Our ‘inner clock’ functions in every cell, but is orchestrated by a master clock in the brain. Maintenance of this system is critical for our well-being; disturbances often lead to psychiatric and other disorders.
Although fruit flies and mice are traditionally used in circadian rhythm studies, zebrafish are becoming more and more popular amongst chronobiologists as a model organism. Zebrafish boast many advantages, including an early-developing, light-entrained circadian system. Like humans, zebrafish are a diurnal species.
Zebrafish pineal gland and light entrainment
In vertebrates, the pineal gland hormone melatonin plays a crucial role in the inner clock system. Produced at night, it regulates daily rhythms, but also annual rhythms such as seasonal changes in reproduction and energy balance. The activity of melatonin is modulated by external photic signals and, in mammals, driven by the suprachiasmatic nucleus (SCN) in the hypothalamus.
In fish, scientists have found no SCN or any structure homologous to it. The fish pineal gland, however, seems to have all the elements for the generation of circadian rhythmicity. The Yoav Gothilf Lab (Tel Aviv University, Israel) has done some very in-depth research on the mechanism behind the light entrainment of the circadian system in zebrafish.
Casein kinase and rhythmicity
For example, Sima Smadja Storz and her colleagues at this lab investigated the role of casein kinase 1 (CK1), which are enzymes that function as regulators of circadian rhythmicity (research published in PLoS ONE, 2013). They studied both cell cultures and zebrafish at different developmental stages. To determine rhythmicity, the researchers looked at the activity patterns of zebrafish larvae in 48-well plates in DanioVision. Before the experiment, larvae were kept under 12h light/12h dim light conditions. The effects of specific CK1 inhibitors were measured by comparing the affected larvae’s total distance moved during a 10 minute dim light experiment to the activity of a control group.
Zebrafish larvae normally develop circadian rhythmicity when exposed to light/dark cycles, after which the animals are able to maintain this rhythmicity, even under continuous light conditions. In larvae that were exposed to the CK1δ inhibition, rhythmicity was completely diminished. Specific CK1ε inhibition did not have as significant an effect. Read more about this research here.
More recently, Dr. Zohar Ben-Moshe from the Yoav Gothilf Lab identified 14 light-entrained molecular clock genes in the zebrafish system using messenger RNA sequencing and microarray analysis. In this study (Nucleic Acids Research, 2014), genes dec1 and per2 were tested further and found to be essential for the light entrainment of the rhythmic locomotor activity of larvae.
The activity of larvae (dec1 knockdown, per2 knockdown, and control groups) was monitored in a DanioVision system. To entrain rhythms of locomotor activity, 5dpf larvae were exposed to 3 hours of light, after which they were monitored under dim light conditions. The total distance moved per 10 minute time-bin was automatically measured with EthoVision XT and used to determine the activity level of the larvae. Knockdown of both dec1 and per2 genes resulted in a noticeably disrupted circadian locomotor activity pattern.
Want to see more studies that used DanioVision? Click here for a list of recent publications.