<i>Bax</i> Deficiency Prevents Glaucomatous RGC Death

<div><p>To determine the effects of BAX deficiency on RGC death in glaucoma, we analyzed RGC layer cells at stages with and without glaucomatous optic nerve damage (see <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.0010004#s4" target="_blank">Materials and Methods</a>). All shown images are from a similar region of the superior, peripheral retina.</p> <p>(A and B) In both <i>Bax^+/+</i> (A) and <i>Bax</i>^−/− (B) mice without glaucomatous optic nerve damage, the retinas appear healthy. The retinas of both genotypes are similar except that <i>Bax</i>^−/− mice have extra RGCs (since BAX is important in normal developmental RGC death [<a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.0010004#pgen-0010004-b39" target="_blank">39</a>]).</p> <p>(C and D) In contrast, an obvious difference was evident between the retinas of <i>Bax</i>^+/+ and <i>Bax</i>^−/− mice that had all suffered severe glaucomatous damage with 95% or more axon degeneration. As expected, for <i>Bax</i>^+/+ retinas (C) from eyes with 95% or more optic nerve axon loss, there was a noticeable decrease in RGC layer cells (compare [C] to [A]). In contrast, retinas from <i>Bax</i>^−/− mice with correspondingly damaged optic nerves (D) had suffered no obvious loss of RGC layer cells (compare [D] to [B]). This suggests that BAX is required for RGC death in DBA/2J glaucoma. As is well established for both RGCs and other neurons, <i>Bax</i>^−/− RGCs that survive without axons have a shrunken morphology [<a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.0010004#pgen-0010004-b38" target="_blank">38</a>,<a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.0010004#pgen-0010004-b82" target="_blank">82</a>]. This is clearly evident in the <i>Bax</i>^−/− glaucomatous mice (D).</p> <p>(E) RGC layer cell counts for eyes with 95% or more axon degeneration confirmed that BAX is necessary for RGC death in this glaucoma. To allow comparison between genotypes, the percent of surviving cells is shown (% soma in mice with 95% or more axon loss compared to mice of the same genotype without glaucomatous damage). At 12 mo of age, <i>Bax</i>^+/+ mice had 61.4% ± 3.8% of their RGC layer cells remaining, <i>Bax</i>^−/− mice had no appreciable cell loss (101% ± 5.3%). The RGC layer cells of <i>Bax</i>^+/− mice were also protected (89.2% ± 5.7%). The <i>p</i> values comparing differences in cell counts between nonglaucomatous and very severely glaucomatous (≥ 95% axon loss) eyes of the same genotype were: <i>Bax</i>^+/+, <i>p</i> < 0.001; <i>Bax</i>^+/−, <i>p</i> = 0.207; <i>Bax</i>^−/−, <i>p</i> = 0.426. No cell loss was seen in <i>Bax</i>^−/− mice even out to 18 mo (94.8% ± 4.4% cells surviving, <i>p</i> = 0.524 compared to nonglaucomatous <i>Bax</i>^−/− mice). These findings show that BAX gene dosage has an important effect on the susceptibility of RGCs to glaucomatous death. Scale bar, 50 μm.</p>