Purchase this article with an account.
Xiaoqin Liu, Zhijing Zhang, Christophe Ribelayga; Immunocytochemical Analysis of Circadian Clock Protein Expression in Mouse Photoreceptors. Invest. Ophthalmol. Vis. Sci. 2011;52(14):3456.
Download citation file:
© ARVO (1962-2015); The Authors (2016-present)
The vertebrate retina contains autonomous circadian clocks that regulate many aspects of its physiology and function. The core machinery of circadian clocks is a self-sustained cell-based mechanism that relies on a specific set of clock genes and their protein products interlocked in recurrent feedback loops. Although the presence of a clock that controls the rhythmic synthesis of melatonin in mammalian photoreceptors has been conclusively demonstrated, clock gene mRNA detection requires amplification, and the expression of clock proteins has not been established. In addition, it is not known whether both types of photoreceptor, namely rods and cones, contain a clock mechanism. Here we used an immunocytochemical approach to examine clock protein expression in rod and cone photoreceptors.
CBA/CaJ mice were sacrificed at the end of the day of a 12 h light/12 h dark cycle (ZT11). Sections of paraformaldehyde-fixed retinas were processed with specific antibodies against PERIOD (PER) 1 (PER1), PER2, PER3, CLOCK, BAML1, CRY1 and CRY2 (all from Chemicon) and double labeled with a cone arrestin antibody (LUMIj; a gift from Dr. C.M. Craft, USC).
Intense CLOCK, BMAL1, PER1 and PER2 antibody labeling were found in the somata of cones. In contrast, only weak BMAL1, PER1 and PER2 antibody labeling were detected in the somata of rods. Interestingly, PER2, PER3 and BMAL1 antibody labeling were intense in photoreceptor terminals. No CRY1 or CRY2 immunosignal was detected in the outer retina, even though many amacrine cells and ganglion cells exhibited CRY immunoreactivity.
The results indicate that 1) only a few clock proteins are expressed in photoreceptor cells; and 2) clock protein expression is much stronger in cone somata than in rod somata. The strong expression of bona fide circadian clock components in cone somata suggests that cones are more likely than rods to contain a functional circadian clock. The accumulation of BMAL1 and PER3 in photoreceptor terminals is surprising but may be linked to the influence of the clock on the structure of photoreceptor synaptic ribbons.
This PDF is available to Subscribers Only