Distribution of cholesterol and galactosylceramide in rat cerebellar white matter.

White matter and the inner granular layer of rat cerebellum was analysed by imaging time-of-flight secondary-ion mass spectrometry (TOF-SIMS) equipped with a Bi+ ion cluster gun. Samples were prepared by high pressure freezing, freeze-fracturing and freeze drying or by plunge freezing and cryostat sectioning. The identified and localized chemical species were: sodium, potassium, phosphocholine, cholesterol and galactosylceramide (GalC) with carbon chain lengths C18:0 (N-stearoyl-galactosylceramide) and C24:0 (N-lignoceroylgalactosylceramide) with CH24:0 (hydroxy-lignoceroylgalactosylceramide). We report new findings regarding the organization of myelin in white matter. One is cholesterol-rich, ribbon-shaped 10-20 microm areas excluding Na+ and K+. The second finding is the different distribution of GalC C18 and GalC C24 in relation to these areas, where GalC C18 was localized in cholesterol-rich areas and GalC C24 was localized in Na/K-enriched areas. The distribution of GalC was in small spots, homogeneous in size, of 0.8-1.5 microm. Sample preparation with high pressure freezing allowed separate localization of sodium and potassium in tissue samples.