Drosophila exhibits a circadian rest-activity cycle, but it is not known whether fly rest constitutes sleep or is mere inactivity. It is shown here that, like mammalian sleep, rest inDrosophila is characterized by an increased arousal threshold and is homeostatically regulated independently of the circadian clock. As in mammals, rest is abundant in young flies, is reduced in older flies, and is modulated by stimulants and hypnotics. Several molecular markers modulated by sleep and waking in mammals are modulated by rest and activity in Drosophila, including cytochrome oxidase C, the endoplasmic reticulum chaperone protein BiP, and enzymes implicated in the catabolism of monoamines. Flies lacking one such enzyme, arylalkylamine N-acetyltransferase, show increased rest after rest deprivation. These results implicate the catabolism of monoamines in the regulation of sleep and waking in the fly and suggest that Drosophila may serve as a model system for the genetic dissection of sleep.
REFERENCES AND NOTES
Campbell S. S., Tobler I., Neurosci. Biobehav. Rev. 8, 269 (1984);
Zepelin H., Rechtschaffen A., Brain Behav. Evol. 10, 425 (1984);
Rechtschaffen A., Perspect. Biol. Med. 41, 359 (1998).
Tobler I., Behav. Brain Res. 8, 351 (1983);
___, Stalder J., J. Comp. Physiol. A 163, 227 (1988) ;
Tobler I., Neuner-Jehle M., J. Sleep Res. 1, 231 (1992);
Kaiser W., Steiner-Kaiser J., Nature 301, 707 (1983);
Kaiser W., J. Comp. Physiol. A 163, 565 (1988).
Cirelli C., Tononi G., Mol. Brain Res. 56, 293 (1998);
; C. Cirelli, P. J. Shaw, G. Tononi, Sleep 22 (suppl.), 113 (1999).
Flies were cultured at 25°C, 50 to 60% humidity, 12 hour:12 hour light/dark cycle, on yeast, dark corn syrup, and agar food. We obtained per01 flies from J. C. Hall (Brandeis University) and Datlo and Df(2R)Px1/In(2LR)SM5, al2 Cy ltv sn2 sp2 flies from the Bloomington Drosophila Stock Center. For details about the ultrasound monitoring system, see Science Online (www.sciencemag.org/feature/data/1047207.shl).
Five behaviors were visually scored in 2-s bins by an observer blind to the output of the ultrasound system on 18 independent trials for a total of 8 hours during the light period. The correspondence rates were as follows: locomoting, 99%; inactive, 97%; grooming anterior limbs, 94%; grooming posterior limbs, 98%; and eating, 97%.
Rest was defined as uninterrupted behavioral quiescence lasting for at least 5 min.
Drosophila Activity Monitoring System (Trikinetics) [
Hamblen M., et al., J. Neurogenet. 3, 249 (1986);
]. The system was validated by visual observation for 17.75 hours (n = 7). Flies were awake but did not cross the infrared beam in 5 of 213 bins (miss rate = 2.35%).
For procedures for arousal thresholds, procedures for automated rest deprivation, and additional controls used to validate the infrared system, see Science Online (www.sciencemag.org/feature/data/1047207.shl).
Mistlberger R. E., Bergmann B. M., Waldenar W., Rechtschaffen A., Sleep 6, 217 (1983);
Tobler I., Borbely A. A., Groos G., Neurosci. Lett. 42, 49 (1983);
Edgar D. M., Dement W. C., Fuller C. A., J. Neurosci. 13, 1065 (1993) .
Stone W. S., Clin. Geriatr. Med. 5, 363 (1989);
Dijk D.-J., Duffy J. F., Riel E., Shanahan T. L., Czeisler C. A., J. Physiol. 516, 611 (1999).
Yanik G., Glaum S., Radulovacki M., Brain Res. 403, 177 (1987).
Methods were as in (3), with modifications: 0.5 μg of pooled total RNA (n = 20) was reverse-transcribed (two independent pools per condition). Polymerase chain reactions were performed in duplicate for each pool (104 primer combinations). For RPA, 1 to 2 μg of total RNA from pooled fly heads (n = 60) was used. The amount of sample RNA was normalized using a riboprobe specific for ribosomal protein rp49.
The behavioral state was determined individually for each fly; only flies that satisfied specific criteria were selected for analysis. A fly was considered awake if it was active for at least 90% of the 3-hour light period and 100% of the hour before killing. A fly was resting if it was inactive for at least 66% of the 3-hour dark period and 100% of the hour before killing. Only about 60 to 70% of the flies examined satisfied these criteria. Failure to specifically identify rest and waking results in samples containing a mixture of behavioral states.
The sequence matched a Drosophila P1 clone (AC005554). Analysis using Genescan indicated that the proposed peptide has a 49% homology with rat Fas.
In situ hybridization was performed as described [
Aronstein K., Auld V., Ffrench-Constant R., Invert. Neurosci. 2, 115 (1996);
]. Sense riboprobes gave no specific hybridization.
Yehuda S., et al., Peptides 19, 407 (1998).
Dunkov B. C., Rodriguez-Arnaiz R., Pittendrigh B., Ffrench-Constant R. H., Feyereisen R., Mol. Gen. Genet. 251, 290 (1996).
Kuhl D., Kennedy T. E., Barzilai A., Kandel E., J. Cell Biol. 119, 1069 (1992);
Rubin D. M., et al., Gene 128, 155 (1993).
Brodbeck D., et al., DNA Cell Biol. 17, 621 (1998).
McGinty D. J., Harper R. M., Brain Res. 101, 569 (1976);
Aston-Jones G., Bloom F. E., J. Neurosci. 1, 876 (1981).
E. Hartmann, Functions of Sleep (Yale Univ. Press, New Haven, CT, 1973);
Siegel J. M., Rogawksi M. A., Brain Res. Rev. 13, 213 (1988).
C. B. Bridges, Cytologia Fujii Jubil., 745 (1937).
Hendricks J., et al., Neuron 25, 129 (2000).
We thank D. F. Robinson, G. A. Davis, M. J. Gallina, J. M. Salbaum, J. Snook, N. Almassy, and E. Balaban for his conception of the ultrasound system. The Neurosciences Institute is supported by the Neurosciences Research Foundation and receives major support for this program from Novartis. C.C. was a Joseph Drown Foundation Fellow.
Volume 287 | Issue 5459
10 March 2000
10 March 2000
Received: 15 November 1999
Accepted: 8 February 2000
Published in print: 10 March 2000
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