Melatonin synthesis in retina: circadian regulation of arylalkylamine N-acetyltransferase activity in cultured photoreceptor cells of embryonic chicken retina

May 6, 2003Brain research

Daily rhythm controls melatonin production enzyme in light-sensing cells of developing chicken eye

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Abstract

AANAT activity in cultured retinal cells from 8-day-old chicken embryos displays circadian rhythms in both light/dark and continuous darkness conditions.

In retinal cells from 6-day-old chicken embryos, AANAT activity fluctuated prominently under a light/dark cycle but did not exhibit a daily pattern when transferred to continuous darkness.
AANAT activity in cells from 8-day-old embryos demonstrated circadian control, showing rhythmic patterns in both light/dark and continuous darkness conditions.
After 8 days of incubation in light/dark cycles followed by 2 days in darkness, AANAT activity in E6 cells became rhythmic, suggesting maturation over time.
AANAT activity was acutely suppressed by a light pulse during the subjective night, indicating sensitivity to light similar to in vivo conditions.
The ability to regulate AANAT activity according to circadian rhythms appears to develop intrinsically in retinal cells as they mature.

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The key regulatory enzyme in melatonin synthesis is arylalkylamine N-acetyltransferase (AANAT). In vivo, AANAT activity in chicken retinal photoreceptor cells exhibits a circadian rhythm that peaks at night. The purpose of the present study was to investigate the temporal development of light/dark and circadian oscillations of AANAT activity in cultured retinal cells prepared from 6- and 8-day-old chicken embryos (E6, E8, respectively). Photoreceptor cells prepared from E6 retinas and incubated under a 14-h light/10-h dark (LD) cycle of illumination for 5-7 days displayed prominent daily fluctuations in AANAT activity on days 5 and 6 in vitro. However, when E6 cells, incubated for 5 days under LD, were transferred to continuous (24 h/day) darkness (DD) on day 6, no daily pattern of activity was observed. This result indicates that AANAT fluctuations were light-driven and not circadian at this stage. In contrast, cells prepared from E8 embryos and incubated under conditions identical to those for E6 cells displayed prominent rhythms of AANAT activity in both LD and DD, indicative of circadian control. To determine if circadian control of AANAT activity would develop in E6 cells incubated for a longer period of time to allow maturation, cells were incubated for 8 days in LD followed by 2 days in DD. AANAT activity in these cells was rhythmic in both LD and DD. In cells incubated in this manner, a 2-h light pulse in the middle of the subjective night suppressed AANAT activity, indicating that the enzyme activity in the cultured cells is acutely suppressed by light, as it is in vivo. These results indicate that the ability to express circadian regulation of AANAT activity is an intrinsic property of retinal cells that can develop in vitro. Development of light-dark regulation of AANAT activity appears to precede the circadian clock-control of enzyme activity.

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Melatonin synthesis in retina: circadian regulation of arylalkylamine N-acetyltransferase activity in cultured photoreceptor cells of embryonic chicken retina

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