A thermogenic circuit that maintains sensitivity to leptin in obesity

We are experiencing an epidemic in the prevalence of obesity; a disease which develops after the body's protective systems become overwhelmed in an environment which is full of sugary and fatty foods. Normally when we put weight on, our fat tissues release a hormone called leptin, that travels to the brain to help us to reduce eating, but also to increase the amount of energy we burn off by a process called thermogenesis (literally, heat production). One of the reasons we find it difficult to lose the extra weight is because the parts of the brain that control eating, become resistant to the effects of leptin, so it no longer works. We have learned a lot about the part of the brain controlling eating, because we have identified the types of nerve cells there which possess the capability to respond to leptin (i.e. those that have receptors for the hormone). Importantly, a different part of the brain controls the thermogenic response to leptin, and the cells here do not become resistant to leptin. This makes them of great interest to scientists and doctors wishing to find way of reducing the obesity problem. Unfortunately, until now we did not know what types of cells these were.

In our preliminary work leading up to this project, we have now identified three different cell types which all appear to play a role in thermogenesis. However, only one of them has leptin receptors. The cells that are directly sensitive to leptin contain a signaling chemical called PrRP. Our hypothesis is that the PrRP-containing nerves control the other cell types and also the messages from the brain to the organs that actually cause thermogenesis. We have bred different types of mice which will allow us to study the different cell types very carefully. We will be able to see how they respond to leptin, how they make connections, and how they communicate with each other. Perhaps most excitingly, for the first time ever, we can switch on or off the different nerves selectively, just by giving the mice a harmless drug. Thus, we can switch on the nerves to drive thermogenesis or switch the nerves off to stop them responding to leptin.

A complete understanding of the different types of nerves will allow the selective targeting of either the nerves themselves or their connections with the organs that control thermogenesis. This knowledge will help the discovery of drugs which, one day, could help prevent the development of obesity.

Leptin acts on its receptor (Lepr) in the brain to reduce food intake and increase energy expenditure. If positive energy balance continues, the circuits regulating food intake become resistant to leptin, leading to obesity. However, the dorsomedial hypothalamus (DMH) maintains its sensitivity, and can still increase energy expenditure by activating thermogenic brown adipose tissue (BAT) via the sympathetic nervous system. This makes the DMH an attractive target for pharmacological manipulation. However, a precise functional description of the DMH has lagged behind that of other hypothalamic nuclei for which the resident neurones have been identified and subsequently targeted. Although NPY and orexin neurones in the DMH have been implicated in thermogenesis, they do not possess Lepr. Instead, we have identified a population which contains Lepr and prolactin-releasing peptide (gene Prrh). Prrh-cre-Leprflox/flox mice, which lack Lepr only in DMH PrRP neurones, cannot respond to leptin's thermogenic signal and are obese.

Electrophysiology in transgenic reporter mice, will allow recording from the three cell types to determine how they respond to leptin and which, if any, project to premotor sympathetic neurones in the brainstem raphe pallidus (RPa), the major relay from the DMH to BAT. The connections of each neurone will be visualised by retrograde tracing from the RPa and by filling the neurones with adeno-associated viruses (AAVs) encoding fluorescent labels. These will include a novel AAV in which GFP is tagged to synaptophysin, to allow identification of terminal fields. We will use the latest "Designer Receptor Exclusively Activated by Designer Drug" technology to show a definitive role for DMH neurones in regulating leptin-induced thermogenesis. More broadly, the circuitry and the functions of the DMH in physiology and behavior are poorly understood, and this application will address a key gap in our knowledge of the hypothalamic regulation of energy balance.

The increase in the incidence of obesity has reached epidemic proportions. Current estimates are that there are 2 billion overweight people globally. 600 million of these are clinically obese. Without effective therapeutic interventions for the treatment of associated co-morbidities (type 2 diabetes, cardiovascular disease, lipodystrophy, etc.), rising care costs could bankrupt the National Health Service and other health providers worldwide. Due to stabilising feedback and the development of hormonal "resistance", attempts to reduce energy intake either through dieting or pharmacological intervention have proved difficult or even dangerous. Importantly, the brain does not appear to become resistant to the modulation of energy expenditure. All the major academic groups and pharmaceutical companies have active programs to understand how the brain induces thermogenesis (and hence increases energy expenditure). Our identification of the specific cell type in the brain, which mediates these effects, is a major step forward, and this new information will be disseminated at international conferences and by publication in high-impact journals. The work has already led to new collaborations which have involved sharing of genetically-engineered viruses and mice. Following publication, each of the mouse models we develop will be made available.

A conservative commercial estimate of the annual market opportunity for anti-obesity drugs is $60 bn. This project will guide future development of drugs for body weight regulation and provide a sound knowledge environment to understand the potential side effects of other drugs which, for example, target peripheral organs that feed back onto the brain. The applicant has been involved previously in successful collaborative projects with a number of industrial partners, providing evidence for several novel targets for drug development. Before the end of the project, we will be in a position to approach companies which may be interested in making mimetic drugs, which could provide composition of matter filings on novel therapeutics comprising long-lasting peptide derivatives.

During the lifetime of the grant, the basic research will be discussed at meetings organised by the Child Health Research Network, the Diabetes and Obesity Research Network and the Association for the Study of Obesity. These annual meetings are forums for basic researchers, psychologists, clinicians, community nurses and other health professionals, patient group representatives and policy makers. Outreach work will be encouraged at all levels within the laboratory. Over the three years, the applicant will lecture at two local schools and at a local Café Scientifique-type meeting. The PDRA will be strongly encouraged to follow the example set by previous lab members, to tutor for the Manchester Access and STEM programmes (aimed at helping under-privileged children into further education), and to complete both a Wellcome Trust Researchers in Residence Scheme and a UK GRADschool.

This grant will provide strong training in both in vivo skills and specialist techniques in electrophysiology and metabolic research. In the last twelve years, the applicant has supervised thirteen PhD students, nineteen masters students and eleven PDRAs, all of whom remained in science (some have their own independent research groups and others have moved into the commercial sector). The PI directs a cross-University IMB initiative to promote and expand research and training in in vivo biology. This problematic area is crucial to the European economy and to the ambitions of Manchester to be a world-leading university. He is program director for the MRes in Integrative Biology at Manchester (which will benefit from three MRes projects on work derived from this grant) and external examiner on Integrative Biology courses at the Universities of Liverpool and Edinburgh.