Monitoring circadian time in rat plasma using a secreted Cypridina luciferase reporter

Abstract

A firefly luciferase reporter enabled us to monitor promoter activity in vivo as well as ex vivo; however, this requires a sufficient supply of the substrate luciferin and specific monitoring devices. To overcome these disadvantages, we developed transgenic rats carrying a secreted enzyme Cypridina luciferase (CLuc) reporter under the promoter of clock gene Per2 (Per2–CLuc). Per2–CLuc activity in serially sampled blood from freely moving rats exhibited robust circadian rhythms with a peak at early morning. The Per2–CLuc bioluminescence could be quantified even with approximately 100pl of plasma. Plasma Per2–CLuc rhythms were phase reversed, and the level was reduced by restricting food access for 2h during the light phase, suggesting that the plasma Per2–CLuc rhythms reflect the phase of peripheral clocks entrained to feeding cues as well as fuel metabolism. Fasting for 2days did not alter the circadian Per2–CLuc rhythms in rats, suggesting that feeding per se did not affect the circadian Per2–CLuc rhythms. Tissue-specific Per2–CLuc rhythms were observed in culture medium of peripheral tissues. The Per2–CLuc reporter is a powerful tool to access gene expression in vivo as well as ex vivo with ordinary laboratory equipment.