(Image caption: These are mature nerve cells generated from human cells using enhanced transcription factors. Credit: Fahad Ali)

Functional nerve cells from skin cells

A new method of generating mature nerve cells from skin cells could greatly enhance understanding of neurodegenerative diseases, and could accelerate the development of new drugs and stem cell-based regenerative medicine.

The nerve cells generated by this new method show the same functional characteristics as the mature cells found in the body, making them much better models for the study of age-related diseases such as Parkinson’s and Alzheimer’s, and for the testing of new drugs.

Eventually, the technique could also be used to generate mature nerve cells for transplantation into patients with a range of neurodegenerative diseases.

By studying how nerves form in developing tadpoles, researchers from the University of Cambridge were able to identify ways to speed up the cellular processes by which human nerve cells mature. The findings are reported in the May 27th edition of the journal Development.

Stem cells are our master cells, which can develop into almost any cell type within the body. Within a stem cell, there are mechanisms that tell it when to divide, and when to stop dividing and transform into another cell type, a process known as cell differentiation. Several years ago, researchers determined that a group of proteins known as transcription factors, which are found in many tissues throughout the body, regulate both mechanisms.

More recently, it was found that by adding these proteins to skin cells, they can be reprogrammed to form other cell types, including nerve cells. These cells are known as induced neurons, or iN cells. However, this method generates a low number of cells, and those that are produced are not fully functional, which is a requirement in order to be useful models of disease: for example, cortical neurons for stroke, or motor neurons for motor neuron disease.

In addition, for age-related diseases such as Parkinson’s and Alzheimer’s, both of which affect millions worldwide, mature nerve cells which show the same characteristics as those found in the body are crucial in order to enhance understanding of the disease and ultimately determine the best way to treat it.

"When you reprogramme cells, you’re essentially converting them from one form to another but often the cells you end up with look like they come from embryos rather than looking and acting like more mature adult cells," said Dr Anna Philpott of the Department of Oncology, who led the research. "In order to increase our understanding of diseases like Alzheimer’s, we need to be able to work with cells that look and behave like those you would see in older individuals who have developed the disease, so producing more ‘adult’ cells after reprogramming is really important."

By manipulating the signals which transcription factors send to the cells, Dr Philpott and her collaborators were able to promote cell differentiation and maturation, even in the presence of conflicting signals that were directing the cell to continue dividing.

When cells are dividing, transcription factors are modified by the addition of phosphate molecules, a process known as phosphorylation, but this can limit how well cells can convert to mature nerves. However, by engineering proteins which cannot be modified by phosphate and adding them to human cells, the researchers found they could produce nerve cells that were significantly more mature, and therefore more useful as models for disease such as Alzheimer’s.

Additionally, very similar protein control mechanisms are at work to mature important cells in other tissues such as pancreatic islets, the cell type that fails to function effectively in type 2 diabetes. As well as making more mature nerves, Dr Philpott’s lab is now using similar methods to improve the function of insulin-producing pancreas cells for future therapeutic applications.

"We’ve found that not only do you have to think about how you start the process of cell differentiation in stem cells, but you also have to think about what you need to do to make differentiation complete - we can learn a lot from how cells in developing embryos manage this," said Dr Philpott.

Breakthrough: Nasal spray may soon replace the pill

When the doctor gives us medicine, it is often in the shape of a pill. But when it comes to brain diseases, pills are actually an extremely ineffecient way to deliver drugs to the brain, and according to researchers from University of Southern Denmark we need to find new and more efficient ways of transporting drugs to the brain. Spraying the patient’s nose could be one such way.

Every time we have an infection or a headache and take a pill, we get a lot more drugs than our body actually needs. The reason is that only a fraction of the drugs in a pill reaches the right places in the body; the rest never reaches its destination and may cause unwelcome side effects before they are flushed out of the body again. This kind of major overdosing is especially true when doctors treat brain diseases, because the brain does not easily accept entering drugs.

"People with brain diseases are often given huge amounts of unnecessary drugs. During a long life, or if you have a chronic disease, this may become problematic for your health", says Massimiliano Di Cagno, assistant professor at the Department of Physics, Chemistry and Pharmacy, University of Southern Denmark.

He is concerned with finding more efficient ways of delivering drugs to the brain. He and his colleagues at University of Southern Denmark and Aalborg University have turned their attention to the nose - specifically the nasal wall and the slimy mucosa that covers it.

As we know from e.g. cocaine addicts, substances can be assimilated extremely quickly and directly through the nose. But many medical substances, however, need help to be transported through the nasal wall and further on to the relevant places in the brain.

Researchers have long struggled with this challenge and have come up with different kinds of transport vehicles that are very good at transporting the active ingredients through the nasal wall into the brain. The problem with these vehicles, though, is that they cannot release their cargo of drugs once they have reached the inside of the brain. The drugs stay locked inside the strong vehicles.

“If the drugs cannot get out of their vehicles, they are no help to the patient. So we needed to develop a vehicle that does not lock the drug in”, explains Massimiliano Di Cagno.

The vehicles for drug delivery through the nose are typically made of so called polymers. A polymer is a large molecule composed of a large number of repeats of one or more types of atoms or groups of atoms bound to each other. Polymers can be natural or synthetic, simple or complex.

Direct track to the brain

Massimiliano Di Cagno and his colleagues tested a natural sugar polymer and they now report that this particular polymer is not only capable of carrying the drugs through the nasal wall but also – and most importantly – releasing the drug where it is needed.

"This is an important breakthrough, which will bring us closer to delivering brain drugs by nasal spray", says Massimiliano Di Cagno.

With this discovery two out of three major challenges in nasal delivery of brain drugs have been met:

“We have solved the problem of getting the drug through the nose, and we have solved the problem of getting the drug released once it has entered the brain. Now there is a third major challenge left: To secure a steady supply of drugs over a long period. This is especially important if you are a chronic patient and need drug delivery every hour or so”, says Massimiliano Di Cagno.

When a patient sprays a solution with active drugs into his nose cavity, the solution will hit the nasal wall and wander from here through the nasal wall to the relevant places in the brain.

“But gravity also rules inside the nose cavity and therefore the spray solution will start to run down as soon as it has been sprayed up the nose. We need it to cling to the nasal wall for a long time, so we need to invent some kind of glue that will help the solution stick to the nasal wall and not run down and out of the nose within minutes”, says Massimiliano Di Cagno.

Genes discovered linking circadian clock with eating schedule

For most people, the urge to eat a meal or snack comes at a few, predictable times during the waking part of the day. But for those with a rare syndrome, hunger comes at unwanted hours, interrupts sleep and causes overeating.

Now, Salk scientists have discovered a pair of genes that normally keeps eating schedules in sync with daily sleep rhythms, and, when mutated, may play a role in so-called night eating syndrome. In mice with mutations in one of the genes, eating patterns are shifted, leading to unusual mealtimes and weight gain. The results were published in Cell Reports today.

"We really never expected that we would be able to decouple the sleep-wake cycle and the eating cycle, especially with a simple mutation," says senior study author Satchidananda Panda, an associate professor in Salk’s Regulatory Biology Laboratory. "It opens up a whole lot of future questions about how these cycles are regulated."

More than a decade ago, researchers discovered that individuals with an inherited sleep disorder often carry a particular mutation in a protein called PER2. The mutation is in an area of the protein that can be phosphorylated—the ability to bond with a phosphate chemical that changes the protein’s function. Humans have three PER, or period, genes, all thought to play a role in the daily circadian clock and all containing the same phosphorylation spot.

The Salk scientists joined forces with a Chinese team led by Ying Xu of Nanjing University to test whether mutations in the equivalent area of PER1 would have the same effect as those in PER2 that caused the sleep disorder. So they bred mice to lack the mouse period genes, and added in a human PER1 or PER2 with a mutation in the phosphorylation site. As expected, mice with a mutated PER2 had sleep defects, dozing off earlier than usual. The same wasn’t true for PER1 mutations though.

"In the mice without PER1, there was no obvious defect in their sleep-wake cycles," says Panda. "Instead, when we looked at their metabolism, we suddenly saw drastic changes."

Mice with the PER1 phosphorylation defects ate earlier than other mice—causing them to wake up and snack before their sleep cycle was over—and ate more food throughout their normal waking period. When the researchers looked at the molecular details of the PER1 protein, they found that the mutated PER1 led to lower protein levels during the sleeping period, higher levels during the waking period, and a faster degradation of protein whenever it was produced by cells.

Panda and his colleagues hypothesize that normally, PER1 and PER2 are kept synchronized since they have identical phosphorylation sites—they are turned on and off at the same times, keeping sleep and eating cycles aligned. But a mutation in one of the genes could break this link, and cause off-cycle eating or sleeping.

"For a long time, people discounted night eating syndrome as not real," says Panda. "These results in mice suggest that it could actually be a genetic basis for the syndrome." The researchers haven’t yet tested, however, whether any humans with night eating syndrome have mutations in PER1.

When Panda and Xu’s team restricted access to food, providing it only at the mice’s normal meal times, they found that even with a genetic mutation in PER1, mice could maintain a normal weight. Over a 10-week follow-up, these mice—with a PER1 mutation but timed access to food—showed no differences to control animals. This tells the researchers that the weight gain caused by PER1 is entirely caused by meal mistiming, not other metabolic defects.

Next, they hope to study exactly how PER1 controls appetite and eating behavior—whether its molecular actions work through the liver, fat cells, brain or other organs.

A New Target for Alcoholism Treatment: Kappa Opioid Receptors

The list of brain receptor targets for opiates reads like a fraternity: Mu Delta Kappa. The mu opioid receptor is the primary target for morphine and endogenous opioids like endorphin, whereas the delta opioid receptor shows the highest affinity for endogenous enkephalins. The kappa opioid receptor (KOR) is very interesting, but the least understood of the opiate receptor family.

Until now, the mu opioid receptor received the most attention in alcoholism research. Naltrexone, a drug approved by the U.S. Food and Drug Administration for the treatment of alcoholism, acts by blocking opiate action at brain receptors and is most potent at the mu opioid receptor. In addition, research has suggested that a variant of the gene that codes for the mu opioid receptor (OPRM1) may be associated with the risk for alcoholism and the response to naltrexone treatment.

However, naltrexone also acts at the kappa opioid receptor and it has not been clear whether this effect of naltrexone is relevant to alcoholism treatment.

A growing body of research in animals implicates the KOR in alcoholism. Stimulation of the KOR, which occurs with alcohol intake, is thought to produce unpleasant and aversive effects. This receptor is hypothesized to play a role in alcohol dependence, at least in part, by promoting negative reinforcement processes. In other words, the theory postulates that during development of alcohol dependence, the KOR system becomes overstimulated, producing dysphoria and anhedonia, which then leads to further alcohol seeking and escalation of alcohol intake that serves to self-medicate those negative symptoms.

A new study in Biological Psychiatry, led by Dr. Brendan Walker at Washington State University, used a rat model of alcohol dependence to directly investigate the KOR system following chronic alcohol exposure and withdrawal.

They found that the KOR system is dysregulated in the amygdala of alcohol-dependent rats, a vital brain region with many functions, including regulation of emotional behavior and decision-making. Chronic alcohol consumption is known to cause neuroadaptations in the amygdala. In this study specifically, they found increased dynorphin A and increased KOR signaling in the amygdala of alcohol-dependent rats.

When the rats were in acute alcohol withdrawal, the researchers administered different drugs, each of which target the KOR system in precise ways, directly into the amygdala. Using this site-specific antagonism, they observed that alcohol dependence-related KOR dysregulation directly contributes to the excessive alcohol consumption that occurs during withdrawal.

“These data provide important new support for the hypothesis that kappa opioid receptor blockers might play a role in the treatment of alcoholism,” said Dr. John Krystal, Editor of Biological Psychiatry. “This study suggests that one role might be to prevent a relapse to alcohol use among patients recently withdrawn from alcohol.”

“This dataset demonstrates the extensive nature of the neuroadaptations the brain undergoes when chronically exposed to alcohol. The implications of these results are far reaching and should help guide pharmacotherapeutic development efforts for the treatment of alcohol use disorders,” said Walker. “Pharmacological compounds that alleviate the negative emotional / mood states that accompany alcohol withdrawal, by attenuating the excessive signaling in the dynorphin / kappa-opioid receptor system, should result in enhanced treatment compliance and facilitate the transition away from alcohol dependence.”

Additional extensive research will be necessary to identify and test the effectiveness of specific drugs that act on the KOR system, but these findings provide researchers with a potentially successful path forward to developing new drugs for the treatment of alcoholism.

How the gut feeling shapes fear

An unlit, deserted car park at night, footsteps in the gloom. The heart beats faster and the stomach ties itself in knots. We often feel threatening situations in our stomachs. While the brain has long been viewed as the centre of all emotions, researchers are increasingly trying to get to the bottom of this proverbial gut instinct.

It is not only the brain that controls processes in our abdominal cavity; our stomach also sends signals back to the brain. At the heart of this dialogue between the brain and abdomen is the vagus nerve, which transmits signals in both directions – from the brain to our internal organs (via the so called efferent nerves) and from the stomach back to our brain (via the afferent nerves). By cutting the afferent nerve fibres in rats, a team of scientists led by Urs Meyer, a researcher in the group of ETH Zurich professor Wolfgang Langhans, turned this two-way communication into a one-way street, enabling the researchers to get to the bottom of the role played by gut instinct. In the test animals, the brain was still able to control processes in the abdomen, but no longer received any signals from the other direction.

Less fear without gut instinct

In the behavioural studies, the researchers determined that the rats were less wary of open spaces and bright lights compared with controlled rats with an intact vagus nerve. “The innate response to fear appears to be influenced significantly by signals sent from the stomach to the brain,” says Meyer.

Nevertheless, the loss of their gut instinct did not make the rats completely fearless: the situation for learned fear behaviour looked different. In a conditioning experiment, the rats learned to link a neutral acoustic stimulus – a sound – to an unpleasant experience. Here, the signal path between the stomach and brain appeared to play no role, with the test animals learning the association as well as the control animals. If, however, the researchers switched from a negative to a neutral stimulus, the rats without gut instinct required significantly longer to associate the sound with the new, neutral situation. This also fits with the results of a recently published study conducted by other researchers, which found that stimulation of the vagus nerve facilitates relearning, says Meyer.

These findings are also of interest to the field of psychiatry, as post-traumatic stress disorder (PTSD), for example, is linked to the association of neutral stimuli with fear triggered by extreme experiences. Stimulation of the vagus nerve could help people with PTSD to once more associate the triggering stimuli with neutral experiences. Vagus nerve stimulation is already used today to treat epilepsy and, in some cases, depression.

Stomach influences signalling in the brain

“A lower level of innate fear, but a longer retention of learned fear – this may sound contradictory,” says Meyer. However, innate and conditioned fear are two different behavioural domains in which different signalling systems in the brain are involved. On closer investigation of the rats’ brains, the researchers found that the loss of signals from the abdomen changes the production of certain signalling substances, so called neurotransmitters, in the brain.

“We were able to show for the first time that the selective interruption of the signal path from the stomach to the brain changed complex behavioural patterns. This has traditionally been attributed to the brain alone,” says Meyer. The study shows clearly that the stomach also has a say in how we respond to fear; however, what it says, i.e. precisely what it signals, is not yet clear. The researchers hope, however, that they will be able to further clarify the role of the vagus nerve and the dialogue between brain and body in future studies.