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S Hattar, HW Liao, M Takao, S Carlson, R Richardson, DM Berson, KW Yau; Possible Role for Melanopsin in Photoentrainment of the Circadian Clock in Rodents . Invest. Ophthalmol. Vis. Sci. 2002;43(13):1364.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: It is known that the circadian clock in the suprachiasmatic nucleus (SCN) can be photoentrained by light signals from the eyes via the retinohypothalamic tract. Previous results by others have suggested that rods and cones are not required for photoentrainment and that other retinal neurons may act as circadian photoreceptors. We would like to address the question whether melanopsin, a recently identified photopigment-like protein in the retina, plays a role in this process. Methods: Peptide antibodies (Covance, Denver, PA) were raised against the N-terminus of melanopsin and affinity-purified. Also, a tau-lacZ construct was targeted to the mouse melanopsin gene locus by homologous recombination. X-gal was then used for labeling melanopsin-positive cells in tau-lacZ +/-, melanopsin +/- animals. Rat ganglion cells innervating the SCN were labeled by retrograde transport of fluorescent beads, recorded by whole-cell current clamp in isolated retinas, and filled with Lucifer Yellow. Intrinsic photosensitivity of these cells was confirmed by persistence of their light responses during bath application of 2mM Co2+. Results: Antibody- and X-gal-labelings both demonstrated that a very small subset of retinal ganglion cells (RGCs) contain melanopsin. Immunocytochemistry shows that the protein is present in the cell body, the dendrites and at least the initial segments of the axons of the RGCs. X-gal labeling shows that these RGCs project to the SCN as well as the intergeniculate leaflet (IGL), another region thought to be involved in photoentrainment. RGCs retrograde-labeled from the SCN exhibited robust light responses in the presence of 2mM Co2+ and were invariably melanopsin immunopositive (n=9). Control RGCs lacking Co2+-insensitive light responses lacked melanopsin immunostaining (n=4). Conclusion: These results indicate that melanopsin is present in the intrinsically photosensitive RGCs that project to the SCN, and it is probably the non-rod/non-cone photopigment responsible for photoentrainment.
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