New Insight Into How Brain ‘Learns’ Cocaine Addiction

A team of researchers says it has solved the longstanding puzzle of why a key protein linked to learning is also needed to become addicted to cocaine. Results of the study, published in the Aug. 1 issue of the journal Cell, describe how the learning-related protein works with other proteins to forge new pathways in the brain in response to a drug-induced rush of the “pleasure” molecule dopamine. By adding important detail to the process of addiction, the researchers, led by a group at Johns Hopkins, say the work may point the way to new treatments.

“The broad question was why and how cocaine strengthened certain circuits in the brain long term, effectively re-wiring the brain for addiction,” says Paul Worley, M.D., a professor in the Solomon H. Snyder Department of Neuroscience at the Johns Hopkins University School of Medicine. “What we found in this study was how two very different types of systems in the brain work together to make that happen.” Cocaine addiction, experts say, is among the strongest of addictions.

Worley did not come to the problem as an addiction researcher, but as an expert in a group of genes known as immediate early genes, which rapidly ramp up production in neurons when the brain is exposed to new information. In 2001, he said, a European group led by François Conquet of GlaxoSmithKline reported that deleting mGluR5, a protein complex that responds to the common brain-signaling molecule glutamate, made mice unresponsive to cocaine. “That finding came out of the blue,” says Worley, who knew mGluR proteins for their interactions with immediate early genes. “I never would have thought this type of protein was linked to dopamine and addiction, because the functions for it that we knew about up to that point were completely unrelated. That’s what scientists love: when you’re pretty sure something is right, but you don’t have a clue why.”

The finding set Worley’s research group on a long search for an explanation. Eventually, in addition to studying the effects of altering genes for the relevant proteins in mice, they partnered with experts in measuring the brain’s electrical signals and in a biophysical technique that detects when chemical bonds are rotated within protein molecules. Using different types of experiments, they pieced together a complex story of how dopamine released in response to cocaine works together with mGluR5 and immediate early genes to switch cells into synapse-strengthening mode.

“The process we identified explains how cocaine exposure can co-opt normal mechanisms of learning to induce addiction,” Worley says. Knowing the details of the mechanism may help researchers identify targets for potential drugs to treat addiction, he adds.

(Image: Milos Jokic)

Researchers discover how brain cells change their tune

Brain cells talk to each other in a variety of tones. Sometimes they speak loudly but other times struggle to be heard. For many years scientists have asked why and how brain cells change tones so frequently. Today National Institutes of Health researchers showed that brief bursts of chemical energy coming from rapidly moving power plants, called mitochondria, may tune brain cell communication.

"We are very excited about the findings," said Zu-Hang Sheng, Ph.D., a senior principal investigator and the chief of the Synaptic Functions Section at the NIH’s National Institute of Neurological Disorders and Stroke (NINDS). "We may have answered a long-standing, fundamental question about how brain cells communicate with each other in a variety of voice tones."

The network of nerve cells throughout the body typically controls thoughts, movements and senses by sending thousands of neurotransmitters, or brain chemicals, at communication points made between the cells called synapses. Neurotransmitters are sent from tiny protrusions found on nerve cells, called presynaptic boutons. Boutons are aligned, like beads on a string, on long, thin structures called axons. They help control the strength of the signals sent by regulating the amount and manner that nerve cells release transmitters.

Mitochondria are known as the cell’s power plant because they use oxygen to convert many of the chemicals cells use as food into adenosine triphosphate (ATP), the main energy that powers cells. This energy is essential for nerve cell survival and communication. Previous studies showed that mitochondria can rapidly move along axons, dancing from one bouton to another.

In this study, published in Cell Reports, Dr. Sheng and his colleagues show that these moving power plants may control the strength of the signals sent from boutons.

"This is the first demonstration that links the movement of mitochondria along axons to a wide variety of nerve cell signals sent during synaptic transmission," said Dr. Sheng.

The researchers used advanced microscopic techniques to watch mitochondria move among boutons while they released neurotransmitters. They found that boutons sent consistent signals when mitochondria were nearby.

"It’s as if the presence of mitochondria causes a bouton to talk in a monotone voice," said Tao Sun, Ph.D., a researcher in Dr. Sheng’s laboratory and the first author of the study.

Surprisingly, when the mitochondria were missing or moving away from boutons, the signal strength fluctuated. The results suggested that the presence of stationary power plants at synapses controls the stability of the nerve signal strength.

To test this idea further, the researchers manipulated mitochondrial movement in axons by changing levels of syntaphilin, a protein that helps anchor mitochondria to the nerve cell’s skeleton found inside axons. Removal of syntaphilin resulted in faster moving mitochondria and electrical recordings from these neurons showed that the signals they sent fluctuated greatly. Conversely, elevating syntaphilin levels in nerve cells arrested mitochondrial movement and resulted in boutons that spoke in monotones by sending signals with the same strength.

"It’s known that about one third of all mitochondria in axons move. Our results show that brain cell communication is tightly controlled by highly dynamic events occurring at numerous tiny cell-to-cell connection points," said Dr. Sheng.

In separate experiments the researchers watched ATP energy levels in these tiny boutons as they sent nerve messages.

"The levels fluctuated more in boutons that did not have mitochondria nearby," said Dr. Sun.

The researchers also found that blocking ATP production in mitochondria with the drug oligomycin reduced the size of the signals boutons sent even if a mitochondrial power plant was nearby.

"Our results suggest that local ATP production by nearby mitochondria is critical for consistent neurotransmitter release," said Dr. Sheng. "It appears that variability in synaptic transmission is controlled by rapidly moving mitochondria which provide brief bursts of energy to the boutons they pass through."

Problems with mitochondrial energy production and movement throughout nerve cells have been implicated in Alzheimer’s disease, Parkinson’s disease, amyotrophic lateral sclerosis, and other major neurodegenerative disorders. Dr. Sheng thinks these results will ultimately help scientists understand how these problems can lead to disorders in brain cell communication.

"Our findings reveal the cellular mechanisms that tune brain communication by regulating mitochondrial mobility, thus advancing our understanding of human neurological disorders," said Dr. Sheng.

What do memories look like?

Oscar Wilde called memory “the diary that we all carry about with us.” Now a team of scientists has developed a way to see where and how that diary is written.

Led by Don Arnold and Richard Roberts of USC, the team engineered microscopic probes that light up synapses in a living neuron in real time by attaching fluorescent markers onto synaptic proteins — all without affecting the neuron’s ability to function.

The fluorescent markers allow scientists to see live excitatory and inhibitory synapses for the first time and, importantly, how they change as new memories are formed.

The synapses appear as bright spots along dendrites (the branches of a neuron that transmit electrochemical signals). As the brain processes new information, those bright spots change, visually indicating how synaptic structures in the brain have been altered by the new data.

“When you make a memory or learn something, there’s a physical change in the brain. It turns out that the thing that gets changed is the distribution of synaptic connections,” said Arnold, associate professor of molecular and computational biology at the USC Dornsife College of Letters, Arts and Sciences, and co-corresponding author of an article about the research that appears in Neuron on June 19.

The probes behave like antibodies, but they bind more tightly and are optimized to work inside the cell — something that ordinary antibodies can’t do. To make these probes, the team used a technique known as “mRNA display,” which was developed by Roberts and Nobel laureate Jack Szostak.

“Using mRNA display, we can search through more than a trillion different potential proteins simultaneously to find the one protein that binds the target the best,” said Roberts, co-corresponding author of the article and professor of chemistry and chemical engineering with joint appointments at USC Dornsife and the USC Viterbi School of Engineering.

Arnold and Roberts’ probes (called “FingRs”) are attached to green fluorescent protein (GFP), a protein isolated from jellyfish that fluoresces bright green when exposed to blue light. Because FingRs are proteins, the genes encoding them can be put into brain cells in living animals, causing the cells themselves to manufacture the probes.

The design of FingRs also includes a regulation system that cuts off the amount of FingR-GFP that is generated after 100 percent of the target protein is labeled, effectively eliminating background fluorescence — generating a sharper, clearer picture.

These probes can be put in the brains of living mice and then imaged through cranial windows using two-photon microscopy.

The new research could offer crucial insight for scientists responding to President Barack Obama’s Brain Research Through Advancing Innovative Neurotechnologies (BRAIN) Initiative, which was announced in April.

Modeled after the Human Genome Project, the objective of the $100 million initiative is to fast-track research that maps out exactly how the brain works and “better understand how we think, learn and remember,” according to the BRAIN Initiative website.

A turbocharger for nerve cells

Locating a car that’s blowing its horn in heavy traffic, channel-hopping between football and a thriller on TV without losing the plot, and not forgetting the start of a sentence by the time we have read to the end – we consider all of these to be normal everyday functions. They enable us to react to fast-changing circumstances and to carry out even complex activities correctly. For this to work, the neuron circuits in our brain have to be very flexible. Scientists working under the leadership of neurobiologists Nils Brose and Erwin Neher at the Max Planck Institutes of Experimental Medicine and Biophysical Chemistry in Göttingen have now discovered an important molecular mechanism that turns neurons into true masters of adaptation.

Neurons communicate with each other by means of specialised cell-to-cell contacts called synapses. First, an emitting neuron is excited and discharges chemical messengers known as neurotransmitters. These signal molecules then reach the receiving cell and influence its activation state. The transmitter discharge process is highly complex and strongly regulated. Its protagonists are synaptic vesicles, small blisters surrounded by a membrane, which are loaded with neurotransmitters and release them by fusing with the cell membrane. In order to be able to respond to stimulation at any time by releasing transmitters, a neuron must have a certain amount of vesicles ready to go at each of its synapses. Brose has been studying the molecular foundations of this stockpiling for years.

The problem is not merely academic. “The number of immediately releasable vesicles at a synapse determines its reliability,” explains Brose. “If there are too few and they are replenished too slowly, the corresponding synapse becomes tired very quickly in conditions of repeated activation. The opposite applies when a synapse can quickly top up its immediately available vesicles under pressure. In fact, such a synapse may even improve with constant activation.”

This synaptic adaptability can be observed in practically all neurons. It is known as short-term plasticity and is indispensable for a large number of extremely important brain processes. Without it, we would not be able to localise sounds, mental maths would be impossible, and the speed and flexibility with which we can alter our behaviour and turn our attention to new goals would be lost.

Some years ago, Brose and his team discovered a protein with the cryptic name of Munc13. Not only is this protein indispensable for the replenishment of vesicles for immediate release at synapses; neuron activity regulates it in such a way that the fresh supply of vesicles can be adjusted in line with demand. This regulation occurs by means of a complex consisting of the signal protein calmodulin and calcium ions that build up in the synapses during intense neuron activity.

“Our earlier work on individual neurons in culture dishes showed that the calcium-calmodulin complex activates Munc13 and consequently ensures that immediately releasable vesicles are replenished faster,” says Noa Lipstein, an Israeli guest scientist in Brose’s lab. “But many colleagues were not convinced that this process also played a role in neurons in the intact brain.”

So Lipstein and her Japanese colleague Takeshi Sakaba created a mutant mouse with genetically altered Munc13 proteins that could not be activated by calcium-calmodulin complexes. The two neurophysiologists first studied the effects of this genetic manipulation on synapses involved in the localisation of sound, which are typically activated several hundred times every second. “Our study shows that the sustained efficiency of synapses in intact neuron networks is critically dependent on the activation of Munc13 by calcium-calmodulin complexes,” explains Lipstein.

The Göttingen-based scientists are convinced of the significance of their study. After all, leading neuroscientists of the past described the calcium sensor responsible for synaptic short-term plasticity and its target protein as the Holy Grail. “I am confident that we have discovered a key molecular mechanism of short-term plasticity that plays a role in all synapses in the brain, and not only in cultivated neurons, as many colleagues believed,” affirms Lipstein. And if she is, in fact, proved right about the interpretation of her findings, Munc13 could even be an ideal pharmacological target for drugs that influence brain function.

Scientists Map Process by Which Brain Cells Form Long-Term Memories

Scientists at the Gladstone Institutes have deciphered how a protein called Arc regulates the activity of neurons – providing much-needed clues into the brain’s ability to form long-lasting memories.

These findings, reported Sunday in Nature Neuroscience, also offer newfound understanding as to what goes on at the molecular level when this process becomes disrupted.

Led by Gladstone senior investigator Steve Finkbeiner, MD, PhD, this research delved deep into the inner workings of synapses. Synapses are the highly specialized junctions that process and transmit information between neurons. Most of the synapses our brain will ever have are formed during early brain development, but throughout our lifetimes these synapses can be made, broken and strengthened. Synapses that are more active become stronger, a process that is essential for forming new memories.

However, this process is also dangerous, as it can overstimulate the neurons and lead to epileptic seizures. It must therefore be kept in check.

Neuroscientists recently discovered one important mechanism that the brain uses to maintain this important balance: a process called “homeostatic scaling.” Homeostatic scaling allows individual neurons to strengthen the new synaptic connections they’ve made to form memories, while at the same time protecting the neurons from becoming overly excited. Exactly how the neurons pull this off has eluded researchers, but they suspected that the Arc protein played a key role.

“Scientists knew that Arc was involved in long-term memory, because mice lacking the Arc protein could learn new tasks, but failed to remember them the next day,” said Finkbeiner, who is also a professor of neurology and physiology at UC San Francisco, with which Gladstone is affiliated. “Because initial observations showed Arc accumulating at the synapses during learning, researchers thought that Arc’s presence at these synapses was driving the formation of long-lasting memories.”

But Finkbeiner and his team thought there was something else in play.

The Role of Arc in Homeostatic Scaling

In laboratory experiments, first in animal models and then in greater detail in the petri dish, the researchers tracked Arc’s movements. And what they found was surprising.

“When individual neurons are stimulated during learning, Arc begins to accumulate at the synapses – but what we discovered was that soon after, the majority of Arc gets shuttled into the nucleus,” said Erica Korb, PhD, the paper’s lead author who completed her graduate work at Gladstone and UCSF.

“A closer look revealed three regions within the Arc protein itself that direct its movements: one exports Arc from the nucleus, a second transports it into the nucleus, and a third keeps it there,” she said. “The presence of this complex and tightly regulated system is strong evidence that this process is biologically important.”

In fact, the team’s experiments revealed that Arc acted as a master regulator of the entire homeostatic scaling process. During memory formation, certain genes must be switched on and off at very specific times in order to generate proteins that help neurons lay down new memories.  From inside the nucleus, the authors found that it was Arc that directed this process required for homeostatic scaling to occur. This strengthened the synaptic connections without overstimulating them – thus translating learning into long-term memories. 

Implications for a Variety of Neurological Diseases

“This discovery is important not only because it solves a long-standing mystery on the role of Arc in long-term memory formation, but also gives new insight into the homeostatic scaling process itself – disruptions in which have already been implicated in a whole host of neurological diseases,” said Finkbeiner. “For example, scientists recently discovered that Arc is depleted in the hippocampus, the brain’s memory center, in Alzheimer’s disease patients. It’s possible that disruptions to the homeostatic scaling process may contribute to the learning and memory deficits seen in Alzheimer’s.”

Dysfunctions in Arc production and transport may also be a vital player in autism. For example, the genetic disorder Fragile X syndrome – a common cause of both mental retardation and autism, directly affects the production of Arc in neurons.

“In the future,” added Dr. Korb, “we hope further research into Arc’s role in human health and disease can provide even deeper insight into these and other disorders, and also lay the groundwork for new therapeutic strategies to fight them.”

(Image: Wikimedia)

Common gene known to cause inherited autism now linked to specific behaviors

The genetic malady known as Fragile X syndrome is the most common cause of inherited autism and intellectual disability. Brain scientists know the gene defect that causes the syndrome and understand the damage it does in misshaping the brain’s synapses — the connections between neurons. But how this abnormal shaping of synapses translates into abnormal behavior is unclear.

Now, researchers at UCLA believe they know. Using a mouse model of Fragile X syndrome (FXS), they recorded the activity of networks of neurons in a living mouse brain while the animal was awake and asleep. They found that during both sleep and quiet wakefulness, these neuronal networks showed too much activity, firing too often and in sync, much more than a normal brain.

This neuronal excitability, the researchers said, may be the basis for symptoms in children with FXS, which can include disrupted sleep, seizures or learning disabilities. The findings may lead to treatments that could quiet the excessive activity and allow for more normal behavior.

The study results are published in the June 2 online edition of the journal Nature Neuroscience.

According to the National Fragile X Foundation, approximately one in every 3,600 to 4,000 males has the disorder, as does one in 4,000 to 6,000 females. FXS is caused by a mutation in the gene FMR1, which encodes the fragile X mental retardation protein, or FMRP. That protein is believed to be important for the formation and regulation of synapses. Mice that lack the FMR1 gene — and therefore lack the FMRP protein — show some of the same symptoms of human FXS, including seizures, impaired sleep, abnormal social relationships and learning defects.

"We wanted to find the link between the abnormal structure of synapses in the FXS mouse and the behavioral abnormalities at the level of brain circuits. That had not been previously established," said senior author Dr. Carlos Portera-Cailliau, an associate professor in the departments of neurology and neurobiology at UCLA. " So we tested the signaling between different neurons in Fragile X mice and indeed found there was abnormally high firing of action potentials — the signals between neurons — and also abnormally high synchrony — that is, too many neurons fired together. That’s a feature that is common in early brain development, but not in the adult."

"In essence, this points to a relative immaturity of brain circuits in FXS," added Tiago Gonçalves, a former postdoctoral researcher in Portera-Cailliau’s laboratory and the first author of the study.

The researchers used two-photon calcium imaging and patch-clamp electrophysiology — two sophisticated technologies that allowed them to record the signals from individual brain cells. Abnormally high firing and network synchrony, said Portera-Cailliau, is evidence of the fact that neuronal circuits are overexcitable in FXS.

"That likely leads to aberrant brain function or impairments in the normal computations of the brain," he said. "For example, high synchrony could lead to seizures; more neurons firing together could cause entire portions of the brain to fire synchronously, which is the basis of seizures."

And epilepsy, Portera-Cailliau said, is seen in up to 20 percent of children with FXS. High firing rates could also impair the ability of the brain to decode sensory stimuli by causing an overwhelming response to even simple sensory stimuli; this could lead to autism and the withdrawal from social interactions, he noted.

"Interestingly, we found that the high firing and synchrony were especially apparent at times when the animals were asleep," said Portera-Cailliau. "This is curious because a prominent symptom of FXS is disrupted sleep and frequent awakenings."

And, he noted, since sleep is important for encoding memories and consolidating learning, this hyperexcitability of brain networks in FXS may interfere with the process of laying down new memories, and perhaps explain the learning disability in children with FXS.

"Because brain scientists know a lot about the factors that regulate neuronal excitability, including inhibitory neurons, they can now try to use a variety of strategies to dampen neuronal excitation," he said. "Hopefully, this may be helpful to treat symptoms of FXS."

The next step, said Portera-Cailliau, is to explore whether Fragile X mice indeed exhibit exaggerated responses to sensory stimuli.

"An overwhelming reaction to a slight sound or caress, or hyperarousal to sensory stimuli, could be common to different types of autism and not just FXS," he said. "If hyperexcitability is the brain-network basis for these symptoms, then reducing neuronal excitability with certain drugs that modulate inhibition could be of therapeutic value in these devastating neurodevelopmental disorders."

Neuroscientists Discover New Phase of Synaptic Development

Breakthrough Could Lead to Better Understanding of Learning and Memory

Students preparing for final exams might want to wait before pulling an all-night cram session — at least as far as their neurons are concerned. Carnegie Mellon University neuroscientists have discovered a new intermediate phase in neuronal development during which repeated exposure to a stimulus shrinks synapses. The findings are published in the May 8 issue of the Journal of Neuroscience.

It’s well known that synapses in the brain, the connections between neurons and other cells that allow for the transmission of information, grow when they’re exposed to a stimulus. New research from the lab of Carnegie Mellon Associate Professor of Biological Sciences Alison L. Barth has shown that in the short term, synapses get even stronger than previously thought, but then quickly go through a transitional phase where they weaken.

"When you think of learning, you think that it’s cumulative. We thought that synapses started small and then got bigger and bigger. This isn’t the case," said Barth, who also is a member of the joint Carnegie Mellon/University of Pittsburgh Center for the Neural Basis of Cognition. "Based on our data, it seems like synapses that have recently been strengthened are peculiarly vulnerable — more stimulation can actually wipe out the effects of learning.

"Psychologists know that for long-lasting memory, spaced training - like studying for your classes after very lecture, all semester long — is superior to cramming all night before the exam," Barth said. "This study shows why. Right after plasticity, synapses are almost fragile — more training during this labile phases is actually counterproductive."

Previous research from Barth’s lab established the biochemical mechanisms responsible for the strengthening of synapses in the neocortex, the part of the brain responsible for thought and language, but only measured the synapses after 24 hours. In the current study, post-doctoral student Jing A. Wen investigated how the synapses developed throughout the first 24 hours of exposure to a stimulus using a specialized transgenic mouse model created by Barth. The model senses its surroundings using only one whisker, which alters its ability to sense its environment and creates a sensory imbalance that increases plasticity in the brain. Since each whisker is linked to a specific area of the cortex, researchers can easily track neuronal changes.

Wen found that during this first day of learning, synapses go through three distinct phases. In the initiation phase, synaptic plasticity is spurred on by NMDA receptors. Over the next 12 hours or so, the synapses get stronger and stronger. As the stimulus is repeated, the NDMA receptors change their function and start to weaken the synapses in what the researchers have called the labile phase. After a few hours of weakening, another receptor, mGluR5, initiates a stabilization phase during which the synapses maintain their residual strength.

Furthermore, the researchers found that they could maintain the super-activated state found at the beginning of the labile phase by stopping the stimulus altogether or by injecting a glutamate receptor antagonist drug at an optimal time point. The findings are analogous to those seen in many psychological studies that use space training to improve memory.

"While synaptic changes can be long lasting, we’ve found that in this initial period there are a number of different things we could play with," Barth said. "The discovery of this labile phase suggests there are ways to control learning through the manipulation of the biochemical pathways that maintain memory."