Functional Development of the Zebrafish Pineal Gland: Light‐Induced Expression of Period2 is Required for Onset of the Circadian Clock

Abstract

In zebrafish, the pineal gland is a photoreceptive organ that contains an intrinsic circadian oscillator and exhibits rhythmic arylalkylamine-N-acetyltransferase (zfaanat2) mRNA expression. In the present study, we investigated the role of light and of a clock gene, zperiod2 (zper2), in the development of this rhythm. Analysis of zfaanat2 mRNA expression in the pineal gland of 3-day-old zebrafish embryos after exposure to different photoperiodic regimes indicated that light is required for proper development of the circadian clock-controlled rhythmic expression of zfaanat2, and that a 1-h light pulse is sufficient to initiate this rhythm. Analysis of zper2 mRNA expression in zebrafish embryos exposed to different photoperiodic regimes indicated that zper2 expression is transiently up-regulated by light but is not regulated by the circadian oscillator. To establish the association between light-induced zper2 expression and light-induced clock-controlled zfaanat2 rhythm, zPer2 knock-down experiments were performed. The zfaanat2 mRNA rhythm, induced by a 1-h light pulse, was abolished in zPer2 knock-down embryos. These experiments indicated that light-induced zper2 expression is crucial for establishment of the clock-controlled zfaanat2 rhythm in the zebrafish pineal gland.

Novel functions for Period 3 and Exo-rhodopsin in rhythmic transcription and melatonin biosynthesis within the zebrafish pineal organ

“…For instance, expression of aanat2 is reduced by 58-68% when embryos are moved to constant darkness shortly after fertilization [64]. Here, we provide some of the first evidence that Exorh is involved in this light response in developing embryos.…”

“…Previous work demonstrates that expression of exorh and aanat2 in the pineal does not initiate normally when embryos are raised in constant darkness [60,64]. For instance, expression of aanat2 is reduced by 58-68% when embryos are moved to constant darkness shortly after fertilization [64].…”

“…Previous work on the aanat2 regulation by Yoav Gothilf and colleagues gives some insight into how this might occur. The light-dependent onset of aanat2 transcription in the pineal requires Per2 [63,64]. Transcription of the per2 gene itself is induced by light [12,63,64].…”

“…The early pineal-specific expression of Exorh makes it an excellent candidate to function as a major light-sensing photopigment in the developing pineal [19,64]. Towards testing the function of Exorh, we designed a MO that binds to the exon 1/intron 1 junction of the exorh genomic sequence (exorh sp MO).…”

“…In zebrafish, pineal photoreceptors have been implicated in several responses to light, including entrainment of the circadian clock, triggering the onset of pineal rhythms during development, and mediating acute suppression of melatonin by a light pulse during the dark period [11,60,[63][64][65]. The pineal opsin protein Exorh is expressed in pineal photoreceptors from early embryogenesis to adulthood and is thus an excellent candidate to be mediating these light responses [5,38,60].…”