An official website of the United States government
Sleep is a fundamental feature of life for virtually all multicellular animals, but many questions remain about how sleep is regulated by circadian rhythms, homeostatic sleep drive that builds up with wakefulness, and modifying factors such as hunger or social interactions, as well as about the biological functions of sleep. Substantial headway has been made in the study of both circadian rhythms and sleep in the fruit flyDrosophila melanogaster, much of it through studies of individual fly activity usingDrosophilaactivity monitors (DAMs). Here, we describe approaches for the activation of specific neurons of interest using optogenetics (involving genetic modifications that allow for light-based neuronal activation) and thermogenetics (involving genetic modifications that allow for temperature-based neuronal activation) so that researchers can evaluate the roles of those neurons in controlling rest and activity behavior. In this protocol, we describe how to set up a rig for simultaneous optogenetic or thermogenetic stimulation and activity monitoring for analysis of sleep and circadian rhythms inDrosophila, how to raise appropriate flies, and how to perform the experiment. This protocol will allow researchers to assess the causative role in the regulation of sleep and activity rhythms of any genetically tractable subset of cells.
more »
« less
Analysis of Sleep and Circadian Rhythms from Drosophila Activity-Monitoring Data Using SCAMP
Sleep is a fundamental feature of life for virtually all multicellular animals, but many questions remain about how sleep is regulated and what biological functions it plays. Substantial headway has been made in the study of both circadian rhythms and sleep in the fruit flyDrosophila melanogaster, much of it through studies of individual fly activity using beam break counts fromDrosophilaactivity monitors (DAMs). The number of laboratories worldwide studying sleep inDrosophilahas grown from only a few 20 years ago to hundreds today. The utility of these studies is limited by the quality of the metrics that can be extracted from the data. Many software options exist to help analyze DAM data; however, these are often expensive or have significant limitations. Therefore, we describe here a method for analyzing DAM-based data using the sleep and circadian analysis MATLAB program (SCAMP). This user-friendly software has an advantage of combining several analyses of both sleep and circadian rhythms in one package and produces graphical outputs as well as spreadsheets of the outputs for further statistical analysis. The version of SCAMP described here is also the first published software package that can analyze data from multibeam DAM5Ms, enabling determination of positional preference over time.
more »
« less
- Award ID(s):
- 2042873
- PAR ID:
- 10539785
- Publisher / Repository:
- Cold Spring Harbor Laboratory Press
- Date Published:
- Journal Name:
- Cold Spring Harbor Protocols
- ISSN:
- 1940-3402
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
More Like this
-
-
The circadian system coordinates multiple behavioral outputs to ensure proper temporal organization. Timing information underlying circadian regulation of behavior depends on a molecular circadian clock that operates within clock neurons in the brain. In Drosophila and other organisms, clock neurons can be divided into several molecularly and functionally discrete subpopulations that form an interconnected central clock network. It is unknown how circadian signals are coherently generated by the clock network and transmitted across output circuits that connect clock cells to downstream neurons that regulate behavior. Here, we have exhaustively investigated the contribution of clock neuron subsets to the control of two prominent behavioral outputs in Drosophila: locomotor activity and feeding. We have used cell-specific manipulations to eliminate molecular clock function or induce electrical silencing either broadly throughout the clock network or in specific subpopulations. We find that clock cell manipulations produce similar changes in locomotor activity and feeding, suggesting that overlapping central clock circuitry regulates these distinct behavioral outputs. Interestingly, the magnitude and nature of the effects depend on the clock subset targeted. Lateral clock neuron manipulations profoundly degrade the rhythmicity of feeding and activity. In contrast, dorsal clock neuron manipulations only subtly affect rhythmicity but produce pronounced changes in the distribution of activity and feeding across the day. These experiments expand our knowledge of clock regulation of activity rhythms and offer the first extensive characterization of central clock control of feeding rhythms. Despite similar effects of central clock cell disruptions on activity and feeding, we find that manipulations that prevent functional signaling in an identified output circuit preferentially degrade locomotor activity rhythms, leaving feeding rhythms relatively intact. This demonstrates that activity and feeding are indeed dissociable behaviors, and furthermore suggests that differential circadian control of these behaviors diverges in output circuits downstream of the clock network.more » « less
-
Abstract Honey bees are critical pollinators in ecosystems and agriculture, but their numbers have significantly declined. Declines in pollinator populations are thought to be due to multiple factors including habitat loss, climate change, increased vulnerability to disease and parasites, and pesticide use. Neonicotinoid pesticides are agonists of insect nicotinic cholinergic receptors, and sub-lethal exposures are linked to reduced honey bee hive survival. Honey bees are highly dependent on circadian clocks to regulate critical behaviors, such as foraging orientation and navigation, time-memory for food sources, sleep, and learning/memory processes. Because circadian clock neurons in insects receive light input through cholinergic signaling we tested for effects of neonicotinoids on honey bee circadian rhythms and sleep. Neonicotinoid ingestion by feeding over several days results in neonicotinoid accumulation in the bee brain, disrupts circadian rhythmicity in many individual bees, shifts the timing of behavioral circadian rhythms in bees that remain rhythmic, and impairs sleep. Neonicotinoids and light input act synergistically to disrupt bee circadian behavior, and neonicotinoids directly stimulate wake-promoting clock neurons in the fruit fly brain. Neonicotinoids disrupt honey bee circadian rhythms and sleep, likely by aberrant stimulation of clock neurons, to potentially impair honey bee navigation, time-memory, and social communication.more » « less
-
ABSTRACT Seasonal changes in sleep/wake cycles and behaviors related to reproduction often co‐occur with seasonal fluctuations in sex hormones. Experimental studies have established that fluctuations in circulating testosterone mediate circadian rhythms. However, most studies are performed under constant lighting conditions and fail to investigate the effects of testosterone on the phenotypic output of circadian rhythms, that is, chronotype (daily activity patterns under light:dark cycles). Here, we experimentally elevated testosterone with implants during short nonbreeding daylengths in male house sparrows (Passer domesticus) to test if observed seasonal changes in chronotype are directly in response to photoperiod or to testosterone. We fitted individuals with accelerometers to track activity across treatment periods. Birds experienced three treatments periods: short day photoperiods before manipulation (SD), followed by testosterone implants while still on short days (SD + T). Implants were then removed. After a decrease in cloacal protuberance size, an indicator of low testosterone levels, birds were then photostimulated on long days (LD). Blood samples were collected at night, when testosterone peaks, to compare testosterone levels to daily onset/offset activity for experimental periods. Our results indicate that experimentally elevated testosterone under short nonbreeding photoperiods significantly advanced daily onset of activity and total daily activity relative to daylength. This suggests that testosterone, independent of photoperiod, is responsible for seasonal shifts in chronotypes and daily activity rhythms. These findings suggest that sex steroid hormone actions regulate timing of daily behaviors, likely coordinating expression of reproductive behaviors to appropriate times of the day.more » « less
-
Sleep is important for survival, and the need for sleep is conserved across species. In the past two decades, the fruit flyDrosophila melanogasterhas emerged as a promising system in which to study the genetic, neural, and physiological bases of sleep. Through significant advances in our understanding of the regulation of sleep in flies, the field is poised to address several open questions about sleep, such as how the need for sleep is encoded, how molecular regulators of sleep are situated within brain networks, and what the functions of sleep are. Here, we describe key findings, open questions, and commonly used methods that have been used to inform existing theories and develop new ways of thinking about the function, regulation, and adaptability of sleep behavior.more » « less
- Free Publicly Accessible Full Text
-
Accepted Manuscript1.0
- Journal Article:
- https://doi.org/10.1101/pdb.prot108182
