Abstract
ABSTRACTAnalysis of circadian rhythm behavior alteration in fish for toxicity assessment usually requires expensive commercial equipment and laborious and complicated tweaking. Here, we report a simple setup that consists of a custom-made light box equipped with white and 940 nm light-emitting diode (LED) light strips as light sources, where the locomotion activities of zebrafish or catfish are captured using an infrared-sensitive coupled charged device (CCD). The whole setup was housed in a temperature-controlled incubator to isolate external noise and to maintain consistent experimental conditions. The video recording and light triggering were synchronized using Total Recorder, a recording scheduling software. By using the setup mentioned above and open source software such as ImageJ or idTracker, the locomotion activities of diurnal (e.g. zebrafish) and nocturnal (e.g. catfish) fish during day and night cycles can be quantitatively analyzed. We used simple image-based methods and a cost-effective instrument to assess the circadian rhythm of multiple fish species, as well as other parameters such as age, ambient temperature and chemical toxicology with high precision and reproducibility. In conclusion, the instrument setting and analysis methods established in this study provide a reliable and easy entry point for toxicity assessment on circadian rhythm dysregulation in fish.
Highlights
Circadian rhythms are based on intracellular time-tracking systems that play a central role in adapting the physiology and behavior of living organisms to anticipate daily environmental changes (Lahiri et al, 2005)
Measurement of the circadian rhythm in zebrafish and catfish To explore the potential utility of these two protocols, we validated the method by using diurnal and nocturnal fish species with distinct locomotion activities during the light and dark cycles
Zebrafish locomotor activity intensified during the light periods, showing that zebrafish are diurnal (Kasai et al, 2009). For both fish species with distinct circadian rhythms, we cross-examined the locomotion activity curves acquired by idTracker- and ImageJ-based methods
Summary
Circadian rhythms are based on intracellular time-tracking systems that play a central role in adapting the physiology and behavior of living organisms to anticipate daily environmental changes (Lahiri et al, 2005). Circadian rhythms are driven by endogenous circadian clocks that are comprised of. Circadian clocks involve clock genes that interact to produce a molecular oscillator adjusting output clock-controlled genes. Some clock genes have been identified in vertebrates and most of the studies on the molecular mechanisms of circadian rhythm generation have come from research of clock genes that were originally identified by mutations, which altered or abolished circadian rhythms (Delaunay et al, 2000). Only a handful of circadian genes have been discovered in vertebrates
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