Characterization of zPer3-luc rhythmicity in zebrafish

HERNANDEZ NE

ProQuest

Lugar: Ann Harbor, Michigan; Año: 2008 p. 112

9780549392552

The zebrafish is a unique vertebrate model for the study of circadian clocks, because cultured peripheral tissues are not only rhythmic, but are also directly entrainably by light. Furthermore, cell lines generated from early-stage zebrafish embryos contain clocks that are also directly light-responsive, demonstrating that zebrafish cells contain not only a circadian oscillator, but also the photoreceptive apparatus sufficient for light entrainment. Herein, I characterize the oscillator properties in various peripheral tissues and brain regions from the period3-luciferase (per3-luc) transgenic zebrafish. Tissue-dependent differences were found in free-running period, response to light, and temperature compensation. Rhythms of per3-luc can free run in both constant dark (DD) and constant light (LL) with similar amplitudes and periods. I further characterized the oscillator properties at the cellular level by establishing a bioluminescent cell preparation derived from 24 h post-fertilization transgenic zebrafish embryos that carry the luciferase gene driven by the zper3 promoter (ZP3L cells). I used this novel reporter system to identify candidate photopigments that may be involved in the photoentrainment. Real-time monitoring of per3-luc circadian rhythms revealed that this cell system displays the essential oscillator properties observed in peripheral tissues and brain regions,  such as sustained rhythmicity in DD and LL conditions, resetting circadian phase by brief light pulses, and entrainment to an oscillatory light-dark cycle. The action spectrum for light-induced resetting of the zper3-luc cell clock is consistent with the involvement of at least one cryptochrome photopigment and a more long wavelength-sensitive photopigments.