Shh effects on ventral spinal cultures.A, At concentrations of 25 ng/ml and higher, Shh promotes neuronal survival as gauged by staining for tubulin βIII. The majority of the cells stain positively for GABA, whereas a subset of cells stains for the nuclear marker of spinal interneurons Lim-1/2 (error bars represent SEM) (tubulin βIII, p < 0.001 at 25 and 50 ng/ml; lim 1/2, p < 0.001 at 5, 10, 25, and 50 ng/ml; GABA, p < 0.001 at 25 and 50 ng/ml). Typical staining for Lim-1/2 in the E14 rat spinal cord (B) (scale bar, 100 μm), and spinal neurons cultured in the presence of 50 ng/ml Shh (C). Scale bar, 20 μm.