Powerful Function of Single Protein That Controls Neurotransmission Discovered

Scientists at Weill Cornell Medical College have discovered that the single protein -- alpha 2 delta -- exerts a spigot-like function, controlling the volume of neurotransmitters and other chemicals that flow between the synapses of brain neurons. The study, published online in Nature, shows how brain cells talk to each other through these signals, relaying thoughts, feelings and action, and this powerful molecule plays a crucial role in regulating effective communication.

In the study, the investigators also suggest how the widely used pain drug Lyrica might work. The alpha 2 delta protein is the target of this drug and the new work suggests an approach to how other drugs could be developed that effectively twist particular neurotransmitter spigots on and off to treat neurological disorders. The research findings surprised the research team, which includes scientists from University College London.

"We are amazed that any single protein has such power," says the study's lead investigator Dr. Timothy A. Ryan, professor of Biochemistry and associate professor of Biochemistry in Anesthesiology at Weill Cornell Medical College. "It is indeed rare to identify a biological molecule's function that is so potent, that seems to be controlling the effectiveness of neurotransmission."

The researchers found that alpha 2 delta determines how many calcium channels will be present at the synaptic junction between neurons. The transmission of chemical signals is triggered at the synapse by the entry of calcium into these channels, so the volume and speed of neurotransmission depends on the availability of these channels.

Researchers discovered that taking away alpha 2 delta from brain cells prevented calcium channels from getting to the synapse. "But if you add more alpha 2 delta, you can triple the number of channels at synapses," Dr. Ryan says. "This change in abundance was tightly linked to how well synapses carry out their function, which is to release neurotransmitters."

Before this study, it was known that Lyrica, which is used for neuropathic pain, seizures and fibromyalgia, binds to alpha 2 delta, but little was understood about how this protein works to control synapses.

Lifting up the Hood

Dr. Ryan is building what he calls a "shop manual" of neurological function, much of which centers on synaptic neurotransmission. In 2007 and 2008, he discovered crucial clues to how neurons repackage the chemicals used to signal across synapses. In 2011, Dr. Ryan discovered that distinct neurons differently tune the speed by which they package these chemicals. And in a recent study published April 29 in Nature Neuroscience, he described, for the first time, the molecular mechanisms at the synapse that control the release of dopamine, a crucial neurotransmitter.

"We are looking under the hood of these machines for the first time," he says. "Many neurological diseases are considered to arise from pathologies of synaptic function. The synapse is so complex; at least a few thousand genes control how they work. Repairing them through treatment requires that we understand how they work."

Dr. Ryan and his team often use two tools to conduct these studies -- they pin fluorescent tags on to molecules involved in synaptic function, and use ultra sensitive microscopy technology to watch these molecules up close and in real-time.

The researchers used the same toolkit to examine the function of calcium channels, which triggers neurotransmission. "At all synapses, the secretion of a neurotransmitter is driven by the arrival of an electric impulse, initiated by another neuron," Dr. Ryan says. When this impulse arrives at the nerve terminal it triggers the opening of calcium channels. The calcium that rushes in is the key trigger that drives a synapse to secrete its neurotransmitter.

"We have known for the past half century that calcium is a key controller of neurotransmission," he says. "Any small change in calcium influx has a big impact on neurotransmission."

Protein Acts like a Shipping Label

But the number of calcium channels at the synapse is not static. Neurons constantly replace worn out channels, and to do this, they build the channels in the neuron's cell body and then package them up and ship them to the nerve terminal. In some cases, that is a very long journey -- as much as a few feet, such as the distance between the brain and the base of the spinal cord or the length of a leg.

In the study, researchers tagged fluorescent proteins onto a gene that encodes protein that makes a calcium channel and delivered it to neurons. They then watched the progress of the newly formed channels as they made their way, from day four to day seven, from the bodies of neurons to the synapse.

They also manipulated the levels of alpha 2 delta, a suspected calcium channel partner, and discovered that when the protein was increased, more calcium channels were moved to the synapse. Less alpha 2 delta reduced the flow. "We discovered that alpha 2 delta made the decision of how many calcium channels should be shipped the length of the neuron to the synapse," Dr. Ryan says. "It's like the channels couldn't be transported without an alpha 2 delta shipping label."

The research team found however that alpha 2 delta must work in at least two steps. When they impaired a piece of alpha 2 delta that resembles proteins that are involved in how cells bind to each other, they found that this broken alpha 2 delta could still help get calcium channels shipped down to synapses. But once there, they no longer helped drive neurotransmitter release. "This means that not only does alpha 2 delta help to get calcium channels shipped out, but it also implies that something at the synapse has to sign-off on receiving the calcium channels, putting them in the right place for them to do their job," Dr. Ryan says.

The researchers suggest that Lyrica might work by interfering with this final step since the piece of alpha 2 delta they "broke" that prevents the signing-off resembles parts of proteins that allows them to stick to each other in a kind of handshake.

These findings suggest that future therapies designed to manipulate neurotransmission could try to target this handshaking process, Dr. Ryan says. To do this will require that researchers identify the missing partner in the handshake.

"We hope these exciting findings are providing a new direction in how to make better drugs to control communication between brain cells," Dr. Ryan says.

The study was funded by the National Institutes of Mental Health and the Welcome Trust. Co-authors of the study include Dr. Michael B. Hoppa from Weill Cornell Medical College, and Dr. Beatrice Lana, Dr. Wojciech Margas, and Dr. Annette C. Dolphin from University College London.

Source: NewYork-Presbyterian Hospital/Weill Cornell Medical Center/Weill Cornell Medical College [May 13, 2012]

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Genes for learning, remembering and forgetting

Certain genes and proteins that promote growth and development of embryos also play a surprising role in sending chemical signals that help adults learn, remember, forget and perhaps become addicted, University of Utah biologists have discovered. 

"We found that these molecules and signaling pathways [named Wnt] do not retire after development of the organism, but have a new and surprising role in the adult. They are called back to action to change the properties of the nervous system in response to experience," says biology Professor Andres Villu Maricq, senior author of the new study in the March 30 issue of the journal Cell. 

The study was performed in C. elegans -- the millimeter-long roundworm or nematode -- which has a nervous system that serves as a model for those of vertebrate animals, including humans. 

Because other Wnt pathways in worms are known to work in humans too, the researchers believe that Wnt genes, the Wnt proteins they produce and so-called "Wnt signaling" also are involved in human learning, memory and forgetting. 

"Almost certainly what we have discovered is going on in our brain as well," Maricq says. And because a worm nerve-signal "receptor" in the study is analogous to a human nicotine receptor involved in addiction, schizophrenia and some other mental disorders, some of the genes identified in the worm study "represent possible new targets for treatment of schizophrenia and perhaps addiction," he adds. 

Wnt genes and their proteins already were known to "pattern the development and distribution of organs in the body" during embryo development, and to be responsible for various cancers and developmental defects when mutated, he says. 

Maricq conducted the study with these Utah biologists: doctoral students Michael Jensen and Dane Maxfield; postdoctoral researchers Michael M. Francis, Frederic Hoerndli and Rui Wang; undergraduate Erica Johnson; Penelope Brockie, a research associate professor; and David M. Madsen, a senior research specialist. 

Synapse Plasticity is the Basis of Learning and Memory 

Synapses are the connections between nerve cells (neurons). Nerve signals are transmitted through synapses. Learning and memory concern how these connections are made, broken, strengthened or weakened. Proteins known as receptors are delivered to the synapses or removed from them to strengthen or weaken the connection. 

In the new study, Maricq and colleagues identified a "Wnt signaling pathway" -- a series of genes and the proteins they produce -- that controls the strength of nerve signal transmission from one neuron through a synapse to the next neuron. This allows "plasticity" of synapses -- a key factor in learning, retaining memories and forgetting. 

"The adult nervous system is not a stagnant tissue, but rather dynamic and plastic, with the strength of synapses -- specialized neuron-to-neuron connections -- changing with experience, learning and memory," Maricq says. "It's not a fixed thing, like when you're done making the heart, you're done." 

When synapses and thus incoming nerve signals are strengthened by adding receptors, an organism learns and remembers; when the opposite occurs, the organism forgets, he adds. 

How is that connection strengthened or weakened? When one neuron sends a nerve signal to another neuron, the first neuron releases a chemical known as a neurotransmitter, which moves through the synapse connecting the two cells and attaches or binds to receptors on the surface of the second neuron. 

"You can think of the receptors like amplifiers, like hearing aids," Maricq says. 

The volume of the received nerve signal depends on the number of receptors, which are stored in a supply depot just below the nerve cell's surface. 

The Wnt signaling identified in the new study "tells the depot to put more receptors into the synapse -- or not," says Maricq. 

He emphasizes that the Wnt chemical signal is different than the actual nerve signal carried by a neurotransmitter chemical, which in the new study was acetylcholine. The Wnt signal "is a secondary signal that controls the volume of the neurotransmitter signal," Maricq says. 

Worms Reveal Details of Nerve Signal Volume Control 

By crippling various genes in the worms, the researchers identified the "signaling pathway" by which a Wnt protein in one nerve cell sends a chemical signal to another cell telling it to increase the number of receptors on its surface, thus increasing the strength or volume of nerve signals between the cells. 

This is a microscope image of the roundworm or nematode C. elegans with its nervous system glowing green due to labeling with a green jellyfish protein [Credit: Penelope Brockie, University of Utah]

The type of nerve-signal receptor in the study is an acetylcholine receptor named ACR-16. When researchers crippled the gene that makes the ACR-16 receptor protein, there were not enough receptors, so nerve signals were disrupted and the worms "had uncoordinated movement," Maricq says. "They were semi-paralyzed." 

The scientists found mutations of other genes that also resulted in inadequate ACR-16 receptors and impaired the worms' movement. They discovered such genes belong to the "Wnt signaling pathway" that puts enough receptors on the cell surface so signals can be received. 

Besides ACR-16, genes in that pathway produced proteins named CWN-2 -- which is a Wnt protein -- LIN-17, CAM-1 and DSH-1. 

Here is how that pathway controls the volume of incoming nerve signals: 

1. A neuron releases CWN-2, which binds onto a receptor protein on the signal-receiving cell. That protein is a newly discovered combination of the LIN-17 and CAM-1 proteins. 

2. The LIN-17/CAM-1 protein sends a signal to a protein called disheveled, or DSH-1. 

3. "DSH-1 somehow sends the volume-control signal" that dispatches more ACR-16 receptors from depots inside the second neuron to that cell's surface, thus boosting the volume of the received nerve signal, Maricq says. 

The researchers used a green jellyfish protein to mark the ACR-16 receptors so they were visible under a microscope. When any of the genes in the Wnt signaling pathway were mutant, the scientists could see the green-labeled receptors accumulate under the surfaces of nerve cells instead of moving to the surface. 

Another experiment recorded electrical currents in worm nerve synapses and found it was smaller when any of the Wnt pathway genes were mutated. The smaller current -- reflecting impaired nerve-signal transmission -- explains why the mutant worms were partially paralyzed. 

Human Version of Worm Receptor Tied to Mental Disorders 

The ACR-16 acetylcholine receptor is the worm version of the alpha-7 nicotinic acetylcholine receptor in humans and other vertebrates. Both are similar in structure and function in animals from worms to fruit flies, mice and people. 

The alpha-7 receptor "is important in schizophrenia and a number of different mental disorders, and may have a role in addiction, but we don't understand how it's regulated," Maricq says. 

Many existing psychiatric drugs modify synapse strength. The new study suggests research should be done to show if the same Wnt signaling genes in worms also control alpha-7 receptor levels on human brain cells. If so, new drugs might be developed to target those genes as a way to treat mental disorders, including addiction. 

"Addiction is like learning at a primitive level," Maricq says. "Addiction means that somewhere in your brain, synapses are too strong. So you want more." 

The study was funded by the National Institutes of Health and the American Heart Association. 

Source: University of Utah [March 29, 2012]

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How pregnancy changes a woman's brain

We know a lot about the links between a pregnant mother’s health, behavior, and moods and her baby’s cognitive and psychological development once it is born. But how does pregnancy change a mother’s brain? “Pregnancy is a critical period for central nervous system development in mothers,” says psychologist Laura M. Glynn of Chapman University. 

“Yet we know virtually nothing about it.” Glynn and her colleague Curt A. Sandman, of University of the California Irvine, are doing something about that. Their review of the literature in Current Directions in Psychological Science, a journal published by the Association for Psychological Science, discusses the theories and findings that are starting to fill what Glynn calls “a significant gap in our understanding of this critical stage of most women’s lives.” 

At no other time in a woman’s life does she experience such massive hormonal fluctuations as during pregnancy. Research suggests that the reproductive hormones may ready a woman’s brain for the demands of motherhood—helping her becomes less rattled by stress and more attuned to her baby’s needs. Although the hypothesis remains untested, Glynn surmises this might be why moms wake up when the baby stirs while dads snore on. Other studies confirm the truth in a common complaint of pregnant women: “Mommy Brain,” or impaired memory before and after birth. “There may be a cost” of these reproduction-related cognitive and emotional changes, says Glynn, “but the benefit is a more sensitive, effective mother.” 

The article reviews research that refines earlier findings on the effects of the prenatal environment on the baby. For instance, evidence is accumulating to show that it’s not prenatal adversity on its own—say, maternal malnourishment or depression—that presents risks for a baby. Congruity between life in utero and life on the outside may matter more. A fetus whose mother is malnourished adapts to scarcity and will cope better with a dearth of food once it’s born—but could become obese if it eats normally. Timing is critical too: maternal anxiety early in gestation takes a toll on the baby’s cognitive development; the same high levels of stress hormones late in pregnancy enhance it. 

Just as Mom permanently affects her fetus, new science suggests that the fetus does the same for Mom. Fetal movement, even when the mother is unaware of it, raises her heart rate and her skin conductivity, signals of emotion—and perhaps of pre-natal preparation for mother-child bonding. Fetal cells pass through the placenta into the mother’s bloodstream. “It’s exciting to think about whether those cells are attracted to certain regions in the brain” that may be involved in optimizing maternal behavior, says Glynn. 

Glynn cautions that most research on the maternal brain has been conducted with rodents, whose pregnancies differ enormously from women’s; more research on human mothers is needed. But she is optimistic that a more comprehensive picture of the persisting brain changes wrought by pregnancy will yield interventions to help at-risk mothers do better by their babies and themselves. 

Source: Association for Psychological Science [December 20, 2011]

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New Insights Into How the Nervous System Becomes Wired During Early Development

Thanks to a new study of the retina, scientists at UC Santa Barbara have developed a greater understanding of how the nervous system becomes wired during early development. 

Confocal micrograph of a single cone bipolar cell (green) that extends its dendrites to receive synaptic inputs (red dots) from a local population of cone photoreceptor terminals (violet) [Credit: Sammy Lee]

The findings reflect the expansion of developmental neurobiology and vision research at UCSB. The work is described in a recent publication of The Journal of Neuroscience. 

The research team examined the connectivity of nerve cells, called neurons, in mice. Neurons communicate with one another via synapses where the dendrites and axon terminals of different cells form contacts. This is where nerve signals are transmitted from one neuron to another. 

Scientists have understood for some time how neuronal activation at developing synapses contributes to the patterns of connectivity observed in maturity, explained Ben Reese, senior author and professor in UCSB's Neuroscience Research Institute and the Department of Psychological & Brain Sciences. 

Incoming activity plays a critical role in sculpting neuronal form and the elaboration of synaptic connections. The new research shows, by contrast, how relationships between neighboring cells of the same type independently regulate neuronal size and connectivity. 

The researchers circumvented the difficulty of visualizing the three-dimensional relationships between neurons within the brain by working within the retina. The retina is an outgrowth of the brain during embryonic development, and is a precisely layered structure in which the cells, their dendrites and their axons are restricted to discrete strata. "This makes the visualization and analysis of neuronal morphology and connectivity far simpler," said Reese. 

The scientists used two genetically modified mouse models to modulate the density of one particular type of retinal neuron, a class of cone bipolar cell. Cone bipolar cells relay information from the population of cone photoreceptors to the retinal ganglion cells. The latter are neurons that in turn project information to locations within the brain where further visual processing of the retinal image takes place. 

The lead author on the study, Sammy Lee, was a postdoctoral researcher working in Reese's lab and supported by a C.J. Martin National Health & Medical Research Council fellowship from Australia during the course of the study. Lee labeled individual cone bipolar cells with a fluorescent dye through a new microinjection procedure developed by Patrick Keeley, a graduate student in the Reese lab. 

"What Dr. Lee has shown is that cone bipolar cells modulate the size of their dendritic fields (branched extensions of the neuron) in association with the local density of like-type neurons," said Reese. "One line of mice has conspicuously fewer cone bipolar cells, each now with a larger dendritic territory, while the other line shows heightened densities and correspondingly smaller dendritic fields." 

Other studies have suggested such homotypic (like-type) modulation of dendritic field size, but the current study directly shows this modulation following genetic manipulation of neuronal density, according to Reese. 

Additionally, the researchers found that connectivity with the afferent population of cone photoreceptors is impacted directly, with the larger dendritic fields being innervated by more cones, and the smaller dendritic fields connecting with fewer cones. At any individual cone, the number of dendritic endings associating with that cone was not observed to change, so that the total number of connections made by a cone bipolar cell was remarkably plastic, defined solely by the number of cone contacts formed. 

"This developmental plasticity in dendritic growth and synapse number may be well-suited to ensure uniform coverage and connectivity between two populations of neurons -- afferents and their targets -- when the number of cells in each population is specified independently," said Reese. 

Other studies from Reese's lab, recently reported in The Proceedings of the National Academy of Sciences and Investigative Ophthalmology and Visual Science, showed how neuronal number is tightly specified genetically, yet is highly variable between different strains of mice. "Wiring together two populations, each of which may vary nearly two-fold in size, yet independent of each other, might best be served by such homotypic plasticity during early development," he said. 

Studies like these may prove relevant for re-establishing connectivity following nerve cell re-specification or replacement in degenerative diseases, particularly as advances in stem cell biology make this an increasing possibility, said Reese. 

Source: University of California - Santa Barbara [December 05, 2011]

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