neurology – Page 4 – Drug Discovery Opinion

Amitriptyline Directly Stimulates Trk Receptors

Neurotrophins – a family of proteins essential for the development, survival and function of neurons – exert their actions through two classes of receptor, Trk tyrosine kinase receptors and p75NTR. There has been considerable interest in using neurotrophins such as nerve growth factor (NGF) for the treatment of neuronal damage as well as for conditions such as stress and depression, but the use of NGF itself has been limited by poor CNS penetration and side effects such as hyperalgesia. This has led to a search for stable small molecules with neurotrophic activity and specificity for TrkA or TrkB receptors although, so far, none of the mimetics can fully reproduce the effects of NGF in animals.

Writing in the journal Chemistry & Biology, researchers at Emory University School of Medicine have now shown that the tricyclic antidepressant, amitriptyline, which had been thought to act predominantly by blocking serotonin and noradrenaline transporters, interacts directly with the extracellular domain of both TrkA and TrkB receptors. Amitriptyline induced TrkA and TrkB homo- and heterodimerization and activation in mouse brain, but heterodimerization was found not to be required for Trk receptor activation. Truncation of the amitriptyline binding motif on TrkA, but not the corresponding region on TrkB, abolished the receptor homo- and heterodimerization. Amitriptyline, but not other tricyclic antidepressants or selective serotonin reuptake inhibitors, promoted amitriptyline and imipramine structure TrkA autophosphorylation in primary neurons and induced neurite outgrowth in PC12 cells. In mice, amitriptyline was further shown to suppress neuronal apoptosis caused by the neuroexitotoxin, kainic acid, in a TrkA-dependent manner. Inhibition of TrkA, but not TrkB, abolished the neuroprotective effect of amitriptyline without affecting its antidepressant activity.

Amitriptyline was found to bind to a motif in the first leucine-rich motif of the extracellular domain of the TrkA receptor with a Kd of 3 µM, which approximates to the brain concentration achieved when used to treat depression or neuropathic pain. Although significantly lower than the affinity of NGF for TrkA, it suggests that amitriptyline affinity for TrkA might be sufficient to explain at least some of its biological activity.

Rhes Protein Linked to Cell Death in Huntington’s Disease

Huntington’s disease (HD) is an inherited neurological disorder that causes a wide range of symptoms including involuntary movements, clumsiness, lack of concentration, memory lapses, mood swings, and depression. Although the disease mechanism is not fully understood, it is known that sufferers have a defect on the short arm of chromosome 4 and produce abnormal versions of the protein, huntingtin. One of the puzzles surrounding the progression of HD has been that, although abnormal huntingtin proteins accumulate in cells everywhere in the body, they predominantly kills cells in the corpus striatum, the part of the brain that controls movement.

Writing in the journal Science, scientists at John Hopkins School of Medicine now report that the answer may lie with the small G protein, Rhes (Ras homologue enriched in striatum), which is found almost exclusively in the corpus striatum. In cell culture experiments using both mouse and human cells, Rhes was found to bind much more tightly to mutant huntingtin than to normal protein. When added singly to cell cultures, neither Rhes nor mutant huntingtin had any effect on survival but, when they were added together, half of the cells died within 48 hours.

Abnormal huntingtin proteins aggregate and form clumps, but there are fewer of these clumps in the corpus striatum of HD patients than in other brain regions or elsewhere in the body, suggesting that clumping of the protein may actually protect the cells. The team found that, in their cell culture experiments, adding Rhes to cells with abnormal huntingtin led to fewer clumps, although the cells died. The results suggest clumping of abnormal huntingtin may prevent it from causing cell death and that Rhes might be responsible for preventing abnormal protein from clumping. Rhes was also shown to promote sumoylation of mutant huntingtin. The team are currently exploring whether removing Rhes from mice with HD will prevent cell death, and hope that it may eventually be possible to design drugs which will specifically target Rhes to treat HD.

Toxoplasmosis Linked to Traffic Accidents

Toxoplasmosis is a parasitic disease caused by infection with the protozoan, Toxoplasma gondii, whose primary hosts are members of the cat family. Infection of humans usually results from consumption of contaminated water, unwashed vegetables or undercooked meat, or from contact with cat faeces. The incidence of human infection varies widely in different countries and, in both acute and latent infections, symptoms are typically mild and often go unnoticed. The latent phase in humans has, however, been associated with behavioural changes and neurological disorders, including increased reaction times. Recent studies have suggested a protective effect of the rhesus factor (RhD antigen) against the negative effects of latent T. gondii infection on reaction times, especially in heterozygous individuals. Uninfected RhD negative men have faster reaction times than uninfected RhD positive men, but heterozygous men with both positive and negative alleles are protected from prolongation of reaction times caused by T. gondii infection.

A study published in the journal BMC Infectious Diseases now suggests that drivers who are rhesus factor (RhD antigen) negative and infected with T. gondii are more likely to be involved in road traffic accidents. Although this is not the first time that an association between toxoplasmosis and driving impairment has been suggested, it is the first prospective study to examine the link, and to consider the effect of RhD phenotype. Almost 4000 Czech male military drivers were tested for T. gondii infection and RhD phenotype at the beginning of a 12-18 months period of compulsory military service. The men were not told whether they were infected or not and were also not told that the incidence of traffic accidents would be monitored. By correlating infection data and RhD phenotype with subsequent traffic accidents reports from military police records, the authors confirmed an increased risk of accidents in infected men, and also demonstrated a protective effect of being RhD positive against this risk. When corrected for factors such as the age of the driver and date of military service, the probability of a traffic accident was found to be higher for RhD negative drivers with high titres of antibodies against T. gondii than for uninfected drivers or for drivers who were RhD positive. Whilst recognising the absence of genotypic data as a limitation of the present study – earlier studies had shown that protection against the deleterious effects of T. gondii infection is largely restricted to heterozygotes – the authors suggest that RhD negative individuals with occupations that need fast reaction times, such as pilots and air traffic controllers, should be regularly tested for infection with T. gondii.

The RhD protein, found on the surface of red blood cells in around 85% of North Americans and Europeans, is highly immunogenic and used to be responsible for a complication of pregnancy. If an RhD negative woman with RhD antibodies – as a result of a blood transfusion or earlier pregnancy –is carrying a baby which is RhD positive as a result of a gene inherited from the father, maternal antibodies will mount an immune response against the red blood cells of the baby. Structural homology data suggest that the RhD protein may act as an ion pump although its function is largely unknown. The authors suggest that the incidence of RhD alleles in human populations may be linked to the advantages conferred to heterozygotes in the presence of endemic T. gondii.

New Target to Enhance Cognitive Function

Histones are basic proteins that interact with negatively charged phosphate groups on DNA, compacting and protecting the DNA, and controlling gene expression. Histones are subject to a number of post-translational modifications, including methylation and acetylation. The balance between acetylation by histone acetyltransferases (HAT) and deacylation by histone deacetylases (HDAC) alters the strength of DNA interactions and plays a key role in regulating gene expression.

Collaborators led by scientists at the Picower Institute for Learning and Memory have identified a promising target that could enable the development of therapeutics to improve memory and learning in patients with neurodegenerative disorders such as Alzheimer’s. The team demonstrated that treatment with HDAC inhibitors enhanced memory and learning ability in normal mice and mouse models of neurodegeneration.

Histone and DNA are the major components of chromatin, the complex that packages genetic information into the chromosomes and inhibitors of the HDAC family have received much attention in recent years as potential treatments for various cancers. Chromatin modification, particularly via deacetylation, has also been implicated in memory formation.

Although HDACs are a family of 11 members, the team has shown that neuron-specific overexpression of HDAC2, but not HDAC1, in mice decreased synaptic plasticity, synapse number and memory formation. This effect was ameliorated by treatment with HDAC inhibitors. Conversely, Hdac2 deficient mice displayed enhancement of synapse number and memory facilitation.

The results, published in full in the journal Nature, suggest exploration of selective HDAC2 inhibitors for treatment of human neurodegenerative diseases involving memory impairment.

Is the Cat Driving you Mad?

Cat's eye Toxoplasmosis is a parasitic infection caused by the protozoan, Toxoplasma gondii. The parasite is able to infect most warm-blooded animals, including humans, although the primary host is the cat family. Infection is via ingestion of infected meat, cat faeces or through mother-to-child transmission. The majority of infected humans display no symptoms, but those with suppressed immune systems, such as AIDS patients or pregnant women, are at greater risk of developing serious illness. It has been estimated that up to 30% of the World’s population is infected with the parasite.

Following the initial acute phase of infection, a latent phase ensues with the parasite forming cysts in muscle and brain. It is known that infection can modify an animal host’s behaviour and experiments in mice have demonstrated a reduced fear of predators. An increase in levels of the neurotransmitter, dopamine, has been observed in the brains of infected mice. Given the tropism of the parasite for brain tissue, it has long been speculated that Toxoplasmosis in humans may be linked with schizophrenia although no causal relationship has been established.

Scientists at Leeds University have now identified two nearly identical genes in the T. gondii genome with similarity to metazoan tyrosine hydroxylase. Unlike metazoan enzymes of this class, the T. gondii enzymes are bifunctional as they are able to utilize tyrosine and phenylalanine as substrates. Whilst the biological role(s) of a bifunctional tyrosine hydroxylase in T. gondii remain unclear, it is possible that the enzyme is required for supply of tyrosine for protein synthesis. Based on the higher substrate selectivity for tyrosine, however, synthesized tyrosine will be converted to L-DOPA. L-DOPA is the precursor to dopamine and it is plausible that the parasite tyrosine hydroxylase is responsible for the increased dopamine levels detected in the brains of infected rodents.

T. gondii’s ability to synthesise dopamine suggests a potential link with neurological conditions such as Schizophrenia, Parkinson’s disease, Tourette’s syndrome and attention deficit disorders.

The full paper is published in the March 11^th issue of PLoS One.

More on the Mechanism of Glatiramer Acetate in Multiple Sclerosis

Approved in 1995, glatiramer acetate (Copaxone®, copolymer-1) is a disease-modifying drug that has been demonstrated to reduce the relapse rate and progression of disability in relapsing-remitting multiple sclerosis (RRMS) patients. The compound is a mixture of synthetic peptides (50-90 amino acids) composed of alanine, glutamic acid, lysine and tyrosine. Originally developed to mimic myelin basic protein, a major component of the neuronal myelin sheath, it was intended for use as an inducer of experimental autoimmune encephalitis (EAE). The unexpected inhibition of EAE that was observed with glatiramer acetate led to clinical trials and subsequent approval for RRMS.

The efficacy of glatiramer acetate has been ascribed to an effect on the adaptive immune response, shifting towards a Th2 polarisation of myelin-specific T-cells. Further studies have demonstrated an immunomodulatory effect on monocytes, macrophages and dendritic cells. However, the full mechanistic picture is still unclear.
neuron
Collaborating scientists from the University of Geneva, Technische Universität München and University of California, San Francisco, have now demonstrated an effect of glatiramer acetate on the IL-1 system. Their research has shown that treatment with the polymer increases blood levels of secreted IL-1 receptor antagonist (sIL-1Ra), a natural inhibitor of IL-1β, both in RRMS patients and in EAE mice. In the same subjects, levels of IL-1β were undetectable. Additional in vitro experiments with T-cell contact-activated monocytes, a model relevant to chronic inflammation, showed that glatiramer acetate strongly reduced expression of IL-1β, whilst enhancing expression of sIL-1Ra. This is in contrast to effects in monocytes subjected to acute inflammatory conditions (stimulation with LPS), where glatiramer acetate increased production of both sIL-1Ra and IL-1β. The authors conclude that the effects on the IL-1 system in chronic inflammatory conditions contribute to the therapeutic effects of glatiramer acetate in RRMS.

The study is published in the online early edition of the journal PNAS.

Selective nNOS Inhibitors may Prevent Cerebral Palsy

The conditions grouped together as cerebral palsy are more common than many people realize and affect about 2 babies in every 1000 in the industrialized world. All types of cerebral palsy are caused by damage to one or more specific areas of the brain which control body movement, posture and muscle coordination. Hypoxia-ischemia (HI)-induced neurological damage before, during, or soon after birth has been implicated as a leading cause of cerebral palsy, and a study in rabbits carried out by researchers at Northwestern University suggests a preventive strategy that may, one day, be feasible for use in humans. In the model, a reduced supply of oxygen from mother to foetus causes an increase in nitric oxide levels in the brain which leads to brain damage and characteristics of cerebral palsy in newly born kittens. Nitric oxide plays an important role in a variety of physiological pathways and is produced from the terminal atom of arginine by a family of nitric oxide synthase enzymes (NOSs), including neuronal (nNOS), macrophage or inducible (iNOS), and endothelial (eNOS) isozymes. The team wanted to develop selective inhibitors of nNOS which they believed would protect the kittens from ischemic damage but have none of the unwanted effects of non-selective NOS inhibitors. Using structure-based design, two potent and selective inhibitors of nNOS were identified and found to be effective in protecting the kittens from neurological damage caused by hypoxia – none of the kittens born to dams treated with either compound died whereas more than half of those born to untreated mothers died. More than 80% of kittens born to rabbits treated with one of the compounds were normal and almost 70% of kittens born to rabbits treated with the other compound were also free from symptoms. The researchers hope that such compounds could, one day, be used as a preventative strategy for reducing the risk of cerebral palsy in babies born to high-risk mothers. The study is published in full in the journal Annals of Neurology.

Natural Sigma-1 Receptor Ligand Indentified

Sigma-1 receptors are widely distributed throughout the body, especially within some areas of the brain, and have been linked to a number of pharmacological activities. Although the endogenous ligand for the sigma-1 receptor had not been identified, synthetic agonists have been linked to beneficial effects on cognition and learning, whilst antagonists have been shown to be effective in some models of pain and to induce apoptosis in susceptible cell types.

Mapuche Machis Scientists have long been searching for naturally occurring ligands for the sigma-1 receptor and a team led by researchers at the University of Wisconsin has now identified a component of plants traditionally used in shamanic rituals in South America as a natural ligand. The active ingredient, N,N-dimethyltryptamine (DMT), is hallucinogenic and also occurs naturally in the body. Although its function is not understood, higher levels have been found in the urine of schizophrenics. n,n-dimethyltryptamine Biochemical, physiological and behavioural studies proved that DMT acts as an agonist of the receptor and, although it is not known whether the sigma-1 receptor is connected to the hallucinogenic activity of DMT, the authors hope that their discovery could have implications for the treatment of drug abuse and depression. The study is published in the February 13^th issue of the journal Science.

Sex, Lobes and Videogames

male female symbols There are many subtle differences in the ways that men and women see the world around them. Women tend to be better at recognizing emotional overtones in others and in language skills whereas men are generally better at solving spatial problems. A team at the University of Iowa has now shown a connection between an ability to mentally rotate objects and the structure of the parietal lobe, the region of the brain involved in spatial ability and mental rotation. Although it was already known that parietal lobes in women have thicker cortexes than those in men, this anatomical difference had not previously been linked to performance differences in mental rotation tests. The surface area of the parietal lobe in men is greater than in women and this appears to be directly linked to improved performance. Magnetic resonance imaging (MRI) showed an approximately 10 % difference between men and women in the overall surface area of the parietal lobe. In mental rotation tests involving 38 men and 38 women, the men averaged 66% correct answers whilst the women averaged 53% correct answers.

The study is published in full in the journal Brain and Cognition.

It is not yet known whether the differences are caused by nature or nurture, although a small study carried out by researchers at the University of Toronto has shown that action video gaming can improve women’s spatial ability. After 10 hr of training with an action video game, both men and women showed substantial improvements in both spatial attention and mental rotation, with women benefiting more than men.

Dream Catcher

dream catcher It is now possible to use computer software to recreate pictures that the eye is seeing using only signals from the visual cortex. Researchers from ATR Computational Neuroscience Laboratories in Japan have described using functional magnetic resonance imaging (fMRI) signals from the visual cortex to reconstruct letters and symbols seen by a subject. They asked subjects to look at 400 random black and white images on a 10 x 10 grid and processed the fMRI signals to map blood flow changes in the brain associated with individual pixels. They were then able to use these maps to reconstruct new images that the subjects looked at. In one test, the researchers were able to reconstruct the letters N-E-U-R-O-N presented to the subjects. This is the first example where such images have been created without using a previously defined set of ‘training’ images.

Although the present study focussed only on the reconstruction of relatively simple black and white patterns, the researchers believe the approach could be extended to reconstruct visual images based on colour, motion or texture. The technique could eventually be used to improve neural prosthetics or to enhance brain-machine interfaces. Other applications could include the reconstruction of mental images that a subject is thinking about, but not actually looking at, such as illusions and dreams which are believed to take place in the early visual cortex.

The study is published in the journal Neuron.