title: Light-dark and circadian melatonin rhythms are established de novo in re-aggregates of the embryonic chicken retina.
abstract: A central function of chicken photoreceptors is the rhythmic synthesis of the neurohormone melatonin; however, it is unclear how they can achieve--already before hatching (i.e. without light exposure)--a circadian pacemaker system. Here we studied melatonin synthesis and secretion in rosetted spheroids, which are de novo histotypic spheres, re-aggregated from dissociated retinal cells of chicken embryos. Spheroids were cultured during a 12-hour light/12-hour dark cycle. Shortly after the onset of photoreceptor differentiation in spheroids near day 5 in culture, melatonin secretion set in and, by day 8 in culture, showed distinct dark-light oscillations, with high values during the dark and lower ones during the light period. Concomitantly, expression of two key enzymes of melatonin synthesis, arylalkylamine-N-acetyltransferase (AA-NAT) and hydroxyindole-O-methyltransferase (HIOMT) was detected by northern blot analysis from day 6 onwards, increasing until day 8. Only HIOMT, but not AA-NAT levels were 2- to 3-fold higher at the end of the light compared with the dark phase. Even under permanent darkness, a slight melatonin oscillation was revealed after 6 days. In conclusion, photoreceptors in de novo regenerated retinal spheroids become light-responsive, establish appropriate pathways for melatonin synthesis and secrete more melatonin in the dark. The underlying circadian pacemaker seems to depend on photoreceptor differentiation, not on previous light-dark entrainment.
authors.affiliation: Institut für Zoologie, Technische Universität Darmstadt, Darmstadt, Germany. ewillbold@aol.com
authors.forename: Elmar
authors.initials: E
authors.lastname: Willbold
authors.forename: Jutta
authors.initials: J
authors.lastname: Huhn
authors.forename: Horst-Werner
authors.initials: HW
authors.lastname: Korf
authors.forename: Pierre
authors.initials: P
authors.lastname: Voisin
authors.forename: Paul G
authors.initials: PG
authors.lastname: Layer
mesh_terms: D000121:Acetylserotonin O-Methyltransferase / Q000096:biosynthesis; D000818:Animals; D001191:Arylamine N-Acetyltransferase / Q000096:biosynthesis; D015152:Blotting, Northern; D002454:Cell Differentiation / Q000502:physiology; D002478:Cells, Cultured; D002642:Chick Embryo; D002940:Circadian Rhythm / Q000502:physiology*; D008550:Melatonin / Q000378:metabolism*; D012160:Retina / Q000196:embryology / Q000502:physiology*; D018874:Spheroids, Cellular
chemical_list: D000121:Acetylserotonin O-Methyltransferase; D001191:Arylamine N-Acetyltransferase; D008550:Melatonin