Localization and imaging of sialylated glycosphingolipids in brain tissue sections by MALDI mass spectrometry

Abstract

In this study, we describe a simple and efficient method for mapping the distribution and localization of all sialylated sphingoglycolipids present in coronal mouse brain sections using a conventional axial matrix-assisted laser desorption/ionization time of flight. A single scan of a histological tissue section gives a complete profile of ganglioside species without derivatization or labeling. We have developed and tested a new matrix preparation (2,6-dihydroxyacetophenone [DHA]/ammonium sulfate/heptafluorobutyric acid [HFBA]) to maximize the detection of all ganglioside species; the ammonium sulfate limits the formation of salt adducts, while the addition of HFBA increases the stability of DHA in a vacuum, thus facilitating imaging applications. Our results, in both extracted samples and whole tissue sections using negative ion reflectron and linear modes, show differences in localization in several brain regions depending on the sialic acids and the ceramide-associated core gangliosides.

Fig. 1

Scheme of ganglioside biosynthesis (Svennerholm 1964). Mouse brain ganglioside species that are detected using DHA/ammonium sulfate/HFBA 0.05% matrix are represented by structural forms with d18:1/C18:0 ceramide moiety. All sialylated ganglioside species of the a- and can be detected in mouse brain. Asialoganglioside species and the other minor sialylated ganglioside species cannot be detected because they are not present or do not contain sialic acid. Cer, ceramide; GlcCer, glucosylceramide; LacCer, lactosylceramide. ▲, glucose; ●, galactose; ▪, N-acetylgalactosamine; ◊, sialic acid.

Fig. 2

Mass spectra of ganglioside species by MALDI/MS in negative ion mode using DHB in 0.1% TFA matrix in linear mode (A) and reflectron mode (C).Detected ganglioside species by saturated DHA/ammonium sulfate 3 mM/HFBA 0.05% were represented using linear mode (B) and reflectron mode (D).

Fig. 3

MALDI imaging mass spectrometry of gangliosides using saturated DHA/ammonium sulfate 125 mM/HFBA 0.05% in negative ion reflectron mode. Major gangliosides species are detected such as GM3, GM1, GD1, GT1 and GQ1. These results show differences in localization in several brain regions depending on the sialic acids and the ceramide core associated gangliosides. All other sialylated glycosphingolipids present in these mice brain sections can be mapped (data not shown).

Fig. 4

MALDI imaging of ganglioside species detected in mouse brain section using negative linear ion mode. The use of linear ion mode for mapping GM1, GD1 and GT1 ganglioside species gave the same results obtained using reflectron ion mode for the two species d18:1/C18:0 and d20:1/C18:0. Fragmentation of gangliosides was also observed. However, the comparison of the mapped areas confirms the presence of GM1 in the corpus callosum.