4.4 Article

Single-cell Resolution Fluorescence Live Imaging of Drosophila Circadian Clocks in Larval Brain Culture

期刊

出版社

JOURNAL OF VISUALIZED EXPERIMENTS
DOI: 10.3791/57015

关键词

Neuroscience; Issue 131; Neuroscience; circadian rhythms; neuronal communication; time-lapse imaging; ex vivo brain culture; fluorescent reporter; Drosophila

资金

  1. JST PRESTO program
  2. Swiss National Science Foundation [31003A_149893, 31003A_169548]
  3. European Research Council [ERC-StG-311194]
  4. Novartis Foundation for Medical-Biomedical Research [13A39]
  5. University of Geneva
  6. Swiss National Science Foundation (SNF) [31003A_169548, 31003A_149893] Funding Source: Swiss National Science Foundation (SNF)

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The circadian pacemaker circuit orchestrates rhythmic behavioral and physiological outputs coordinated with environmental cues, such as day/night cycles. The molecular clock within each pacemaker neuron generates circadian rhythms in gene expression, which underlie the rhythmic neuronal functions essential to the operation of the circuit. Investigation of the properties of the individual molecular oscillators in different subclasses of pacemaker neurons and their interaction with neuronal signaling yields a better understanding of the circadian pacemaker circuit. Here, we present a time-lapse fluorescent microscopy approach developed to monitor the molecular clockwork in clock neurons of cultured Drosophila larval brain. This method allows the multi-day recording of the rhythms of genetically encoded fluorescent circadian reporters at single-cell resolution. This setup can be combined with pharmacological manipulations to closely analyze real-time response of the molecular clock to various compounds. Beyond circadian rhythms, this multipurpose method in combination with powerful Drosophila genetic techniques offers the possibility to study diverse neuronal or molecular processes in live brain tissue.

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