Transient receptor potential (TRP) ion channels mediate a variety of sensations including thermal stimuli. The channels also react to multiple ligands: for example, TRPV1 channels respond to heat as well as to ligands which elicit a hot sensation such as capsaicin, whereas TRPM8 channels respond to cold and also to ligands which elicit a cool sensation such as menthol. Since TRP channels are expressed on sensory neurons, a number of groups are developing selective channel modulators for the treatment of pain.
Researchers led by a team at the Instituto de Neurociencias de Alicante have now shown that TRPM8 channels in the eye also regulate basal tear secretion. In studies in mice, the researchers found that cooling of the eye surface by 1-2°C by evaporation of the tear film can induce production of more tears by stimulation of TRPM8 in nerve endings in the cornea. In mice lacking TRPM8, the cornea was insensitive to cooling and the basal rate of tear production was greatly reduced. Tear production caused by exposure to irritants which is mediated by other channels such as TRPV1 was, however, unaffected in the TRPM8 deficient mice. In normal mice, tear secretion could also be decreased by raising the temperature of the cornea to 33-36°C. Tear production in humans is also regulated by cold – the basal rate of tear production is significantly lower at 43°C than at 18-20°C.
The study, which is published in the journal Nature Medicine, indicates that TRPM8 contributes to the regulation of basal tear flow and opens new possibilities for the treatment of dry eye syndrome by increasing tear secretion. Dry mucosal surfaces, including dry eye syndrome, are a common problem, particularly for the elderly with up to one third of people over the age of sixty five estimated to have dry eyes.
Transient Receptor Potential (TRP) ion channels are a large superfamily of transmembrane proteins which, amongst other functions, help an organism to sense its environment. The TRPV1 channel –which is activated by heat and also by capsaicin, the pungent component of hot chilli peppers – has been linked to the development of epithelial cancers.
A new study by researchers at the University of Michigan and Children’s Hospital Boston shows that the TRPV3 receptor – a molecular sensor for warm temperatures – plays a key role in maintenance of the skin barrier and hair growth and could perhaps also be linked to the development of skin cancer. Epidermal growth factor receptor (EGFR) and one of its ligands, transforming growth factor-α (TGF-α), were known to regulate skin and hair growth and the new study shows that TRPV3 is a key component of the EGFR signalling pathway. Activation of TRPV3 was found to lead to release of TGF-α, which activates EGFR. Activation of EGFR, in turn, increases TRPV3 channel activity, forming a positive feedback loop. Like animals with naturally occurring loss-of-function mutations in the genes for EGFR or TGF-α, TRPV3 knockout mice were found to have curly coats and whiskers.
In the TRPV3 knockout mice, the outer layer of the skin was found to be thinner than normal and to have a dry scaly texture which resulted from impaired terminal differentiation of keratinocytes. The study suggests that small molecule activators of TRPV-3 could be used to treat a variety of skin injuries and diseases such as burns, bed sores, eczema, psoriasis, itch, and fungal infections.
One cautionary note is that mice with overactive EGFR or TGF-α are hairless and develop skin cancers. Upregulation of EGFR has also been linked to a number of human cancers and drugs targeting EGFR are an expanding class of cancer treatments. Although mice overexpressing TRPV3 have not been shown to have an increased number of tumours, this is something that would need to be investigated further. The study is published in the journal Cell.
Ion channels, proteins that regulate the transfer of ions across the cell membrane, can be broadly classified according to ‘gating mechanism’ – what makes the channel open and close. Voltage-gated channels respond to a voltage gradient across the plasma membrane whereas ligand-gated channels are activated by binding of extracellular ligands or intracellular second messengers. Recent detailed studies of ion channels are showing, however, that things are not quite so simple.
Researchers in the US and Germany have now shown that they can confer significant voltage dependence to the inwardly rectifying K+ channel, Kir6.2, by introducing a point mutation, L157E. Kir6.2 is a ligand-gated channel that lacks a canonical voltage-sensing domain (VSD). In classical models of voltage-dependent gating, the VSD strongly influences opening and closing of the pore-forming domain so that the channel open probability is reduced to virtually zero at sufficiently negative voltages and increased to near unity upon depolarization. Previous observations have shown that such ‘tight coupling’ between the VSD and the pore does not apply to all channel types and the new study shows that substitution of charged residues at pore-lining positions can affect channel gating in very unexpected ways. Comparing Kir6.2[L157E] with wild type Kir6.2, the team found that the probability of opening under conditions of low intracellular K+ was much greater for the mutant channel. The presence of a natural ligand, phosphatidyl inositol bis-phosphate (PIP2), removed the voltage dependence of the mutant channel. Both voltage- and ligand-dependent gating of Kir6.2[L157E] were highly sensitive to intracellular [K+], indicating an interaction between ion permeation and gating.
The authors of the study, which is published in PLoS Biology, propose that ions flowing through the ion channel pore can significantly affect channel activity and that the mechanisms of voltage-gating and ligand-gating may be more closely linked than previously supposed. They further suggest that such interactions are likely to be a general, if latent, feature of the superfamily of cation channels.
Energy-conserving mechanisms that evolved as protective measures in an environment of restricted food supply and high demand for physical activity promote obesity in times of abundant food and low physical activity. ATP-sensitive potassium (KATP) channels in heart and skeletal muscle act as safety valves that limit action potentials to prevent energy depletion and are essential for survival and stress adaptation, but researchers at the Mayo Clinic, the University of Iowa, New York University School of Medicine and the University of Connecticut have now found that the channels also regulate cellular energy use under non-stressed physiological conditions and contribute to fat deposition and obesity.
Both when the animals were at rest or normally active, heart and skeletal muscles of mice lacking the KATP channel dissipated more energy as heat than those of wild type mice and the animals were resistant to increases in body weight caused by a Western-style high fat diet. However, since the animals’ muscles are also less efficient when exercising, they show lower endurance and are less capable of maintaining physical performance than wild type animals.
The authors hope that therapies that reduce the activity of KATP channels in a tissue-specific manner may have the potential to reduce obesity by making muscles more thermogenic at rest and less fuel efficient during exercise.
Image: Wikipedia The body has no way to detect the absence of oxygen and feelings of suffocation are triggered instead by high levels of carbon dioxide. The ‘false-suffocation-alarm theory’ proposes that this alarm is triggered inappropriately in patients with panic disorder but, although several studies have linked high carbon dioxide levels with panic attacks in susceptible individuals, the reasons for this have not been clear. Researchers at the University of Iowa have now shown that inhaled carbon dioxide increases acidity in the brain and evokes fear by activating an acid-sensing ion channel (ASIC1a) in the amygdala. The amygdala plays a key role in the processing and memory of emotional reactions, including fear, but it was not known whether it also directly senses fear-evoking stimuli. ASICs are activated in vitro when extracellular pH falls, and although acidic pH modifies the activity of many receptors and proteins, few others are activated by extracellular acidosis, and few are as exquisitely pH sensitive as ASICs.
In tests in mice, increased carbon dioxide levels led to exaggerated innate and learned fear responses which could be blunted either by disrupting the ASIC1a gene or by pharmacological inhibition of ASIC1a using either the tarantula toxin, psalmotoxin, or A-317567.
The finding that ASIC1a channels in the amygdala act as chemosensors provides a molecular mechanism by which carbon dioxide can trigger fear and anxiety and suggests that targeting brain pH or ASIC channels could lead to new therapies for panic and anxiety disorders.
Smokers who use nicotine replacement products to help them quit smoking can experience local irritation at the site of application. It had been generally assumed that the irritation was caused by nicotine stimulation of nicotinic acetylcholine receptors (nAChRs) in sensory neurons, but new research by scientists in Belgium and Spain has shown that the irritation is caused instead by activation of the transient receptor potential cation channel, TRPA1. TRP cation channels are an ancient family of receptors that act as diverse cellular sensors, responding to temperature, taste, touch, pain and other stimuli, both at the level of the entire multicellular organism and also at the level of single cells. TRPA1 is expressed in nociceptive neurons and is activated by a variety of noxious stimuli, including extreme cold and pungent compounds such as mustard oil, as well as playing a role in inflammatory pain.
In the present study, the researchers showed that nicotine activates both human and mouse TRPA1 channels expressed in Chinese hamster ovary cells. In contrast to TRPA1 activation by electrophiles such as mustard oil, during which N-terminal cysteine residues are covalently modified, activation by nicotine does not involve covalent modification (as would be expected from the structure of nicotine). A significantly lower proportion of nociceptive mouse trigeminal neurons from Trpa1 knockout mice responded to nicotine exposure than did neurons from wild-type mice. In neurons from wild-type mice, nicotine was found to evoke two separate responses: rapid and quickly desensitising responses mediated by nAChRs and slower, more sustained responses mediated by TRPA1. To test whether TRPA1 contributes to irritant effects of nicotine in vivo, the team compared the airway constriction reflexes of wild-type and Trpa1 knockout mice to stimulation of the nasal mucosa. Airway constriction increased significantly after nicotine application in wild-type but not Trpa1 knockout mice.
The study, which is published in the journal Nature Neuroscience, suggests that inhibitors of TRPA1 could be used to develop smoking cessation therapies with fewer side effects and better compliance. Although relatively high nicotine concentrations are needed to activate TRPA1, the concentrations needed are within the range delivered by nicotine nasal sprays, the most effective form of nicotine replacement therapy, but also the one with the highest dropout rate because of local irritation.
Globally, hepatitis C virus (HCV) infects almost 200 million people and is a leading cause of cirrhosis and hepatocellular carcinoma – albeit several decades after initial infection. In a majority of cases, the virus is able to establish a persistent infection and, even with current gold standard treatments, sustained cure rates once the infection has become established are only around 50%. Researchers at the University of Leeds have now uncovered a previously unrecognized mechanism that allows the virus to evade the immune system and establish a chronic infection.
Establishment of persistent infection means that HCV-infected cells must be resistant to pro-apoptotic stimuli and the team found that one of the viral proteins, NS5A, is able to block apoptosis in human hepatoma cells either infected with HCV or harbouring an HCV subgenomic replicon. Amplification of an outward K+ current mediated by voltage-gated Kv2.1 channels normally precedes apoptosis triggered by oxidative stress and NS5A was found to block this process by inhibiting phosphorylation of Kv2.1 by p38 MAP kinase. Inhibition of a host cell ion channel by a viral protein as a means of preventing apoptosis has not previously been described, and the researchers hope that their findings could lead to new strategies for antiviral therapy. The study is published in the August 26th online edition of PNAS.
A number of groups are developing nicotinic α-7 receptor agonists or partial agonists for the treatment of Alzheimer’s disease. It is believed that α-7 agonists may contribute to symptomatic treatment through cholinergic mechanisms and may also protect vulnerable neurons from the neurotoxic effects of β-amyloid peptides. Although α-7 agonists have shown positive effects on cognition in both animal models and Alzheimer’s disease patients, researchers at the Salk Institute have suggested that ‘Nicotinic Receptor May Help Trigger Alzheimer’s Disease’, and propose that nicotinic α-7 receptor antagonists may be a better target for the treatment of Alzheimer’s disease.
The team found that, whereas transgenic mice that overexpress a mutated form of human amyloid precursor protein (APP) perform poorly in learning and memory tests, mice that overexpress APP but also lack nicotinic α-7 receptors perform as well as wild type mice. β-Amyloid disrupts the function of nicotinic α-7 receptors and is believed to accumulate in neurons via high affinity binding to the receptors. Agonist stimulation of α-7 receptors could thus restore impaired or altered intracellular signalling caused by β-amyloid and, by desensitisation or competitive binding, could also prevent internalisation of β-amyloid. Although it is likely that α-7 receptor antagonists would also prevent internalisation of β-amyloid, there is currently little other information to support the development of antagonists, especially given the toxicities of known α-7 receptor antagonists such as α-bungarotoxin and methyllycaconitine.
Acute crush injury to the spinal cord is immediately followed by secondary tissue damage, linked to massive release of ATP and activation of high-affinity P2X7 receptors. Researchers at the University of Rochester Medical Center have previously shown that intraspinal injection of the P2X7 antagonist, adenosine 5′-triphosphate-2′,3′-dialdehyde (OxATP), improves outcomes of spinal injury in rats, but intraspinal injection – together with cardiovascular toxicity – makes this treatment unattractive for human trauma victims. In a new study, the team have shown that outcomes in rats can also be improved by systemic treatment with a different P2X7 antagonist, the Coomassie dye, Brilliant Blue G.
Writing in the journal PNAS, they show that iv administration of Brilliant Blue G (10 or 50 mg/kg) 15 minutes after injury, and for three consecutive days, protected spinal cord neurons from purinergic excitotoxicity and also reduced local inflammatory responses, resulting in reduced spinal cord anatomic damage and improved motor recovery. After 6 weeks, treated animals recovered sufficiently to walk with a limp whereas untreated animals did not walk again. Although it seems unlikely that Brilliant Blue G would efficiently cross the intact blood brain barrier, the dye was found to accumulate in the lesions in the injured animals.
There is currently no effective treatment to prevent secondary damage in patients with acute spinal cord injury and the team hope that their work will lead to safe practical treatments that could be administered soon after injury to improve outcomes for spinal injury victims.
The team chose to use Brilliant Blue G for their experiments because they saw structural and functional similarities with a food additive, FD&C blue dye No 1 (E133), used in a variety of processed foods and generally considered to be safe. A number of groups have now designed selective P2X7 antagonists – some of which have entered the clinic – and it would be interesting to see the effect of these newer compounds in the rat spinal injury model. Because of the differing affinities of antagonists for rat and human receptors, care will be needed in the choice of appropriate molecules for study, and in extrapolation of results from rodents to humans.
Asthma involves an immune response to inhaled allergens and chemical irritants, but the limited efficacy of existing treatments aimed at modifying this response suggest that additional physiological mechanisms may be at work in the disease process. In a report published in the May 19th online Early Edition of PNAS, US researchers have now found that the ion channel, TRPA1, plays a key role in allergic asthma. TRPA1 is a ‘chemosensor’ that is activated by mustard oil as well as by a number of endogenous and exogenous stimuli known to be triggers of asthmatic inflammation. The team found that, compared with wild type mice, animals that did not express TRPA1 showed fewer asthma symptoms, with reduced inflammation, airway mucus and bronchoconstriction. Although the exact role of TRPA1 in asthmatic inflammation is not yet understood, the ion channel is known to be activated by allergens such as cigarette smoke that can trigger asthma attacks. TRPA1 is found in airway nerves and the researchers believe that blocking TRPA1 may prevent infiltration of the lung by the inflammatory cells responsible for asthma symptoms such as wheezing and overproduction of mucus. In further studies, the team went on to show that treatment with the TRPA1 antagonist, HC-030031, reduced the symptoms of allergic asthma in mice. TRPA1 antagonists have previously been shown to reduce chronic inflammatory and neuropathic pain. The discovery of a role for TRPA1 as a neuronal mediator of allergic airway inflammation could lead to new treatments for allergic asthma and Hydra Biosciences, whose scientists contributed to the study, hope to start human clinical trials with a novel TRPA1 inhibitor within 12 months.