Assessment of gene and protein expression of cholinergic markers.

(A, D & G) Quantitative real-time PCR (qRT-PCR) assessed expression levels of VAChT, ChAT and CHT1 mRNA from retinal tissue of VAChT^{Six3-Cre-flox/flox} (n = 5) and littermate control mice (VAChT^flox/flox, n = 6). (B, E & H) Immunoblotting followed by (C, F & I) densitometry assessed protein levels of cholinergic proteins (A) Expression of VAChT mRNA in the retina of VAChT^{Six3-Cre-flox/flox} mice is significantly reduced relative to control. Expression of VAChT protein ChAT increased. (B) Immunoblotting. (B, E, H) Expression levels of VAChT, ChAT and CHT1 protein in the retina of VAChT^{Six3-Cre-flox/flox} and littermate control mice were quantified using densitometry. Expression of VAChT protein in the retina of VAChT^{Six3-Cre-flox/flox} mice (n = 5) is reduced relative to littermate control (n = 6) (C). Synaptophysin protein was used as a loading control for VAChT, while actin protein was used for ChAT and CHT1. Values in (A) and (C) represent mean ± SEM, γ, P < 0.01; β, P < 0.001 versus control mice.