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Richard A Stone, Wenjie Wei, Shanta Sarfare, Wei Pan, Karl C Engelhart, Tejvir S. Khurana, Maureen G. Maguire, P. Michael Iuvone, Debora L. Nickla; Diurnal Cycling of Clock and Melanopsin Genes in Chick Choroid. Invest. Ophthalmol. Vis. Sci. 2018;59(9):5048. doi: https://doi.org/.
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© ARVO (1962-2015); The Authors (2016-present)
The choroid undergoes diurnal thickness oscillations in chick and mammals, including humans. These fluctuations seem to have a role in regulating refractive development; other roles for them are unknown. Addressing the potential control of these fluctuations, we assayed the gene expression levels in the choroid for five clock genes (ARNTL, CLOCK, CRY1, NPAS2, PER3), for the photopigment melanopsin (OPN4m) and for one melatonin receptor (MTNR1a), using retina/RPE from the same eyes for comparisons. Previously, we found altered expression of most of these genes in the retina/RPE of chick eyes with experimental myopia.
Chicks with intact visual input bilaterally (n=8/time) and under a 12hr light:dark cycle (4100K fluorescent light, ~300 lux in cage) were euthanized at 0, 4, 8, 12, 16 or 20 hours zeitgeber time (ZT), with lights on ZT 0. Choroid and retina/RPE were dissected from each eye and snap-frozen without pooling; RNA was extracted with Purelink RNA mini kit, and cDNA was synthesized with Superscript SS IV VILO mastermix (Life Technologies). Using TaqMan® Gene Expression Assays with GAPDH and between-plate controls, relative expression levels of all genes were measured. General linear models (GEE approach), correlations and post-hoc tests assessed this complex data set.
Relative gene expression levels in choroid were lower than those in retina. MTNR1a gene expression was evident only in retina. For both choroid and retina, expression levels of OPN4m and clock genes varied significantly over time for each gene, often in cyclical patterns. Choroidal expression levels of OPN4m were highest at ZT 8; PER3, at ZT 0; and the other clock genes, around ZT 8-12. Comparing the choroid to the retina for each of the genes, the patterns of diurnal gene expressions tended to be similar though not precisely identical.
Melanopsin expression in the chicken choroid suggests that some choroidal cells, possibly trigeminal sensory nerve fibers or other choroidal neurons, are intrinsically photoreceptive. As in retina, the expression levels of clock and melanopsin genes in choroid undergo diurnal oscillations, indicative of autonomous circadian clocks. The roles of the diurnal changes of clock and melanopsin genes in daily choroidal thickness oscillations, refractive development and other aspects of ocular physiology require further study.
This is an abstract that was submitted for the 2018 ARVO Annual Meeting, held in Honolulu, Hawaii, April 29 - May 3, 2018.
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