Peripheral clocks in Drosophila

For several decades, biorhythm researchers in the two main animal model system, the fruitfly and the mouse have believed that behavioural rhythms are determined exclusively by central brain clock neurons. These neurons act as pacemakers and can generate the 24 hour locomotor cycles that are so important for allowing the animal to interact with its environment at the optimum time of day. In the fly, the pacemaker cells are called the sLNvs and in the mouse, the SCN. In the fly, the peripheral nervous system and non-neural organs have independent clocks, whereas in the mammal, the SCN synchronises the rhythms in the periphery via a number of signalling factors. Recently we have observed that clocks in the antennae or the eyes, which lie outside the central brain of the fruitfly, can nevertheless generate behavioural rhythms even though the central clock neurons are made genetically arrhythmic. This means there are pacemaker cells in these two peripheral tissues that can mediate rhythmic behaviour, so it is not all about the sLNvs. However, the clock cells in the antennae need the central brain network to be intact, even though it is clockless, in order for these behavioural rhythms to be expressed. These results will come as something of a shock to fly clock researchers as it overturns current dogma.

We shall carefully examine the antennae and identify the clock cells that act as behavioural pacemakers. We know that these cells are involved in temperature, wind and gravity sensing as well as hearing, but is it particular subsets of these or is it all of them ? The antennae as also full of smell receptors, so we shall ask whether these too can act as pacemaker cells. We shall manipulate these cells genetically in a number of ways, for example by stopping their clocks, hyperexciting these cells, or speeding them up or slowing them down to see what happens to the rhythmic behaviour. We will also examine how 'normal' these rhythms are, do they respond to light and temperature in the usual ways that clocks do, or are they abnormal? We shall also find out how the antennal pacemaker cells might connect to the central brain circuit that is required for rhythms to be expressed. We will extend our studies to the eye in a similar way and finally we shall ask whether non-neural tissue, the fly's liver called the fat body, can also drive behavioural rhythms. While this might seem unlikely, given the surprising results that we have already obtained, we take nothing for granted.

The fruitfly circadian pacemaker that mediates rhythmic locomotor behaviour under constant darkness is located in the central brain, specifically the sLNv neurons that express the neuropeptide PDF. Other clock neurons within the brain also contribute to the 24 hour period and the robustness of the rhythm. Recently, we have observed that locomotor rhythms can be generated in genetically clockless flies (per0) by expressing PER only in the antennae. Surgical removal of the antennae restores the arrhythmic behavioural phenotype. These antenally driven rhythms are PDF but not CRY dependent, so CRY does not work as a clock component in this peripheral tissue. However, the antennal clock needs the intact, but clockless central brain PDF circuit to mediate behavioural rhythmicity. Furthermore, in wild-type flies, driving dbtS or dbtL in the antennae shortens or lengthens the locomotor period respectively, suggesting that this antennal clock contributes to normal behavioural rhythmicity. A similar situation exists when PER is driven only in the eye. These results are as remarkable as they are unexpected. Clearly our sLNv-centric view of the pacemaker needs significant revision.

We shall perform an anatomical study of clock gene expression in the antennae and using available Gal4 drivers reveal whether it is the mechanoceptors themselves that act as clocks. We shall discover which type of mechanoreceptors are mediating the behavioural clock and we shall further genetically manipulate the clock and the excitability of these receptors to see what effects they have on behavioural rhythms. We shall extend this analysis to olfactory receptors on the antennae and consolidate our preliminary results in a similar manner with the eye clock. We will attempt to identify the neuronal connections between the antennae and the clock circuit. Finally we shall ask whether a non neural peripheral organ, the fat body, can generate behavioural rhythms

There is considerable public anxiety about what jet-lag or more importantly, chronic shift-work imposes on human and animal welfare. Recent work has revealed how jet lagging the periphery in mammals, eg the pancreas, can result in arrhythmic insulin signalling and the onset of diabetes. The fly provides an almost perfect model for mammalian rhythms so it is highly relevant for understanding normal rhythmic behaviour and physiology. Our work will certainly interest the medical profession, the pharmaceutical industry who are always on the look out for neuronal and peripheral targets to combat jet-lag and shift work problems, and entomologists who use biological methods to control pests. With respect to the latter we have a collaboration with Oxitec in Oxford and BioFly in Israel who are part of our Marie Curie ITN to study clocks in model insects as well as pests. Some of our work on cryptochrome, the circadian photoreceptor for example, has recently interested BioFly, who traditionally have used wavelengths of light for mass rearing billions of insects that are inappropriate for entraining the insect's circadian clock. A small change in their rearing practises may have enormous commercial repercussions and they are considering making the relevant changes. Consequently our fly research has and will continue to have impact in applied entomology. Circadian clock research that was initiated in flies and extended in mammals has also convinced policy makers in Oxfordshire to start the school day a little later for 30,000 teenagers, because on puberty, the adolescent clock delays by about an hour. Our work and what it might stimulate in mammalian research is relevant for reviewing the peripheral-central axis of the clock that has become a dogma in our field. Our results, when consolidated and extended to mammals may lead to the enhancement of healthier circadian environments. Finally our work has impact for the general public who are always interested in biorhythms and body clocks as it is a subject to which they can easily relate.
While policymakers might take note of our results, empowering the public by educating them on the benefits of natural circadian entrainment, allows them to modify their own lifestyle accordingly. Perhaps this is the greatest immediate tangible benefit we can provide through public communication. The main vehicle to reach and engage our beneficiaries will be GENIE (Genetics, Education, Networking, Innovation and Excellence) a national CETL (Centre of Excellence in Teaching and Learning) that is part of our Genetics Department. GENIE is significantly engaged in outreach, and within its website is a Virtual Genetics Education Centre for schools and colleges, higher education centres, the general public, as well as health professionals and policymakers. The Genie website has more than 22,000 hits per month, so it is a major vehicle for scientific dissemination to the general public. Moreover, GENIE is proactive in getting in touch with different sectors of society through courses, seminars and workshops organised all year round; this will assure ample visibility for our research. In the past year for example we have contributed a number of body clock outreach events.