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Christiana Katti, Rachel Butler, Sumathi Sekaran; Connexin Expression in the Mouse Retina is Controlled by Light and the Circadian Clock. Invest. Ophthalmol. Vis. Sci. 2012;53(14):2719.
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© ARVO (1962-2015); The Authors (2016-present)
The dynamic regulation of gap junctional coupling between retinal neurons is one mechanism by which the eye adapts to changes in ambient lighting levels. Connexins are the transmembrane proteins that form gap junctions which enable electrical coupling in the retina. Phosphorylation of connexins has been identified as a mechanism by which coupling is modulated. We investigated whether retinal connexins are also regulated at the translational and transcriptional levels by assessing Cx36 protein and transcript expression during the diurnal and circadian cycle.
Wildtype C57BL6/FVB/NCR mice were kept in a controlled 12:12 hr light/dark cycle or in complete darkness (for a maximum of 24 hrs). Retinae were isolated and flash frozen or eyes were fixed in 4% paraformaldehyde. Cx36 protein levels were analyzed using western blotting. The Cx36 antibody (Invitrogen) recognised 2 bands in western blots corresponding to the unphosphorylated (36 kDa) and phosphorylated (38 kDa) forms of the protein. Cx36 distribution was assessed by immunohistochemistry in retinal sections. Cx36 transcript levels were quantified using qPCR. Transcript levels were also quantified in retinally degenerate C3Hrd/rd retinae.
In a normal diurnal cycle, the phosphorylated form of the Cx36 protein was rhythmic peaking in the late night phase. Surprisingly we also observed a similar rhythm in unphosphorylated Cx36 protein levels. In the circadian cycle, the phosphorylated Cx36 rhythm was absent but the unphosphorylated Cx36 rhythm was maintained. Using immunohistochemistry in retinal sections, rhythmic Cx36 expression was localised to the outer plexiform layer but not the inner plexiform layer. Cx36 transcript levels were also found to be rhythmic in the diurnal and circadian cycles. Interestingly both transcript rhythms were abolished in C3Hrd/rd mice in which the outer retina is degenerated.
In addition to post-translational control, Cx36 is regulated at the translational and transcriptional level by light and the circadian clock. These rhythms are dependent on a functional outer retina.
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