Mouse models of retinal ganglion cell death and glaucoma

Abstract

Once considered too difficult to use for glaucoma studies, mice are now becoming a powerful tool in the research of the molecular and pathological events associated with this disease. Often adapting technologies first developed in rats, ganglion cell death in mice can be induced using acute models and chronic models of experimental glaucoma. Similarly, elevated IOP has been reported in transgenic animals carrying defects in targeted genes. Also, one group of mice, from the DBA/2 line of inbred animals, develops a spontaneous optic neuropathy with many features of human glaucoma that is associated with IOP elevation caused by an anterior chamber pigmentary disease. The advent of mice for glaucoma research is already having a significant impact on our understanding of this disease, principally because of the access to genetic manipulation technology and genetics already well established for these animals.

Fig. 1

Acute induction of retinal ganglion cell loss in mice. (A) Histograph showing the reduction of cells in the ganglion cell layer after a crush lesion of the optic nerve. Quantification is measured as a percentage of the cells counted in the untreated fellow eyes of each mouse examined. The data shown here represents the decrease in cells observed in BALB/cByJ mice after crush. (B) Histograph showing the reduction of cells in the ganglion cell layer 4 days after intravitreal injection of the glutamate analog N-methyl-D-aspartate (NMDA). Cells are lost in a dose-dependent fashion in response to NMDA injection (total nmoles injected are shown). The data shown here represents cell loss in CB6F1 hybrid mice (F1 mice from a cross of a BALB/c female with a C57BL/6 male). (C, D) Nissl-stained flat-mounted retinas of a mouse 2 weeks after optic nerve crush in an SJL/J mouse. The untreated fellow eye (C) shows a high density of cells in the ganglion cell layer, which is mounted facing up on the slide. The crushed eye (D) shows a dramatic loss of cells. Scale bar = 80 μm (E, F) Nissl-stained flat-mounted retinas of a C57BL/6J mouse 2 days after injection with NMDA. The uninjected eye (E) shows a typical compliment of cells in the ganglion cell layer. In the fellow injected eye (F), however, fewer cells remain and there is evidence of apoptotic fragments from dead and dying cells. Scale bar = 10 μm.

Fig. 2

Iris transillumination defects in aging DBA/2J mice. As these mice age, the deterioration of the iris stroma and the dispersion of sloughed pigment is readily detected as transillumination of light reflected off the lens. These defects first become evident by 6–8 months of age. Scale bar = 0.2 mm.

Fig. 3

Optic nerve disease and retinal ganglion cell loss in DBA/2J mice. (A) DiI-labeled optic nerves of a young (3 months) DBA/2J mouse. In this method, nerves are labeled postmortem by introducing crystals of DiI into the optic nerve head of each eye. The dye is able to diffuse along the axons of the optic nerve, but since the dye does not transfer from one cell to another, only contiguous axons become labeled. In young mice, it is possible to label nerves all the way to the optic chiasm (arrow) by this technique. (OD – right optic nerve; OS – left optic nerve). (B) DiI-labeled optic nerves of an old (10 months) DBA/2J mouse, that shows signs of glaucomatous damage. In this mouse, the left nerve (OS) has yet to show significant signs of damage, while the right (OD) nerve is nearly completely degenerated. Scale bar = 250 μm (arrow – optic chiasm). (C–F) Nissl-stained flat-mounted retinas from DBA/2J mice at different ages. A similar region of the retina from each eye is shown. Typically, as these mice age, cell density in the ganglion cell layer is diminished. The majority of eyes in mice between 10 and 13 months clearly show signs of cell loss. Scale bar = 10 μm. (G) Flat-mounted retina from a 10.5 month old DBA/2J mouse carrying the Fem1c^Rosa3 marker transgene. This promoter trap marker fusion protein is inserted into the first intron of the mouse Fem1c gene and is predominantly expressed by ganglion cells in the retina (Schlamp et al., 2004). Healthy ganglion cells express the fusion protein, which contains β-galactosidase, and thus can be stained blue using X-gal as a substrate. During glaucomatous retinal damage, DBA/2J mice exhibit a loss of blue cell staining, typically in wedge-shaped patterns. (H) Computer enhanced image of the retina shown in (G), in which X-gal stain deposits are recognized on the basis of color parameters. Large to small cells are depicted as different colored spots on the computer two-dimensional plot. This retina clearly exhibits two distinct wedges of cell loss in different stages of completion. Scale bar = 100 μm.