Seven distinct cDNAs encoding functional subunits of the AMPA/kainate-type glutamate receptors have been recently cloned. This in situ hybridization study was done to determine which subunits are expressed in the retina and, where possible, which neurons express them. Hybridization of 35S-UTP-labeled cRNA probes with transverse sections revealed that mRNAs for all seven receptor subunits (GluR1-GluR7) are expressed in both cat and rat retinas. GluR1 and GluR2 produced labeling over the entire inner nuclear layer (INL) and ganglion cell layer (GCL). GluR3-GluR7 have more limited distributions, indicative of expression by only a subset of neurons. All of the subunits are expressed by the cells at the inner edge of the INL, where amacrine cells reside, yet the layers with the horizontal, bipolar, and ganglion cells contain different subsets of subunits. These findings suggest that these glutamate receptor subunits are employed at many of the retinal synapses, including the photoreceptor input to the outer plexiform layer and the bipolar cell's contacts with the processes at the INL. It is also possible that some glial cells in the INL express some of the subunits. Since different combinations of GluR1-GluR3 have been shown to play an important role in the calcium permeability in response to glutamate, we investigated whether single cells coexpressed those subunits. By hybridizing adjacent semithin (1 micron) sections of the cat retina with probes for GluR1-GluR3, it was possible to observe coexpression of all three subunits, or of pairs of these subunits, in cells within the INL and GCL.
GT protected the retina against glutamate toxicity via an antioxidant mechanism. These findings reveal a novel mechanism by which GT protects the retina against neurodegeneration in disorders such as diabetic retinopathy.
In birds, displaced ganglion cells (DGCs) constitute the exclusive source of retinal input to the nucleus of the basal optic root (nBOR) of the accessory optic system. Tyrosine-hydroxylase (TH) immunoreactivity was examined in the pigeon retina after injections of rhodamine-labeled microspheres into the nBOR. A population of about 400 DGCs was observed in each case to exhibit both TH immunoreactivity and rhodamine bead fluorescence. This corresponded to about 10-15% of the total number of identified DGCs in each retina. Double-labeled cells were medium- to large-size (12 to 20 microns in the largest axis) and were always located at the border between the inner nuclear and the inner plexiform layers. Their dendrites could be followed horizontally in lamina 1 of the inner plexiform layer for up to 300 microns from the cell body. The distribution of double-labeled DGCs appeared to be mostly peripheral, matching the overall distribution of identified DGCs. Larger DGCs (21-28 microns) were never seen to contain TH immunoreactivity. Examination of brain sections revealed plexuses of thin varicose TH-positive axons in all subdivisions of the nBOR. Unilateral enucleation produced an almost complete elimination of TH immunoreactivity in the contralateral nucleus. Such results suggest the existence of a population of catecholaminergic DGCs projecting into the accessory optic system of the pigeon. They also support the emerging hypothesis concerning the neurotransmitter heterogeneity of ganglion cells in the vertebrate retina.
Gap junction channels formed by connexins (Cx) may play essential roles in some processes that occur during retinal development, such as apoptosis and calcium wave spread. The present study was undertaken to determine the distribution pattern of Cx36, Cx43, and Cx45 by immunofluorescence, as well as their gene expression levels by quantitative PCR during postnatal development of the mouse retina. Our results showed an increased expression of neuronal Cx36 from P1 until P10, when this Cx reached adult levels, and it was mainly distributed in the outer and inner plexiform layers. In turn, Cx43 was almost absent in retinal progenitor cells at P1, it became more prominent in glial cell processes about P10, and did not change until adulthood. Double-labeling studies in situ and in vitro with antivimentin, a Müller cell marker, confirmed that Cx43 was expressed by these cells. In addition, quantitative PCR showed that Cx43 and vimentin shared very similar temporal expression patterns. Finally, in contrast to Cx36 and Cx43, Cx45 mRNA was strongly down-regulated during development. In early postnatal days, Cx45 was seen ubiquitously distributed throughout the retina in cells undergoing proliferation and differentiation, as well in differentiated neurons. In adult retina, this protein had a more restricted distribution both in neurons and glial cells, as confirmed in situ and in vitro. In conclusion, we observed a distinct temporal expression pattern for Cx36, Cx43, and Cx45, which is probably related to particular roles in retinal function and maintenance of homeostasis during development of the mouse retina.
In the retina, ambient light levels influence the cell coupling provided by gap junction (GJ) channels, to compensate the visual function for various lighting conditions. However, the effects of ambient light levels on expression of connexins (Cx), the proteins that form the GJ channels, are poorly understood. In the present study, we first determined whether gene expression of specific Cx (Cx26, Cx31.1, Cx36, Cx37, Cx40, Cx43, Cx45, Cx50, and Cx57) was affected by prolonged dark adaptation. Cx mRNA relative levels were determined in mouse retinas dark adapted for 3 hr, 1 day, and 7 days by using quantitative real-time PCR. Transcript levels of some Cx were repressed after 3 hr (Cx57), 1 day (Cx45), or 7 days (Cx36 and Cx43) of dark adaptation; others were increased after 1 day (Cx50) or 7 days (Cx31.1 and Cx37); and two of them (Cx26 and Cx40) were not significantly altered. The second aim was to determine whether prolonged dark adaptation affects protein expression of two important Cx in retina: neuronal Cx36 and glial Cx43. We were able to demonstrate that important changes in protein distribution and expression also took place in retina during long-term dark adaptation. Given their localization, the specific alterations in Cx expression may reflect their distinct response to ambient light levels.
Electrical coupling provided by connexins (Cx) in gap junctions (GJ) plays important roles in both the developing and the mature retina. In mammalian nocturnal species, Cx36 is an essential component in the rod pathway, the retinal circuit specialized for night, scotopic vision. Here, we report the expression of Cx36 in a species (Gallus gallus) that phylogenetic development endows with an essentially rodless retina. Cx36 gene is very highly expressed in comparison with other Cxs previously described in the adult retina, such as Cx43, Cx45, and Cx50. Moreover, real-time PCR, Western blot, and immunofluorescence all revealed that Cx36 expression massively increased over time during development. We thoroughly examined Cx36 in the inner and outer plexiform layers, where this protein was particularly abundant. Cx36 was observed mainly in the off sublamina of the inner plexiform layer rather than in the on sublamina previously described in the mammalian retina. In addition, Cx36 colocalized with specific cell markers, revealing the expression of this protein in distinct amacrine cells. To investigate further the involvement of Cx36 in visual processing, we examined its functional regulation in retinas from dark-adapted animals. Light deprivation markedly up-regulates Cx36 gene expression in the retina, resulting in an increased accumulation of the protein within and between cone synaptic terminals. In summary, the developmental regulation of Cx36 expression results in particular circuitry-related roles in the chick retina. Moreover, this study demonstrated that Cx36 onto- and phylogenesis in the vertebrate retina simultaneously exhibit similarities and particularities.
Rac1 may play a role in the apoptosis of light-damaged photoreceptors. The increased expression of PAK4 after light stimulus possibly functions as a protective mechanism against apoptosis.
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