shRNA-mediated knockdown of Six3 and Six3OS in the developing retina alters bipolar cell and Muller glial development. (A) Six3 knockdown and Six3OS knockdown both lead to an increase in Muller glia and a decrease in rod bipolar cells as measured by immunostaining of dissociated electroporated retinal cells. Simultaneous knockdown of both Six3 and Six3OS resulted in a normal number of rod bipolar and Muller glia, but a decrease in syntaxin (Syn)-positive amacrine cells was observed. *P < 0.05. GS, glutamine synthetase. (B-E) Section immunohistochemistry demonstrates a decrease in GFP-positive bipolar cells following knockdown of either Six3OS or Six3, but not both in combination (white arrowheads). Chx10 (ceh-10 homeodomain containing homolog (C. elegans)) was used as pan-bipolar cell marker. GCL, ganglion cell layer; INL, inner nuclear layer; ONL, outer nuclear layer; OS, outer segment.