Endurance exercise modifies the circadian clock in zebrafish (Danio rerio) temperature independently M. Egg, A. Tischler, T. Schwerte, A. Sandbichler, C. Folterbauer and B. Pelster Institut fu ¨r Zoologie, Universita ¨t Innsbruck, Technikerstr, Austria Received 15 September 2011, revision requested 13 October 2011, revision received 24 October 2011, accepted 31 October 2011 Correspondence: Dr. Margit Egg, Institut fu ¨r Zoologie, Leopold- Franzens-Universita ¨t Innsbruck, Technikerstr.25, A-6020 Innsbruck, Austria. E-mail: margit.egg@uibk.ac.at Abstract Aim: Several rodent and human studies revealed that physical exercise acts as a non-photic zeitgeber for the circadian clock. The intrinsic entraining mechanism is still unknown, although it was assumed that the exercise- mediated increase in core temperature could be the underlying zeitgeber. As the homoeostatic control of mammalian core temperature interferes strongly with the investigation of this hypothesis, the present study used the poiki- lotherm zebrafish to answer this question. Methods: Gene transcription levels of the two circadian core clock genes period1 and clock1 were quantified using real-time qPCR of whole animal zebrafish larvae. Results: Long-term endurance exercise of zebrafish larvae aged 9–15 days post-fertilization (dpf) or 21–32 dpf at a constant water temperature of 25 °C caused significantly altered transcription levels of the circadian genes period1 and clock1. Cosinor analysis of diurnal transcription profiles obtained after 3 days of swim training revealed significant differences regarding acrophase, mesor and amplitude of period1, resulting in a phase delay of the gene oscillation. After termination of the exercise bout, at 15 dpf, oscillation amplitudes of both circadian genes were significantly reduced. Conclusion: The results showed that physical exercise is able to affect the transcription of circadian genes in developing zebrafish larvae. Considering the poikilothermy of zebrafish, an exercise-mediated change in body core temperature could be excluded as the underlying intrinsic zeitgeber. How- ever, the day-active zebrafish arises as a useful model to address the syn- chronizing effect of exercise on the circadian clock. Keywords circadian clock, gene transcription, temperature, training, zebrafish, zeitgeber. Endogenous circadian timing systems of physiological, metabolic and behavioural processes are known from all light-dependent organisms, from bacteria to humans. Synchronization of these internal ‘body clocks’ to the environmental day and night cycles is achieved through various zeitgebers, the strongest of them being light. Other non-photic stimuli such as temperature, food intake, social cues, oxygen availability or physical activity have been widely discussed, but often they are not yet properly defined. The entrainment of the circadian clock by activity has initially been shown more than two decades ago. Thus, hamsters running on a wheel during the middle of their normal rest period re-adjusted to a new light– dark (LD) cycle twice as fast as non-exercised control animals (Mrosovsky & Salmon 1987). In mice, the circadian clock was shown to be synchronized to voluntary exercise, which could only be performed for certain hours during a day. Magnitude and direction of the observed phase shifts depended on the time of day, at which the exercise was performed (Edgar & Dement 1991). Not only the time of day but also the intensity and the duration of exercise appeared to have differ- ential effects on the circadian pacemaker of mice Acta Physiol 2012, 205, 167–176 Ó 2011 The Authors Acta Physiologica Ó 2011 Scandinavian Physiological Society, doi: 10.1111/j.1748-1716.2011.02382.x 167