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Fig. 3. Abnormal development of parasympathetic, but not sympathetic, ganglia in Ret^DN/+ mice. (A) The number, size and target organ innervation of sphenopalatine ganglia (SPG) parasympathetic neurons was decreased in Ret^DN/+ mice, when compared with wild-type (wt), Ret^9/+ and Ret^TGM/+ mutant animals. (Top left) Thionin-stained SPG neuron number per animal was decreased in Ret^DN/+ mice (P0). (Bottom left) The size of SPG neurons in Ret^DN/+ mice was 40-50% smaller than wild-type SPG neurons (n=3, ^*=P<0.01, mean± s.e.m.). (Top right) Thionin-stained Ret^DN/+ SPG neurons appear smaller than wild type (wt) (P21; scale bar: 50 µm). (Bottom right) The SPG target organ, the intraorbital harderian gland, showed a decrease of PGP9.5-stained nerve fibers in Ret^DN/+ mice compared with wild-type littermates where nerve fibers surround almost every acinus (arrowheads) Scale bar: 70 µm. (B) The sympathetic nervous system developed normally in Ret^DN/+ mice, as revealed by whole-mount tyrosine hydroxylase (TH) immunostaining (P0). (Top) The superior cervical ganglion (SCG) was normally located, and had normal projections to the eye (vertical arrowheads) and submandibular gland (sm, horizontal arrowheads) in Ret^DN/+ and wild-type (wt) mice (scale bar: 400 µm). (Bottom) The sympathetic chain ganglia and their neurite outgrowths (arrowheads) also appeared normal in Ret^DN/+ mice (scale bar: 300 µm). Abnormal development of the SCG (located more caudally near the stellate ganglion, stg; black circle indicates the expected SCG location), SCG projections (top right; scale bar: 800 µm), and sympathetic chain (bottom right; scale bar: 600 µm) in Ret-null mice (Ret^TGM/TGM) is shown for reference.

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