TNFα mediates extrinsic apoptosis of retinal ganglion cells in a dose-dependent manner. (A) A single intraocular injection of TNFα was delivered and the total number of cells in the ganglion cell layer (GCL) was obtained from 4’,6-diamidino-2-phenylindole-stained retinal whole mounts after 8 weeks. No cell loss was detected at 6 weeks (data not shown). Wild-type (black bars) and Bax -/- mice (gray bars) exhibited dose-dependent loss of total cells, indicating cell death occurred via a Bax-independent mechanism. The percent change was calculated by comparing the injected eye to the control eye. Statistics were calculated relative to the PBS-injected control group and are presented as mean ± SE (*P <0.005, **P <0.001); n ≥3 for all genotypes and conditions. (B-E) Retinal whole mounts were also stained for the retinal ganglion cell-specific transcription factor BRN3A. Approximately 40% of the neurons in the GCL stain for BRN3A in control retinas of both wild-type (B) and Bax -/-
(D) mice ,. Retinas exposed to a single injection of TNFα, 8 weeks prior, exhibit a complete absence of BRN3A staining (wild-type mice in C, and Bax-deficient mice in E). Scale bar =15 μm.