolidation”). A prominent and competing theory is that sleep is important for re-balancing activity in brain networks that have been perturbed during learning while awake. Such “rebalancing” of brain activity involves homeostatic plasticity mechanisms that were first discovered at Brandeis University, and have been thoroughly studied by a number of Brandeis labs including the Turrigiano lab. Now, a study from the Turrigiano lab just published in the journal Cell shows that these homeostatic mechanisms are indeed gated by sleep and wake, but in the opposite direction from that theorized previously: homeostatic brain rebalancing occurs exclusively when animals are awake, and is suppressed by sleep. These findings raise the intriguing possibility that different forms of brain plasticity – for example those involved in memory consolidation and those involved in homeostatic rebalancing – must be temporally segregated from each other to prevent interference.

sleeprats

The requirement that neurons carefully maintain an average firing rate, much like the thermostat in a house senses and maintains temperature, has long been suggested by computational work. Without homeostatic (“thermostat-like”) control of firing rates, models of neural networks cannot learn and drift into states of epilepsy-like saturation or complete quiescence. Much of the work in discovering and describing candidate mechanisms continues to be conducted at Brandeis. In 2013, the Turrigiano Lab provided the first ­in vivo evidence for firing rate homeostasis in the mammalian brain: lab members recorded the activity of individual neurons in the visual cortex of freely behaving rat pups for 8h per day across a nine-day period during which vision through one eye was occluded. The activity of neurons initially dropped, but over the next 4 days, firing rates came back to basal levels despite the visual occlusion. In essence, these experiments confirmed what had long been suspected – the activity of neurons in intact brains is indeed homeostatically governed.

Due to the unique opportunity to study a fundamental mechanism of brain plasticity in an unrestrained animal, the lab has been probing the possibility of an intersection between an animal’s behavior and homeostatic plasticity. In order to truly evaluate possible circadian and behavioral influences on neuronal homeostasis, it was necessary to capture the entire 9-day experiment, rather than evaluate snapshots of each day. For this work, the Turrigiano Lab had to find creative computational solutions to recording many terabytes of data necessary to follow the activity of single neurons without interruption for more than 200 hours. Ultimately, these data revealed that the homeostatic regulation of neuronal activity in the cortex is gated by sleep and wake states. In a surprising and unpredicted twist, the homeostatic recovery of activity occurred almost exclusively during periods of activity and was inhibited during sleep. Prior predictions either assumed no role for behavioral state, or that sleeping would account for homeostasis. Finally, the lab established evidence for a causal role for active waking by artificially enhancing natural waking periods during the homeostatic rebound. When animals were kept awake, homeostatic plasticity was further enhanced.

This finding opens doors onto a new field of understanding the behavioral, environmental, and circadian influences on homeostatic plasticity mechanisms in the brain. Some of the key questions that immediately beg to be answered include:

  • What it is about sleep that precludes the expression of homeostatic plasticity?
  • How is it possible that mechanisms requiring complex patterns of transcription, translation, trafficking, and modification can be modulated on the short timescales of behavioral state-transitions in rodents?
  • And finally, how generalizable is this finding? As homeostasis is bidirectional, does a shift in the opposite direction similarly require wake or does the change in sign allow for new rules in expression?

Authors on the paper include postdoctoral fellow Keith Hengen, Neuroscience grad student Alejandro Torrado Pachedo, and Neuroscience undergraduate James McGregor ’14 (now in grad school at Emory).

Hengen KB, Torrado Pacheco A, McGregor JN, Van Hooser SD, Turrigiano GG. Neuronal Firing Rate Homeostasis is Inhibited by Sleep and Promoted by Wake. Cell. 2016.

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