Retinal Degeneration In Rd Mouse Research Articles

Microglial NADPH oxidase activation mediates rod cell death in the retinal degeneration in rd mice

Accumulating evidence supports that nicotinamide adenine dinucleotide phosphate (NADPH) oxidase contributes to microglia-mediated neurotoxicity in the CNS neurodegenerative diseases. Several studies, including ours, suggest that microglial activation is involved in the retinal degeneration in the animal models of retinitis pigmentosa (RP). In the present study, we investigated the activation of NADPH oxidase in the rod degeneration in rd mice and further explored its role in the microglia-mediated photoreceptor apoptosis. Expression of gp91phox protein, a major subunit of NAPDH oxidase in the whole retina of rd mice at postnatal days (P) 8, 10, 12, 14, 16 and 18 was assessed by western blot analysis. Location of gp91phox in the rd retina at each age group and its cellular source were studied by immunohistochemical analysis and double labeling respectively. The generation of superoxide radicals in the rd retinas was demonstrated by intraperitoneal injection of hydroethidine. Apocynin was applied intraperitoneally in the rd mice from P8 to P14 to inhibit the activity of NAPDH oxidase and the outer nuclear layer (ONL) thickness was measured before and after apocynin treatment. Our results demonstrated that during the rod degenerative process, the expression of gp91phox started to increase in the outer part of rd retina at P10 and reached a peak at P14. Double labeling of gp91phox with CD11b showed co-localization of gp91phox in the retinal microglial cells. Increasing generation of superoxide radicals visualized by hydroethidine was noted at P8 and reached a peak at P14. Apocynin markedly reduced the production of superoxide radicals and preserved the rod cells. The results suggested that NADPH oxidase might play an important role in the rod degeneration in the rd mice. Inhibition of NAPDH oxidase could be a possible approach to treat RP in the early degenerative stage.

The Role of CCR1 Expression in the Retinal Degeneration in rd Mice

Purpose: Chemokine receptors are reported to be involved in neuronal cell death and CNS neurodegenerative diseases. The aim of the current study was to investigate the expression of CCR1, a major chemokine receptor for CC chemokines in retinal dystrophy in rd (retinal degeneration) mice and further explore its role in photoreceptor degeneration.Materials and Methods: The expression levels of CCR1 mRNA in the whole control and rd retinas at postnatal days (P) 8, 10, 12, 14, 16, and 18 were determined by RT-PCR assay. Location of CCR1 in the retina of rd mice at each age group was studied by immunohistochemical analysis. Expression of CCR1 in the photoreceptor cells and apoptotic cells was determined by double labeling.Results: Expression of CCR1 mRNA was noted in both control and rd retinas at each age group. CCR1-positive cells started to emerge in the outer nuclear layer (ONL) in rd retinas at P8 and reached a peak at P12 and P14. Double labeling of CCR1 with rhodopsin, CD11b, or TUNEL staining showed expression of CCR1 in the photoreceptor cells, rather than in the microglial cells. Partial CCR1 expression was observed in some of the apoptotic photoreceptor cells.Conclusions: Expression of CCR1 in the photoreceptor cells was increased with the progress of retinal degeneration in rd mice. Activation of CCR1 may play a role in the photoreceptor apoptosis.

Identification of Sequential Events and Factors Associated with Microglial Activation, Migration, and Cytotoxicity in Retinal Degeneration inrdMice

To elucidate the role of activated microglia in the photoreceptor apoptosis of rd mice by identifying sequential events and factors associated with microglial activation, migration, and cytotoxicity during retinal degeneration. Photoreceptor apoptosis in rd mice at postnatal days (P)8, 10, 12, 14, 16, and 18 was detected by terminal dUTP transferase nick end labeling (TUNEL). Retinal microglia were identified by CD11b antibody. Expression of chemokine mRNA, including monocyte chemoattractant protein (MCP)-1, MCP-3, macrophage inflammatory protein (MIP)-1alpha, MIP-1beta, regulated on activation normal T-cell expressed and secreted (RANTES), interferon-gamma-inducible 10-kDa protein (IP-10), and fractalkine in the retina were examined by reverse transcription-polymerase chain reaction (RT-PCR) assay. Production of tumor necrosis factor (TNF)-alpha in the dystrophic retina was studied by enzyme-linked immunosorbent assay (ELISA) and immunohistochemistry analysis. Microglial expression of TNF-alpha was determined by double immunolabeling. Whereas photoreceptor apoptosis in the rd mice started at P10 and reached a peak at P16, activation and migration of microglial cells were observed at P10 and peaked at P14. The expression of MCP-1, MCP-3, MIP-1alpha, MIP-1beta, and RANTES transcripts were noted at P8 and reached a peak at P12. Production of TNF-alpha was noted in the outer nuclear layer (ONL) of the rd mice at P8 and reached a peak at P12. At the peak of microglial activity, TNF-alpha was predominantly expressed in the activated microglial cells in the ONL. Activation of microglia, as well as expression of their signaling molecules (chemokines) and microglia-derived toxic factor (TNF-alpha), coincides with or precedes the occurrence of photoreceptor apoptosis, suggesting activated microglia play a major role in retinal degeneration in rd mice. The chemokines MCP-1, MCP-3, MIP-1alpha, MIP-1beta, and RANTES are involved in activation and recruitment of the microglia to the degenerating photoreceptor cell layer. TNF-alpha, produced by the activated microglia, may accentuate the photoreceptor cell death.

BCL-2 and P53 could be involved in retinal degeneration in RD mice

BCL-2 and P53 could be involved in retinal degeneration in RD mice

3332 BCL-2 and P53 could be involved in retinal degeneration in RD mice

3332 BCL-2 and P53 could be involved in retinal degeneration in RD mice

Opsin, G-protein and 48-kDa protein in normal and rd mouse retinas: Developmental expression of mRNAS and proteins and light/dark cycling of mRNAs

Retinal degeneration in rd mice is manifested during the most rapid period of postnatal photoreceptor differentiation and is hypothesized to be caused by a lesion in cGMP metabolism. We have studied the sequence of developmental expression of three proteins involved in the cGMP cascade and the mRNAs from which they are translated, in rd and control mouse retinas. Slot blot analysis of retinal RNAs indicates that the mRNAs coding for opsin, the α, β and γ subunits of G-protein and 48-kDa protein each has the same time for onset of expression in normal and diseased retinas. Gβ and 48-kDa protein mRNAs are already detectable at birth, opsin mRNA appears by postnatal day 5 (P5), Gγ mRNA at P6 and Gα mRNA by P8. The levels of all these mRNAs decrease in the diseased retinas after P11–P12, correlating with the reduction in photoreceptor cell number that characterizes the rd disease. Immunocytochemistry indicates that the 48-kDa protein is present at birth, Gγ and opsin are detectable at P4 and Gα at P7. After P7, opsin and G-protein immunoreactivity are localized throughout the photoreceptor cell in the rd retinas but they are found only in the outer segment in control retinas. The 48-kDa protein immunoreactivity, which is observed in the whole photoreceptor layer both in rd and control retinas throughout development, is the only one of all immunoreactivities analysed that remains at 2 months of age in the rd retina and is probably localized in cones. However, at 6 months of age, 48-kDa protein immunoreactive cells are no longer present in the rd retina. We have also investigated whether there is a daily rhythm for the levels of mRNA present at different times during the light/dark periods in developing rd rd and rd/+ retinas and in adult normal (+/+) retinas. We find that the levels of each mRNA analysed appear to cycle in the +/+ adult retina, with the greatest amount of opsin and the three subunits of G-protein mRNAs occurring just before light onset and the greatest amount of 48-kDa protein mRNA occurring just before lights off. Cycling in the developing diseased or control retinas (P0–P12) is not observed and may be masked by the pronounced cell growth that occurs during this period.

MRNAs coding for proteins of the cGMP cascade in the degenerative retina of the rd mouse

A lesion in cGMP metabolism has been hypothesized to cause retinal degeneration in rd mice. Available cloned cDNAs coding for proteins involved in the cGMP cascade have been used to compare the corresponding retinal RNAs in the degenerative (rd/rd) mouse at 8-11 days with those in the 8-11-day-old morphologically normal (rd/+) and adult normal (+/+) mice. Northern analysis of these RNAs hybridized to the specific 32P-labeled cDNA probes for G-protein, 48,000 MW protein and opsin, indicates in each case, that the corresponding transcripts are made in the rd/rd mouse retina and that there are no overt differences in their size compared to the transcripts hybridized in the rd/+ or +/+ mouse retinas. Although a defect in the phosphorylation of opsin has been described in rd mice, no difference was found in the transcripts hybridized by a cDNA probe corresponding to the region of the opsin molecule on which phosphorylation occurs. We do find, however, that labeled bovine-derived opsin cDNA recognizes five different RNA size classes in mouse and bovine retinas. Control experiments were performed to confirm that the RNA hybridized by opsin cDNA was not due to non-specific hybridization to unrelated RNAs.

Enhanced sensory convergence to the visual cortex in the rodless (rd/rd) mouse.

The rd/rd mutation provokes a specific degeneration of the photoreceptive cells in the mouse retina, without affecting other neuronal elements. This degeneration starts 8 to 10 days after birth. Discriminative learning experiments have shown the degenerated retina to remain light-sensitive, the sensitivity being reduced to 10(-5) of normal with a peak still at 500 nm. An ontogenetic study showed a progressive vanishing of visual potentials (electroretinogram and visual evoked potentials) during the third week of life. Potentials of auditory origin, which can be recorded from the visual cortex in normal mice, were found to be clearly facilitated by the absence of the visual input either because of retinal degeneration in rd mouse or provoked by enucleation at birth.

Assessment of possible transneuronal changes in the retina of rats with inherited retinal dystrophy: cell size, number, synapses, and axonal transport by retinal ganglion cells.

In pigmented RCS rats with inherited retinal dystrophy, most photoreceptor cells disappear between postnatal days 20 and 100. We have examined the time course of the degeneration of photoreceptor nuclei and synapses and determined whether transneuronal changes occur in the inner nuclear layer (INL), inner plexiform layer (IPL), and retinal ganglion cells following loss of photoreceptor cells in these animals. Electron microscopic photomontages of the entire thickness of the IPL of dystrophic (RCS-p+) and control (RCS-rdy+ p+) rats 334 to 515 days old were prepared, and synapses were counted and identified as either conventional (amacrine) or ribbon (bipolar) types. Neither the incidence of synapses in the IPL nor the ratio of conventional to ribbon synapses differed in the dystrophic and control retinas. Ganglion cell diameter, perimeter, area, and density were measured from drawings of wholemount preparations of dystrophic and control rats 105 days and older. Diameter, perimeter, area and number of ganglion cells were not significantly different in the two genotypes. Anterograde axonal transport was measured by studying the displacement of labeled material as it traveled along ganglion cell axons and accumulated in the superior colliculus. The normal and dystrophic rats showed no significant difference in (1) the rates of rapidly moving components (approximately 110-180 mm/day) and slowly moving components (1.7-2.5 mm/day) or (2) the amount of radioactive material transported to the superior colliculus. The absence of transneuronal changes in retinal ganglion cells of RCS rats contrasts with results obtained earlier in rd mice (Graftstein et al., '72). Unlike the RCS rat, retinal degeneration in rd mice occurs before the maturation of the retina. We hypothesize that the ganglion cells may be more affected by loss of input early in development, and, therefore, ganglion cells of retinal dystrophic rats are less affected despite little or no synaptic input for several months. Furthermore, any reduction in the electrical activity of retinal ganglion cells that might follow loss of photoreceptor cells does not result in a significantly decreased rate of axonal transport.