Positive Feedback: Researchers have found a new role for mTOR in autism-related disorders

Researchers have found a novel role for a protein that has been implicated in an autism-related disorder known as tuberous sclerosis complex (TSC).

The disease, which affects 1 in about 8,000 children, manifests itself in the form of mental retardation in addition to severe epileptic episodes. The disease is caused by mutations in two tumor-suppressing proteins, TSC1 and TSC2.

“Kids with this condition have benign tumors that grow all over the body,” said Bernardo Sabatini, the Takeda Professor of Neurobiology at Harvard Medical School and senior author of the study, “but we wanted to know what happened in the brain.”

The researchers found that when mutations in TSC1 and TSC2 adversely affected a third protein, mTOR, this mutation increased brain activity, which can result in epileptic seizures.

The findings were published in the May 8 issue of Neuron.

A protein kinase, mTOR is responsible for controlling cell growth in many parts of the body and has been widely implicated in epilepsy and autism. TSC1 and TSC2 normally repress the activity of mTOR to keep cell growth in check. In the case of TSC, there are mutations in TSC1 or TSC2, and mTOR’s ability to promote cell growth goes unchecked, resulting in tumors in regularly dividing cells.

“But neurons don’t divide,” said Sabatini. “So it was important to note the changes in these non-dividing cells.”  

The researchers hypothesized that mTOR’s function in the brain related to homeostasis, the brain’s ability to maintain a controlled level of electrical activity. When there’s a lot of electrical activity, a negative feedback system switches on to suppress activity. Conversely, when levels are too low, other positive feedback pathways are engaged that bring the activity level back up.

“We went into this study with the specific hypothesis that mTOR would be part of the homeostatic loop in the brain,” explained Sabatini.  

In the case of TSC patients, they thought that mTOR was incapable of maintaining homeostasis and kept adding to the level of electrical activity, leading to seizures. 

“But we were wrong,” he added.

“What we actually found was that mTOR is part of a positive feedback pathway,” said Helen Bateup, HMS research fellow in neurobiology and first author on the study. “When a cell is active, mTOR gets turned on more frequently and makes the cell even more active by reducing the amount of inhibition that the neuron receives.”

In cells where TSC proteins are mutated, this positive feedback gets out of control, and the neuronal circuit remains overactive despite all the pathways that normally shut down activity being turned on.

“It’s like the circuit is trying to keep itself quiet, but it can’t,” said Sabatini. “The out-of-control mTOR causes some cells to loss all inhibition, something that can’t be compensated for by turning down excitation.”                                        

The researchers think this key difference in how mTOR operates, in working to promote electrical activity, is important for the disease because patients end up with high levels of dysfunctional mTOR that makes for highly active circuits prone to epileptic fits. Furthermore, “we know that once a person has one seizure, they’re much more likely to have more, a concept known as kindling,” said Sabatini.

These findings are among the first to show that contrary to scientific consensus, mTOR does not play a part in everything.

“We have shown that one of the few things that mTOR does not seem to partake in is this negative feedback pathway,” said Sabatini.

Working in both in vitro and in vivo mouse models, the researchers think the next step would be tease out the molecular pathway of mTOR’s involvement in this positive feedback loop. “It’s also important to compare how this pathway works in normal brains versus a diseased model,” added Bateup.

“A huge challenge when studying the brain is that there are so many feedback pathways that a mutation in one gene can result in a hundred other secondary changes,” said Sabatini.

Rapamycin, a drug currently used to prevent organ rejection following transplants, targets mTOR and brings activity levels back to normal.

“We could use the drug to restore this excitatory-inhibitory balance in the brain,” said Bateup. “A lot of drugs that treat epilepsy try to make inhibition more powerful but given that the primary problem here is that a group of cells has lost inhibition, that approach won’t work,” she added. “What we might need is to target the excitation side. Or find ways of changing the biochemistry of the cells to make inhibitory synapses again.” 

“For this disease, this is the right time to start looking at human cells,” said Sabatini. “We have really good data from the mouse model and it would be a really nice test to see if the mouse model is really predictive of human disorder and if it’s worth being continued.” 

Brain capable of making its own version of Valium

The oral drug Valium – also known by its generic name, diazepam – was once popular with doctors in the 1970s as a treatment for seizures brought on by epilepsy. However, the drug, also used to treat anxiety, has fallen out of favor in recent years as it is prone to abuse and often dangerous if taken in high doses.

Now, in light of a recent study, the need for Valium to treat epilepsy may be even further diminished. Researchers from Stanford University School of Medicine have discovered a naturally occurring protein in the brains of mammals that acts like Valium, stopping certain types of seizures from occurring. Researchers hope that if they are able to discover a way to boost this protein naturally, doctors would no longer have a need to prescribe Valium.

The protein, identified as diazepam binding inhibitor (DBI), essentially acts like the brain’s very own brake system, sensing when a seizure is about to occur and arresting the process before it can spiral out of control.

“Our thinking on brain circuits and epilepsy has been that our brains have their own ways to control seizures, and this is why most of us aren’t having seizures every day,” study author John Huguenard, professor of neurology and neurological sciences at Stanford, told FoxNews.com. “But what happens as a seizure starts, a few cells in the brain may get too active, and you get an avalanche of activity that eventually can take up most of the brain circuitry. The brain’s own ‘Valium’ is acting as an anti-avalanche method, checking things when they’re first starting.”

According to Huguenard, the brain has two main groups of nerve cells. The first type of cells – excitatory cells – are responsible for stimulating other cells and sending messages from one area of the brain to another. This messaging process, known as excitation, is responsible for communicating what we see, what we smell, what we do, etc.

The other key type of cells are known as inhibitory cells, which are responsible for keeping the brain circuitry under control. If one area of the brain gets too excited and starts to receive too many signals at once, the inhibitory cells kick into gear and slow the process in order to restore balance.

“In terms of this form of epilepsy we’ve been studying, if a certain group of brain cells can’t communicate well through this inhibitory process, then (the animals) have seizures,” Huguenard said.

The protein DBI is a crucial component of the inhibitory process, as it boosts the actions of an important neurotransmitter called gamma-aminobutyric acid (GABA). Roughly one-fifth of the inhibitory nerve cells in the brain operate by secreting GABA, which binds to receptors located on excitatory cells, rendering them temporarily unable to fire any more electrical signals.  

Without DBI, GABA cannot be enhanced, and the excitatory cells ultimately don’t get the message telling them to calm down. However, up until now, this function of DBI was not well understood by researchers.

To determine exactly how DBI operates in the brains of mammals, Huguenard and his team analyzed a group of bioengineered mice with the DBI gene mutation, meaning their brains were incapable of producing DBI.

“When we tested seizures in these animals and tested communication, we found that (the inhibitory process) was ineffective and that the animals had more seizures,” Huguenard said. “It told us that this gene is producing a product in the brain that is controlling the seizures.”

When they re-introduced the DBI-gene back into the brains of these mice, GABA-induced inhibition was restored and the mice suffered from fewer seizures.

Benzodiazepine drugs, like Valium, work in a very similar way to DBI by also enhancing GABA-induced inhibition. But they often come at a high cost. Many who take these medications long-term develop a physical dependence on the drug, experiencing serious withdrawal symptoms if they cease taking it. Some studies have also found Valium to have an adverse effect on both short-term and long-term cognition.

While the researchers only examined the brains of mice, they are optimistic DBI exists similarly in the brains of humans as well. If the results end up translating to the human mind, Huguenard hopes to find a way to naturally boost DBI in the brain, negating the need for Valium to help control seizures.

“The ultimate goal would be to develop new lines of therapy that would take this general approach – taking the brain’s mechanism for dealing with seizures and making them even more effective,” Huguenard said.

The research was published May 30 in the journal Neuron.

Scientists discover the origin of a giant synapse

Humans and most mammals can determine the spatial origin of sounds with remarkable acuity. We use this ability all the time—crossing the street; locating an invisible ringing cell phone in a cluttered bedroom. To accomplish this small daily miracle, the brain has developed a circuit that’s rapid enough to detect the tiny lag that occurs between the moment the auditory information reaches one of our ears, and the moment it reaches the other. The mastermind of this circuit is the “Calyx of Held,” the largest known synapse in the brain. EPFL scientists have revealed the role that a certain protein plays in initiating the growth of these giant synapses.

The discovery, published in Nature Neuroscience, could also help shed light on a number of neuropsychiatric disorders.

Enormous synapses enable faster communication

Ordinarily, neurons have thousands of contact points – known as synapses - with neighboring neurons. Within a given time frame, a neuron has to receive several signals from its neighbors in order to be able to fire its own signal in response. Because of this, information passes from neuron to neuron in a relatively random manner.

In the auditory part of the brain, this is not the case. Synapses often grow to extremely large sizes, and these behemoths are known as “Calyx of Held” synapses. Because they have hundreds of contact points, they are capable of transmitting a signal singlehandedly to a neighboring neuron. “It’s almost like peer-to-peer communication between neurons,” explains EPFL professor Ralf Schneggenburger, who led the study. The result is that information is processed extremely quickly, in a few fractions of a millisecond, instead of the slower pace of more than 10 milliseconds that occurs in most other neuronal circuits.

Identifying the protein

To isolate the protein responsible for controlling the growth of this gigantic synapse, the scientists had to perform painstaking research. Using methods for analyzing gene expression in mice, they identified several members of the “BMP” family of proteins from among more than 20,000 possible candidates.

To verify that they had truly identified the right protein, the researchers disabled BMP protein receptors in the auditory part of a mouse brain. “The resulting electrophysiological signal of the Calyx of Held was significantly altered,” explains Le Xiao, first author on the study. “This would suggest a large anatomical difference.”

The scientists then reconstructed the synapses in three dimensions from slices that were observed under an electron microscope. Instead of a single, massive Calyx of Held, which would encompass nearly half the neuron, the 3D image of the neuron clearly shows several, smaller synapses. “This shows that the process involving the BMP protein not only causes that one synapse to grow, but also performs a selection, by eliminating the others,” says Schneggenburger.

Synaptic connectivity, the key to many psychiatric puzzles

The impact of this study will go well beyond increasing our understanding of the auditory system. The results suggest that the BMP protein plays an important role in developing connectivity in the brain. Schneggenburger and his colleagues are currently investigating its role elsewhere in the brain. “Some neuropsychiatric disorders, such as schizophrenia and autism, are characterized by the abnormal development of synaptic connectivity in certain key parts of the brain,” explains Schneggenburger. By identifying and explaining the role of various proteins in this process, the scientists hope to be able to shed more light on these poorly understood disorders.

‘Should I stay or should I go?’ CSHL scientists link brain cell types to behavior

You are sitting on your couch flipping through TV channels trying to decide whether to stay put or get up for a snack. Such everyday decisions about whether to “stay” or to “go” are supported by a brain region called the anterior cingulate cortex (ACC), which is part of the prefrontal cortex. Neuroscientists from Cold Spring Harbor Laboratory (CSHL) have now identified key circuit elements that contribute to such decisions in the ACC.

CSHL Associate Professor Adam Kepecs and his team publish results that, for the first time, link specific brain cell types to a particular behavior pattern in mice – a “stay or go” pattern called foraging behavior. The paper, published online in Nature, shows that the firing of two distinct types of inhibitory neurons, known as somatostatin (SOM) and parvalbumin (PV) neurons, has a strong correlation with the start and end of a period of foraging behavior.

Linking specific neuronal types to well-defined behaviors has proved extremely difficult. “There’s a big gap in our knowledge between our understanding of neuron types in terms of their physical location and their place in any given neural circuit, and what these neurons actually do during behavior,” says Kepecs.

Part of the problem is the technical challenge of doing these studies in live, freely behaving mice. Key to solving that problem is a mouse model developed in the laboratory of CSHL Professor Z. Josh Huang. The mouse has a genetic modification that allows investigators to target a specific population of neurons with any protein of interest.

Kepecs’ group, led by postdocs Duda Kvitsiani and Sachin Ranade, used this mouse to label specific neuron types in the ACC with a light-activated protein – a technique known as optogenetic tagging. Whenever they shone light onto the brains of the mice they were recording from, only the tagged PV and SOM neurons responded promptly with a ‘spike’ in their activity, enabling the researchers to pick them out from the vast diversity of cellular responses seen at any given moment.

The team recorded neural activity in the ACC of these mice while they engaged in foraging behavior. They discovered that the PV and SOM inhibitory neurons responded around the time of the foraging decisions — in other words whether to stay and drink or go and explore elsewhere. Specifically, when the mice entered an area where they could collect a water reward, SOM inhibitory neurons shut down and entered a period of low-level activity, thereby opening a ‘gate’ for information to flow in to ACC. When the mice decided to leave that area and look elsewhere, PV inhibitory neurons fired and abruptly reset cell activity.

“The brain is complex and continuously active, so it makes sense that these two types of inhibitory interneurons define the boundaries of a behavior such as foraging, opening and then closing the ‘gate’ within a particular neural circuit through changes in their activity,” says Kepecs.

This is an important advance, addressing a problem in behavioral neuroscience that scientists call “the cortical response zoo.” When researchers record neural activity in cortex during behavior, and they don’t know which type of neurons they are recording from, a bewildering array of responses is seen. This greatly complicates the task of interpretation. Hence the significance of the Kepecs team’s results, for the first time showing that specific cortical neuron types can be linked to specific aspects of behavior.

“We think about the brain and behavior in terms of levels; what the cell types are and the circuits or networks they form; which regions of the brain they are in; and what behavior is modulated by them,” explains Kepecs. “By observing that the activity of specific cell types in the prefrontal cortex is correlated with a behavioral period, we have identified a link between these levels.”

New neuron formation could increase capacity for new learning, at the expense of old memories

Cause of infantile amnesia revealed

New research presented today shows that formation of new neurons in the hippocampus - a brain region known for its importance in learning and remembering - could cause forgetting of old memories by causing a reorganization of existing brain circuits. Drs. Paul Frankland and Sheena Josselyn, both from the Hospital for Sick Children in Toronto, argue this reorganization could have the positive effect of clearing old memories, reducing interference and thereby increasing capacity for new learning. These results were presented at the 2013 Canadian Neuroscience Meeting, the annual meeting of the Canadian Association for Neuroscience - Association Canadienne des Neurosciences (CAN-ACN).

Researchers have long known of the phenomenon of infantile amnesia: This refers to the absence of long-term memory of events occurring within the first 2-3 years of life, and little long-term memories for events occurring until about 7 years of age. Studies have shown that though young children can remember events in the short term, these memories do not persist. This new study by Frankland and Josselyn shows that this amnesia is associated with high levels of new neuron production - a process called neurogenesis - in the hippocampus, and that more permanent memory formation is associated with a reduction in neurogenesis.

Dr. Frankland and Dr. Josselyn’s approach was to look at retention of memories in young mice in which they suppressed the usual high levels of neurogenesis in the hippocampus (thereby replicating the circuit stability normally observed in adult mice), but also in older mice in which they stimulated increased neurogenesis (thereby replicating the conditions normally seen in younger mice). Dr. Frankland was able to show a causal relationship between a reduction in neurogenesis and increased remembering, and the converse, decreased remembering when neurogenesis increased.

Dr. Frankland concludes: ” Why infantile amnesia exists has long been a mystery. We think our new studies begin to explain why we have no memories from our earliest years.”

Scientists Discover Cinnamon Compounds’ Potential Ability to Prevent Alzheimer’s

Cinnamon: Can the red-brown spice with the unmistakable fragrance and variety of uses offer an important benefit? The common baking spice might hold the key to delaying the onset of –– or warding off –– the effects of Alzheimer’s disease.

That is, according to Roshni George and Donald Graves, scientists at UC Santa Barbara. The results of their study, “Interaction of Cinnamaldehyde and Epicatechin with Tau: Implications of Beneficial Effects in Modulating Alzheimer’s Disease Pathogenesis,” appears in the online early edition of the Journal of Alzheimer’s Disease, and in the upcoming Volume 36, issue 1 print edition.

Alzheimer’s disease is the most common form of dementia, a neurodegenerative disease that progressively worsens over time as it kills brain cells. No cure has yet been found, nor has the major cause of Alzheimer’s been identified.

However, two compounds found in cinnamon –– cinnamaldehyde and epicatechin –– are showing some promise in the effort to fight the disease. According to George and Graves, the compounds have been shown to prevent the development of the filamentous “tangles” found in the brain cells that characterize Alzheimer’s.

Responsible for the assembly of microtubules in a cell, a protein called tau plays a large role in the structure of the neurons, as well as their function.

"The problem with tau in Alzheimer’s is that it starts aggregating," said George, a graduate student researcher. When the protein does not bind properly to the microtubules that form the cell’s structure, it has a tendency to clump together, she explained, forming insoluble fibers in the neuron. The older we get the more susceptible we are to these twists and tangles; Alzheimer’s patients develop them more often and in larger amounts.

The use of cinnamaldehyde, the compound responsible for the bright, sweet smell of cinnamon, has proven effective in preventing the tau knots. By protecting tau from oxidative stress, the compound, an oil, could inhibit the protein’s aggregation. To do this, cinnamaldehyde binds to two residues of an amino acid called cysteine on the tau protein. The cysteine residues are vulnerable to modifications, a factor that contributes to the development of Alzheimer’s.

"Take, for example, sunburn, a form of oxidative damage," said Graves, adjunct professor in UCSB’s Department of Molecular, Cellular, and Developmental Biology. "If you wore a hat, you could protect your face and head from the oxidation. In a sense this cinnamaldehyde is like a cap." While it can protect the tau protein by binding to its vulnerable cysteine residues, it can also come off, Graves added, which can ensure the proper functioning of the protein.

Oxidative stress is a major factor to consider in the health of cells in general. Through normal cellular processes, free radical-generating substances like peroxides are formed, but antioxidants in the cell work to neutralize them and prevent oxidation. Under some conditions however, the scales are tipped, with increased production of peroxides and free radicals, and decreased amounts of antioxidants, leading to oxidative stress.

Epicatechin, which is also present in other foods, such as blueberries, chocolate, and red wine, has proven to be a powerful antioxidant. Not only does it quench the burn of oxidation, it is actually activated by oxidation so the compound can interact with the cysteines on the tau protein in a way similar to the protective action of cinnamaldehyde.

"Cell membranes that are oxidized also produce reactive derivatives, such as Acrolein, that can damage the cysteines," said George. "Epicatechin also sequesters those byproducts."

Studies indicate that there is a high correlation between Type 2 diabetes and the incidence of Alzheimer’s disease. The elevated glucose levels typical of diabetes lead to the overproduction of reactive oxygen species, resulting in oxidative stress, which is a common factor in both diabetes and Alzheimer’s disease. Other research has shown cinnamon’s beneficial effects in managing blood glucose and other problems associated with diabetes.

"Since tau is vulnerable to oxidative stress, this study then asks whether Alzheimer’s disease could benefit from cinnamon, especially looking at the potential of small compounds," said George.

Although this research shows promise, Graves said, they are “still a long way from knowing whether this will work in human beings.” The researchers caution against ingesting more than the typical amounts of cinnamon already used in cooking.

If cinnamon and its compounds do live up to their promise, it could be a significant step in the ongoing battle against Alzheimer’s. A major risk factor for the disease –– age –––– is uncontrollable. In the United States, Alzheimer’s presents a particular problem as the population lives longer and the Baby Boom generation turns gray, leading to a steep rise in the prevalance of the disease. It is a phenomenon that threatens to overwhelm the U.S. health care system. According to the Alzheimer’s Association, in 2013, Alzheimer’s disease will cost the nation $203 billion.

"Wouldn’t it be interesting if a small molecule from a spice could help?" commented Graves, "perhaps prevent it, or slow down the progression."

(Image: iStockphoto)

The Secret Lives (and Deaths) of Neurons

As the human body fine-tunes its neurological wiring, nerve cells often must fix a faulty connection by amputating an axon — the “business end” of the neuron that sends electrical impulses to tissues or other neurons. It is a dance with death, however, because the molecular poison the neuron deploys to sever an axon could, if uncontained, kill the entire cell.

Researchers from the University of North Carolina School of Medicine have uncovered some surprising insights about the process of axon amputation, or “pruning,” in a study published May 21 in the journal Nature Communications. Axon pruning has mystified scientists curious to know how a neuron can unleash a self-destruct mechanism within its axon, but keep it from spreading to the rest of the cell. The researchers’ findings could offer clues about the processes underlying some neurological disorders.

“Aberrant axon pruning is thought to underlie some of the causes for neurodevelopmental disorders, such as schizophrenia and autism,” said Mohanish Deshmukh, PhD, professor of cell biology and physiology at UNC and the study’s senior author. “This study sheds light on some of the mechanisms by which neurons are able to regulate axon pruning.”

Axon pruning is part of normal development and plays a key role in learning and memory. Another important process, apoptosis — the purposeful death of an entire cell — is also crucial because it allows the body to cull broken or incorrectly placed neurons. But both processes have been linked with disease when improperly regulated.

The research team placed mouse neurons in special devices called microfluidic chambers that allowed the researchers to independently manipulate the environments surrounding the axon and cell body to induce axon pruning or apoptosis.

They found that although the nerve cell uses the same poison — a group of molecules known as Caspases — whether it intends to kill the whole cell or just the axon, it deploys the Caspases in a different way depending on the context.

“People had assumed that the mechanism was the same regardless of whether the context was axon pruning or apoptosis, but we found that it’s actually quite distinct,” said Deshmukh. “The neuron essentially uses the same components for both cases, but tweaks them in a very elegant way so the neuron knows whether it needs to undergo apoptosis or axon pruning.”

In apoptosis, the neuron deploys the deadly Caspases using an activator known as Apaf-1. In the case of axon pruning, Apaf-1 was simply not involved, despite the presence of Caspases. “This is really going to take the field by surprise,” said Deshmukh. “There’s very little precedent of Caspases being activated without Apaf-1. We just didn’t know they could be activated through a different mechanism.”

In addition, the team discovered that neurons employ other molecules as safety brakes to keep the “kill” signal contained to the axon alone. “Having this brake keeps that signal from spreading to the rest of the body,” said Deshmukh. “Remarkably, just removing one brake makes the neurons more vulnerable.”

Deshmukh said the findings offer a glimpse into how nerve cells reconfigure themselves during development and beyond. Enhancing our understanding of these basic processes could help illuminate what has gone wrong in the case of some neurological disorders.

Clouds in the Head: New Model of Brain’s Thought Processes

A new model of the brain’s thought processes explains the apparently chaotic activity patterns of individual neurons. They do not correspond to a simple stimulus/response linkage, but arise from the networking of different neural circuits. Scientists funded by the Swiss National Science Foundation (SNSF) propose that the field of brain research should expand its focus.