(Image caption: Three-dimensional reconstruction of a synapse in the mouse brain. Readily releasable fusionable synaptic vesicles (blue, around 45 millionths of a millimetre in diameter) are docked at the cell membrane. Credit: © MPI f. Experimental Medicine/ Benjamin H. Cooper)

Synapses always on the starting blocks

While neurons rapidly propagate information in their interior via electrical signals, they communicate with each other at special contact points known as the synapses. Chemical messenger substances, the neurotransmitters, are stored in vesicles at the synapses. When a synapse becomes active, some of these vesicles fuse with the cell membrane and release their contents. To ensure that valuable time is not lost, synapses always have some readily releasable vesicles on standby. With the help of high-resolution, three-dimensional electron microscopy, scientists at the Max Planck Institute of Experimental Medicine in Göttingen succeeded in demonstrating that these fusionable vesicles have a very special characteristic: they already have close contact with the cell membrane long before the actual fusion occurs. In addition, the research team also decoded the molecular machinery that facilitates the operation of this docking mechanism.

The fusion of the neurotransmitter vesicles with the cell membrane involves close cooperation between numerous protein components, which monitor each other and ensure that every single ‘participant’ is always in the right place. This is referred to as the fusion machinery and the comparison is an apt one: if a cogwheel in a clock mechanism is broken, the hands do not move. In a similar way, faulty or missing molecules impair synaptic operations.

In research studies carried out some years ago, Nils Brose and his colleague JeongSeop Rhee from the Max Planck Institute of Experimental Medicine in Göttingen already demonstrated that the transmission of information at the synapses in genetically modified mice, in which all known genes of the Munc13 or CAPS proteins had been switched off, is severely defective. Although the neurons of the genetically modified mice do not differ from those of healthy mice when examined under an optical microscope, if Munc13 is missing, the release of neurotransmitters actually grinds to a halt completely. Brose and Rhee’s findings showed that to be able to react immediately to signals at all times, each synapse must keep a small number of ‘readily releasable’ fusionable vesicles on standby.

But how do Munc13 and CAPS convert the vesicles to this kind of fusionable state? To answer this question, the Göttingen-based scientists studied the synaptic contacts in the minutest possible detail. To do this, neurobiologists Cordelia Imig and Ben Cooper, who have been working with Brose and Rhee for many years, used a high-pressure freezing process. This involves the rapid freezing of neurons in the brain tissue under high pressure so that no disruptive ice crystals are formed and the fine structure of the cells is particularly well conserved. The samples obtained in this way were then analysed using electron tomography. Using this method, electron microscope images of a structure are recorded from many different angles, in a similar way to the process used in medical computed tomography. The individual images can then be combined on the computer to give a high-resolution three-dimensional image – of a synapse in this case (see image).

“Our results showed that readily releasable vesicles in healthy synapses touch the cell membrane,” explains Cooper. “However, if Munc13 and CAPS proteins are missing, the vesicles do not reach the active zone and accumulate a few nanometres away from it.” To their astonishment, the researchers also observed that SNARE proteins, which collaborate with Munc13 and CAPS in the nerve endings, are also involved in this docking process. SNARE proteins are found in the cell and vesicle membranes of healthy synapses and control the fusion of the two membranes during neurotransmitter release. When a vesicle approaches the cell membrane, the individual SNARE molecules line up opposite each other like the sides of a zip and pull the membranes close to each other in this way. The vesicles await the starting gun for their fusion in this state – in the starting blocks, so to speak.

The findings of the neurobiologists in Göttingen prove that Munc13, CAPS and SNARE proteins closely align the vesicle and cell membrane in the synapse, long before the signal for fusion is given. This is the only way that the fast and controlled transmission of information at the synapse can be guaranteed, thanks to which we can react specifically to information from our environment. “It had long been clear that synapses have to be extremely fast to carry out all of the many complex brain functions. Our study shows for the first time how this is managed at the molecular level and on the level of the synaptic vesicles,” says Brose. Because almost all of the protein components involved in this process also play a role in neurological and psychiatric diseases, the Göttingen-based scientists believe that their discovery will soon benefit medical research.

(Image caption: Pictured is a mouse hippocampal neuron studded with thousands of synaptic connections (yellow). The number and location of synapses — not too many or too few — is critical to healthy brain function. The researchers found that MHCI proteins, known for their role in the immune system, also are one of the only known factors that ensure synapse density is not too high. The protein does so by inhibiting insulin receptors, which promote synapse formation. Credit: Lisa Boulanger)

Immune proteins moonlight to regulate brain-cell connections

When it comes to the brain, “more is better” seems like an obvious assumption. But in the case of synapses, which are the connections between brain cells, too many or too few can both disrupt brain function.

Researchers from Princeton University and the University of California-San Diego (UCSD) recently found that an immune-system protein called MHCI, or major histocompatibility complex class I, moonlights in the nervous system to help regulate the number of synapses, which transmit chemical and electrical signals between neurons. The researchers report in the Journal of Neuroscience that in the brain MHCI could play an unexpected role in conditions such as Alzheimer’s disease, type II diabetes and autism.

MHCI proteins are known for their role in the immune system where they present protein fragments from pathogens and cancerous cells to T cells, which are white blood cells with a central role in the body’s response to infection. This presentation allows T cells to recognize and kill infected and cancerous cells.

In the brain, however, the researchers found that MHCI immune molecules are one of the only known factors that limit the density of synapses, ensuring that synapses form in the appropriate numbers necessary to support healthy brain function. MHCI limits synapse density by inhibiting insulin receptors, which regulate the body’s sugar metabolism and, in the brain, promote synapse formation.

Senior author Lisa Boulanger, an assistant professor in the Department of Molecular Biology and the Princeton Neuroscience Institute (PNI), said that MHCI’s role in ensuring appropriate insulin signaling and synapse density raises the possibility that changes in the protein’s activity could contribute to conditions such Alzheimer’s disease, type II diabetes and autism. These conditions have all been associated with a complex combination of disrupted insulin-signaling pathways, changes in synapse density, and inflammation, which activates immune-system molecules such as MHCI.

Patients with type II diabetes develop “insulin resistance” in which insulin receptors become incapable of responding to insulin, the reason for which is unknown, Boulanger said. Similarly, patients with Alzheimer’s disease develop insulin resistance in the brain that is so pronounced some have dubbed the disease “type III diabetes,” Boulanger said.

"Our results suggest that changes in MHCI immune proteins could contribute to disorders of insulin resistance," Boulanger said. "For example, chronic inflammation is associated with type II diabetes, but the reason for this link has remained a mystery. Our results suggest that inflammation-induced changes in MHCI could have consequences for insulin signaling in neurons and maybe elsewhere."

MHCI levels also are “dramatically altered” in the brains of people with Alzheimer’s disease, Boulanger said. Normal memory depends on appropriate levels of MHCI. Boulanger was senior author on a 2013 paper in the journal Learning and Memory that found that mice bred to produce less functional MHCI proteins exhibited striking changes in the function of the hippocampus, a part of the brain where some memories are formed, and had severe memory impairments.

"MHCI levels are altered in the Alzheimer’s brain, and altering MHCI levels in mice disrupts memory, reduces synapse number and causes neuronal insulin resistance, all of which are core features of Alzheimer’s disease," Boulanger said.

Links between MHCI and autism also are emerging, Boulanger said. People with autism have more synapses than usual in specific brain regions. In addition, several autism-associated genes regulate synapse number, often via a signaling protein known as mTOR (mammalian target of rapamycin). In their study, Boulanger and her co-authors found that mice with reduced levels of MHCI had increased insulin-receptor signaling via the mTOR pathway, and, consequently, more synapses. When elevated mTOR signaling was reduced in MHCI-deficient mice, normal synapse density was restored.

Thus, Boulanger said, MHCI and autism-associated genes appear to converge on the mTOR-synapse regulation pathway. This is intriguing given that inflammation during pregnancy, which alters MHCI levels in the fetal brain, may slightly increase the risk of autism in genetically predisposed individuals, she said.

"Up-regulating MHCI is essential for the maternal immune response, but changing MHCI activity in the fetal brain when synaptic connections are being formed could potentially affect synapse density," Boulanger said.

Ben Barres, a professor of neurobiology, developmental biology and neurology at the Stanford University School of Medicine, said that while it is known that both insulin-receptor signaling increases synapse density, and MHCI signaling decreases it, the researchers are the first to show that MHCI actually affects insulin receptors to control synapse density.

"The idea that there could be a direct interaction between these two signaling systems comes as a great surprise," said Barres, who was not involved in the research. "This discovery not only will lead to new insight into how brain circuitry develops but to new insight into declining brain function that occurs with aging."

Particularly, the research suggests a possible functional connection between type II diabetes and Alzheimer’s disease, Barres said.

"Type II diabetes has recently emerged as a risk factor for Alzheimer’s disease but it has not been clear what the connection is to the synapse loss experienced with Alzheimer’s disease," he said. "Given that type II diabetes is accompanied by decreased insulin responsiveness, it may be that the MHCI signaling becomes able to overcome normal insulin signaling and contribute to synapse decline in this disease."

Research during the past 15 years has shown that MHCI lives a prolific double-life in the brain, Boulanger said. The brain is “immune privileged,” meaning the immune system doesn’t respond as rapidly or effectively to perceived threats in the brain. Dozens of studies have shown, however, that MHCI is not only present throughout the healthy brain, but is essential for normal brain development and function, Boulanger said. A 2013 paper from her lab published in the journal Molecular and Cellular Neuroscience showed that MHCI is even present in the fetal-mouse brain, at a stage when the immune system is not yet mature.

"Many people thought that immune molecules like MHCI must be missing from the brain," Boulanger said. "It turns out that MHCI immune proteins do operate in the brain — they just do something completely different. The dual roles of these proteins in the immune system and nervous system may allow them to mediate both harmful and beneficial interactions between the two systems."

An enzyme and synaptic plasticity: Study reveals novel role for the Pin1

Synapses are “dynamic” things: they can regulate their action in neural processes related to learning, for example, but also as a consequence of diseases. A research team –led by SISSA– has demonstrated the role of a small enzyme (Pin1) in synaptic plasticity. The study has just been published in the journal Nature Communications.

A small, “empty” space teeming with activity: a synapse is a complex structure where the neural (electrical) signal from the presynaptic neuron, as it travels towards its target –a muscle, a gland or another neuron– turns into a chemical signal capable of crossing the synaptic space before becoming electrical again once on the other side. A synapse is a “dynamic” space not only because of the endless work that goes on there, but also for its ability to change its action over time (synaptic plasticity) as a result of either normal physiological processes (e.g., during learning) or because of disorders due to pathological conditions. A study, mainly carried out by SISSA researchers (which also involved the University of Zurich, LNCIB in Trieste, and EBRI in Rome), showed that a small enzyme (Pin1, peptidylprolyl isomerase) that plays a mediating role in signal transmission has an effect on synaptic plasticity.

The synapse we studied is of the inhibitory kind. The signal it transmits hinders activation of the postsynaptic neuron, making it less likely for it to become activated and emit its action potential”, explains Paola Zacchi, a SISSA researcher who coordinated the study. “When Pin1 is absent from the synapse, signal transmission occurs “at full strength”, but also without control. Instead, when it is present, it regulates signal strength, making it weaker. We observed that Pin1 is able to modify the number of postsynaptic receptors”. The larger the number of receptors capable of binding to the neurotransmitter, the stronger the signal that reaches the postsynaptic membrane. “This also means that Pin1 plays a role in plasticity” explains Zacchi.

How does a synapse work? “A chemical synapse, the most common in vertebrates, is a small gap between nerve cells where the passage of a neural signal occurs”, explains Zacchi. In chemical synapses the two neurons are not in contact but they are separated by a distance of about 20 nanometres. For this reason, the electrical signal travelling along the presynaptic nerve ending is interrupted before resuming on the neuron on the other side of the gap. In between the two nerve cells the electrical signal is translated into a chemical signal (which then becomes electrical again).

“Arrival of the action potential on the presynaptic button causes release, into the interneural space, of molecules of neurotransmitter, which are picked up by receptors on the postsynaptic membrane”, says Zacchi. “If the synapse is excitatory, this leads to postsynaptic activation which, if sufficiently intense, triggers another action potential. If the synapse is inhibitory, as in our studies, the signal suppresses postsynaptic activation and inhibits firing of the electrical potential. In the process of neurotransmitter release and binding, other molecules come into play, such as scaffold proteins, which assemble receptors at the right place on the membrane in front of the neurotransmitter release sites, and neuroligins which act as bridges between the two ends of the synapse as well as interacting with the scaffold proteins. Pin1, the enzyme in the study, interacts with both neuroligins and scaffold proteins.

The Pin1 enzyme has long been known for its role in cancer and the development of neurodegenerative diseases such as Alzheimer’s and Parkinson’s (whereas neuroligins seem to be involved in autism). “Studies like this enhance our understanding of the biochemical mechanisms of synaptic plasticity, extending our knowledge of healthy mechanisms, but also helping those who are trying to understand what can be done in a wide range of pathological conditions”.

Protein pairing builds brain networks

Neural networks are formed by the interconnection of specific neurons in the brain. The molecular mechanisms involved in creating these connections, however, have so far eluded scientists. Research led by Jun Aruga from the RIKEN Brain Science Institute has now identified an interaction between two proteins that is crucial for making connections between specific types of neurons, with implications for some neurological disorders.

Connections between neurons are made via synapses—small gaps across which chemicals called neurotransmitters pass, relaying signals from a presynaptic neuron to a postsynaptic neuron. Aruga and his colleagues focused on a protein called mGluR7, which is found only at synapses with a specific type of postsynaptic neuron in an area of the brain involved in forming memories.

“mGluR7 is located on the presynaptic side of connections made with hippocampal local inhibitory neurons,” explains Aruga. “Previous studies have proposed that this protein prevents neurotransmitter release from the presynaptic neuron when the neurotransmitter glutamate binds to it.”

The researchers discovered that the localization of mGluR7 to specific synapses is determined by the presence of another protein called Elfn1. This protein is found on the other side of the same synapses, directly opposite mGluR7. When the researchers artificially introduced Elfn1 into cultured cells, mGluR7 became associated with the same cells, and they showed that this was due to a physical interaction between the two proteins. Conversely, deleting Elfn1 in the brains of mice reduced the amount of mGluR7 at the synapses.

These changes interfered with the process of strengthening connections at synapses, which takes place during memory formation, and caused patterns of brain waves that indicated abnormally high levels of electrical activity. Genetically altered mice also exhibited other symptoms that resembled human conditions.

“Deleting Elfn1 increased the susceptibility of mice to seizures,” explains Aruga. “It also enhanced behaviors similar to attention deficit hyperactivity disorder (ADHD).”

Indeed, the researchers found that humans with epilepsy and ADHD also had a faulty version of the gene encoding Elfn1, suggesting that a deficit in the ability of Elfn1 to localize mGluR7 and form specific connections in neural networks is important in some neurological conditions.

“In combination, the human and mouse results implicate the Elfn1–mGluR7 complex in the pathophysiology of epilepsy and ADHD, at least in part,” explains Aruga, although he remains cautious at this early stage of research. “Because of sample size limitations, the human genetics result is not conclusive, but we are now awaiting the results of follow-up studies with additional subjects.”

The Yin and Yang of Overcoming Cocaine Addiction

Yaoying Ma says that biology, by nature, has a yin and a yang—a push and a pull.

Addiction, particularly relapse, she finds, is no exception.

Ma is a research associate in the lab of Yan Dong, assistant professor of neuroscience in the University of Pittsburgh’s Kenneth P. Dietrich School of Arts and Sciences. She is the lead author of a paper published online today in the journal Neuron that posits that it may be possible to ramp up an intrinsic anti-addiction response as a means to fight cocaine relapse and keep the wolves of relapse at bay.

This paper is the first to establish the existence of a brain circuitry that resists a relapse of cocaine use through a naturally occurring neural remodeling with “silent synapses.”

The work is a follow-up on a recent study conducted by Dong and his colleagues, which was published in Nature Neuroscience last November. The team used rat models to examine the effects of cocaine self-administration and withdrawal on nerve cells in the nucleus accumbens, a small region in the brain that is commonly associated with reward, emotion, motivation, and addiction. Specifically, they investigated the roles of synapses—the structures at the ends of nerve cells that relay signals.

The team reported in its Nature Neuroscience study that when a rat uses cocaine, some immature synapses are generated, which are called “silent synapses” because they are semifunctional and send few signals under normal physiological conditions. After that rat stops using cocaine, these “silent synapses” go through a maturation phase and acquire their full function to send signals. Once they can send signals, the synapses will send craving signals for cocaine if the rat is exposed to cues previously associated with the drug.

The current Neuron paper shows that there’s another side of “silent synapse” remodeling. Silent synapses that are generated in a specific cortical projection to the nucleus accumbens by cocaine exposure become “unsilenced” after cocaine withdrawal, resulting in a profound remodeling of this cortical projection. Additional experiments show that silent synapse-based remodeling of this cortical projection decreases cocaine craving. Importantly, this anti-relapse circuitry remodeling is induced by cocaine exposure itself, suggesting that our body has its own way to fight addiction.

Dong, the paper’s senior author, says that the pro-relapse response is predominant after cocaine exposure. But since the anti-relapse response exists inside the brain, it could possibly be clinically tweaked to achieve therapeutic benefits.

Ma notes that this finding “may provide insight into ways to manipulate this yin-yang balance and hopefully provide new neurobiological targets for interventions designed to decrease relapse.”

“The story won’t stop here,” Ma adds. “Our ongoing study is exploring some unusual but simple ways to beef up the endogenous anti-relapse mechanism.”

(Image: PA)

Memory in silent neurons

When we learn, we associate a sensory experience either with other stimuli or with a certain type of behaviour. The neurons in the cerebral cortex that transmit the information modify the synaptic connections that they have with the other neurons. According to a generally-accepted model of synaptic plasticity, a neuron that communicates with others of the same kind emits an electrical impulse as well as activating its synapses transiently. This electrical pulse, combined with the signal received from other neurons, acts to stimulate the synapses. How is it that some neurons are caught up in the communication interplay even when they are barely connected? This is the crucial chicken-or-egg puzzle of synaptic plasticity that a team led by Anthony Holtmaat, professor in the Department of Basic Neurosciences in the Faculty of Medicine at UNIGE, is aiming to solve. The results of their research into memory in silent neurons can be found in the latest edition of Nature.

Learning and memory are governed by a mechanism of sustainable synaptic strengthening. When we embark on a learning experience, our brain associates a sensory experience either with other stimuli or with a certain form of behaviour. The neurons in the cerebral cortex responsible for ensuring the transmission of the relevant information, then modify the synaptic connections that they have with other neurons. This is the very arrangement that subsequently enables the brain to optimise the way information is processed when it is met again, as well as predicting its consequences.

Neuroscientists typically induce electrical pulses in the neurons artificially in order to perform research on synaptic mechanisms.

The neuroscientists from UNIGE, however, chose a different approach in their attempt to discover what happens naturally in the neurons when they receive sensory stimuli. They observed the cerebral cortices of mice whose whiskers were repeatedly stimulated mechanically without an artificially-induced electrical pulse. The rodents use their whiskers as a sensor for navigating and interacting; they are, therefore, a key element for perception in mice.

An extremely low signal is enough

By observing these natural stimuli, professor Holtmaat’s team was able to demonstrate that sensory stimulus alone can generate long-term synaptic strengthening without the neuron discharging either an induced or natural electrical pulse. As a result – and contrary to what was previously believed – the synapses will be strengthened even when the neurons involved in a stimulus remain silent.In addition, if the sensory stimulation lasts over time, the synapses become so strong that the neuron in turn is activated and becomes fully engaged in the neural network. Once activated, the neuron can then further strengthen the synapses in a forwards and backwards movement. These findings could solve the brain’s “What came first?” mystery, as they make it possible to examine all the synaptic pathways that contribute to memory, rather than focusing on whether it is the synapsis or the neuron that activates the other.

The entire brain is mobilised

A second discovery lay in store for the researchers. During the same experiment, they were also able to establish that the stimuli that were most effective in strengthening the synapses came from secondary, non-cortical brain regions rather than major cortical pathways (which convey actual sensory information). Accordingly, storing information would simply require the co-activation of several synaptic pathways in the neuron, even if the latter remains silent. These findings may also have important implications both for the way we understand learning mechanisms and for therapeutic possibilities, in particular for rehabilitation following a stroke or in neurodegenerative disorders. As professor Holtmaat explains: “It is possible that sensory stimulation, when combined with another activity (motor activity, for example), works better for strengthening synaptic connections”. The professor concludes: “In the context of therapy, you could combine two different stimuli as a way of enhancing the effectiveness.”

(Image caption: In a study of brains from children with autism, neurons in brains from autistic patients did not undergo normal pruning during childhood and adolescence. The images show representative neurons from unaffected brains (left) and brains from autistic patients (right); the spines on the neurons indicate the location of synapses. Credit: Guomei Tang, PhD and Mark S. Sonders, PhD/Columbia University Medical Center)

Children with Autism Have Extra Synapses in Brain

Children and adolescents with autism have a surplus of synapses in the brain, and this excess is due to a slowdown in a normal brain “pruning” process during development, according to a study by neuroscientists at Columbia University Medical Center (CUMC). Because synapses are the points where neurons connect and communicate with each other, the excessive synapses may have profound effects on how the brain functions. The study was published in the August 21 online issue of the journal Neuron.

A drug that restores normal synaptic pruning can improve autistic-like behaviors in mice, the researchers found, even when the drug is given after the behaviors have appeared.

“This is an important finding that could lead to a novel and much-needed therapeutic strategy for autism,” said Jeffrey Lieberman, MD, Lawrence C. Kolb Professor and Chair of Psychiatry at CUMC and director of New York State Psychiatric Institute, who was not involved in the study.

Although the drug, rapamycin, has side effects that may preclude its use in people with autism, “the fact that we can see changes in behavior suggests that autism may still be treatable after a child is diagnosed, if we can find a better drug,” said the study’s senior investigator, David Sulzer, PhD, professor of neurobiology in the Departments of Psychiatry, Neurology, and Pharmacology at CUMC.

During normal brain development, a burst of synapse formation occurs in infancy, particularly in the cortex, a region involved in autistic behaviors; pruning eliminates about half of these cortical synapses by late adolescence. Synapses are known to be affected by many genes linked to autism, and some researchers have hypothesized that people with autism may have more synapses.

To test this hypothesis, co-author Guomei Tang, PhD, assistant professor of neurology at CUMC, examined brains from children with autism who had died from other causes. Thirteen brains came from children ages two to 9, and thirteen brains came from children ages 13 to 20. Twenty-two brains from children without autism were also examined for comparison.

Dr. Tang measured synapse density in a small section of tissue in each brain by counting the number of tiny spines that branch from these cortical neurons; each spine connects with another neuron via a synapse.

By late childhood, she found, spine density had dropped by about half in the control brains, but by only 16 percent in the brains from autism patients.

“It’s the first time that anyone has looked for, and seen, a lack of pruning during development of children with autism,” Dr. Sulzer said, “although lower numbers of synapses in some brain areas have been detected in brains from older patients and in mice with autistic-like behaviors.”

Clues to what caused the pruning defect were also found in the patients’ brains; the autistic children’s brain cells were filled with old and damaged parts and were very deficient in a degradation pathway known as “autophagy.” Cells use autophagy (a term from the Greek for self-eating) to degrade their own components.

Using mouse models of autism, the researchers traced the pruning defect to a protein called mTOR. When mTOR is overactive, they found, brain cells lose much of their “self-eating” ability. And without this ability, the brains of the mice were pruned poorly and contained excess synapses. “While people usually think of learning as requiring formation of new synapses, “Dr. Sulzer says, “the removal of inappropriate synapses may be just as important.”

The researchers could restore normal autophagy and synaptic pruning—and reverse autistic-like behaviors in the mice—by administering rapamycin, a drug that inhibits mTOR. The drug was effective even when administered to the mice after they developed the behaviors, suggesting that such an approach may be used to treat patients even after the disorder has been diagnosed.

Because large amounts of overactive mTOR were also found in almost all of the brains of the autism patients, the same processes may occur in children with autism.

“What’s remarkable about the findings,” said Dr. Sulzer, “is that hundreds of genes have been linked to autism, but almost all of our human subjects had overactive mTOR and decreased autophagy, and all appear to have a lack of normal synaptic pruning. This says that many, perhaps the majority, of genes may converge onto this mTOR/autophagy pathway, the same way that many tributaries all lead into the Mississippi River. Overactive mTOR and reduced autophagy, by blocking normal synaptic pruning that may underlie learning appropriate behavior, may be a unifying feature of autism.”

Alan Packer, PhD, senior scientist at the Simons Foundation, which funded the research, said the study is an important step forward in understanding what’s happening in the brains of people with autism.

“The current view is that autism is heterogeneous, with potentially hundreds of genes that can contribute. That’s a very wide spectrum, so the goal now is to understand how those hundreds of genes cluster together into a smaller number of pathways; that will give us better clues to potential treatments,” he said.

“The mTOR pathway certainly looks like one of these pathways. It is possible that screening for mTOR and autophagic activity will provide a means to diagnose some features of autism, and normalizing these pathways might help to treat synaptic dysfunction and treat the disease.”

(Image Caption: Human neurons differentiated from induced pluripotent stem cells, with cell nuclei shown in blue and synapses in red and green. Credit: Credit: Zhexing Wen/Johns Hopkins Medicine)

Stem Cells Reveal How Illness-Linked Genetic Variation Affects Neurons

A genetic variation linked to schizophrenia, bipolar disorder and severe depression wreaks havoc on connections among neurons in the developing brain, a team of researchers reports. The study, led by Guo-li Ming, M.D., Ph.D., and Hongjun Song, Ph.D., of the Johns Hopkins University School of Medicine and described online Aug. 17 in the journal Nature, used stem cells generated from people with and without mental illness to observe the effects of a rare and pernicious genetic variation on young brain cells. The results add to evidence that several major mental illnesses have common roots in faulty “wiring” during early brain development.

“This was the next best thing to going back in time to see what happened while a person was in the womb to later cause mental illness,” says Ming. “We found the most convincing evidence yet that the answer lies in the synapses that connect brain cells to one another.”

Previous evidence for the relationship came from autopsies and from studies suggesting that some genetic variants that affect synapses also increase the chance of mental illness. But those studies could not show a direct cause-and-effect relationship, Ming says.

One difficulty in studying the genetics of common mental illnesses is that they are generally caused by environmental factors in combination with multiple gene variants, any one of which usually could not by itself cause disease. A rare exception is the gene known as disrupted in schizophrenia 1 (DISC1), in which some mutations have a strong effect. Two families have been found in which many members with the DISC1 mutations have mental illness.

To find out how a DISC1 variation with a few deleted DNA “letters” affects the developing brain, the research team collected skin cells from a mother and daughter in one of these families who have neither the variation nor mental illness, as well as the father, who has the variation and severe depression, and another daughter, who carries the variation and has schizophrenia. For comparison, they also collected samples from an unrelated healthy person. Postdoctoral fellow Zhexing Wen, Ph.D., coaxed the skin cells to form five lines of stem cells and to mature into very pure populations of synapse-forming neurons.

After growing the neurons in a dish for six weeks, collaborators at Pennsylvania State University measured their electrical activity and found that neurons with the DISC1 variation had about half the number of synapses as those without the variation. To make sure that the differences were really due to the DISC1 variation and not to other genetic differences, graduate student Ha Nam Nguyen spent two years making targeted genetic changes to three of the stem cell lines.

In one of the cell lines with the variation, he swapped out the DISC1 gene for a healthy version. He also inserted the disease-causing variation into one healthy cell line from a family member, as well as the cell line from the unrelated control. Sure enough, the researchers report, the cells without the variation now grew the normal amount of synapses, while those with the inserted mutation had half as many.

“We had our definitive answer to whether this DISC1 variation is responsible for the reduced synapse growth,” Ming says.

To find out how DISC1 acts on synapses, the researchers also compared the activity levels of genes in the healthy neurons to those with the variation. To their surprise, the activities of more than 100 genes were different. “This is the first indication that DISC1 regulates the activity of a large number of genes, many of which are related to synapses,” Ming says.

The research team is now looking more closely at other genes that are linked to mental disorders. By better understanding the roots of mental illness, they hope to eventually develop better treatments for it, Ming says.