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Long-term preservation of cone photoreceptors and visual acuity in rd10 mutant mice exposed to continuous environmental enrichment.

Barone I, Novelli E, Strettoi E - Mol. Vis. (2014)

Bottom Line: In any case, the final outcome is near-blindness without a conclusive cure yet.No major differences were detected in the morphology of the neurons of the inner retina between the two experimental groups.We therefore confirm and extend previous findings that showed an EE is an effective, minimally invasive tool for promoting long-lasting retinal protection in experimental models of RP.

View Article: PubMed Central - PubMed

Affiliation: CNR Neuroscience Institute, Pisa, Italy.

ABSTRACT

Purpose: In human patients and animal models of retinitis pigmentosa (RP), a gradual loss of rod photoreceptors and decline in scotopic vision are the primary manifestations of the disease. Secondary death of cones and gradual, regressive remodeling of the inner retina follow and progress at different speeds according to the underlying genetic defect. In any case, the final outcome is near-blindness without a conclusive cure yet. We recently reported that environmental enrichment (EE), an experimental manipulation based on exposure to enhanced motor, sensory, and social stimulation, when started at birth, exerts clear beneficial effects on a mouse model of RP, by slowing vision loss. The purpose of this study was to investigate in the same mouse the long-term effects of chronic exposure to an EE and assess the outcome of this manipulation on cone survival, inner retinal preservation, and visual behavior.

Methods: Two groups of rd10 mutant mice were maintained in an EE or standard (ST) laboratory conditions up to 1 year of age. Then, retinal preservation was assessed with immunocytochemistry, confocal microscopy examination, cone counts, and electron microscopy of the photoreceptor layer, while visual acuity was tested behaviorally with a Prusky water maze.

Results: rd10 mice are a model of autosomal recessive RP with a typical rod-cone, center to the periphery pattern of photoreceptor degeneration. They carry a mutation of the rod-specific phosphodiesterase gene and undergo rod death that peaks at around P24, while cone electroretinogram (ERG) is extinct by P60. We previously showed that early exposure to an EE efficiently delays photoreceptor degeneration in these mutants, extending the time window of cone viability and cone-mediated vision well beyond the phase of maximum rod death. Here we find that a maintained EE can delay the degeneration of cones even in the long term. Confocal and electron microscopy examination of the retinas of the rd10 EE and ST mice at 1 year of age showed major degeneration of the photoreceptor layer in both experimental groups, with small clusters of photoreceptors persisting in the peripheral retina. These vestigial cells were positive for L and M opsins and cone arrestin and represented the residual population of cones. In the retinas of the EE mice, cones were more numerous and less remodeled than in the ST counterparts, albeit virtually devoid of outer segments, as confirmed with electron microscopy (EM) observations. Cone counting in retinal whole mounts showed that rd10 EE mice at 1 year had almost three times as many surviving cones (34,000±4,000) as the ST control mice (12,700±1,800), t test p=0.003. Accordingly, the rd10 EE mice at 1 year of age were still capable of performing the visual water task in photopic conditions, showing a residual visual acuity of 0.138±0 cycles/degree. This ability was virtually absent in the rd10 ST age-matched mice (0.063±0.014), t test, p=0.029. No major differences were detected in the morphology of the neurons of the inner retina between the two experimental groups.

Conclusions: The approaches used to test the effects of an EE were consistent in showing significantly better preservation of cones and measurable visual acuity in 1-year-old rd10 EE mice. We therefore confirm and extend previous findings that showed an EE is an effective, minimally invasive tool for promoting long-lasting retinal protection in experimental models of RP.

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Ultrastructure of the outer retina in EE and ST mice. Electron micrographs of the outer retina of EE (A, C) and ST (B) rd10 mice. Asterisks in the enriched environment (EE) sample label nuclei of residual cells in the outer retina, adjacent to a well-organized outer limiting membrane (OLM). These nuclei have different densities and presumably belong to surviving cones and misplaced bipolar cells. The outer plexiform layer (OPL) is still present in A and C, while in the ST sample (D), the cell bodies of the inner nuclear layer (INL) cells are in close proximity to the RPE.
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f5: Ultrastructure of the outer retina in EE and ST mice. Electron micrographs of the outer retina of EE (A, C) and ST (B) rd10 mice. Asterisks in the enriched environment (EE) sample label nuclei of residual cells in the outer retina, adjacent to a well-organized outer limiting membrane (OLM). These nuclei have different densities and presumably belong to surviving cones and misplaced bipolar cells. The outer plexiform layer (OPL) is still present in A and C, while in the ST sample (D), the cell bodies of the inner nuclear layer (INL) cells are in close proximity to the RPE.

Mentions: Analysis of the vertical semithin sections consistently confirmed the presence of a single, discontinuous layer of elongated cells in the outer retinas of the EE mice but not in the ST mice (Figure 4). Electron microscopy showed that these cells were positioned in the outermost retinal tier, abutting a clearly identifiable outer limiting membrane (Figure 5), and were largely occupied by a nucleus with a variable ultrastructure: Some nuclei had highly condensed chromatin (similar to that of normal rods), while others were more homogeneous (Figure 5A,D). Most likely, they included the residual population of cones, as well as the cell bodies of some bipolar cells, which are known to move toward the outer retina when photoreceptors die out [18], as shown in Figure 2 (asterisks). No outer segments were evident in individual EM sections. The outer plexiform layer, instead, although thinned, was clearly visible and occupied by processes with an irregular course (Figure 5A,D). The examination of sections from retinal samples with matching eccentricity and location showed that rare small cells persisted in the outer retinas of the rd10 mice kept in ST conditions. The OPL was not recognizable, and the cells of the inner nuclear layer (INL) were adjacent to the retinal pigment epithelium (Figure 5B).


Long-term preservation of cone photoreceptors and visual acuity in rd10 mutant mice exposed to continuous environmental enrichment.

Barone I, Novelli E, Strettoi E - Mol. Vis. (2014)

Ultrastructure of the outer retina in EE and ST mice. Electron micrographs of the outer retina of EE (A, C) and ST (B) rd10 mice. Asterisks in the enriched environment (EE) sample label nuclei of residual cells in the outer retina, adjacent to a well-organized outer limiting membrane (OLM). These nuclei have different densities and presumably belong to surviving cones and misplaced bipolar cells. The outer plexiform layer (OPL) is still present in A and C, while in the ST sample (D), the cell bodies of the inner nuclear layer (INL) cells are in close proximity to the RPE.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4225138&req=5

f5: Ultrastructure of the outer retina in EE and ST mice. Electron micrographs of the outer retina of EE (A, C) and ST (B) rd10 mice. Asterisks in the enriched environment (EE) sample label nuclei of residual cells in the outer retina, adjacent to a well-organized outer limiting membrane (OLM). These nuclei have different densities and presumably belong to surviving cones and misplaced bipolar cells. The outer plexiform layer (OPL) is still present in A and C, while in the ST sample (D), the cell bodies of the inner nuclear layer (INL) cells are in close proximity to the RPE.
Mentions: Analysis of the vertical semithin sections consistently confirmed the presence of a single, discontinuous layer of elongated cells in the outer retinas of the EE mice but not in the ST mice (Figure 4). Electron microscopy showed that these cells were positioned in the outermost retinal tier, abutting a clearly identifiable outer limiting membrane (Figure 5), and were largely occupied by a nucleus with a variable ultrastructure: Some nuclei had highly condensed chromatin (similar to that of normal rods), while others were more homogeneous (Figure 5A,D). Most likely, they included the residual population of cones, as well as the cell bodies of some bipolar cells, which are known to move toward the outer retina when photoreceptors die out [18], as shown in Figure 2 (asterisks). No outer segments were evident in individual EM sections. The outer plexiform layer, instead, although thinned, was clearly visible and occupied by processes with an irregular course (Figure 5A,D). The examination of sections from retinal samples with matching eccentricity and location showed that rare small cells persisted in the outer retinas of the rd10 mice kept in ST conditions. The OPL was not recognizable, and the cells of the inner nuclear layer (INL) were adjacent to the retinal pigment epithelium (Figure 5B).

Bottom Line: In any case, the final outcome is near-blindness without a conclusive cure yet.No major differences were detected in the morphology of the neurons of the inner retina between the two experimental groups.We therefore confirm and extend previous findings that showed an EE is an effective, minimally invasive tool for promoting long-lasting retinal protection in experimental models of RP.

View Article: PubMed Central - PubMed

Affiliation: CNR Neuroscience Institute, Pisa, Italy.

ABSTRACT

Purpose: In human patients and animal models of retinitis pigmentosa (RP), a gradual loss of rod photoreceptors and decline in scotopic vision are the primary manifestations of the disease. Secondary death of cones and gradual, regressive remodeling of the inner retina follow and progress at different speeds according to the underlying genetic defect. In any case, the final outcome is near-blindness without a conclusive cure yet. We recently reported that environmental enrichment (EE), an experimental manipulation based on exposure to enhanced motor, sensory, and social stimulation, when started at birth, exerts clear beneficial effects on a mouse model of RP, by slowing vision loss. The purpose of this study was to investigate in the same mouse the long-term effects of chronic exposure to an EE and assess the outcome of this manipulation on cone survival, inner retinal preservation, and visual behavior.

Methods: Two groups of rd10 mutant mice were maintained in an EE or standard (ST) laboratory conditions up to 1 year of age. Then, retinal preservation was assessed with immunocytochemistry, confocal microscopy examination, cone counts, and electron microscopy of the photoreceptor layer, while visual acuity was tested behaviorally with a Prusky water maze.

Results: rd10 mice are a model of autosomal recessive RP with a typical rod-cone, center to the periphery pattern of photoreceptor degeneration. They carry a mutation of the rod-specific phosphodiesterase gene and undergo rod death that peaks at around P24, while cone electroretinogram (ERG) is extinct by P60. We previously showed that early exposure to an EE efficiently delays photoreceptor degeneration in these mutants, extending the time window of cone viability and cone-mediated vision well beyond the phase of maximum rod death. Here we find that a maintained EE can delay the degeneration of cones even in the long term. Confocal and electron microscopy examination of the retinas of the rd10 EE and ST mice at 1 year of age showed major degeneration of the photoreceptor layer in both experimental groups, with small clusters of photoreceptors persisting in the peripheral retina. These vestigial cells were positive for L and M opsins and cone arrestin and represented the residual population of cones. In the retinas of the EE mice, cones were more numerous and less remodeled than in the ST counterparts, albeit virtually devoid of outer segments, as confirmed with electron microscopy (EM) observations. Cone counting in retinal whole mounts showed that rd10 EE mice at 1 year had almost three times as many surviving cones (34,000±4,000) as the ST control mice (12,700±1,800), t test p=0.003. Accordingly, the rd10 EE mice at 1 year of age were still capable of performing the visual water task in photopic conditions, showing a residual visual acuity of 0.138±0 cycles/degree. This ability was virtually absent in the rd10 ST age-matched mice (0.063±0.014), t test, p=0.029. No major differences were detected in the morphology of the neurons of the inner retina between the two experimental groups.

Conclusions: The approaches used to test the effects of an EE were consistent in showing significantly better preservation of cones and measurable visual acuity in 1-year-old rd10 EE mice. We therefore confirm and extend previous findings that showed an EE is an effective, minimally invasive tool for promoting long-lasting retinal protection in experimental models of RP.

Show MeSH
Related in: MedlinePlus