Characterization of phosphonoglycosphingolipids containing pyruvate: localization in Aplysia nerve bundles

A phosphonoglycosphingolipid, designated as FGL-IIb, was first identified in nerve fibers of Aplysia kurodai by two-dimensional TLC (Abe, S. et al. (1986) Biomed. Res. 7, 47-51), and its chemical structure has been determined to be 3,4-O-(1-carboxyethylidene)]Gal beta 1----3GalNac alpha 1----3(Fuc alpha 1----2)(2-aminoethylphosphonyl----6)Gal beta 1----4Glc beta 1----1ceramide (Araki, S. et al., submitted). Cryostat and paraffin sections of the nervous tissue and skin of Aplysia were examined immunohistochemically with antiserum against FGL-IIb. With this antiserum, only nerve bundles were stained distinctly: nerve cells in ganglia and in subcutaneous and muscular tissues and other cell elements were not stained. From histochemical findings in cryostat sections pretreated with chloroform-methanol (2 : 1, v/v) and from the results of Western blot analysis of the nervous tissue, the staining was concluded to be due to glycolipid antigens. The antiserum reacted with FGL-IIb and other phosphonoglycosphingolipids named FGL-I, FGL-IIa, FGL-V, and F-9 on TLC plates. This reactivity of FGL-IIb was abolished by mild acid-methanol treatment, and the lost reactivity was recovered by alkaline hydrolysis. These findings suggest that the free carboxyl group of the pyruvic acid of FGL-IIb is essential for the immunological reaction and that all the glycolipids listed above have the same epitope as that of FGL-IIb. Immunohistochemical findings indicated that these glycolipids including FGL-IIb are localized specifically in nerve bundles of Aplysia.