Please can you give a brief introduction to itching?
Itch was actually defined by a German physician more than 350 years ago. His name was Samuel Hafenreffer. He defined itch as an unpleasant sensation that makes people want to scratch. You probably think this is a very simple definition but itch is really a very complex sensory modality.
There are acute and chronic forms of itch. An example of something that causes acute itching is touching poisoned ivy. When you contact poisoned ivy it activates the sensory nerve in the skin which gives you an itch sensation.
There are many forms of chronic itch including the neurogenic itch for instance in kidney disease and other diseases. People believe this may involve central mechanisms – not the primary sensory fibre but maybe the spinal cord or brain in the itch pathway which somehow get activated.
Also nerve damage can cause itch such as in certain cases of shingles which can cause a neuropathic itch so that the nerve becomes damaged or compressed. There are other types of itch such as psychiatric conditions such as Obsessive Compulsive Disorder where you keep scratching and that can generate psychogenic itch.
So itching is very complex and we are only just starting to learn the very basic mechanism of the itch right now. You can imagine there are many stimuli (chemical and mechanical) that can induce itch including chemicals in the environment or itch-mediators inside the body.
What was previously known about the causes of itching sensations?
We know quite a bit. A well-known itch mediator or pruritogen is called histamine. Histamine is mainly secreted from skin mast cells. Mast cells are immune cells that migrate from bone marrow to many places in the body including the skin. Under injury, inflammation or insult mast cells will degranulate, in other words they secrete many substances from their granules including histamine. Histamine, serotonin, ATP, and proteases are the main contents secreted from mast cell granules. Once histamine has been released it can excite the sensory nerve endings which terminate in the skin epidermis or dermis.
People have shown - in many cases using antihistamine, which targets and inhibits the histamine receptor - you can almost completely block histamine-induced itch in humans and in animals. But the problem is these antihistamines are mainly ineffective in many chronic itch conditions, such as atopic dermatitis. This is strong evidence to suggest that there is a histamine-independent pathway for neurons mediating other types of itch.
Of course there is some other experimental data which suggests this is the case too. These include nerve recordings or pharmacological studies using different antagonists and agonists for certain receptors. The majority (about 2/3rd’s) of the itch conditions are histamine independent. Those itches are hard to treat and people give them treatments to alleviate inflammation but that does not inhibit the itch nerve endings, it just reduces the inflammation. So far there is no effective drug to block the itch subsidiary fibres.
This led to our study: we need to find the receptors or itch-mediators that mediate histamine-independent itch. Once we’ve got those molecules then we can figure out how the molecules become abnormal during chronic itch conditions and then we can potentially develop drugs to block those signal pathways.
Why is itching an important sensation?
That is a big debate in the field. It is generally believed that itch provides a protective mechanism. When we feel itch we want to scratch, there are several purposes for that. The first purpose is that scratch-induced mechanical itch and mechanical pain can suppress itch – so we feel good about it, but this is not the main purpose. The main purpose is scratching to remove irritants from the skin. The irritants could include poisoned ivy, an insect crawling on your skin or biting you, such as mosquitoes.
Other people argue that this kind of scratching response is too late – the itching sensation comes a couple of minutes after the initial insult. But also the unpleasant sensation will warn you to avoid this irritant in the future.
Pain is very essential for organisms because they want to avoid the immediate danger of the environment like avoiding touching a hot stove, acid, extremely hot or extremely cold conditions you want to avoid. Humans without the pain sensing fibres cannot live long as they are going to injure themselves. I am not sure what the quality of life would be for individuals that only lack itch. This hasn’t been reported yet.
In our paper we ablated those itch specific fibres and the animals still did fine, but that is only in the laboratory setting, I don’t know how they’d survive in the wild. It needs to be tested why we have itch in the first place, but it is generally believed to be a protective mechanism.
When does itching stop being beneficial and start being problematic?
This is the same as for chronic pain. Pain is a beneficial tool: when pain is acute you avoid danger. This is the same for itch, but when the itch becomes chronic this becomes very problematic there is no beneficial value for chronic itch.
People probably define chronic itch as an itch that lasts at least 6 weeks, but that can be debated. Chronic itch can really affect quality of life, especially for children who can lose sleep as they need to keep scratching. Also, sometimes scratching cannot relieve the itch and it can also damage the skin, so you have this vicious scratch-itching cycles.
How did your research into itch-specific nerve cells originate?
That is actually a long story! There is a debate in the field of somatosensory research on whether itch-specific nerve cells exist. People have known for a long time that pain-sensing neurons and itch-sensing neurons are alike. Itch is often described as pain’s little brother. So people think they may use the same set of neurons to conduct the two different sensations of pain versus itch.
Other theories suggest this is not the case – maybe they don’t use the same population of cells, rather there is a completely different set of neurons for itch and pain, i.e. there is a dedicated population just to conduct itch sensations.
This has been a hot debate for close to 100 years now. The reason we haven’t solved the problem is we cannot do the vigorous tests in humans. So you do need some genetic tricks to answer this question.
Three years ago we published a paper in Cell where we identified a novel receptor for mediating histamine-independent itch. We found the receptor for chloroquine. Chloroquine is an anti-malarial drug and it is widely taken in the world including in Africa. It is a very important drug. The problem in Africa is most of the black Africans that take chloroquine develop a severe itch as a side effect. Many of these patients therefore refuse to take this drug because it makes them so itchy.
Researchers in Africa did a lot of work to try to figure out what’s going on in chloroquine-induced itch in humans. They found it is not an allergic response because the patients when they first take chloroquine immediately develop itching sensations. When these patients are given antihistamines the itching does not go away – they still feel itch – so it appears to be histamine-independent.
A few years ago we found a receptor called MrgA3 which is a receptor for chloroquine. The interesting this about this receptor is that it only expresses a subset of primary sensory neurons. The neurons expressing this receptor we think are good candidates for itch-specific nerve cells. With that result in mind, we started testing that hypothesis using the receptor as a molecular probe to do three sets of experiments to prove we identified for the first time the itch-specific nerve fibres.
What did your research involve?
It involved mouse genetics, molecular biology, electrophysiology and also animal behaviour. Using genetics we can make specific mice and label the specific nerves to see what they look like. That is important because it can answer several important questions. We probably all know that we feel itch mainly from the skin, but why is this the case? The sensory fibres are not only in the skin but they are also in the deeper tissues like muscle, bone. We can feel pain from deeper tissues, such as stomach ache. So why do we only feel itch from the skin? If you don’t see those itch-specific fibres you cannot answer this question.
So now we use a genetic trick we can label those specific fibres with fluorescent protein and we find those itch-specific nerves only go to the skin. They don’t go to the deeper tissues such as muscle, vital organs. Hence you don’t feel itch from deep tissues. This provides a cellular basis for why we only feel itch in the skin.
We can also listen to those nerve fibres using electrophysiology and see whether they can really be activated by itchy compounds, such as histamine or chloroquine – the anti-malarial drug. We found that they responded robustly if you add the itchy compounds into the skin and leave them to the nerve fibres and those itch-specific fibres actually fire action potentials to get activated in response in those itchy compounds. So that’s the first criteria we set on how to consider itch-specific nerve fibres.
The second criteria also involves genetic tricks. We want to kill or ablate those itch fibres specifically not touching other cells – including the pain-sensing fibres. After we killed the itch fibres, the animals didn’t feel itch as normal mice did. We reviewed many types of itch sensation, including histamine-induced itch, chloroquine-induced itch and several other types of itch. Chronic itch conditions in those itch-fibre-ablated mice also show significantly reduced scratching.
Those chronic conditions include dry skin condition – when the weather becomes dry you can feel itch because moisture is lost from the skin. We can mimic these conditions in mice and the mice develop spontaneous scratching. That is significantly reduced in the mice where we have killed the itch-specific fibres. Allergic itch is also significantly reduced. This suggests the fibres we killed are very essential for itch sensations not only the acute itch but also chronic itch.
We are not ruling out the existence of other itch-specific fibres, because we are not completely losing itch sensation yet. Only certain itch sensations were lost completely. Histamine induced itch and allergic itch still around a third or a half remain. So other types of itch fibre definitely exist and we need to identify these in the future.
For the ablation test, although the itch sensation was significantly reduced the pain sensation was completely normal. That suggests these neurons are important for itch but they are dispensable for the pain sensation to get normal pain sensitivity. We still cannot conclude that these fibres that we deleted were not mediating pain signals, because it is possible that after we killed them other nerve fibres transmitted pain signals instead so the animal did not show any loss of pain sensitivity. How can we rule out this possibility? We had to do a third set of experiments.
In the third set of experiments instead of killing these fibres we specifically activated these fibres, so it is the opposite of the second experiment – instead of ablating them we activated them. We again used some genetic manipulation to express a capsaicin receptor only in the itch fibres. Capsaicin is the key component of a chilli pepper; it gives you a hot sensation because it activates a channel called TrpV1. This Trp channel is expressed in many pain sensing fibres, so that’s why when you activate this channel with capsaicin you get a pain sensation.
So now we want to ask the question if we express this pain receptor into itch specific fibres then treat the animal with capsaicin: do they feel pain or itch? They only feel itch, they never respond with a pain behaviour – they only respond with scratching. This suggests that no matter how you activate this fibre we are interested in, even if you use a normally painful stimulus like capsaicin, you still get an itch response.
Because of this final experiment we can conclusively say this Mrg expressing neurons are the itch-specific fibres
Are these findings also likely to be confirmed in humans?
It is theoretically possible. Actually in humans there is a hint of the existence of itch-specific fibres already, but it is not shown conclusively as we did in mice.
The first hint is that when you apply capsaicin – that painful stimulant – very deeply by microinjection, 100% of people feel pain. But if you apply capsaicin very topically, it turns out capsaicin generates an itch sensation. So that result tells us one thing: the itch fibre will terminate very superficially in the epidermis, whereas the pain fibre will go deeper in the epidermis. And so, depending on how you apply capsaicin you will get two different results.
We can label those itch fibres in mice and see that they only go to the skin but if you look closely you see they only go to the apical surface, or the superficial layer of epidermis.
How do we really prove there are itch-specific fibres in humans? One particular experiment that can be done is to record the skin fibres and apply itchy compounds like histamine and see the itchy fibres fire action potential. At the same time humans can report what they feel whether it be itch or pain. After capsaicin injection they do feel pain, but they haven’t done those experiments applied capsaicin in the itch fibres and then ask what humans really feel. You need to find a way to deliver capsaicin to the itch-fibres only and not to the pain fibres. Potentially that can really prove the existence of itch-fibres in humans. I strongly believe that itch-fibres exist in humans but we do need an experiment to prove that.
Did your research shed any light on why chronic itchiness is sometimes caused by the suppression of chronic pain with morphine?
Our current study doesn’t have any implications on that question. People believe that morphine-induced itch is more associated with the central mechanism. When people are given morphine into the spinal cord to suppress chronic pain, the majority of them have an itch as a side effect. However, more recent work has shown that morphine applied to the skin can also induce an itch sensation.
There are some previous studies that suggest the one of the human Mrg receptors can be activated by morphine, so potentially the Mrg (the receptor we worked on) may be involved in morphine-induced itch but we don’t have any data currently to support that hypothesis. But the current study may facilitate further studies to show how morphine induces itch because by identifying those itch fibres we can really study that kind of mechanism.
Do you think your work will help in developing treatments for chronic itch, including itch caused by life-saving medications?
Yes it is possible. Right now we have a research project to see whether we can come up with any inhibitors for those receptors - whether we can block chronic itch. We have animal models and we also look for inhibitors for human receptors. So I think within a few years maybe we can have some candidates to test on animals and eventually in human trials.
Since we labelled the neurons now we know they only go into the skin that suggests we should only focus on topic applications of those drugs on the skin, not on oral routes.
Would you like to make any further comments?
I think it is a very exciting time as there has been a lot of progress in the itch research field. There are many labs other than ours. We used mouse genetics and molecular biology combined with better animals models now to study itch sensation that can lead to identification of many novel receptors and cell types and also novel itchy compounds. With these novel tools in hand we really can study more detailed mechanism for chronic itch.
Where can readers find more information?
They can find our paper here:
http://www.nature.com/neuro/journal/vaop/ncurrent/full/nn.3289.html
About Associate Professor Xinzhong Dong
Xinzhong Dong is an associate professor in the Department of Neuroscience at Johns Hopkins University School of Medicine and also an Early Career Scientists of Howard Hughes Medical Institute. He received his PhD degree from University of California Los Angeles and worked as a postdoctoral fellow at California Institute of Technology. In 2004, Dr. Dong started his own laboratory at Johns Hopkins University where they have been taken a multi-disciplinary approach including molecular biology, genetics, electrophysiology, imaging, behavioural tests, and biochemistry to study different types of somatosensation such as pain and itch.
His major contribution to the sensory biology field includes identification of molecular markers to define specific subtypes of sensory neurons, identification of novel itch receptors, itch mediators, and itch nerve fibers, and characterization of novel TRP channel modulators. He has won several awards and fellowships including Alfred P. Sloan Research Fellow and Young Investigator Award from Chinese Biological Investigator Society. His works are supported by funding from National Institute of Health, Howard Hughes Medical Institute, and other private foundations.
Mitochondrial fusion critical for adult neurogenesis and brain circuit refinement