Figure 2. Optic nerve crush decreased AQP4 protein and mRNA levels. Following retinal injuries with optic nerve crush, retinas were dissected and plasma membrane proteins were isolated or total RNA was isolated and transcribed into cDNA. Thirty microgram protein was loaded into each lane. Immunoreactive bands for AQP4 and β-tubulin 2, 7, and 14 days after optic nerve crush showing a significant reduction in AQP4 protein levels. Quantitative measurement using western blot showed that optic nerve crush decreased AQP4 protein levels by ~40% (60±10, 63±10, 38±10, at 2, 7, and 14 day, respectively, n=7, A). Densitometric quantification is shown in B. Data are expressed as a ratio of the treated to control or sham value and each measurement represents mean±SEM *Denote statistical significance of AQP4 protein levels in optic nerve crushed retinas versus sham (p<0.005) as determined by one-way ANOVA and Tukey multiple comparison test. Optic nerve crush also decreased AQP4 transcripts significantly (61±5%, 60±8%, 58±6, at 2, 7, and 14 day, respectively, n=7), as determined by quantitative real-time PCR (B). Gene expression data of AQP4 is calculated after normalizing with ACTB. **Denote significant differences compared with sham-retinas at p<0.05. Abbreviations: sham eye (C), and crushed (R).