Inhibition of histone deacetylase activity prevents the silencing of a reporter gene after optic nerve crush (ONC). (A) Western blot analysis of whole retinal acetylated histone H4 (AcH4) following intraperitoneal injection of trichostatin A (TSA) or vehicle control (DMSO). The times indicated are the time elapsed between injection of TSA and collection of the retinas. TSA resulted in hyperacetylation of H4 at all time points after treatment. (B) Retinal ganglion cell (RGC) specific gene expression was monitored as a function of βGalactosidase activity from the βGeo gene trap cassette inserted into Fem1c (see Methods). Histograph showing total retinal βGalactosidase activity 5 days post ONC. Mean ± SE is shown (n = 8-12 retinas per condition). Retinas with no injection or after DMSO vehicle injection (i.p.) exhibited a 50-75% decrease in βGEO activity after ONC. Conversely, TSA treated mice exhibited significantly higher βGEO activity following ONC than with crush alone or crush with DMSO injection (*P = 0.015 and P = 0.003, respectively). (C) Retinal wholemounts showing histochemical staining of βGEO enzyme activity, 5 days after ONC. The midperipheral region of the superior quadrant of each retina is shown. In this experiment, eyes were treated by intravitreal injections of DMSO vehicle or TSA at the time of surgery. Control retinas (control) typically showed prominent labeling of RGCs. Retinas from eyes that underwent crush alone (crush) or crush with DMSO treatment (DMSO) both showed a marked loss of staining of RGCs. βGEO activity remained robust in eyes that received TSA injections. Scale bar = 25 μm.