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S. R. Pulivarthy, N. Tanaka; Photoentrainable Circadian Oscillator in Cultured Mammalian Cells. Invest. Ophthalmol. Vis. Sci. 2007;48(13):59.
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© ARVO (1962-2015); The Authors (2016-present)
Melanopsin expressed in the intrinsically photosensitive retinal ganglion cells (ipRGC) functions as the dominant photopigment for entrainment of the circadian oscillator to the daily light dark cycle. The ipRGCs transmit the photic information to entrain the master oscillator of the hypothalamic suprachiasmatic nucleus (SCN) which, in turn, appropriately resets the phase of peripheral tissue autonomous oscillators. To better understand the molecular mechanism of circadian entrainment we developed a cell based model by ectopic expression of melanopsin in immortalised fibroblasts that harbour a functional oscillator. Photoactivation of melanopsin with brief light pulse led to membrane depolarization, increased intracellular calcium levels and phosphorylation of CREB at serine 133. Maximum binding of phosphorylated CREB to the CRE box of Per promoter was detected within 30minutes of light pulse followed by transient induction of rhythmically expressed Period (Per) genes. Transient induction of Per genes ultimately resulted in change in phase of the oscillator. This approach allows for precise dose dependent, and non-invasive resetting of the oscillator. Intriguingly, the magnitude of transient Per induction, phase shift and amplitude of the oscillator following light pulse exhibited clear circadian modulation. Light triggered phase shifts and amplitude changes were inversely related, such that light pulse with minimum effect on phase exhibited robust amplitude of Per oscillation in the subsequent days and vice versa. Results from these experiments and a model for entrainment and oscillator synchronization will be discussed.
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