Alcohol's effects on gene expression in the central nervous system

Alcohol's primary target is the central nervous system (CNS), where it influences neurotransmission to produce intoxication. Scientists can now use microarray technology to study brain function gene by gene. Symposium proceedings published in the February issue of Alcoholism: Clinical & Experimental Research address the effects of alcohol on what is called "gene expression" in the CNS regions of animal models.
"All of our cells have exactly the same deoxyribonucleic acid (DNA), which means they all have the same genes," explained William J. McBride, symposium organizer and professor of neurobiology at the Indiana University School of Medicine. "The reason that different cells can appear and work so differently with the same genes – giving us, for example, unique eyes, skin, or hair – is that only some genes are used or 'turned on' in each cell. This is called gene expression."
McBride said that researchers now know that alcohol can change gene expression in the brain, and that these changes are likely responsible for many of the 'symptoms' of addiction, such as tolerance, physical dependence, and craving, as well as the 'consequences' of alcoholism, such as brain damage.
"The challenge has been to find out which genes – out of more than 50,000 – are turned on or off in the brains of alcoholics," he said. "Microarray studies – the examination of a small glass microscope slide that has thousands of different DNA samples attached to it – that are applied to brain function are just beginning in the field of alcoholism. Several years ago, it was impossible to analyze more than a handful of these genes, however, microarray technology has changed that."
Symposium speakers at the June 2004 Research Society on Alcoholism meeting in Vancouver, B.C. presented the following findings from recent studies that used genetic animal models:


# Gene expression profiling in the nucleus accumbens, prefrontal cortex, and ventral tegmental areas show that distinct biological pathways are associated with alcohol's action in specific brain regions and certain mouse strains.


"We were able to use gene expression profiling to determine that alcohol produces multiple effects on different biological processes," said McBride, "and that these changes are different in several brain regions which may be involved in alcohol addiction."


# Researchers have identified individual genes and gene networks that may play an important role in determining the behavioral responses to alcohol as well as possibly influencing drinking behavior.


"Thus far, genes that appear to be responsive to alcohol include genes involved in the intracellular signaling process (which can alter how the neuron functions), neuropeptide signaling (which modulates nerve cell activity), and myelin structure (which is needed for communication between nerve cells)," said McBride. "Gene expression profiling has also been used to identify chromosomes and chromosomal regions that influence alcohol drinking and response to alcohol."


# Intracranial self-administration of ethanol into the posterior ventral tegmental area (VTA) of alcohol-preferring rats produced results suggesting that the reinforcing effects of alcohol are activating VTA dopamine neurons and producing changes in synaptic connections that resemble those that occur in memory and learning.


"Learning and memory require enhanced synaptic function between neurons," explained McBride. "Enhanced synaptic function is characterized by increased formation of synaptic proteins. The stimulation of VTA dopamine neurons by alcohol increases the expression of genes involved in the synthesis of synaptic proteins in target regions of the VTA. In short, these results suggest that alcohol can produce changes in the brain reward system that can further increase the rewarding effects of alcohol."


# Microarray techniques confirmed earlier reports indicating that chronic alcohol exposure/withdrawal differentially alters gene expression in the prefrontal cortex of mice. More than 300 genes were found to be altered by acute alcohol treatment.


"The prefrontal cortex is involved in motivated behaviors," noted McBride. "Studies with humans indicate that this brain region is sensitive to the effects of heavy alcohol drinking and repeated withdrawals. The microarray technique provides insight into cellular changes that occur over time with chronic alcohol drinking and repeated withdrawals."


Collectively speaking, added McBride, findings presented at the symposium demonstrate the quantitative and qualitative applications of microarrays to studying the genetic and biological bases of alcoholism and alcohol abuse within discrete brain regions.
"For researchers, microarray technology has the potential of studying the genetic and biological bases of alcohol's rewarding effects, sensitivity to the effects of alcohol, development of tolerance to the effects of alcohol, development of alcohol dependence, and alcohol withdrawal severity," he said. "For the average reader, knowing which genetic profiles might contribute to excessive alcohol drinking could be used to identify risk factors that contribute to alcoholism and alcohol abuse, and could aid in the development of selective treatment strategies for different subgroups of alcoholics."
McBride added that, despite recent advances, researchers need further developments in microarray technologies and bioinformatic approaches to better understand the complex neurobiological mechanisms underlying alcohol addiction. "Future research will need to determine changes in gene expression in very discrete neuronal pathways that may be involved in mediating the effects of alcohol that lead to addiction," he said. "Future studies will also require the integrative efforts of many investigators working with different animal models in order to identify the multiple genetic factors that contribute to the risk for alcoholism and alcohol abuse."

Source : Indiana University School of Medicine

Memories may skew visual perception

Taking a trip down memory lane while you are driving could land you in a roadside ditch, new research indicates. Vanderbilt University psychologists have found that our visual perception can be contaminated by memories of what we have recently seen, impairing our ability to properly understand and act on what we are currently seeing.


"This study shows that holding the memory of a visual event in our mind for a short period of time can 'contaminate' visual perception during the time that we're remembering," Randolph Blake, study co-author and Centennial Professor of Psychology, said.


"Our study represents the first conclusive evidence for such contamination, and the results strongly suggest that remembering and perceiving engage at least some of the same brain areas."


The study, led by research associate Min-Suk Kang, was recently published in the journal Psychonomic Bulletin & Review.


"There are numerous instances where we engage in visually guided activities, such as driving, while rehashing visual events in our mind's eye. Common sense tells us that this mental replay is harmless in that it does not interfere with our ability to register and react to objects within our visual field," Kang and his co-authors wrote. "Evidently, however, that is not always true when the contents of our working memories overlap with the contents of our perceptual world."


In this study, the researchers used a visual illusion called motion repulsion to learn whether information held in working memory affects perception. This illusion is produced when two sets of moving dots are superimposed, with dots in one set moving in a different direction from those in the other set. Under these conditions, people tend to misperceive the actual directions of motion, and perceive a larger difference between the two sets of motions than actually exists.


Ordinarily this illusion is produced by having people view both sets of motion at the same time. Kang and colleagues set out to determine if the illusion would occur when one set of motions, rather than being physically present, was held in working memory.


In the experiment, participants were shown a random pattern of dots and were asked to remember the direction in which the dots were moving. They were then were shown a second pattern of moving dots. They were asked to report on the direction of second dots' movement.


The research subjects' reports of the second dots' movement was exaggerated and influenced by what they had previously seen. If they were first shown dots moving in one direction and later shown dots moving in a slightly counterclockwise direction relative to the first presented dots, they reported the counterclockwise movement to be more dramatic than it had actually been.


"We find that observers misperceive the actual direction of motion of a single motion stimulus if, while viewing that stimulus, they are holding a different motion direction in visual working memory," the authors wrote.


The results provide further support for previous findings by Vanderbilt researchers Frank Tong and Stephanie Harrison that the contents of working memory may be represented in early visual areas in the brain, including the primary visual cortex, that were previously thought to play no role in higher cognitive functions such as memory.


"Our findings provide compelling evidence that visual working memory representations directly interact with the same neural mechanisms involved in processing basic sensory events," Kang and his colleagues wrote.

Source : Vanderbilt University

Stem cell study could aid motor neurone disease research

Scientists have discovered a new way to generate human motor nerve cells in a development that will help research into motor neurone disease.



A team from the Universities of Edinburgh, Cambridge and Cardiff has created a range of motor neurons – nerves cells that send messages from the brain and spine to other parts of the body – from human embryonic stem cells in the laboratory.


It is the first time that researchers have been able to generate a variety of human motor neurons, which differ in their make-up and display properties depending on where they are located in the spinal cord.


The research, published in the journal Nature Communications, could help scientists better understand motor neurone disease. The process will enable scientists to create different types of motor neurons and study why some are more vulnerable to disease than others.


Motor neurons control muscle activity such as speaking, walking, swallowing and breathing. However, in motor neurone disease – a progressive and ultimately fatal disorder – these cells break down leading to paralysis, difficulty speaking, breathing and swallowing.


Previously scientists had only been able to generate one particular kind of motor neuron, which they did by using retinoic acid, a vitamin A derivative.


In the latest study, scientists have found a way to generate a wider range of motor neurons using a new process without retinoic acid.


Professor Siddharthan Chandran, Director of the Euan MacDonald Centre for Motor Neurone Disease Research at the University of Edinburgh, said: "Motor neurons differ in their make-up, so understanding why some are more vulnerable than others to disease is important for developing treatment for this devastating condition."


Dr Rickie Patani, of the University of Cambridge, said: "Although motor neurons are often considered as a single group, they represent a diverse collection of neuronal subtypes. The ability to create a range of different motor neurons is a key step in understanding the basis of selective subtype vulnerability in conditions such as motor neuron disease and spinal muscular atrophy."

Source : University of Edinburgh

Bodyguard for the brain

Humans are getting older and older, and the number of people with dementia is increasing. The factors controlling degeneration of the brain are still mostly unknown. However, researchers assume that factors such as stress, accumulation of toxic waste products as well as inflammation accelerate aging. But, vice versa, there are also mechanisms that can - like a bodyguard - protect the brain from degenerating, or repair defective structures.


Researchers from the Universities of Bonn and Mainz have now discovered a hitherto unknown function of the cannabinoid-1 receptor (CB1). A receptor is a protein that can bind to other substances, triggering a chain of signals. Cannabinoids such as THC – the active agent in cannabis sativa – and endocannabinoids formed by the body bind to the CB1 receptors. The existence of this receptor is also the reason for the intoxicating effect of hashish and marijuana.


Not only does the CB1 receptor have an addictive potential, but it also plays a role in the degeneration of the brain. "If we switch off the receptor using gene technology, mouse brains age much faster," said Önder Albayram, principal author of the publication and a doctoral student on the team of Professor Dr. Andreas Zimmer from the Institut für Molekulare Psychiatrie at the University of Bonn. "This means that the CB1 signal system has a protective effect for nerve cells."


Mice prove their brain power in a pool


The researchers studied mice in different age categories – young six week old animals, middle-aged ones at five months, and those of an advanced age at 12 months. The animals had to master various tasks – first, they had to find a submerged platform in the pool. Once the mice knew its location, the platform was moved, and the animals had to find it again. This was how the researchers tested how well the rodents learned and remembered.


The animals in which the CB1 receptor had been switched off (the knock-out mice) clearly differed from their kind. "The knock-out mice showed clearly diminished learning and memory capacity," said Privatdozent Dr. Andras Bilkei-Gorzo from Professor Zimmer's team, who led the study. So, animals that did not have the receptor were less successful in their search for the platform. "In addition, they showed a clear loss of nerve cells in the hippocampus," he explained further. This part of the brain is the central area for forming and storing information. In addition, the researchers found inflammation processes in the brain. As the mice advanced in age, the degenerative processes became increasingly noticeable.


Amazing parallels with the human brain


The animals with the intact CB1 receptor, to the contrary, did clearly better with regard to their learning and memory capabilities, as well as the health of their nerve cells. "The root cause of aging is one of the secrets of life," commented Albayram. This study has begun to open the door to solving this enigma. The processes in the mouse brains have a surprising number of parallels with age-related changes in human brains. So, the endocannabinoid system may also present a protective mechanism in the aging of the human brain.


The principal author cautioned, "This will require additional research." The scientists would like to better understand the mechanism by which CB1 receptors protect the brain from inflammation processes. And based on these signal chains, it might then be possible to develop substances for new therapies.


Source : University of Bonn

Genome code cracked for most common form of pediatric brain cancer

Scientists at the Johns Hopkins Kimmel Cancer Center have deciphered the genetic code for medulloblastoma, the most common pediatric brain cancer and a leading killer of children with cancer. The genetic "map" is believed to be the first reported of a pediatric cancer genome and is published online in the December 16 issue of Science Express.


Notably, the findings show that children with medulloblastoma have five- to tenfold fewer cancer-linked alterations in their genomes compared with their adult counterparts, the scientists say.


"These analyses clearly show that genetic changes in pediatric cancers are remarkably different from adult tumors. With fewer alterations, the hope is that it may be easier to use the information to develop new therapies for them," says Victor Velculescu, M.D., Ph.D., associate professor of oncology at the Johns Hopkins Kimmel Cancer Center.


"We now know what many pieces of the medulloblastoma puzzle are," adds Bert Vogelstein, M.D., Clayton Professor of Oncology and co-director of the Ludwig Center at Johns Hopkins. "Now, we must figure out how to put the puzzle together and zero in on parts of the puzzle to develop new therapies. This is what scientists will be focused on for the next decade."


The Johns Hopkins team used automated tools to sequence hundreds of millions of individual chemicals called nucleotides, which pair together in a preprogrammed fashion to build DNA and, in turn, a genome. Combinations of these nucleotide letters form genes, which provide instructions that guide cell activity. Alterations in the nucleotides, called mutations, can create coding errors that transform a normal cell into a cancerous one. The scientists at Johns Hopkins have previously mapped genome sequences for pancreatic, adult brain, breast and colon cancers with similar methods.


For the study, scientists sequenced nearly all protein-encoding genes in 22 samples of pediatric medulloblastoma and compared these sequences with normal DNA from each patient to identify tumor-specific changes or mutations. Each tumor sample had an average of 11 mutations. There were 225 mutations in all.


Then, the investigators searched through a second set of 66 medulloblastomas, including some samples from adults, to find how these mutations altered the proteins made by the genes.


The team found that most of the mutations congregate within a few gene families or pathways. The most prevalent pathway ordered the way long strands of DNA, that make up chromosomes, are twisted and shaped into dense packets that open and close depending on when genes need to be activated. Such a process is regulated by chemicals that operate outside of genes, termed "epigenetic" by scientists.


Within the epigenetic pathway, two commonly mutated genes were both involved in how molecules called histones wrap around DNA.


"These epigenetic changes may be more important than we thought in childhood cancers," says Will Parsons, M.D., Ph.D., formerly of Johns Hopkins and now an assistant professor at Texas Children's Cancer Center and Baylor College of Medicine.


Mutations in MLL2 and MLL3 were identified in 16 percent of the entire set of 88 medulloblastoma samples. Add to this three other epigenetic alterations found by the scientists in the genome scan, and the total set accounts for 20 percent of mutations in all the brain cancer samples.


Second to epigenetic pathways were gene mutations in pathways such as Hedgehog and Wnt that control tissue and organ development in humans and other animals. Both pathways have previously been linked to childhood medulloblastoma.


Cancer is the leading cause of death by disease in children in the U.S., and more children die of brain tumors than any other type of cancer. Medulloblastoma is the most common malignant brain tumor in children, occurring in about 400 children per year in the U.S.


"It's a particular challenge to treat children with brain cancer," says Parsons, "because our most effective treatments, surgery and radiation therapy, can cause significant side effects, including cognitive disabilities and hormone abnormalities. For our youngest patients, the effects can be potentially devastating."


Yet, Parsons is encouraged by the study's findings. "As oncologists, we're working to understand how specific genetic changes found in patients' cancers should guide their treatment. Any information that allows us to understand a patient's prognosis or provides clues about therapies that might work best in a patient is crucial and will help us provide better care."


Source : Johns Hopkins Medical Institutions

1 tiny electron could be key to furture drugs that repair sunburn

Researchers who have been working for nearly a decade to piece together the process by which an enzyme repairs sun-damaged DNA have finally witnessed the entire process in full detail in the laboratory.



What they saw contradicts fundamental notions of how key biological molecules break up during the repair of sunburn – and that knowledge could someday lead to drugs or even lotions that could heal sunburn in humans.


In the Proceedings of the National Academy of Sciences, the Ohio State University researchers and their colleagues confirm what was previously known about the enzyme photolyase, which is naturally produced in the cells of plants and some animals – though not in mammals, including humans. The enzyme repairs DNA by tearing open the misshapen, damaged area of the DNA in two places and reforming it into its original, undamaged shape.


But the enzyme doesn't break up the injury in both places at once, as researchers previously suspected from theoretical calculations. Instead, it's a two-step process that sends an electron through the DNA molecule in a circuitous route from one breakup site to the other, the new study revealed.


The research was led by Dongping Zhong, the Robert Smith Professor of Physics and professor in the departments of chemistry and biochemistry at Ohio State.


Zhong and his team literally shed light on the process in the laboratory using a laser with a kind of strobe effect to take super-fast measurements of the enzyme in action.


What they saw surprised them.


The two key chemical bond sites broke up one after the other – the first in just a few trillionths of a second, and the next after a 90-trillionths-of-a-second delay.


The reason? The single electron ejected from the enzyme – the source of energy for the breakup – took time and energy to travel from one bond site to the other, tunneling along the outer edge of the ring-shaped damage site.


Also, it turns out that for the enzyme taking the long way around is the most efficient way for the electron to do the job, Zhong explained.


"The enzyme needs to inject an electron into damaged DNA -- but how?" he said. "There are two pathways. One is direct jump from the enzyme across the ring from one side to the other, which is a short distance. But instead the electron takes the scenic route. We found that along the way, there is another molecule that acts as a bridge to speed the electron flow, and in this way, the long route actually takes less time."


Now that they have revealed how the enzyme actually works, the researchers hope that others can use this knowledge to create synthetic photolyase for drugs or even lotions that can repair DNA.


Ultraviolet (UV) light damages DNA by exciting the atoms in the DNA molecule, causing accidental bonds to form between the atoms. The bond is called a photo-lesion, and can lead to a kind of molecular injury called a dimer. Dimers prevent DNA from replicating properly, and cause genetic mutations that lead to diseases such as cancer.


The dimer in question is called a cyclobutane pyrimidine dimer, and it is shaped like a ring that juts out from the side of the DNA.


For those organisms lucky enough to have photolyase in their cells, the enzyme absorbs energy from visible light – specifically, blue light – to shoot an electron into the cyclobutane ring to break it up. The result is a perfectly repaired strand of DNA.


That's why photolyase-carrying insects, fish, birds, amphibians, marsupials, and even bacteria, viruses and yeast are all protected from cancer-causing UV rays from the sun. Meanwhile, humans and all other mammals lack the enzyme, and so are particularly vulnerable to UV.


A synthetic form of photolyase could make up for our enzymatic shortfall. But Zhong's group will leave that discovery to other researchers; they have now set their sights on photoreceptors – the proteins that absorb light and initiate signaling for many biological functions.

Source : Ohio State University

Artificial lung mimics real organ's design and efficiency

An artificial lung built by Cleveland researchers has reached efficiencies akin to the genuine organ, using air – not pure oxygen as current man-made lungs require - for the source of the essential element.



Use in humans is still years away, but for the 200 million lung disease sufferers worldwide, the device is a major step toward creating an easily portable and implantable artificial lung, said Joe Potkay, a research assistant professor in electrical engineering and computer science at Case Western Reserve University. Potkay is the lead author of the paper describing the device and research, in the journal Lab on a Chip.


The scientists built the prototype device by following the natural lung's design and tiny dimensions. The artificial lung is filled with breathable silicone rubber versions of blood vessels that branch down to a diameter less than one-fourth the diameter of human hair.


"Based on current device performance, we estimate that a unit that could be used in humans would be about 6 inches by 6 inches by 4 inches tall, or about the volume of the human lung. In addition, the device could be driven by the heart and would not require a mechanical pump," Potkay said.


Current artificial lung systems require heavy tanks of oxygen, limiting their portability. Due to their inefficient oxygen exchange, they can be used only on patients at rest, and not while active. And, the lifetime of the system is measured in days.


The Cleveland researchers focused first on improving efficiency and portability.


Potkay, who specializes in micro- and nano-technology, worked with Brian Cmolik, MD, an assistant clinical professor at Case Western Reserve School of Medicine and researcher at the Advanced Platform Technology Center and the Cardiothoracic Surgery department at the Louis Stokes Cleveland VA Medical Center. Michael Magnetta and Abigail Vinson, biomedical engineers and third-year students at Case Western Reserve University School of Medicine, joined the team and helped develop the prototype during the past two years.


The researchers first built a mould with miniature features and then layered on a liquid silicone rubber that solidified into artificial capillaries and alveoli, and separated the air and blood channels with a gas diffusion membrane.


By making the parts on the same scale as the natural lung, the team was able to create a very large surface-area-to-volume ratio and shrink the distances for gas diffusion compared to the current state of the art. Tests using pig blood show oxygen exchange efficiency is three to five times better, which enables them to use plain air instead of pure oxygen as the ventilating gas.


Potkay's team is now collaborating with researchers from Case Western Reserve's departments of biomedical engineering and chemical engineering to develop a coating to prevent clogging in the narrow artificial capillaries and on construction techniques needed to build a durable artificial lung large enough to test in rodent models of lung disease.


Within a decade, the group expects to have human-scale artificial lungs in use in clinical trials.


They envision patients would tap into the devices while allowing their own diseased lungs to heal, or maybe implant one as a bridge while awaiting a lung transplant – a wait that lasts, on average, more than a year.


Source : Case Western Reserve University