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ScienceWeek
NEUROSCIENCE: ON CIRCADIAN MECHANISMS IN DROSOPHILA
The following points are made by William J. Schwartz (Nature 2004 431:751):
1) Animals have an internal timekeeping mechanism that precisely regulates 24-hour (circadian) rhythms of body function and behavior, and synchronizes them to the day-night cycle. A constellation of "clock" genes lies at the core of this timepiece, and these genes interact in complex intracellular feedback loops to produce oscillations in their own expression(1). But how are such molecular cycles translated into the adaptable temporal programs that are characteristic of whole organisms? In fruitflies (Drosophila melanogaster), for example, how does a daily intracellular molecular oscillation drive a rhythm of rest and activity that is overtly bimodal, with pronounced bouts of activity around morning and evening that can anticipate the times of lights-on and lights-off? Recent work (2,3) has demonstrated that such behavior arises at an intercellular (tissue) level of organization, with discrete sets of clock-gene-expressing brain cells differentially involved in the response to dawn and dusk.
2) One well-known clock gene in fruitflies is period (per), which, in the brains of adult flies, is expressed in photoreceptor cells, glial (non-neuronal) cells and in a few clusters of neurons(4). These neuron clusters lie in specific areas of the brain: there are three groups of dorsal neurons (DN1, DN2 and DN3), and two groups of lateral neurons, on each side of the brain. One group of five to eight lateral neurons lies towards the top of the brain (that is, dorsolaterally; these are called LNd neurons). The other group, the LNv neurons, lies towards the bottom of the brain (ventrolaterally); it includes four to six large cells and five small cells. The LNv neurons (except for one of the five small cells) express the neurotransmitter molecule known as pigment-dispersing factor (PDF), whereas none of the LNd or dorsal neurons does.
3) Attention has focused on the LNv neurons as the essential circadian "pacemaker" cells that set fly activity rhythms, especially so because removing them leads to defective behavioral rhythmicity in constant darkness(5). But data from work on flies lacking PDF and on other mutants, as well as studies in which flies were engineered to express neuronal genes that block electrical activity or synaptic transmission, suggest that a multi-neuronal network is also somehow involved.
4) Stoleru et al(2) and Grima et al(3) sought to dissect this network, using mutant flies and genetic crosses to target specific genes to specific cells. Stoleru et al(2) succeeded in delivering a cell-death gene to the LNv or LNd neurons in flies, killing these cells. Tests of the flies' cycles of rest and activity showed that the insects' behavior was still rhythmic in a light-dark cycle -- but the rhythms were different. The LNv-lacking flies anticipated only lights-off in the evening, not lights-on in the morning. Meanwhile, the flies nominally lacking LNd anticipated lights-on but not lights-off. In constant darkness, the strains showed unimodal evening- or morning-phased rhythms, respectively (although rhythmicity could not be sustained in the LNv-less flies).
5) Grima et al(3) used a different, non-ablative strategy. They started with flies that were deficient in the gene per and therefore arrhythmic, and then forced the re-expression of per only in the LNv neurons, or in both the LNv and LNd neurons. LNv-restricted per expression rescued behavioral rhythmicity, but only lights-on was anticipated. Expression in the small LNv cells seemed to be sufficient for this, even in constant darkness. Lights-off was also anticipated when per was also expressed in about half of the LNd cells. Thus, two independent strategies have led to the same conclusion: morning and evening bouts of activity are differentially controlled by the LNv and LNd cells, respectively.
References (abridged):
1. Hall, J. C. Adv. Genet. 48, 1-280 (2003)
2. Stoleru, D., Peng, Y., Agosto, J. & Rosbash, M. Nature 431, 862-868 (2004)
3. Grima, B., ChÚlot, E., Xia, R. & Rouyer, F. Nature 431, 869-873 (2004)
4. Helfrich-Foerster, C. Microsc. Res. Tech. 62, 94-102 (2003)
5. Renn, S. C., Park, J. H., Rosbash, M., Hall, J. C. & Taghert, P. H. Cell 99, 791-802 (1999)
Nature http://www.nature.com/nature
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NEUROBIOLOGY: ON THE CIRCADIAN CLOCK IN DROSOPHILA
The following points are made by S. Tanoue et al (Current Biology 2004 14:638):
1) The Drosophila circadian clock is controlled by interlocked transcriptional feedback loops that operate in many neuronal and nonneuronal tissues. These clocks are roughly divided into a central clock, which resides in the brain and is known to control rhythms in locomotor activity, and peripheral clocks, which comprise all other clock tissues and are thought to control other rhythmic outputs. The authors previously demonstrated that peripheral oscillators are required to mediate rhythmic olfactory responses in the Drosophila antenna, but the identity and relative autonomy of these peripheral oscillators has not been defined.
2) Circadian oscillators are present in a variety of tissues throughout the head, thorax, and abdomen of Drosophila. Oscillator function in different tissues has been analyzed in more detail by using per-driven luciferase reporter genes, which have the advantage of monitoring rhythmic gene expression in live animals and cultured tissues. These studies demonstrated that oscillators in different tissues operate independently and that they are directly light entrainable. The autonomy of Drosophila oscillator function contrasts with the hierarchical nature of the mammalian circadian system, where light entrains a central oscillator that then entrains oscillators in peripheral tissues. Although circadian oscillators are present in many tissues from Drosophila, relatively little is known about the rhythms that they control.
3) The authors report that targeted ablation of lateral neurons by using apoptosis-promoting factors and targeted clock disruption in antennal neurons with newly developed dominant-negative versions of the transcription factors CLOCK and CYCLE demonstrate that antennal neurons, but not central clock cells, are necessary for olfactory rhythms. Targeted rescue of antennal neuron oscillators in cyc01 flies through wild-type CYCLE shows that these neurons are also sufficient for olfaction rhythms.
4) The authors conclude: Antennal neurons are both necessary and sufficient for olfaction rhythms, which demonstrates for the first time that a peripheral tissue can function as an autonomous pacemaker in Drosophila. These results reveal fundamental differences in the function and organization of circadian oscillators in Drosophila and mammals and suggest that components of the olfactory signal transduction cascade could be targets of circadian regulation.
References (abridged):
1. Glossop, N.R., Lyons, L.C., and Hardin, P.E. (1999). Interlocked feedback loops within the Drosophila circadian oscillator. Science 286, 766-768
2. Allada, R., White, N.E., So, W.V., Hall, J.C., and Rosbash, M. (1998). A mutant Drosophila homolog of mammalian Clock disrupts circadian rhythms and transcription of period and timeless. Cell 93, 791-804
3. Blau, J. and Young, M.W. (1999). Cycling vrille expression is required for a functional Drosophila clock. Cell 99, 661-671
4. Cyran, S.A., Buchsbaum, A.M., Reddy, K.L., Lin, M.C., Glossop, N.R., Hardin, P.E., Young, M.W., Storti, R.V., and Blau, J. (2003). vrille, Pdp1, and dClock form a second feedback loop in the Drosophila circadian clock. Cell 112, 329-341
5. Darlington, T.K., Wager-Smith, K., Ceriani, M.F., Staknis, D., Gekakis, N., Steeves, T.D., Weitz, C.J., Takahashi, J.S., and Kay, S.A. (1998). Closing the circadian loop: CLOCK-induced transcription of its own inhibitors per and tim. Science 280, 1599-1603
Current Biology http://www.current-biology.com
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ON CIRCADIAN RHYTHMS
Notes by ScienceWeek
In biology, a "circadian rhythm" is a daily cyclical process, be it biochemical, or physiological, or behavioral. The human sleep-wake cycle is the most familiar example. Circadian rhythms are often described in terms of endogenous "biological clocks", with the thrust of research to reduce some particular behavioral or physiological circadian rhythm to biochemical events. These clocks are usually set by environmental cues such as the light-dark cycle, and what is characteristic of an endogenous clock is that if one removes the environmental cue, keeps the organism in constant light, for example, the endogenous rhythm will continue, but will tend to drift out of phase with the outdoors environmental light-dark cycle. Restoring the external light-dark cue will reset the clock to its normal intrinsic rhythm.
The following points are made by G.M. Shepherd (citation below):
1) It might be thought that the obvious relations of many plant and animal activities to day and night would have invited close study even in ancient times. However, it appears that the facts were simply too familiar. It was not until 1729 that the French geologist J. de Mairan (1678-1771) did the simple experiment of placing a plant under constant temperature and illumination, and observed that its normal daily period of fluctuations still persisted. This showed that periodic behavior could be a function of the organism itself. Although this finding aroused some interest, people still did not quite know what to make of it; there were suspicions that the experiment might be affected by some undetectable rays or forces.
2) It was not until the 1930s that biologists began to make the connection between photoperiodicity (the changing illumination during the day) and bodily rhythms. A breakthrough came in 1950, in the work of two German biologists, Gustav Kramer and Karl von Frisch (1886-1982). Kramer showed that birds could use the sun as a compass by virtue of the fact that they have an "internal clock" which, in effect, tells them the time of day and how much to correct for the position (azimuth) of the sun in the sky. Von Frisch came to a similar conclusion for bees. A number of biologists then initiated the search for the cellular basis of the "internal clock", a search that has continued to the present. The importance of circadian rhythms has grown in parallel with the increasing understanding of their mechanisms, and the increasing realization of how pervasive they are in the life of the organism.
Adapted from: G.M. Shepherd: Neurobiology. 2nd Edition. Oxford University Press 1988, p.507.
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