Investigative Ophthalmology & Visual Science Cover Image for Volume 48, Issue 11
November 2007
Volume 48, Issue 11
Free
Retina  |   November 2007
Activation of Endoplasmic Reticulum Stress in Degenerating Photoreceptors of the rd1 Mouse
Author Affiliations
  • Li-ping Yang
    From the Peking University Eye Center, Peking University Third Hospital, Peking University, Beijing, People’s Republic of China; and the
  • Le-meng Wu
    From the Peking University Eye Center, Peking University Third Hospital, Peking University, Beijing, People’s Republic of China; and the
  • Xiu-juan Guo
    From the Peking University Eye Center, Peking University Third Hospital, Peking University, Beijing, People’s Republic of China; and the
  • Mark O. M. Tso
    From the Peking University Eye Center, Peking University Third Hospital, Peking University, Beijing, People’s Republic of China; and the
    Wilmer Eye Institute, Johns Hopkins University School of Medicine, Baltimore, Maryland.
Investigative Ophthalmology & Visual Science November 2007, Vol.48, 5191-5198. doi:https://doi.org/10.1167/iovs.07-0512
  • Views
  • PDF
  • Share
  • Tools
    • Alerts
      ×
      This feature is available to authenticated users only.
      Sign In or Create an Account ×
    • Get Citation

      Li-ping Yang, Le-meng Wu, Xiu-juan Guo, Mark O. M. Tso; Activation of Endoplasmic Reticulum Stress in Degenerating Photoreceptors of the rd1 Mouse. Invest. Ophthalmol. Vis. Sci. 2007;48(11):5191-5198. https://doi.org/10.1167/iovs.07-0512.

      Download citation file:


      © ARVO (1962-2015); The Authors (2016-present)

      ×
  • Supplements
Abstract

purpose. Endoplasmic reticulum (ER) stress has been implicated in a wide variety of neurodegenerative disorders of the central nervous system (CNS). This study was designed to elucidate the role of ER stress in photoreceptor apoptosis in the rd1 mouse.

methods. Photoreceptor apoptosis in the rd1 mouse was detected by terminal dUTP transferase nick-end labeling (TUNEL). Protein expressions of ER stress sensors, including glucose-regulated protein-78 (GRP78/BiP), caspase-12, phospho-eukaryotic initiation factor 2α (eIF2α), and phospho-pancreatic ER kinase (PERK), were examined by immunofluorescence and Western blot assays.

results. Accompanying photoreceptor apoptosis in the rd1 mouse, the protein expressions of GRP78/BiP, caspase-12, phospho-eIF2α, and phospho-PERK were upregulated in a time-dependent manner. The upregulation of these proteins coincided with or preceded photoreceptor apoptosis. At the peak of their expression, these proteins were primarily located in the photoreceptor inner segments, the outer nuclear layer, or both.

conclusions. ER stress plays an important role in photoreceptor apoptosis in the rd1 mouse. Therefore, ER stress modulators may be strong candidates as therapeutic agents in the treatment of retinal degenerative diseases.

Retinitis pigmentosa (RP) is a heterogeneous group of inherited retinal degenerative diseases that lead to blindness and typically involve progressive loss of rod and then cone photoreceptor cells (affecting approximately 1 in 3000 humans). Early stages of RP are characterized by night blindness, followed by a loss of peripheral vision. 1 The disease may progress slowly over many years, or progression may be rapid. In the past 15 years, more than 30 genes have been implicated in the genetics of RP (http://www.sph.uth.tmc.edu/retnet). Many of the genes encode photoreceptor-specific proteins, among them the structural protein peripherin, rod outer segment membrane protein-1, rod cGMP phosphodiesterase, and rhodopsin. 2 3 4 5  
The rd1 mouse carries a nonsense mutation in the gene coding for the β subunit of the rod photoreceptor-specific cGMP phosphodiesterase 6 (PDE6-β), rendering the enzyme nonfunctional. 6 7 Defects in the photoreceptor PDE6-β gene have been shown to underlie cases of autosomal recessive RP (arRP), accounting for approximately 1% to 2% of all cases of RP, which makes the rd1 mouse a relevant and useful model of human RP. 8 9 10 The absence of phosphodiesterase activity leads to an increased accumulation of cGMP in the photoreceptors, 6 which in turn leads to an increase in Na+ and Ca2+ influx through the cGMP-gated cation channels. Photoreceptor Ca2+ levels in the rd1 mouse are elevated, starting at postnatal day (P) 5, and have been shown to be increased to approximately 190% by P15 compared with photoreceptors from wild-type mice. 11 Uncontrolled Ca2+-influx triggers apoptosis, and Ca2+ channel blockers have been shown to rescue the rd1 photoreceptors. 12 13 Although there is a consensus on the role of Ca2+ as an initiator of degeneration in the rd1 retina, the subsequent cellular steps leading to degeneration remain unresolved. 
Recently, endoplasmic reticulum (ER) stress has been implicated in a wide variety of human diseases, including diabetes, cancer, and many neurodegenerative disorders such as brain ischemia, Alzheimer disease, Parkinson disease, Huntington disease, and amyotrophic lateral sclerosis. 14 15 16 17 18 19 20 21 22 The ER is a multifunctional organelle involved in the folding and processing of proteins, intracellular calcium homeostasis, and cell death signaling activation. 23 A number of cellular stress conditions, such as perturbed calcium homeostasis or redox status, and an accumulation of unfolded proteins in the ER cause ER stress that activates the unfolded protein response (UPR). 24 Several sensors of ER stress have been identified. These include glucose-regulated protein-78 (GRP78/BiP), pancreatic ER kinase (PERK), eukaryotic initiation factor 2α (eIF2α), and caspase-12. Once the UPR is activated, the cell may eventually return to normal ER homeostasis or, under prolonged ER stress, may continue toward apoptosis. Because neurons are highly susceptible to the toxic effects of misfolded proteins, ER stress–mediated cell death may have an important role in the pathogenesis of this disease. 
Although the rd1 mouse is a relatively well-studied model for RP, the direct causes of photoreceptor death are unclear. Previous studies speculated that Ca2+ overload and the generation of reactive oxygen species (but not mutually independent) are two possible mechanisms underlying the apoptotic pathway. 25 Given that disturbance of calcium homeostasis and oxidative stress may cause ER stress, we hypothesized that ER stress is activated in degenerating photoreceptors in the rd1 mouse. 17 24 26  
Materials and Methods
Animals
All experiments were performed in accordance with the ARVO Statement for the Use of Animals in Ophthalmic and Vision Research. All animals were husbanded in accordance with the guidelines of the Association for the Assessment and Accreditation of Laboratory Animal Care. C3H/FeJ mice homozygous for the rd-1 mutation were examined at four time points: postnatal days (P)10, 12, 14, and 24. Age-matched congenic C3H mice (C3.BliAPdeb-rd1), which are wild-type at the rd-1 locus, were used as controls. Both genotypes were obtained from the Jackson Laboratory (Bar Harbor, ME) and were housed in an air-conditioned room under a 12-hour light/12-hour dark cycle at a light intensity of 20 to 40 lux, under specific pathogen-free conditions at the Animal Facility of Peking University Health Science Center. 
Tissue Preparation
After the mice were humanely killed with an anesthetic overdose of pentobarbital, the eyes were immediately enucleated and immersed in 4% (wt/vol) paraformaldehyde (PFA) in phosphate-buffered saline (PBS) for 1 hour. Anterior segments were removed, and posterior segments were immersed in the same fixative for another 5 hours. Tissue samples were transferred to 20% sucrose buffer solution at 4°C overnight before they were embedded in an OCT compound. Frozen sections were then sectioned at 8 μm through the optic nerve head and the ora serrata with a cryostat. The sections were stored at −80°C until various investigative studies were conducted. 
TUNEL Assays
Photoreceptor apoptosis was determined by using a TUNEL system (DeadEnd Colorimetric TUNEL System; Promega, Madison, WI) according to the manufacturer’s instructions. For quantitative analysis, the number of TUNEL-positive photoreceptor nuclei was counted by a masked observer. Positively stained apoptotic photoreceptors were counted in a standard length of retina (1.2 mm) centered on the optic nerve head using a graticule at 40× objective magnification, as described previously. 27 Nine sections from three mice were used for TUNEL study at each time point. 
Western Blot Assays
The eyes were enucleated and bisected, and the retinas were peeled from the eyecups and immediately homogenized with 0.5 mL ice-cold lysis buffer (50 mM Tris-Cl, pH 8, 0.02% sodium azide, 1 μg/mL aprotinin, 1% NP-40, 100 μg/mL phenylmethylsulfonyl fluoride [PMSF]). The insoluble material was removed by centrifugation at 12,000g at 4°C for 20 minutes. Final protein concentrations were determined using a protein assay kit (BCA; Pierce Biotechnology, Rockford, IL) according to the manufacturer’s specifications. Western blot analysis was then performed as previously described. 28 Polyclonal antibodies specific for GRP78/BiP (1:500, ab21685; Abcam, Cambridge, UK), caspase-12 (1:1000, AB3612; Chemicon, Temecula, CA), phospho-eIF2α (1:1000, 3597; Cell Signaling, Beverly, MA), and phospho-PERK (1:1000, 3179; Cell Signaling) were used for immunodetection. Next, the blots were incubated with donkey anti–rabbit IgG (H+L) horseradish peroxidase (HRP) (Promega), which is an affinity-purified HRP-conjugated secondary antibody. Blots were visualized using an enhanced chemiluminescent technique (SC-2048; Santa Cruz Biotechnology, Santa Cruz, CA). As a control for equal loading of proteins, a β-actin antibody (1:2000, A2228; Sigma-Aldrich, St. Louis, MO) was used. To evaluate β-actin expression, the blot was stripped in stripping buffer (62.5 mM Tris-Cl, pH 6.8, 2% SDS, 100 mM β-mercaptoethanol) for 30 minutes at 50°C and reprobed. For quantitative evaluation of the Western blot studies, the films were scanned and the optical densities were quantified with analysis software (Quantity One 1-D; Bio-Rad, Hercules, CA). The Western blot experiments were repeated four times from separate samples for each time point. 
Immunofluorescence Study
For immunofluorescence, the tissue sections were fixed with chilled fresh acetone for 10 minutes. After they were washed with PBS, the sections were incubated with a blocking buffer (1% bovine serum albumin) for 3 hours at room temperature. The sections were then incubated overnight with primary antibodies at 4°C to GRP78/BiP (1:330, ab21685; Abcam), caspase-12 (1:200, AB3612; Chemicon), phospho-eIF2α (1:300, 3597; Cell Signaling), and phospho-PERK (1:300, 3179; Cell Signaling). After rinsing with PBS, the sections were incubated (Cy3 AffiniPure Goat Anti-Rabbit IgG; H+L; code 111 to 165–003; Jackson ImmunoResearch Laboratory, West Grove, PA) for 45 minutes and were examined under a confocal laser scanning microscope (510 META; Carl Zeiss, Oberkochen, Germany). 
Statistical Analysis
Data are summarized as the mean ± SD. Statistical comparisons were made by a single-factor ANOVA followed by the least-significant difference post hoc test for multiple comparisons. P < 0.01 was considered significant. 
Results
Photoreceptor Death in the rd1 Mouse
As previously reported, the retinas from the rd1 mouse and the control C3B mouse were morphologically comparable until P10, 29 30 when TUNEL-positive photoreceptor cells were first detected in the outer nuclear layer (ONL) and the ONL width was comparable in the rd1 and control mice. At P12, the number of TUNEL-positive photoreceptor cells reached a peak in the rd1 mouse, and the ONL width was reduced by approximately one third the width in the wild-type mouse. At P14, only scattered TUNEL-positive photoreceptor cells were observed, and the ONL width was reduced to two to three layers of photoreceptor cells. At P24, no TUNEL-positive photoreceptor cells were detected in the ONL, and the ONL width was reduced to only one layer of photoreceptor cells. The time course of the incremental alteration of the TUNEL-positive photoreceptor cells in the ONL of the rd1 retina is illustrated in Figure 1
Western Blot Assay of GRP78/BiP, Caspase-12, Phospho-eIF2α, and Phospho-PERK in the rd1 Mouse
Protein expressions of GRP78/BiP, caspase-12, phospho-eIF2α, and phospho-PERK were determined by a Western blot assay. A representative gel image is shown in Figure 2 . The results indicate that the control retina constitutively expressed modest quantities of GRP78/BiP protein. Accompanying photoreceptor degeneration in the rd1 mouse, the expression of GRP78/BiP protein was upregulated in a time-dependent manner. The expression of GRP78/BiP was mildly upregulated at P10 and was markedly upregulated and peaked at P12. Subsequently, expression slowly declined and by P24 had returned to the basal level. 
The control retina constitutively expressed modest quantities of procaspase-12 protein. Accompanying photoreceptor degeneration in the rd1 mouse, the expression of procaspase-12 protein was first downregulated at P10, was markedly upregulated and peaked at P12, and declined and returned to the basal level at P24. The control C3B retina expressed very low levels of cleaved caspase-12. Accompanying photoreceptor degeneration, the expression of cleaved caspase-12 protein was upregulated in a time-dependent manner. It was markedly upregulated and peaked at P12 and then returned to the basal level at P14. 
The control retina constitutively expressed modest quantities of phospho-eIF2α protein. Accompanying photoreceptor degeneration in the rd1 mouse, phospho-eIF2α protein expression was first downregulated at P10, and then it was upregulated. The expression of phospho-eIF2α was markedly upregulated at P12 but returned to the basal level at P14. 
The control retina constitutively expressed modest quantities of phospho-PERK protein. Accompanying photoreceptor degeneration in the rd1 mice, the expression of phospho-PERK protein was upregulated. The expression of phospho-PERK was markedly upregulated at P10 and P12. From P14 onward, its expression declined to the basal level. At P24, the phospho-PERK expression was markedly downregulated. 
Immunofluorescence Study of GRP78/BiP, Caspase-12, Phospho-eIF2α, and Phospho-PERK in the rd1 Mouse
Western blot assays provided strong evidence for the upregulation of endoplasmic reticulum proteins in the retinas of the rd1 mouse. Location of GRP78/BiP, caspase-12, phospho-eIF2α, and phospho-PERK proteins was determined by the immunofluorescence study. In all immunolabeling studies, no positive cells were observed in the samples in which the first antibody was omitted to serve as a negative control (data not shown). 
At P12, positive GRP78/BiP staining of the control retina was mainly distributed in the inner nuclear layer (INL), and no positive staining was seen in the ONL or in the photoreceptor inner segments. At P10, no significant difference was shown in the rd1 retina compared with the control retina. At P12, the GRP78/BiP-positive staining in the rd1 retina was most evident and was located primarily at the photoreceptor inner segments. The INL and the ONL also showed GRP78/BiP-positive labeling. From P14 onward, the intensity of GRP78/BiP immunofluorescence in the rd1 retina decreased dramatically compared with the P12 retina. This was particularly evident in the INL, where the staining was almost entirely undetectable. Only minor positive staining was evident in the ONL and at the photoreceptor inner segments. At P24, no representative positive labeling was seen in the rd1 retina (Fig. 3)
At P12, caspase-12 immunofluorescence was evenly distributed among all the layers of the control retina. At P10, immunofluorescence in the rd1 retina labeled the outlines of the cells in the ONL and the outlines of the photoreceptor inner segments. At P12, the caspase-12–positive labeling in the rd1 retina was most evident and was primarily located at the photoreceptor inner segments and the ONL. The INL also showed some positive labeling. At P14, the positive labeling in the rd1 retina was weaker than at P12. At P24, positive staining was almost undetectable in the rd1 retina (Fig. 4)
At P12, the positive phospho-eIF2α staining of the control retina labeled the outlines of the cells in the ONL and the INL. At P10, the positive labeling in the rd1 retina was weaker and less intense than in the control retina. At P12, the positive phospho-eIF2α staining in the rd1 retina was most evident and was primarily located in the photoreceptor inner segments and the ONL. At P14, staining in the ONL and in the photoreceptor inner segments was markedly decreased. At P24, positive staining throughout the rd1 retina was nearly undetectable (Fig. 5)
At P12, the positive phospho-PERK staining of the control retina was mainly distributed in the photoreceptor inner segments, and the INL also showed scattered positive labeling. At P10 and P12, the phospho-PERK immunofluorescence in the photoreceptor inner segments and in the INL in the rd1 retina was markedly increased compared with the control retina. From P14 onward, the intensity of phospho-PERK labeling in the photoreceptor inner segments was weaker, but the positive labeling in the INL had increased (Fig. 6)
Discussion
We observed that accompanying photoreceptor degeneration in the rd1 mouse, the ER stress proteins, including GRP78/BiP, caspase-12, phospho-eIF2α, and phospho-PERK, were markedly upregulated. The upregulation of ER stress proteins coincided with or preceded the photoreceptor apoptosis, suggesting that ER stress plays an important role in photoreceptor apoptosis in the rd1 mouse. We hypothesized that the initial gene defect in the rd1 mouse resulted in Ca2+ overload, which induced the ER stress pathway, causing the cells to go into apoptosis. 
Calcium is an important intracellular signaling molecule that requires tight regulation of the intracellular concentrations for optimal triggering of signaling cascades and cell survival. 31 In the mouse photoreceptor, normal calcium levels range between approximately 250 nM (in complete darkness) and approximately 60 nM (in the light). 32 In the rd1 mouse photoreceptors, however, calcium levels increased up to approximately 190% over the wild-type mice levels. 11 Consequently, messages for calcium-binding proteins and specific calcium sensors, such as calpains, may be upregulated in the rd1 retina. Calpains are cysteine proteases activated by calcium during the apoptotic processes. 33 Two ubiquitously expressed calpains are the isozymes calpain I (μ-calpain) and calpain II (m-calpain), which are activated in vitro by micromolar and millimolar calcium concentrations, respectively. 34 Calpain I and calpain II are expressed in the retina, and they were activated in the rd1 mouse. 35 36 37 Calpains do not directly cause chromatin condensation, but they are proteases that activate apoptotic factors. Several proteins are known targets of calpain protease activity, such as caspase-12. Caspase-12 is localized on the cytoplasmic side of the ER, which enables direct sensing of ER perturbations. 38 Caspase-12 is activated when the ER undergoes stress, but not by membrane- or mitochondrial-targeted apoptotic signals. Mice deficient for caspase-12 are resistant to inducers of ER stress, suggesting that caspase-12 is significant in ER stress-induced apoptosis. 39 In the present study, accompanying photoreceptor apoptosis in the rd1 mouse, the expression of procaspase-12 and cleaved caspase-12 were markedly upregulated. The expression of procaspase-12 was downregulated at P10, but this might have been because some of the procaspase-12 was cleaved. The upregulation of caspase-12 coincided with the onset and peak of photoreceptor apoptosis, suggesting that caspase-12 is involved in the photoreceptor degeneration in the rd1 mouse. The immunofluorescence study demonstrated that at the peak of its expression, caspase-12 was primarily located at the inner segments of photoreceptor cells, with some positive staining also showing in the ONL. This is consistent with the theory that caspase-12 is localized in the ER, so that the protein is most likely localized in the inner segments of photoreceptor cells containing mitochondria and the ER. The cleaved active form of caspase-12 participates in the apoptotic event by translocation to the nucleus or possibly through other caspases. 40 41 42  
Caspase-12 is phylogenetically one of the inflammatory caspases. 43 It has been reported that murine caspase-12 expression is induced by IFN-γ and that the expected NF-κB and AP-1 binding sites are present in its promoter region. 44 45 As seen in a previous study, ER stress may activate some of the proinflammatory signal transduction pathways associated with innate immunity. 46 The upregulated expression of chemokines and noxious factors in the rd1 mouse retina might have resulted in part from caspase-12 activation. 29  
The UPR is initiated by the binding of the ER chaperone GRP78/BiP to the misfolded proteins. Under normal conditions, GRP78/BiP forms a complex with three key proteins at the ER membrane—PERK, transcriptional factor ATF-6, and endoribonuclease IRE-1. 47 The binding of GRP78/BiP to unfolding proteins releases GRP78/BiP from PERK, ATF-6, and IRE-1. Subsequently, GRP78/BiP is activated and the protein folding capacity of the ER is increased. UPR is also characterized by phosphorylation and activation of the PERK, which is localized on the ER membrane and is phosphorylated when stress is imposed on the ER. In turn, it phosphorylates the eIF2α and elicits various cellular responses, such as protein synthesis inhibition and apoptotic signal activation. 47 48 49 In this study, the expression of GRP78/BiP was upregulated in the rd1 retina from P10 and peaked at P12, coinciding with the onset and peak of photoreceptor apoptosis. The expression of phospho-eIF2α was downregulated at P10, but then it was markedly upregulated and peaked at P12. The expression of phospho-PERK was markedly upregulated and peaked at P10, after which, its expression declined. These results suggest that UPR is activated in the rd1 mouse photoreceptor degeneration process. A large number of dominant mutations throughout rhodopsin cause protein sequestration in the ER of flies. 50 51 The UPR is activated and plays a protective role against the progression of retinal degeneration. 52 Until now, however, no direct evidence has indicated that unfolding protein accumulation was involved in the rd1 mouse retinal degeneration. Further study will be needed to support this suggestion. 
Based on observations in the rd1 mouse, Sharma and Rohrer 25 proposed the pathway of apoptosis in photoreceptor degeneration: an increase in intracellular Ca2+ activates calpain, which may cleave the proapoptotic Bcl-2 family protein bid. Interaction of truncated bid (t-bid) with the mitochondrial permeability transition pore causes mitochondrial membrane potential loss (ΔΨm), leading to the release of cytochrome c. 25 The increase in cytoplasmic cytochrome c causes the assembly of the apoptosome, leading to the activation of caspases that cleave downstream death substrates and activate endonucleases that cleave genomic DNA into fragments resulting in apoptotic nuclear morphology. It is likely that the energetic burden on the rd1 rods is high as their ion pumps attempt to reestablish homeostasis. 53 In accordance with this assumption, increments in the activity or expression of glycolytic enzymes have been reported in the rd1 mouse, and these changes are thought to lead to an increase in the production of ROS and oxidative stress. 53 54 ROS and oxidative damage also are part of the rd1 rod pathology. 55 From this study, we added that disturbance of calcium homeostasis and oxidative stress in the rd1 mouse activated ER stress, which elicited various cellular responses, such as increments in the protein folding capacity and protein synthesis inhibition, intended to induce the cell’s return to normal ER homeostasis. Under prolonged ER stress, however, apoptosis-promoting factors such as cleaved caspase-12 were activated, and the cells progressed to apoptosis. 
In conclusion, our study demonstrated that ER stress played an important role in photoreceptor apoptosis in the rd1 mouse. Therefore, ER stress modulators may be strong candidates as therapeutic agents in the treatment of the retinal degenerative diseases. 
 
Figure 1.
 
TUNEL immunofluorescence of the rd1 retina. (A) No TUNEL-positive cells were seen in the ONL at P8. (B) TUNEL-positive cells were first seen in the ONL at P10. (C) TUNEL-positive cells were most abundant at P12. (D) TUNEL-positive cells were less abundant at P14. (E) No TUNEL-positive cells were detected at P24. (F) The time course of the incremental alteration of the TUNEL-positive photoreceptor cells in the ONL in the rd1 retina. Data are expressed as mean ± SD.
Figure 1.
 
TUNEL immunofluorescence of the rd1 retina. (A) No TUNEL-positive cells were seen in the ONL at P8. (B) TUNEL-positive cells were first seen in the ONL at P10. (C) TUNEL-positive cells were most abundant at P12. (D) TUNEL-positive cells were less abundant at P14. (E) No TUNEL-positive cells were detected at P24. (F) The time course of the incremental alteration of the TUNEL-positive photoreceptor cells in the ONL in the rd1 retina. Data are expressed as mean ± SD.
Figure 2.
 
Retinal expressions of ER stress proteins in the wild-type and the rd1 mouse. (A) Time course for GRP78/BiP, procaspase-12/cleaved caspase-12, phospho-eIF2α, and phospho-PERK protein expression in the control and the rd1 retinas at each age group. (B) The relative level of GRP78/BiP, procaspase-12/cleaved caspase-12, phospho-eIF2α, and phospho-PERK protein expression in the rd1 mouse were quantified and corrected for the levels of β-actin protein expression. (C) P12 wild-type C3B mouse. P < 0.01 was considered significant compared with the control retina.
Figure 2.
 
Retinal expressions of ER stress proteins in the wild-type and the rd1 mouse. (A) Time course for GRP78/BiP, procaspase-12/cleaved caspase-12, phospho-eIF2α, and phospho-PERK protein expression in the control and the rd1 retinas at each age group. (B) The relative level of GRP78/BiP, procaspase-12/cleaved caspase-12, phospho-eIF2α, and phospho-PERK protein expression in the rd1 mouse were quantified and corrected for the levels of β-actin protein expression. (C) P12 wild-type C3B mouse. P < 0.01 was considered significant compared with the control retina.
Figure 3.
 
Localizations of GRP78/BiP protein in the wild-type and the rd1 mouse. (A) At P12, the positive GRP78/BiP staining of the C3B retina was mainly distributed in the INL (arrows). (B) At P10, the positive staining in the rd1 retina was also mainly in the INL (arrows), with no positive staining shown in the ONL or at the photoreceptor inner segments. (C) At P12, the GRP78/BiP-positive staining in the rd1 retina was most evident and was mainly located at the photoreceptor inner segments (arrows). The INL and ONL also showed positive labeling (arrows). (D) At P14, the positive staining in the INL of the rd1 retina was almost undetectable, and the intensity of positive labeling in the photoreceptor inner segments and ONL dramatically decreased, with limited positive staining evident (arrows). (E) At P24, no representative positive labeling was seen in the rd1 retina.
Figure 3.
 
Localizations of GRP78/BiP protein in the wild-type and the rd1 mouse. (A) At P12, the positive GRP78/BiP staining of the C3B retina was mainly distributed in the INL (arrows). (B) At P10, the positive staining in the rd1 retina was also mainly in the INL (arrows), with no positive staining shown in the ONL or at the photoreceptor inner segments. (C) At P12, the GRP78/BiP-positive staining in the rd1 retina was most evident and was mainly located at the photoreceptor inner segments (arrows). The INL and ONL also showed positive labeling (arrows). (D) At P14, the positive staining in the INL of the rd1 retina was almost undetectable, and the intensity of positive labeling in the photoreceptor inner segments and ONL dramatically decreased, with limited positive staining evident (arrows). (E) At P24, no representative positive labeling was seen in the rd1 retina.
Figure 4.
 
Localizations of caspase-12 protein in the wild-type and the rd1 mouse. (A) At P12, the caspase-12 immunofluorescence of the C3B retina was evenly distributed among all layers of the retina. (B) At P10, the immunofluorescence in the rd1 retina also labeled the outlines of cells in the ONL and the outlines of photoreceptor inner segments. (C) At P12, the caspase-12–positive labeling in the rd1 retina was most evident and was mainly located at the photoreceptor inner segments and in the ONL (arrows). (D) At P14, the positive labeling in the rd1 retina was also located at the photoreceptor inner segments and in the ONL (arrows), but it was weaker than at P12. (E) At P24, positive staining in the rd1 retina was almost undetectable.
Figure 4.
 
Localizations of caspase-12 protein in the wild-type and the rd1 mouse. (A) At P12, the caspase-12 immunofluorescence of the C3B retina was evenly distributed among all layers of the retina. (B) At P10, the immunofluorescence in the rd1 retina also labeled the outlines of cells in the ONL and the outlines of photoreceptor inner segments. (C) At P12, the caspase-12–positive labeling in the rd1 retina was most evident and was mainly located at the photoreceptor inner segments and in the ONL (arrows). (D) At P14, the positive labeling in the rd1 retina was also located at the photoreceptor inner segments and in the ONL (arrows), but it was weaker than at P12. (E) At P24, positive staining in the rd1 retina was almost undetectable.
Figure 5.
 
Localizations of phospho-eIF2α protein in the wild-type and the rd1 mouse. (A) At P12, positive phospho-eIF2α staining in the C3B retinas labeled the outlines of the cells in the ONL and INL. (B) At P10, positive labeling in the rd1 retina was weaker and less intense than in the control retina. (C) At P12, the positive phospho-eIF2α staining in the rd1 retina was mainly located in the photoreceptor inner segments and in the ONL (arrows). (D) At P14, the positive staining in the rd1 retina markedly decreased, and limited positive staining showed in the ONL and in the photoreceptor inner segments (arrows). (E) At P24, the positive staining in all parts of the rd1 retina was almost undetectable.
Figure 5.
 
Localizations of phospho-eIF2α protein in the wild-type and the rd1 mouse. (A) At P12, positive phospho-eIF2α staining in the C3B retinas labeled the outlines of the cells in the ONL and INL. (B) At P10, positive labeling in the rd1 retina was weaker and less intense than in the control retina. (C) At P12, the positive phospho-eIF2α staining in the rd1 retina was mainly located in the photoreceptor inner segments and in the ONL (arrows). (D) At P14, the positive staining in the rd1 retina markedly decreased, and limited positive staining showed in the ONL and in the photoreceptor inner segments (arrows). (E) At P24, the positive staining in all parts of the rd1 retina was almost undetectable.
Figure 6.
 
Localizations of phospho-PERK protein in the wild-type and the rd1 mouse. (A) At P12, positive phospho-PERK staining in the C3B retina was primarily distributed in the photoreceptor inner segments; the INL also showed scattered positive labeling. (B) At P10, phospho-PERK immunofluorescence in the photoreceptor inner segments and in the INL of the rd1 retina markedly increased compared with the control retina. (C) At P12, positive staining in the photoreceptor inner segments and the INL of the rd1 retina were still extensive. (D) At P14, positive labeling in the photoreceptor inner segments of the rd1 retina decreased, but positive labeling in the INL increased. (E) At P24, positive labeling in the photoreceptor inner segments of the rd1 retina decreased to one layer, and positive labeling in the INL increased.
Figure 6.
 
Localizations of phospho-PERK protein in the wild-type and the rd1 mouse. (A) At P12, positive phospho-PERK staining in the C3B retina was primarily distributed in the photoreceptor inner segments; the INL also showed scattered positive labeling. (B) At P10, phospho-PERK immunofluorescence in the photoreceptor inner segments and in the INL of the rd1 retina markedly increased compared with the control retina. (C) At P12, positive staining in the photoreceptor inner segments and the INL of the rd1 retina were still extensive. (D) At P14, positive labeling in the photoreceptor inner segments of the rd1 retina decreased, but positive labeling in the INL increased. (E) At P24, positive labeling in the photoreceptor inner segments of the rd1 retina decreased to one layer, and positive labeling in the INL increased.
BundeyS, CrewsSJ. A study of retinitis pigmentosa in the city of Birmingham, I: prevalence. J Med Genet. 1984;21:417–420. [CrossRef] [PubMed]
FarrarGJ, KennaP, JordanSA, et al. Autosomal dominant retinitis pigmentosa: a novel mutation at the peripherin/RDS locus in the original 6p-linked pedigree. Genomics. 1992;14:805–807. [CrossRef] [PubMed]
BascomRA, LiuL, HeckenlivelyJR, et al. Mutation analysis of the ROM1 gene in retinitis pigmentosa. Hum Mol Genet. 1995;4:1895–1902. [CrossRef] [PubMed]
McLaughlinME, SandbergMA, BersonEL, DryjaTP. Recessive mutations in the gene encoding the beta-subunit of rod phosphodiesterase in patients with retinitis pigmentosa. Nat Genet. 1993;4:130–134. [CrossRef] [PubMed]
FarrarGJ, KennaP, RedmondR, et al. Autosomal dominant retinitis pigmentosa: absence of the rhodopsin proline-histidine substitution (codon 23) in pedigrees from Europe. Am J Hum Genet. 1990;47:941–945. [PubMed]
FarberDB, ParkS, YamashitaC. Cyclic GMP-phosphodiesterase of rd retina: biosynthesis and content. Exp Eye Res. 1988;46:363–374. [CrossRef] [PubMed]
BowesC, LiT, DancigerM, et al. Retinal degeneration in the rd mouse is caused by a defect in the beta subunit of rod cGMP-phosphodiesterase. Nature. 1990;347:677–680. [CrossRef] [PubMed]
DancigerM, HeilbronV, GaoYQ, ZhaoDY, JacobsonSG, FarberDB. A homozygous PDE6B mutation in a family with autosomal recessive retinitis pigmentosa. Mol Vis. 1996;2:10. [PubMed]
DancigerM, BlaneyJ, GaoYQ, et al. Mutations in the PDE6B gene in autosomal recessive retinitis pigmentosa. Genomics. 1995;30:1–7. [CrossRef] [PubMed]
McLaughlinME, EhrhartTL, BersonEL, DryjaTP. Mutation spectrum of the gene encoding the beta subunit of rod phosphodiesterase among patients with autosomal recessive retinitis pigmentosa. Proc Natl Acad Sci USA. 1995;92:3249–3253. [CrossRef] [PubMed]
FoxDA, PoblenzAT, HeL. Calcium overload triggers rod photoreceptor apoptotic cell death in chemical-induced and inherited retinal degenerations. Ann NY Acad Sci. 1999;893:282–285. [CrossRef] [PubMed]
LolleyRN, RongH, CraftCM. Linkage of photoreceptor degeneration by apoptosis with inherited defect in phototransduction. Invest Ophthalmol Vis Sci. 1994;35:358–362. [PubMed]
TakanoY, OhguroH, DezawaM, et al. Study of drug effects of calcium channel blockers on retinal degeneration of rd mouse. Biochem Biophys Res Commun. 2004;313:1015–1022. [CrossRef] [PubMed]
OzcanU, CaoQ, YilmazE, et al. Endoplasmic reticulum stress links obesity, insulin action, and type 2 diabetes. Science. 2004;306:457–461. [CrossRef] [PubMed]
BiM, NaczkiC, KoritzinskyM, et al. ER stress regulated translation increases tolerance to extreme hypoxia and promotes tumor growth. EMBO J. 2005;24:3470–3481. [CrossRef] [PubMed]
TajiriS, OyadomariS, YanoS, et al. Ischemia-induced neuronal cell death is mediated by the endoplasmic reticulum stress pathway involving CHOP. Cell Death Differ. 2004;11:403–415. [CrossRef] [PubMed]
HayashiT, SaitoA, OkunoS, et al. Oxidative damage to the endoplasmic reticulum is implicated in ischemic neuronal cell death. J Cereb Blood Flow Metab. 2003;23:1117–1128. [PubMed]
HayashiT, SaitoA, OkunoS, Ferrand-DrakeM, DoddRL, ChanPH. Oxidative injury to the endoplasmic reticulum in mouse brains after transient focal ischemia. Neurobiol Dis. 2004;15:229–239. [CrossRef] [PubMed]
KatayamaT, ImaizumiK, ManabeT, HitomiJ, KudoT, TohyamaM. Induction of neuronal death by ER stress in Alzheimer’s disease. J Chem Neuroanat. 2004;28:67–78. [CrossRef] [PubMed]
SilvaRM, RiesV, OoTF, et al. CHOP/GADD153 is a mediator of apoptotic death in substantia nigra dopamine neurons in an in vivo neurotoxin model of parkinsonism. J Neurochem. 2005;95:974–986. [CrossRef] [PubMed]
PaschenW, MengesdorfT. Endoplasmic reticulum stress response and neurodegeneration. Cell Calcium. 2005;38:409–415. [CrossRef] [PubMed]
TurnerBJ, AtkinJD. ER stress and UPR in familial amyotrophic lateral sclerosis. Curr Mol Med. 2006;6:79–86. [CrossRef] [PubMed]
BaumannO, WalzB. Endoplasmic reticulum of animal cells and its organization into structural and functional domains. Int Rev Cytol. 2001;205:149–214. [PubMed]
KaufmanRJ. Stress signaling from the lumen of the endoplasmic reticulum: coordination of gene transcriptional and translational controls. Genes Dev. 1999;13:1211–1233. [CrossRef] [PubMed]
SharmaAK, RohrerB. Calcium-induced calpain mediates apoptosis via caspase-3 in a mouse photoreceptor cell line. J Biol Chem. 2004;279:35564–35572. [CrossRef] [PubMed]
XueX, PiaoJH, NakajimaA, et al. Tumor necrosis factor alpha (TNFα) induces the unfolded protein response (UPR) in a reactive oxygen species (ROS)-dependent fashion, and the UPR counteracts ROS accumulation by TNFα. J Biol Chem. 2005;280:33917–33925. [CrossRef] [PubMed]
HughesEH, SchlichtenbredeFC, MurphyCC, et al. Generation of activated sialoadhesin-positive microglia during retinal degeneration. Invest Ophthalmol Vis Sci. 2003;44:2229–2234. [CrossRef] [PubMed]
WuM, KusukawaN. SDS agarose gels for analysis of proteins. BioTechniques. 1998;24:676–678. [PubMed]
ZengHY, ZhuXA, ZhangC, YangLP, WuLM, TsoMO. Identification of sequential events and factors associated with microglial activation, migration, and cytotoxicity in retinal degeneration in rd mice. Invest Ophthalmol Vis Sci. 2005;46:2992–2999. [CrossRef] [PubMed]
ZeissCJ, JohnsonEA. Proliferation of microglia, but not photoreceptors, in the outer nuclear layer of the rd-1 mouse. Invest Ophthalmol Vis Sci. 2004;45:971–976. [CrossRef] [PubMed]
NicoteraP, OrreniusS. The role of calcium in apoptosis. Cell Calcium. 1998;23:173–180. [CrossRef] [PubMed]
WoodruffML, SampathAP, MatthewsHR, KrasnoperovaNV, LemJ, FainGL. Measurement of cytoplasmic calcium concentration in the rods of wild-type and transducin knock-out mice. J Physiol. 2002;542:843–854. [CrossRef] [PubMed]
WangKK. Calpain and caspase: can you tell the difference?. Trends Neurosci. 2000;23:59.
CroallDE, DeMartinoGN. Calcium-activated neutral protease (calpain) system: structure, function, and regulation. Physiol Rev. 1991;71:813–847. [PubMed]
ChiuK, LamTT, Ying LiWW, CaprioliJ, Kwong KwongJM. Calpain and N-methyl-d-aspartate (NMDA)-induced excitotoxicity in rat retinas. Brain Res. 2005;1046:207–215. [CrossRef] [PubMed]
DoonanF, DonovanM, CotterTG. Activation of multiple pathways during photoreceptor apoptosis in the rd mouse. Invest Ophthalmol Vis Sci. 2005;46:3530–3538. [CrossRef] [PubMed]
Paquet-DurandF, AzadiS, HauckSM, UeffingM, van VeenT, EkstromP. Calpain is activated in degenerating photoreceptors in the rd1 mouse. J Neurochem. 2006;96:802–814. [CrossRef] [PubMed]
NakagawaT, YuanJ. Cross-talk between two cysteine protease families: activation of caspase-12 by calpain in apoptosis. J Cell Biol. 2000;150:887–894. [CrossRef] [PubMed]
NakagawaT, ZhuH, MorishimaN, et al. Caspase-12 mediates endoplasmic-reticulum-specific apoptosis and cytotoxicity by amyloid-beta. Nature. 2000;403:98–103. [CrossRef] [PubMed]
FujitaE, KourokuY, JimboA, IsoaiA, MaruyamaK, MomoiT. Caspase-12 processing and fragment translocation into nuclei of tunicamycin-treated cells. Cell Death Differ. 2002;9:1108–1114. [CrossRef] [PubMed]
HetzC, Russelakis-CarneiroM, MaundrellK, CastillaJ, SotoC. Caspase-12 and endoplasmic reticulum stress mediate neurotoxicity of pathological prion protein. EMBO J. 2003;22:5435–5445. [CrossRef] [PubMed]
WootzH, HanssonI, KorhonenL, NapankangasU, LindholmD. Caspase-12 cleavage and increased oxidative stress during motoneuron degeneration in transgenic mouse model of ALS. Biochem Biophys Res Commun. 2004;322:281–286. [CrossRef] [PubMed]
MartinonF, TschoppJ. Inflammatory caspases: linking an intracellular innate immune system to autoinflammatory diseases. Cell. 2004;117:561–574. [CrossRef] [PubMed]
KalaiM, LamkanfiM, DeneckerG, et al. Regulation of the expression and processing of caspase-12. J Cell Biol. 2003;162:457–467. [CrossRef] [PubMed]
OubrahimH, WangJ, StadtmanER, ChockPB. Molecular cloning and characterization of murine caspase-12 gene promoter. Proc Natl Acad Sci USA. 2005;102:2322–2327. [CrossRef] [PubMed]
ShiraishiH, OkamotoH, YoshimuraA, YoshidaH. ER stress-induced apoptosis and caspase-12 activation occurs downstream of mitochondrial apoptosis involving Apaf-1. J Cell Sci. 2006;119:3958–3966. [CrossRef] [PubMed]
HardingHP, ZhangY, RonD. Protein translation and folding are coupled by an endoplasmic-reticulum-resident kinase. Nature. 1999;397:271–274. [CrossRef] [PubMed]
HardingHP, CalfonM, UranoF, NovoaI, RonD. Transcriptional and translational control in the mammalian unfolded protein response. Annu Rev Cell Dev Biol. 2002;18:575–599. [CrossRef] [PubMed]
PaschenW. Shutdown of translation: lethal or protective? unfolded protein response versus apoptosis. J Cereb Blood Flow Metab. 2003;23:773–779. [PubMed]
ColleyNJ, CassillJA, BakerEK, ZukerCS. Defective intracellular transport is the molecular basis of rhodopsin-dependent dominant retinal degeneration. Proc Natl Acad Sci USA. 1995;92:3070–3074. [CrossRef] [PubMed]
KuradaP, O’TousaJE. Retinal degeneration caused by dominant rhodopsin mutations in Drosophila. Neuron. 1995;14:571–579. [CrossRef] [PubMed]
RyooHD, DomingosPM, KangMJ, StellerH. Unfolded protein response in a Drosophila model for retinal degeneration. EMBO J. 2007;26:242–252. [CrossRef] [PubMed]
AcostaML, FletcherEL, AzizogluS, FosterLE, FarberDB, KalloniatisM. Early markers of retinal degeneration in rd/rd mice. Mol Vis. 2005;11:717–728. [PubMed]
LohrHR, KuntchithapauthamK, SharmaAK, RohrerB. Multiple, parallel cellular suicide mechanisms participate in photoreceptor cell death. Exp Eye Res. 2006;83:380–389. [CrossRef] [PubMed]
SanzMM, JohnsonLE, AhujaS, EkstromPA, RomeroJ, van VeenT. Significant photoreceptor rescue by treatment with a combination of antioxidants in an animal model for retinal degeneration. Neuroscience. 2007;145:1120–1129. [CrossRef] [PubMed]
Figure 1.
 
TUNEL immunofluorescence of the rd1 retina. (A) No TUNEL-positive cells were seen in the ONL at P8. (B) TUNEL-positive cells were first seen in the ONL at P10. (C) TUNEL-positive cells were most abundant at P12. (D) TUNEL-positive cells were less abundant at P14. (E) No TUNEL-positive cells were detected at P24. (F) The time course of the incremental alteration of the TUNEL-positive photoreceptor cells in the ONL in the rd1 retina. Data are expressed as mean ± SD.
Figure 1.
 
TUNEL immunofluorescence of the rd1 retina. (A) No TUNEL-positive cells were seen in the ONL at P8. (B) TUNEL-positive cells were first seen in the ONL at P10. (C) TUNEL-positive cells were most abundant at P12. (D) TUNEL-positive cells were less abundant at P14. (E) No TUNEL-positive cells were detected at P24. (F) The time course of the incremental alteration of the TUNEL-positive photoreceptor cells in the ONL in the rd1 retina. Data are expressed as mean ± SD.
Figure 2.
 
Retinal expressions of ER stress proteins in the wild-type and the rd1 mouse. (A) Time course for GRP78/BiP, procaspase-12/cleaved caspase-12, phospho-eIF2α, and phospho-PERK protein expression in the control and the rd1 retinas at each age group. (B) The relative level of GRP78/BiP, procaspase-12/cleaved caspase-12, phospho-eIF2α, and phospho-PERK protein expression in the rd1 mouse were quantified and corrected for the levels of β-actin protein expression. (C) P12 wild-type C3B mouse. P < 0.01 was considered significant compared with the control retina.
Figure 2.
 
Retinal expressions of ER stress proteins in the wild-type and the rd1 mouse. (A) Time course for GRP78/BiP, procaspase-12/cleaved caspase-12, phospho-eIF2α, and phospho-PERK protein expression in the control and the rd1 retinas at each age group. (B) The relative level of GRP78/BiP, procaspase-12/cleaved caspase-12, phospho-eIF2α, and phospho-PERK protein expression in the rd1 mouse were quantified and corrected for the levels of β-actin protein expression. (C) P12 wild-type C3B mouse. P < 0.01 was considered significant compared with the control retina.
Figure 3.
 
Localizations of GRP78/BiP protein in the wild-type and the rd1 mouse. (A) At P12, the positive GRP78/BiP staining of the C3B retina was mainly distributed in the INL (arrows). (B) At P10, the positive staining in the rd1 retina was also mainly in the INL (arrows), with no positive staining shown in the ONL or at the photoreceptor inner segments. (C) At P12, the GRP78/BiP-positive staining in the rd1 retina was most evident and was mainly located at the photoreceptor inner segments (arrows). The INL and ONL also showed positive labeling (arrows). (D) At P14, the positive staining in the INL of the rd1 retina was almost undetectable, and the intensity of positive labeling in the photoreceptor inner segments and ONL dramatically decreased, with limited positive staining evident (arrows). (E) At P24, no representative positive labeling was seen in the rd1 retina.
Figure 3.
 
Localizations of GRP78/BiP protein in the wild-type and the rd1 mouse. (A) At P12, the positive GRP78/BiP staining of the C3B retina was mainly distributed in the INL (arrows). (B) At P10, the positive staining in the rd1 retina was also mainly in the INL (arrows), with no positive staining shown in the ONL or at the photoreceptor inner segments. (C) At P12, the GRP78/BiP-positive staining in the rd1 retina was most evident and was mainly located at the photoreceptor inner segments (arrows). The INL and ONL also showed positive labeling (arrows). (D) At P14, the positive staining in the INL of the rd1 retina was almost undetectable, and the intensity of positive labeling in the photoreceptor inner segments and ONL dramatically decreased, with limited positive staining evident (arrows). (E) At P24, no representative positive labeling was seen in the rd1 retina.
Figure 4.
 
Localizations of caspase-12 protein in the wild-type and the rd1 mouse. (A) At P12, the caspase-12 immunofluorescence of the C3B retina was evenly distributed among all layers of the retina. (B) At P10, the immunofluorescence in the rd1 retina also labeled the outlines of cells in the ONL and the outlines of photoreceptor inner segments. (C) At P12, the caspase-12–positive labeling in the rd1 retina was most evident and was mainly located at the photoreceptor inner segments and in the ONL (arrows). (D) At P14, the positive labeling in the rd1 retina was also located at the photoreceptor inner segments and in the ONL (arrows), but it was weaker than at P12. (E) At P24, positive staining in the rd1 retina was almost undetectable.
Figure 4.
 
Localizations of caspase-12 protein in the wild-type and the rd1 mouse. (A) At P12, the caspase-12 immunofluorescence of the C3B retina was evenly distributed among all layers of the retina. (B) At P10, the immunofluorescence in the rd1 retina also labeled the outlines of cells in the ONL and the outlines of photoreceptor inner segments. (C) At P12, the caspase-12–positive labeling in the rd1 retina was most evident and was mainly located at the photoreceptor inner segments and in the ONL (arrows). (D) At P14, the positive labeling in the rd1 retina was also located at the photoreceptor inner segments and in the ONL (arrows), but it was weaker than at P12. (E) At P24, positive staining in the rd1 retina was almost undetectable.
Figure 5.
 
Localizations of phospho-eIF2α protein in the wild-type and the rd1 mouse. (A) At P12, positive phospho-eIF2α staining in the C3B retinas labeled the outlines of the cells in the ONL and INL. (B) At P10, positive labeling in the rd1 retina was weaker and less intense than in the control retina. (C) At P12, the positive phospho-eIF2α staining in the rd1 retina was mainly located in the photoreceptor inner segments and in the ONL (arrows). (D) At P14, the positive staining in the rd1 retina markedly decreased, and limited positive staining showed in the ONL and in the photoreceptor inner segments (arrows). (E) At P24, the positive staining in all parts of the rd1 retina was almost undetectable.
Figure 5.
 
Localizations of phospho-eIF2α protein in the wild-type and the rd1 mouse. (A) At P12, positive phospho-eIF2α staining in the C3B retinas labeled the outlines of the cells in the ONL and INL. (B) At P10, positive labeling in the rd1 retina was weaker and less intense than in the control retina. (C) At P12, the positive phospho-eIF2α staining in the rd1 retina was mainly located in the photoreceptor inner segments and in the ONL (arrows). (D) At P14, the positive staining in the rd1 retina markedly decreased, and limited positive staining showed in the ONL and in the photoreceptor inner segments (arrows). (E) At P24, the positive staining in all parts of the rd1 retina was almost undetectable.
Figure 6.
 
Localizations of phospho-PERK protein in the wild-type and the rd1 mouse. (A) At P12, positive phospho-PERK staining in the C3B retina was primarily distributed in the photoreceptor inner segments; the INL also showed scattered positive labeling. (B) At P10, phospho-PERK immunofluorescence in the photoreceptor inner segments and in the INL of the rd1 retina markedly increased compared with the control retina. (C) At P12, positive staining in the photoreceptor inner segments and the INL of the rd1 retina were still extensive. (D) At P14, positive labeling in the photoreceptor inner segments of the rd1 retina decreased, but positive labeling in the INL increased. (E) At P24, positive labeling in the photoreceptor inner segments of the rd1 retina decreased to one layer, and positive labeling in the INL increased.
Figure 6.
 
Localizations of phospho-PERK protein in the wild-type and the rd1 mouse. (A) At P12, positive phospho-PERK staining in the C3B retina was primarily distributed in the photoreceptor inner segments; the INL also showed scattered positive labeling. (B) At P10, phospho-PERK immunofluorescence in the photoreceptor inner segments and in the INL of the rd1 retina markedly increased compared with the control retina. (C) At P12, positive staining in the photoreceptor inner segments and the INL of the rd1 retina were still extensive. (D) At P14, positive labeling in the photoreceptor inner segments of the rd1 retina decreased, but positive labeling in the INL increased. (E) At P24, positive labeling in the photoreceptor inner segments of the rd1 retina decreased to one layer, and positive labeling in the INL increased.
×
×

This PDF is available to Subscribers Only

Sign in or purchase a subscription to access this content. ×

You must be signed into an individual account to use this feature.

×