Figure 1.
Genes that showed an up- or down-regulation.
Genes up-regulated of at least 1.6 fold at (A) 3 hours and (B) 6 hours of light exposure. The genes that showed an up-regulation in the three replicas are listed and the only gene that appears in both lists is Dio2. Genes that showed a down-regulation less than 0.7 at (C) 3 hours and (D) 6 hours of exposure to light. The only gene that showed a down-regulation at both timings was Adra1b. Each circle represents one replica. The numbers in each set represent genes that are identified, predicted genes or transcribed locus. In the list, only the identified ones are mentioned.
Figure 2.
Expression of Dio2 and Dio3 using real time PCR.
(A) Expression of Dio2 showing an up-regulation due to the time of the day (compared to Dark) and as response to light (Circadian Rhythm and Intense Light). (B) Expression of Dio3 showing a down regulation mainly due to light. Data are reported as mean ± S.E.M (N = 6).
Figure 3.
Western blot corresponding to DIO2 and DIO3 showing an agreement with real time PCR results.
(A) Representative immunoblot corresponding to DIO2 and DIO3. Although DIO2 shows non-specific bands, a protein band of estimated molecular weight is clearly present in rats exposed to light and absent in dark. MW: Molecular Weight. *:band corresponding to DIO2. (B) Histograms of densitometric measurements of the western blots (N = 3) All band intensities of each protein were compared separately with that of β-actin. The specificity of both antibodies can be observed.
Figure 4.
Immunofluorescence of adult rat retina in dark and light conditions (3 hours of steady illumination).
(A) In blue for the nuclei (DAPI) and in green for DIO2. (OS photoreceptor outer segment; IS inner segment; ONL outer nuclear layer; OPL outer plexiform layer; INL inner nuclear layer; IPL inner plexiform layer; GC ganglion cells). Scale bar = 10μm. (B) Inner nuclear layer nucleus in blue, do not colocalize with DIO2 in green, in the right side the y–z plane, and in the lower left the x–z plane. Scale bar = 5 µm.
Figure 5.
Expression of Opn1mw, Opn1sw, Sag, Gcap1, Ccng1, Cpt1a.
Dark columns show the data of rats kept in dark overnight until 7 am (0 ZT). White columns show data after 12 hours of ambient light (12 ZT). Data reported as mean ± S.E.M (N = 3).
Table 1.
Primers used for the real time PCR.