Immunohistochemical evidence of synaptic retraction, cytoarchitectural remodeling, and cell death in the inner retina of the rat model of Oygen-Induced Retinopathy (OIR)

Purpose. Postnatal exposure to hyperoxia destroys the plexiform layers of the neonatal rat retina, resulting in significant electroretinographic anomalies. The purpose of this study was to identify the mechanisms at the origin of this loss. Methods. Sprague-Dawley (SD) and Long Evans (LE) rats were exposed to hyperoxia from birth to postnatal day (P) 6 or P14 and from P6 to P14, after which rats were euthanatized at P6, P14, or P60. Results. At P60, synaptophysin staining confirmed the lack of functional synaptic terminals in SD (outer plexiform layer [OPL]) and LE (OPL and inner plexiform layer [IPL]) rats. Uneven staining of ON-bipolar cell terminals with mGluR6 suggests that their loss could play a role in OPL thinning. Protein kinase C(PKC)-α and recoverin (rod and cone ON-bipolar cells, respectively) showed a lack of dendritic terminals in the OPL with disorganized axonal projections in the IPL. Although photoreceptor nuclei appeared intact, a decrease in bassoon staining (synaptic ribbon terminals) suggests limited communication to the inner retina. Findings were significantly more pronounced in LE rats. An increase in TUNEL-positive cells was observed in LE (inner nuclear layer [INL] and outer nuclear layer [ONL]) and SD (INL) rats after P0 to P14 exposure (425.3%, 102.2%, and 146.3% greater than control, respectively [P < 0.05]). Conclusions. Results suggest that cell death and synaptic retraction are at the root of OPL thinning. Increased TUNEL-positive cells in the INL confirm that cells die, at least in part, because of apoptosis. These findings propose a previously undescribed mechanism of cell death and synaptic retraction that are likely at the origin of the functional consequences of hyperoxia.

Loss of Outer Retinal Neurons and Circuitry Alterations in the DBA/2J Mouse

Purpose. The DBA/2J mouse line develops essential iris atrophy, pigment dispersion, and glaucomatous age-related changes, including an increase of IOP, optic nerve atrophy, and retinal ganglion cell (RGC) death. The aim of this study was to evaluate possible morphological changes in the outer retina of the DBA/2J mouse concomitant with disease progression and aging, based on the reduction of both the a- and b-waves and photopic flicker ERGs in this mouse line. Methods. Vertically sectioned DBA/2J mice retinas were evaluated at 3, 8, and 16 months of age using photoreceptor, horizontal, and bipolar cell markers. Sixteen-month-old C57BL/6 mice retinas were used as controls. Results. The DBA/2J mice had outer retinal degeneration at all ages, with the most severe degeneration in the oldest retinas. At 3 months of age, the number of photoreceptor cells and the thickness of the OPL were reduced. In addition, there was a loss of horizontal and ON-bipolar cell processes. At 8 months of age, RGC degeneration occurred in patches, and in the outer retina overlying these patches, cone morphology was impaired with a reduction in size as well as loss of outer segments and growth of horizontal and bipolar cell processes into the outer nuclear layer. At 16 months of age, connectivity between photoreceptors and horizontal and bipolar cell processes overlying these patches was lost. Conclusions. Retinal degeneration in DBA/2J mice includes photoreceptor death, loss of bipolar and horizontal cell processes, and loss of synaptic contacts in an aging-dependent manner.

Inherited Photoreceptor Degeneration Causes the Death of Melanopsin-Positive Retinal Ganglion Cells and Increases Their Coexpression of Brn3a

Purpose: To study the population of intrinsically photosensitive retinal ganglion cells (melanopsin-expressing RGCs, m+RGCs) in P23H-1 rats, a rat model of inherited photoreceptor degeneration. Methods: At postnatal (P) times P30, P365, and P540, retinas from P23H dystrophic rats (line 1, rapid degeneration; and line 3, slow degeneration) and Sprague Dawley (SD) rats (control) were dissected as whole-mounts and immunodetected for melanopsin and/or Brn3a. The dendritic arborization of m+RGCs and the numbers of Brn3a+RGCs and m+RGCs were quantified and their retinal distribution and coexpression analyzed. Results: In SD rats, aging did not affect the population of Brn3a+RGCs or m+RGCs or the percentage that showed coexpression (0.27%). Young P23H-1 rats had a significantly lower number of Brn3a+RGCs and showed a further decline with age. The population of m+RGCs in young P23H-1 rats was similar to that found in SD rats and decreased by 22.6% and 28.2% at P365 and P540, respectively, similarly to the decrease of the Brn3a+RGCs. At these ages the m+RGCs showed a decrease of their dendritic arborization parameters, which was similar in both the P23H-1 and P23H-3 lines. The percentage of coexpression of Brn3a was, however, already significantly higher at P30 (3.31%) and increased significantly with age (10.65% at P540). Conclusions: Inherited photoreceptor degeneration was followed by secondary loss of Brn3a+RGCs and m+RGCs. Surviving m+RGCs showed decreased dendritic arborization parameters and increased coexpression of Brn3a and melanopsin, phenotypic and molecular changes that may represent an effort to resist degeneration and/or preferential survival of m+RGCs capable of synthesizing Brn3a.