Neuronal circuits for itch in the spinal dorsal horn

Chronic itch is a distressing condition that affects over 20% of the population. It is a feature of many diseases, including conditions affecting skin, kidneys and blood, as well as some types of cancer and HIV infection. It is also a side-effect of certain drugs, such as morphine. For most patients with chronic itch there are no satisfactory treatments, and a quarter of these patients suffer itch for more than 5 years. Despite its clinical importance, we have only a limited knowledge about the nerve circuits in the spinal cord and brain that are responsible for itch. Sensory information enters the spinal cord and is processed through complex circuits involving different types of local nerve cells (interneurons), before being carried to the brain by a type of nerve cell called a projection neuron. Many different types of spinal cord interneuron are involved in itch and pain pathways. Some of these (excitatory interneurons) transmit information through these circuits, while others (inhibitory interneurons) block the passage of information. Painful stimuli can powerfully suppress itch, which is why scratching provides temporary relief. We have identified a type of inhibitory interneuron in the spinal cord that reduces itch, and shown that these cells are responsible for the relief of itch by scratching. These inhibitory cells contain a peptide (a short protein) called dynorphin. Two different types of excitatory interneuron are thought to be part of the itch pathway. One of these contains a different peptide (gastrin-releasing peptide, or GRP) and the other possesses the receptor on which GRP acts (the GRP receptor, or GRPR).
Recent studies have suggested a nerve circuit for transmitting itch, in which sensory fibres from the skin activate GRP cells, which then excite GRPR cells. Information from these is transmitted to the brain via itch-selective projection neurons. We have provided evidence that the inhibitory dynorphin cells reduce itch by blocking transmission at the level of the GRPR cells. In this project, we will test several components of the proposed circuit. We have already found that activating dynorphin cells reduces itch, but it is important to show that reducing their activity increases itch. We will also test whether their inhibitory action is selective for itch, or whether it also reduces pain. The GRP cells are known to receive input from one type of itch-sensitive nerve fibre. However, we recently identified a new class of nerve fibre that is likely to respond to different itch-inducing stimuli, and we will see whether these also target the GRP cells. The GRPR cells are already implicated in itch, because destroying them reduces itching without affecting pain. However, little is known about these cells. Our initial studies suggest that they correspond to a type of excitatory interneuron, known as vertical cells, which provide input to projection neurons, and we will carry out further studies to confirm this. We will determine the types of sensory input they receive, and whether they respond selectively to itchy (but not painful) stimuli. Most projection neurons in this region of the spinal cord carry pain signals, with a subset being activated by itchy stimuli. We will therefore see whether the GRPR cells only target the itch-activated projection neurons. We will also test the prediction that dynorphin cells block itch by inhibiting the GRPR cells, and we will identify the chemical messengers that they use. Finally, we will determine whether the GRPR cells are indeed critical and selective for itch, by increasing or decreasing their activity and testing the effects on itch and pain behaviour.
The project will provide important new information about nerve circuits in the spinal cord that are involved in itch. Taken together with emerging information about the drug receptors that are present on different types of nerve cell, it should assist in the development of new treatments for chronic itch.

Chronic itch (pruritus) is a distressing symptom of many diseases, and is often difficult to treat. However, despite its importance, relatively little is known about the spinal cord circuits that underlie itch. We have shown activating dynorphin-expressing inhibitory interneurons in laminae I-II reduces itch behaviour, and that these cells are involved in the suppression of itch by counterstimuli. In this project we will investigate the roles of 3 different classes of interneurons in spinal itch pathways. We will test the prediction that inactivating the dynorphin interneurons leads to increased itch, and we will determine whether these cells are also involved in gating pain. Excitatory interneurons that express gastrin-releasing peptide (GRP) are thought to be a key component of spinal itch circuits, and are known to receive synaptic input from pruriceptors that express MrgA3. We have recently identified somatostatin-containing afferents as a second type of pruriceptor, and we will test whether these also innervate the GRP cells. The receptor for GRP (GRPR) is expressed by excitatory interneurons in laminae I-II, and ablation of these cells selectively blocks itch. Our preliminary observations indicate that the GRPR neurons include vertical cells, which are known to innervate projection neurons in lamina I. We will further characterise the GRPR cells, by examining their responses to pruritic stimuli, testing whether they selectively innervate pruritogen-activated projection neurons, and determining whether they are the site at which dynorphin cells act to suppress itch. Finally, we will determine whether GRPR cells are critical and selective for itch, by chemogenetically activating/inactivating them, and testing whether this increases/decreases itch (but not pain) behaviour. The project will provide valuable information about spinal cord circuits that are involved in itch, and should assist in the development of new antipruritic drugs.

Itch is a distressing symptom of many diseases and can occur in otherwise healthy individuals. Chronic pruritus (itching that lasts for more than 6 weeks) is very common, for example a lifetime prevalence of 22% was reported in a large sample of the working age population in Germany, with a quarter of those affected having suffered from itching that had lasted for at least 5 years. Chronic pruritus has a detrimental effect on sleep, leading to reduced ability to work. Because of the lack of adequate treatment options for many types of itch, it represents a major unmet clinical need. Recent research has begun to shed light on the types of spinal cord neuron that are involved in itch mechanisms, but there is still considerable uncertainty concerning the neuronal circuits involved, and also the extent to which itch and pain are processed through "labelled lines". Further insight into the organisation of neuronal circuitry in the superficial dorsal horn is also needed to improve our understanding of pain mechanisms. It has been estimated that 20% of adults in Europe suffer from chronic pain of moderate to severe intensity, and this proportion is likely to increase as the population ages. Chronic pain is also often poorly treated, due to the lack of suitable medications.

Who will benefit from this research?
Those who will benefit directly include scientists from other disciplines (e.g. pharmacology, molecular genetics, developmental biology) working on the somatosensory system, as well as the pharmaceutical industry, in particular in relation to the development of new antipruritic and analgesic drugs. In the longer term, beneficiaries include human patients suffering from chronic pruritus and pain, and the clinicians responsible for their treatment. This work should therefore have an impact on health and well-being. Improved treatments for itch and pain would be of considerable economic benefit to society due to the reduction in time lost from work.

How will they benefit from this research?
Improved understanding of the roles of different interneuron populations in somatosensory dorsal horn circuits will facilitate advances in other disciplines, for example by providing a framework for defining the location and function of receptors, ion channels and transcription factors within these circuits. Several laboratories are currently undertaking transcriptomic analyses of dorsal horn neuron populations, and since this project will identify distinct populations of interneurons that are involved in itch and pain mechanisms, our results can be combined with this emerging transcriptomic data to search for potential drug targets that are selectively expressed in itch and pain pathways. This understanding will therefore be of great benefit to the pharmaceutical industry in its search for new antipruritic and analgesic drugs. The project will also provide training in in vivo experimental approaches, which have been identified as a priority for the pharmaceutical industry.
In addition, greater knowledge of dorsal horn neuronal circuits is important for our understanding of the changes that can occur in chronic pain states. Work from our laboratory has already generated important insights into neuropathic pain, by demonstrating that several previously proposed mechanisms (e.g. dorsal sprouting of tactile afferents, death of inhibitory interneurons in the dorsal horn) are not necessary for the development of chronic pain after peripheral nerve injury, thus directing research away from these areas.
The intersectional genetic targeting strategies that we develop will be important for those working in various CNS regions, because it is increasingly apparent that neuronal populations cannot generally be defined (and therefore targeted) based on the expression of single genes.