Linking insulin to learning: Important insights in research with worms

Recent work by Harvard researchers demonstrates how the signaling pathway of insulin and insulinlike peptides plays a critical role in helping to regulate learning and memory.

The research, led by Yun Zhang, associate professor of organismic and evolutionary biology, is described in a Feb. 6 paper in Neuron.

“People think of insulin and diabetes, but many metabolic syndromes are associated with some types of cognitive defects and behavioral disorders, like depression or dementia,” Zhang said. “That suggests that insulin and insulinlike peptides may play an important role in neural function, but it’s been very difficult to nail down the underlying mechanism, because these peptides do not have to function through synapses that connect different neurons in the brain.”

To get at that mechanism, Zhang and colleagues turned to an organism whose genome and nervous system are well described and highly accessible by genetics: C. elegans.

Using genetic tools, researchers altered the transparent worms by removing their ability to create individual insulinlike compounds. These new “mutant” worms were then tested to see whether they would learn to avoid eating a particular type of bacteria that is known to infect the worms. Tests showed that although some worms did learn to steer clear of the bacteria, others didn’t — suggesting that removing a specific insulinlike compound halted the worms’ ability to learn.

Researchers were surprised to find, however, that it wasn’t just removing the molecules that could make the animals lose the ability to learn — some peptides were found to inhibit learning.

“We hadn’t predicted that we would find both positive and negative regulators from these peptides,” Zhang said. “Why does the animal need this bidirectional regulation of learning? One possibility is that learning depends on context. There are certain things you want to learn — for example, the worms in these experiments wanted to learn that they shouldn’t eat this type of infectious bacteria. That’s a positive regulation of the learning. But if they needed to eat, even if it is a bad food, to survive, they would need a way to suppress this type of learning.”

Even more surprising for Zhang and her colleagues was evidence that the various insulinlike molecules could regulate each other.

“Many animals, including humans, have multiple insulinlike molecules, and it appears that these molecules can act like a network,” she said. “Each of them may play a slightly different role in the nervous system, and they function together to coordinate the signaling related to learning and memory. By changing the way the molecules interact, the brain can fine-tune learning in a host of different ways.”

Mouse brain cells live long and prosper

Mouse brain cells scamper close to eternal life: They can actually outlive their bodies. Mouse neurons transplanted into rat brains lived as long as the rats did, surviving twice as long as the mouse’s average life span, researchers report online February 25 in the Proceedings of the National Academy of Sciences.

The findings suggest that long lives might not mean deteriorating brains. “This could absolutely be true in other mammals — humans too,” says study author Lorenzo Magrassi, a neurosurgeon at the University of Pavia in Italy.

The findings are “very promising,” says Carmela Abraham, a neuroscientist at Boston University. “The question is: Can neurons live longer if we prolong our life span?” Magrassi’s experiment, she says, suggests the answer is yes.

One theory about aging, Magrassi says, is that every species has a genetically determined life span and that all the cells in the body wear out and die at roughly the same time. For the neurons his team studied, he says, “We have shown that this simple idea is certainly not true.”

Magrassi’s team surgically transplanted neurons from embryonic mice with an average life span of 18 months into rats. To do so, the researchers slipped a glass microneedle through the abdomens of anesthetized pregnant mice. Then, using a dissecting microscope and a tool to illuminate the corn-kernel-sized mouse embryos, the researchers scraped out tiny bits of brain tissue and injected the neurons into fetal rat brains. After the rat pups were born, Magrassi and colleagues waited as long as three years, until the animals were near death, to euthanize the rats and dissect their brains.

The transplanted mouse cells had linked up with the rat brain cells and developed into mature, working neurons, though they did retain their characteristic small size. Also, because Magrassi’s team had tagged the mouse cells to glow green, the researchers could distinguish between mouse and rat neurons. The mouse cells lived twice as long as they would have in a mouse brain, and they showed signs of aging similar to those of neighboring rat neurons.

Figuring out what’s helping the neurons survive could lead researchers to treatments for human neurodegenerative diseases, such as Parkinson’s and Alzheimer’s, Magrassi says.

BPA May Affect the Developing Brain by Disrupting Gene Regulation

Environmental exposure to bisphenol A (BPA), a widespread chemical found in plastics and resins, may suppress a gene vital to nerve cell function and to the development of the central nervous system, according to a study led by researchers at Duke Medicine.

The researchers published their findings - which were observed in cortical neurons of mice, rats and humans - in the journal Proceedings of the National Academy of Sciences on Feb. 25, 2013.

"Our study found that BPA may impair the development of the central nervous system, and raises the question as to whether exposure could predispose animals and humans to neurodevelopmental disorders," said lead author Wolfgang Liedtke, M.D., PhD, associate professor of medicine/neurology and neurobiology at Duke.

BPA, a molecule that mimics estrogen and interferes with the body’s endocrine system, can be found in a wide variety of manufactured products, including thermal printer paper, some plastic water bottles and the lining of metal cans. The chemical can be ingested if it seeps into the contents of food and beverage containers.

Research in animals has raised concerns that exposure to BPA may cause health problems such as behavioral issues, endocrine and reproductive disorders, obesity, cancer and immune system disorders. Some studies suggest that infants and young children may be the most vulnerable to the effects of BPA, which led the U.S. Food and Drug Administration to ban the use of the chemical in baby bottles and cups in July 2012.

While BPA has been shown to affect the developing nervous system, little is understood as to how this occurs. The research team developed a series of experiments in rodent and human nerve cells to learn how BPA induces changes that disrupt gene regulation.

During early development of neurons, high levels of chloride are present in the cells. These levels drop as neurons mature, thanks to a chloride transporter protein called KCC2, which churns chloride ions out of the cells. If the level of chloride within neurons remains elevated, it can damage neural circuits and compromise a developing nerve cell’s ability to migrate to its proper position in the brain.

Exposing neurons to minute amounts of BPA alters the chloride levels inside the cells by somehow shutting down the Kcc2 gene, which makes the KCC2 protein, thereby delaying the removal of chloride from neurons.

MECP2, another protein important for normal brain function, was found to be a possible culprit behind this change. When exposed to BPA, MECP2 is more abundant and binds to the Kcc2 gene at a higher rate, which might help to shut it down. This could contribute to problems in the developing brain due to a delay in chloride being removed.

These findings raise the question of whether BPA could contribute to neurodevelopmental disorders such as Rett syndrome, a severe autism spectrum disorder that is only found in girls and is characterized by mutations in the gene that produces MECP2.

While both male and female neurons were affected by BPA in the studies, female neurons were more susceptible to the chemical’s toxicity. Further research will dig deeper into the sex-specific effects of BPA exposure and whether certain sex hormone receptors are involved in BPA’s effect on KCC2.

"Our findings improve our understanding of how environmental exposure to BPA can affect the regulation of the Kcc2 gene. However, we expect future studies to focus on what targets aside from Kcc2 are affected by BPA," Liedtke said. "This is a chapter in an ongoing story."

Small groups of brain cells store concepts for memory formation– from Luke Skywalker to your grandmother

Concepts in our minds – from Luke Skywalker to our grandmother - are represented by their own distinct group of neurons, according to new research involving a University of Leicester neuroscientist.

The research, by neuroscientist Professor Rodrigo Quian Quiroga from the University of Leicester Centre for Systems Neuroscience together with Professor Itzhak Fried, of the UCLA David Geffen School of Medicine, Tel Aviv Sourasky Medical Center and Tel Aviv University, and Professor Christof Koch, of the California Institute of Technology and Allen Institute for Brain Science, Seattle, is featured in a recent article of the prestigious Scientific American magazine.

Recent experiments during brain surgeries have shown that small groups of brain cells are responsible for encoding memories of specific people or objects.

These neurons may also represent different variations of one thing – from the name of a person to their appearance from many different viewpoints.

The researchers believe that single concepts may be held in as little as thousands of neurons or less – a tiny fraction of the billion or so neurons contained in the medial temporal lobe, which is a memory related structure within the brain.

The group were able to monitor the brain activity of consenting patients undergoing surgery to treat epilepsy. This allowed the team to monitor the activity of single neurons in conscious patients while they looked at images on laptop screens, creating and recalling memories.

In previous experiments, they had found that single neurons would ‘fire’ for specific concepts – such as Luke Skywalker – even when they were viewing images of him from different angles or simply hearing or reading his name.

They have also found that single neurons can also fire to related people and objects – for instance, the neuron that responded to Luke Skywalker also fired to Yoda, another Jedi from Star Wars.

They argue that relatively small groups of neurons hold concepts like Luke Skywalker and that related concepts such as Yoda are held by some but not all of the same neurons. At the same time, a completely separate set of neurons would hold an unrelated concept like Jennifer Aniston.

The group believes this partially overlapping representation of related concepts are the neural underpinnings of encoding associations, a key memory function.

Professor Quian Quiroga said: “After the first thrill when finding neurons in the human hippocampus with such remarkable firing characteristics, converging evidence from experiments we have been carrying out in the last years suggests that we may be hitting one of the key mechanisms of memory formation and recall.

“The abstract representation of concepts provided by these neurons is indeed ideal for representing the meaning of the sensory stimuli around us, the internal representation we use to form and retrieve memories. These concepts cells, we believe, are the building blocks of memory functions.”

Modeling Alzheimer’s disease using iPSCs reveals stress phenotypes associated with intracellular Aβ and differential drug responsiveness

Working with a group from Nagasaki University, a research group at the Center for iPS Cell Research and Application (CiRA) has successfully modeled Alzheimer’s disease (AD) using both familial and sporadic patient-derived induced pluripotent stem cells (iPSCs), and revealed stress phenotypes and differential drug responsiveness associated with intracellular amyloid β oligomers in AD neurons and astrocytes.

In a study published online in Cell Stem Cell, Associate Professor Haruhisa Inoue and his team at CiRA and a research group led by Professor Nobuhisa Iwata of Nagasaki University generated cortical neurons and astrocytes from iPSCs derived from two familial AD patients with mutations in amyloid precursor protein (APP), and two sporadic AD patients. The neural cells from one of the familial and one of the sporadic patients showed endoplasmic reticulum (ER)-stress and oxidative-stress phenotypes associated with intracellular Aβ oligomers. The team also found that these stress phenotypes were attenuated with docosahexaenoic acid (DHA) treatment. These findings may help explain the variable clinical results obtained using DHA treatment, and suggest that DHA may in fact be effective only for a subset of patients.
Using both familial and sporadic AD iPSCs, the researchers discovered that pathogenesis differed between individual AD patients. For example, secreted Aβ42 levels were depressed in familial AD with APP E693Δ mutation, elevated in familial AD with APP V717L mutation, but normal in sporadic AD.

"This shows that patient classification by iPSC technology may contribute to a preemptive therapeutic approach toward AD," said Inoue, a principal investigator at CiRA who is also a research director for the CREST research program funded by the Japan Science and Technology Agency. "Further advances in iPSC technology will be required before large-scale analysis of AD patient-specific iPSCs is possible."

Fragile X makes brain cells talk too much

The most common inherited form of mental retardation and autism, fragile X syndrome, turns some brain cells into chatterboxes, scientists at Washington University School of Medicine in St. Louis report.

The extra talk may make it harder for brain cells to identify and attend to important signals, potentially establishing an intriguing parallel at the cellular level to the attention problems seen in autism.

According to the researchers, understanding the effects of this altered signaling will be important to developing successful treatments for fragile X and autism.

“We don’t know precisely how information is encoded in the brain, but we presume that some signals are important and some are noise,” says senior author Vitaly Klyachko, PhD, assistant professor of cell biology and physiology. “Our theoretical model suggests that the changes we detected may make it much more difficult for brain cells to distinguish the important signals from the noise.”

The findings appear Feb. 20 in Neuron.

Fragile X is caused by mutations in a gene called Fmr1. This gene is found on the X chromosome, one of the two sex chromosomes. Females have two copies of that chromosome, while males only have one. As a result, males have fragile X syndrome more often than females, and the effects in males tend to be more severe.

Symptoms of fragile X include mental retardation, hyperactivity, epilepsy, impulsive behavior, and delays in the development of speech and walking. Fragile X also affects anatomy, leading to unusually large heads, flat feet, large body size and distinctive facial features. Thirty percent of fragile X patients are autistic.

Scientists deleted the Fmr1 gene many years ago in mice to create a model of fragile X. Without Fmr1, the mice have abnormalities in brain cells and social and behavioral deficits similar to those seen in human fragile X.

According to Klyachko, nearly all fragile X mouse studies in the past two decades have focused on how Fmr1 loss affects dendrites, the branches of nerve cells that receive signals. In contrast, his new study finds significant changes in axons, the branches of nerve cells that send signals.

Normally, signals travel down the axon as surges of electrical energy. These surges only last for tiny fractions of a second, briefly causing the axon to release compounds known as neurotransmitters into the short gap between nerve cells. The neurotransmitters cross the gap and bind to their receptors on the dendrite to convey the signal.

When Klyachko monitored electrical surges along axons in the fragile X mice, though, he discovered that they lasted significantly longer. This caused release of more of neurotransmitters from the axon. When it should have stopped talking, the axon continued to chatter.

“The axons are putting out much more neurotransmitter than they should, and we think this confuses the system and overloads the circuitry,” Klyachko explains. “It may also create problems in terms of brain cells using up their resources much more quickly than they normally would.”

Infusing synthetic copies of the gene’s protein, called FMRP, into brain cells from the mouse model rapidly restored the electrical surges to their normal length.

Additional experiments revealed that FMRP works by interacting with one of the biggest channels on the surfaces of axons. These channels let electrically charged potassium ions into the axons, helping to shape and control the duration of the electrical surge.

In healthy brain cells, the main function of these channels is to prevent the electrical surge from getting too long. With FMRP gone, the channel is active for a shorter time, prolonging the surge and overwhelming the dendrite with too much chatter.

Klyachko and his colleagues are now studying the connections between FMRP and the channel it interacts with in axons. They hope to learn more about how information is encoded and processed at the level of individual brain cells. These insights one day may help clinicians better diagnose and treat many kinds of mental disorders.

Researchers develop tool for reading the minds of mice

If you want to read a mouse’s mind, it takes some fluorescent protein and a tiny microscope implanted in the rodent’s head.

Stanford scientists have demonstrated a technique for observing hundreds of neurons firing in the brain of a live mouse, in real time, and have linked that activity to long-term information storage. The unprecedented work could provide a useful tool for studying new therapies for neurodegenerative diseases such as Alzheimer’s.

The researchers first used a gene therapy approach to cause the mouse’s neurons to express a green fluorescent protein that was engineered to be sensitive to the presence of calcium ions. When a neuron fires, the cell naturally floods with calcium ions. Calcium stimulates the protein, causing the entire cell to fluoresce bright green.

A tiny microscope implanted just above the mouse’s hippocampus – a part of the brain that is critical for spatial and episodic memory – captures the light of roughly 700 neurons. The microscope is connected to a camera chip, which sends a digital version of the image to a computer screen.

The computer then displays near real-time video of the mouse’s brain activity as a mouse runs around a small enclosure, which the researchers call an arena.

The neuronal firings look like tiny green fireworks, randomly bursting against a black background, but the scientists have deciphered clear patterns in the chaos.

"We can literally figure out where the mouse is in the arena by looking at these lights," said Mark Schnitzer, an associate professor of biology and of applied physics and the senior author on the paper, recently published in the journal Nature Neuroscience.

When a mouse is scratching at the wall in a certain area of the arena, a specific neuron will fire and flash green. When the mouse scampers to a different area, the light from the first neuron fades and a new cell sparks up.

"The hippocampus is very sensitive to where the animal is in its environment, and different cells respond to different parts of the arena," Schnitzer said. "Imagine walking around your office. Some of the neurons in your hippocampus light up when you’re near your desk, and others fire when you’re near your chair. This is how your brain makes a representative map of a space."

The group has found that a mouse’s neurons fire in the same patterns even when a month has passed between experiments. “The ability to come back and observe the same cells is very important for studying progressive brain diseases,” Schnitzer said.

Neuronal activity induces tau release from healthy neurons

Researchers from King’s College London have discovered that neuronal activity can stimulate tau release from healthy neurons in the absence of cell death. The results published by Diane Hanger and her colleagues in EMBO reports show that treatment of neurons with known biological signaling molecules increases the release of tau into the culture medium. The release of tau from cortical neurons is therefore a physiological process that can be regulated by neuronal activity.

Tau proteins stabilize microtubules, the long threads of polymers that help to maintain the structure of the cell. However, in Alzheimer’s disease or certain types of dementia, tau accumulates in neurons or glial cells, where it contributes to neurodegeneration.

In addition to intracellular aggregation, recent experiments have shown that tau is released from neuronal cells and taken up by neighboring cells, which allows the spread of aggregated tau across the brain. This release could occur passively from dying neuronal cells, though some evidence suggests it might take place before neuronal cell death and neurodegeneration. The new findings indicate that tau release is an active process in healthy neurons and this could be altered in diseased brains.

“Our findings suggest that altered tau release is likely to occur in response to changes in neuronal excitability in the Alzheimer’s brain. Secreted tau could therefore be involved in the propagation of tau pathology in tauopathies, a group of neurodegenerative diseases associated with the accumulation of tau proteins in the brain,” commented Diane Hanger, Reader in the Department of Neuroscience at King’s College London. In these experiments, Amy Pooler, the lead author, revealed that molecules such as potassium chloride, glutamate or an AMPA receptor agonist could release tau from cortical neurons in an active physiological process that is, at least partially, dependent on pre-synaptic vesicle secretion.

The new findings by the scientists indicate that tau has previously unknown roles in biological signaling between cells, in addition to its well-established role in stabilizing microtubules.

“We believe that targeting the release of tau could be explored as a new therapeutic approach for the treatment of Alzheimer’s disease and related tauopathies,” said Hanger. Additional studies are needed in model organisms to test this hypothesis further.

(Image: Patrick Hoesly)

Hypothalamic control of energy balance: insights into the role of synaptic plasticity

The past 20 years witnessed an enormous leap in understanding of the central regulation of whole-body energy metabolism. Genetic tools have enabled identification of the region-specific expression of peripheral metabolic hormone receptors and have identified neuronal circuits that mediate the action of these hormones on behavior and peripheral tissue functions. One of the surprising findings of recent years is the observation that brain circuits involved in metabolism regulation remain plastic through adulthood. In this review, we discuss these findings and focus on the role of neurons and glial cells in the dynamic process of plasticity, which is fundamental to the regulation of physiological and pathological metabolic events.