GANGLIOSIDES IN DEVELOPING RAT BRAIN: ISOLATION AND COMPOSITION OF SUBCELLULAR MEMBRANES ENRICHED IN GANGLIOSIDES

Abstract

Neurones in the central nervous system of vertebrates contain considerable amounts of a complex group of acidic glycosphingolipids, the gangliosides. Determination of gangliosides, cholesterol, phospholipid phosphorus, neutral glycolipid hexose, and protein in developing rat brain showed that the deposition of gangliosides is predominantly a premyelin event. A light membrane fraction enriched in gangliosides relative to protein was obtained from neonatal rat-brain crude mitochondrial fractions by sucrose density-gradient centrifugations. Adult rat-brain fractions of similar density were not enriched in gangliosides because of the presence of myelin. If the initial homogenization and the separation of the crude mitochondrial fraction were carried out in 0.32 M sucrose at pH 9.2, the ganglioside enrichment of the light membrane fraction from both adult and neonatal rat brain was doubled. By further separation of the light membrane fraction on a second density gradient, particulate material was obtained from neonatal rat brain which consisted almost entirely of vesicular membrane elements. Based on dry weight, it contained gangliosides 7–9%, phospholipids 36–40%, cholesterol 5–7%, neutral glycolipids 1–3%, protein 28–29%, and cations (Na+, K+, Ca2+, Mg2+) 8%. This membrane fraction is likely derived from the neuronal plasma membrane or the endoplasmic reticulum.