Brain Ganglioside Composition Research Articles

Role of gangliosides in brain aging and neurodegeneration

Abstract Gangliosides are membrane glycosphingolipids bearing sialic acid residues. Within membranes, gangliosides are specifically enriched in highly organized domains, lipid rafts, and are attributed with diverse functions such as intercellular interactions, cell recognition, neurotransmission, and signal transduction. The highest concentration and variability of ganglioside structures are found in the human brain. Specific temporal and regional distribution of brain gangliosides has been reported; moreover, gangliosides may serve as markers of neurodevelopmental stages, aging and neurodegeneration. Brain ganglioside content and composition as well as ganglioside metabolism are altered in Alzheimer’s disease. It appears that the alterations of ganglioside metabolism leading to changes in membrane physico-chemical properties are not merely a consequence of primary pathology, but may also be involved in the early pathogenesis of Alzheimer’s disease through documented effects on APP proteolytic processing and amyloid aggregation. Investigations of glycolipid metabolic alterations which accompany neurodegenerative disorders provide insight into pathogenetic mechanisms and enable recognition of diagnostic markers as well as molecular structures acting as therapeutic tools interfering with cascade of pathological events.

Analysis of human hippocampus gangliosides by fully-automated chip-based nanoelectrospray tandem mass spectrometry

Modern microfluidic devices are currently introduced in electrospray (ESI) mass spectrometry (MS), tending to substitute the classical capillary-based ESI infusion. Automated systems using the combination of robotized sample handling and chip-based ESI are significantly increasing the analysis reproducibility, precision, throughput, and efficiency. In the last couple of years our group developed the chip-based ESI–MS approach for glycomics in biomedical research and applied it for oligosaccharide, glycopeptide and ganglioside investigation. Here we report upon the optimization and application of this modern technique for the analysis of differential ganglioside expression patterns in human fetal and adult hippocampus. By this methodology, ganglioside species exhibiting high degree of heterogeneity in the ceramide motifs and biologically-relevant modifications could be identified in human hippocampus. The ultra-high reproducibility of the experiments uniquely provided by the chip-ESI approach allowed for a reliable MS-based ganglioside comparative assay. Moreover, the particular feature of chip ESI-tandem MS to provide structural information at high sensitivity was useful for detailed characterization of hippocampus-associated species. The experimental data presented in this study indicate the benefits of microfluidic/MS for determination of the topospecific brain ganglioside composition and development-related changes in their expression, which might be of high value in clinical investigation and for studies related to ganglioside-based therapy of central nervous system diseases.

New Approaches to Analysis of Variations in Brain Ganglioside Composition Related to Idioadaptations and Aromorphosis of Vertebrates

For objective interpretation of results of comparative studies of lipids, performed for many years at Institute of Evolutionary Physiology and Biochemistry, it is essentially important to use modern methods of analysis, first of all, polythetic methods of cluster analysis. Using the unweighed pair group method of the mean values, it has been shown that the dendrograms constructed on the basis of analysis of characters of molecular organization of the carbohydrate part of the ganglioside molecule correspond on the whole to the tree of classical taxonomy of vertebrates. Carbohydrate components of glycoconjugates, including gangliosides, play an important role in intercellular interactions, in processes of cell adhesion, recognition, and apoptosis. It may be suggested that changes of the carbohydrate component of the ganglioside molecule in the course of evolution of vertebrates contribute to achievement of a qualitatively new level of biochemical organization of organs and tissues, providing a possibility of aromorphic modifications of organization of animals, which are essential for the appearance of new classes. On the contrary, analysis of the data on molecular organization of a fatty acid component of vertebrate brain gangliosides by the same method could not reveal any relation of the species position on the dendrogram to its taxonomic belonging. At the same time, it was found out that the content of various fatty acids (FA) of brain gangliosides of ectothermic vertebrates correlated statistically significantly with the environmental temperature, positively in the case of saturated acids and negatively in the case of unsaturated ones. This agrees with the previously obtained data on the significance of differences in FA composition of warm-water and cold-water stenothermic species of teleosts. Changes of the FA composition of brain gangliosides seem to contribute to processes of idioadaptations to environmental temperature in various groups of ectothermic vertebrates, these adaptations aimed at maintaining properties of cell membranes, which are optimal for their vital activity.

Novel polysialogangliosides of skate brain structural determination of tetra, penta and hexasialogangliosides with a NeuAc-GalNAc linkage.

The gangliosides in the brain of a cartilaginous fish, skate (Bathyraja smirnovi), have been isolated and characterized by means of methylation analysis, antibody binding, enzymatic hydrolysis and MALDI-TOF MS. In addition to gangliosides with known structures (GM2, fucosyl-GM1, GD3, GD2, GT3 and GT2), five polysialogangliosides were isolated and characterized as having the following structures. (1) IV3NeuAc, III6NeuAc, II3NeuAc-Gg4Cer; (2) IV3NeuAc2, III6NeuAc, II3NeuAc-Gg4Cer; (3) IV3NeuAc, III6NeuAc, II3NeuAc2-Gg4Cer; (4) IV3NeuAc, III6NeuAc, II3NeuAc3-Gg4Cer; and (5) IV3NeuAc2, III6NeuAc, II3NeuAc3-Gg4Cer. These structures are 'hybrid-type' which comprise combinations of alpha-series and either a, b or c-series structures. Three gangliosides (2), (4) and (5), were novel. The main features of the ganglioside composition of skate brain were an abundance of gangliotriaosyl species, a lack of gangliotetraosyl species (except fucosyl-GM1), and an abundance of hybrid-types. These characteristics closely resemble those in shark brain which we reported previously [Nakamura, K., Tamai, Y. & Kasama, T. (1997) Neurochem. Int. 30, 593-604]. Two of the hybrid-type gangliosides (1) and (4), were examined for their neuritogenic activity toward cultured neuronal cells (Neuro-2A), and were found to have more potent activity than nonhybrid-type gangliosides such as GM1.

Kidney lipids in galactosylceramide synthase-deficient mice: absence of galactosylsulfatide and compensatory increase in more polar sulfoglycolipids

UDP-galactose:ceramide galactosyltransferase (CGT) catalyzes the final step in the synthesis of galactosylceramide (GalCer). It has previously been shown that CGT-deficient mice do not synthesize GalCer and its sulfated derivative GalCer I(3)-sulfate (galactosylsulfatide, SM4s) but form myelin containing glucosylceramide (GlcCer) and sphingomyelin with 2-hydroxy fatty acids. Because relatively high concentrations of GalCer and SM4s are present also in mammalian kidney, we analyzed the composition of lipids in the kidney of Cgt(-/-) and, as a control, Cgt(+/-) and wild-type mice. The homozygous mutant mice lacked GalCer, galabiaosylceramide (Ga(2)Cer), and SM4s. Yet, they did not show any major morphological or functional defects in the kidney. A slight increase in GlcCer containing 4-hydroxysphinganine was evident among neutral glycolipids. Intriguingly, more polar sulfoglycolipids, that is, lactosylceramide II(3)-sulfate (SM3) and gangliotetraosylceramide II(3),IV(3)-bis-sulfate (SB1a), were expressed at 2 to 3 times the normal levels in Cgt(-/-) mice, indicating upregulation of biosynthesis of SB1a from GlcCer via SM3. Given that SM4s is a major polar glycolipid constituting renal tubular membrane, the increase in SM3 and SB1a in the mice deficient in CGT and thus SM4s appears to be a compensatory process, which could partly restore kidney function in the knockout mice.

Drastically abnormal gluco- and galactosylceramide composition does not affect ganglioside metabolism in the brain of mice deficient in galactosylceramide synthase.

Mice that are genetically deficient in UDP-galactose: ceramide galactosyltransferase are unable to synthesize galactosylceramide. Consequently, sulfatide, which can be synthesized only by sulfation of galactosylceramide, is also totally absent in affected mouse brain. Alpha-hydroxy fatty acid-containing glucosylceramide partially replaces the missing galactosylceramide. A substantial proportion of sphingomyelin, which normally contains only non-hydroxy fatty acids, also contains alpha-hydroxy fatty acids. These findings indicate that alpha-hydroxy fatty acid-containing ceramide normally present only in galactosylceramide and sulfatide is diverted to other compounds because they cannot be synthesized into galactosylceramide due to the lack of the galactosyltransferase. We have examined brain gangliosides in order to determine if alpha-hydroxy fatty acid-containing glucosylceramide present in an abnormally high concentration is also incorporated into gangliosides. The brain ganglioside composition, however, is entirely normal in both the total amount and molecular distribution in these mice. One feasible explanation is that UDP-galactose: glucosylceramide galactosyltransferase does not recognize alpha-hydroxy fatty acid-containing glucosylceramide as acceptor. This analytical finding is consistent with the relative sparing of gray matter in the affected mice and provides an insight into sphingolipid metabolism in the mouse brain.

Polysialylation of brain gangliosides as a possible molecular mechanism for survival of Antarctic ice fish below the freezing point

In order to determine possible adaptation strategies of vertebrates to extreme low-temperature environments, we compared the concentration and composition of gangliosides from the brains of eight species of Antarctic Notothenioid “ice” fishes with those of warm-adapted species and those of fishes from habitats of moderate temperature. The concentration of whole-brain gangliosides in the ice fishes was comparable with that in moderate-temperature species (between 3.36 and 4.31 mg NeuAc/g protein). The composition of brain gangliosides differed, however. In particular, the relative concentrations of polysialogangliosides (= polarity) and alkali-labile gangliosides was higher in all Antarctic species investigated than in warm-adapted fish species. This difference is considered a suitable mechanism for keeping neuronal membranes functional even below the freezing point. This interpretation is supported by additional physicochemical results with artificial monolayer membranes, which give evidence for a high thermosensitivity of ganglioside complexes in connection with calcium.

Brain gangliosides and cold-adaptation in high-antarctic fish

The concentration and composition of gangliosides from the brain of eight species of Antarctic Notothenioid fishes belonging to the class of perciformes and two species of boreal fishes (tropic cichlid fish Oreochromis mossambicus; Codfish Gadus morhua) were investigated. The concentration of whole brain gangliosides in Notothenioid fishes (between 1622 and 2183 μg NeuAc/g dry wt.) was slightly lower than that in the brains of fish species, which live in warm, temperate habitats (2483 μg NeuAc/g dry wt.). The composition of brain gangliosides was completely different from that of warm adapted fish species (e.g. the tropic cichlid fish Oreochromis mossambicus). The relative concentration of polysialogangliosides (GT1b-GH) is strongly increased in all the investigated Antarctic species. They were found to have the most complex and most polar brain ganglioside pattern (high degree of sialylation and alkali-lability) within the teleosts. This may be one of the mechanisms, beside antifreeze proteins, to keep the neuronal membranes functional even below the freezing point.

Influence of ambient temperature on content and composition of brain gangliosides in vertebrates

The concentration and composition of brain gangliosides have been comparatively investigated by means of one- and two-dimensional high performance thin layer chromatography in several European fish species exhibiting a pronounced capacity for thermal adaptation. The values were compared with those of tropical and Antarctic fish as well as with homeo- and heterothermic mammals. The relative proportion of higher sialylated (more polar) ganglioside fractions was elevated in brains of fish (a) whose habitat was naturally cold or (b) that had been experimentally or seasonally adapted to a cold versus a warm habitat. Furthermore, in adaptation to the cold, more alkali-labile brain ganglioside fractions were found with the following sequence: Antarctic fish (53–67%), cold acclimated fish (45–51%), warm acclimated fish (38–44%), tropical species (< 35%), hamsters in torpor (9%), active hamsters (4%) and pig (3%). These results indicate a possible mechanism at a molecular level to keep the neuronal membrane functional, even under changing temperature conditions.

Phylogenetic conservation of ganglioside GD3 expression during early vertebrate ontogeny

Gangliosides were investigated in adult brains and in 5-vesicle stage embryos of representatives belonging to the four vertebrate classes: Chondrichthyes, Amphibia, Aves and Mammalia. Considerable variability in brain ganglioside composition and concentration was observed among the adult vertebrates. The ganglioside patterns of the developmentally matched vertebrate embryos were similar in that each comprised GD3 as the predominant ganglioside. The phylogenetic conservation of abundant GD3 expression during early vertebrate ontogeny is interpreted as biochemical evidence consistent with von Baer's theory of increasing differentiation and suggests that GD3 is of critical importance for normal vertebrate development.

Rat brain gangliosides during hypoxia

The content and composition of rat brain gangliosides in various forms of hypoxia is studied. The content of lipid-bound sialic acid is found to be considerably elevated in the earliest stages of hypoxia combined with hypercapnia and cooling. The content of gangliosides remains elevated over the hypoxia period and returns to control values 48 hours after its discontinuation. Repeated hypoxia results in the same changes in rat brain gangliosides. Hemic sodium nitrite-induced hypoxia also leads to a reliable elevation of these lipids. Hypoxia also causes changes in the content of individual ganglioside fractions: a decreased content of mono- and an increased content of polysialogangliosides.

Ganglioside composition changes in spongiform encephalopathies: Analyses of 263K scrapie-infected hamster brains

Ganglioside composition in brains of terminally ill LVG/LAK golden Syrian hamsters infected with the 263K strain of the scrapie agent was analyzed. Results were compared to those obtained from noninfected animals matched by age, sex, and strain. Gangliosides extracted from scrapie-infected animals showed little change in major components, while an increased number of new alkali-labile species appeared. Additionally, the animal strain employed demonstrated a significant polymorphism in brain ganglioside composition. No significant changes in incubation time, clinical development or pathologic features of scrapie were associated with this polymorphism.

Variability in brain ganglioside content and composition of endothermic mammals, heterothermic hibernators and ectothermic fishes

Content and composition of brain gangliosides were compared among endothermic mammals, heterothermic hibernators and ectothermic fishes from habitats with extreme ambient temperatures (tropic vs. antarctic waters). In general the content of brain gangliosides in fishes is significantly lower and exhibits a greater variability than in mammals. The composition of brain gangliosides was investigated using both one- and two-dimensional High Performance Thin Layer Chromatography (HPTLC). Both techniques showed a remarkable increase in the number of individual ganglioside fractions and an additional increase of higher polar fractions in fishes as compared with mammals. The 2D-HPTLC revealed a significant decrease in the relative proportion of alkali-labile gangliosides in the course of evolution from fish to mammals. Moreover this decrease in alkali-lability is correlated with the state of thermal adaptation (antarctic fishes, 53–66%; tropical cichlid fish, 35%). These results provide additional evidence for the notion that the extremely high polarity of brain gangliosides, especially of cold-blooded vertebrates, reflects a very efficient mechanism on the molecular level to keep the neuronal membrane functional under low temperature conditions.

Age-related alteration of brain gangliosides in senescence-accelerated mouse (SAM)-P/8 9

The senescence-accelerated mouse (SAM)-P/8 was examined with respect to changes in the content and composition of brain gangliosides during aging from juvenile to senescence. The gangliosides were compared with those of control mice, senescence-accelerated resistant mouse(SAM)-R/1. The ganglioside contents in the whole brains of SAM-P/8 and -R/1 were at almost constant level from 0.5 to 6 months, but decreased thereafter until senescence to about 80% of the levels reached at the younger ages. Upon aging, the ganglioside compositions changed with an increase of G GM1, and decreases of G D1a, G T1b in both strains (G T1b G D1a G D1b). A minor component, G M3 was two to four fold higher in the molecular distributions of the whole brain gangliosides of SAM-P/8 than those of -R/1 at any age examined throughout the life span. The regional gangliosides in olfactory bulb, cerebral cortex, hippocampus, hypothalamus, cerebellum, corpora quadrigemina region, brain stem and medulla oblongata were compared between the two strains at the age of three months. The ganglioside contents in the brain stem and medulla oblongata were lower in SAM-P/8 tha -R/1, but there was no significant difference between the two strains in the other regions. As a minor component, G M3 was found to occur in a higher concentration in SAM-P/8 than -R/1 in all brain regions examined, except in the olfactory bulb where G M3 was detected as a major component with no difference in the distribution level between the two strains.

Brain gangliosides and hibernation

1. 1. Region-specific differences in the composition of brain gangliosides were found in homeothermic mammals versus hibernators by means of HP-thinlayer chromatography and a newly developed autoradiographic procedure using 14C N-acetyl-mannosamine as tracer 2. 2. The data support the hypothesis that changes in the composition of Ca 2+-ganglioside-complexes within the neuronal membrane may modulate the synapse, thus guaranteeing the transmission process although temperature had changed.

Seasonal variability of sialo-glycoconjugates in the brain of the Djungarian hamster ( Phodopus sungorus)

1. 1. The influence of season, photoperiod and ambient temperature on the content of proteins, sialo-glycoproteins and gangliosides and on the composition of gangliosides of three different brain regions (cortex, cerebellum and basalbrain) of the Djungarian dwarf hamster ( Phodopus sungorus) had been investigated. Concomittantly changes in body wt and fur colouration were recorded. 2. 2. Dwarf hamsters living under natural photoperiod and ambient temperature conditions (“outside”) showed a distinct annual cycle in body wt (summer: about 45 g; winter: about 25 g) and fur colouration (summer: dark grey; winter: whitish). 3. 3. Among the three brain regions the mean concentration of proteins ranged between 120 and 155 mg protein/g wet wt. The sialo-glycoprotein content varied between 260 and 410 μg NeuAc/g wet wt, and that of gangliosides between 800 and 1650 μg NeuAc/g wet wt. Seasonal fluctuations were not found. 4. 4. The composition of brain gangliosides remained uninfluenced throughout the year in the cerebellum, whereas seasonal variations were observed in cortex and basalbrain. 5. 5. Consequently the concentration ratio of the two major mammalian ganglioside fractions GD1a vs GT1b remained almost stable in cerebellum (0.3). In contrast to this the seasonal values of cortex and basalbrain changed from 0.6 and 0.8 in winter to 0.7 and 1.1 in summer. This indicated a higher polarity of the gangliosides in these brain regions during cold adaptation. 6. 6. The results are discussed with regard to modulatory functions of neuronal gangliosides for the process of synaptic transmission during seasonal adaptation.

Some effects of chronic experimental protein malnutrition and ethanol intake on the ganglioside composition of rat brain during adulthood.

Four groups of rats were used in a nutritionally-controlled study of experimental protein malnutrition and ethanol intake on brain ganglioside composition (GM3, GM2, GM1, GD1a, GD1b, GT1b and GQ1). Male Wistar rats chronically consuming ethanol were fed isoenergetic diets containing 22% (SC-22) or 6% protein (SPD-6), from 60 to 240 days of age. Protein malnutrition initiated at 60 days of age affected monosialoganglioside GM1 composition by 240 days of age. Among the effects of ethanol was an increase of the relative proportion of N-acetylneuraminic acid in the trisialoganglioside GT1b, while there was decrease of the disialoganglioside GD1a. Effects of low protein diet generally did not interfere with ethanol effects except for GD1a and GT1b gangliosides. These changes may be adaptive responses to the fluidizing effect of ethanol on brain membranes.

Changes in developmental profiles of brain gangliosides during ontogeny of a teleost fish (Sarotherodon mossambicus, Cichlidae)

Ontogenetic changes of brain ganglioside concentration and composition have been followed in the teleost fishSarotherodon mossambicus Cichlidae) from the 1st day post hatching to the adult stage, in order to correlate these with findings in higher vertebrates. During the developmental period from hatching to the transition to free swimming, which comes along with maximum rate of synaptogenesis, a sharp rise in the brain ganglioside content occurs, which is mainly due to the trisialoganglioside GT1b. In the following phase of myelination (characterized in birds and mammals by an increase in GM1, GM1′ and GM4) accretion of the novel and so far unidetified fraction (GM2) occurs, which is highly enriched in the brain stem. The results obtained are discussed with respect to gangliosides as useful biochemical markers for brain development and maturation in all vertebrates.

Variability in brain ganglioside composition: A further molecular mechanism beside serum antifreeze-glykoproteins for adaptation to cold in Antarctic and Arctic-boreal fishes

Gangliosides and sialoglycoproteins from brain, liver and muscle have been isolated from 6 Antarctic fish species from the suborder Notothenoids and from 4 Arctic-boreal fish species. In addition freezing and melting points from serum of both groups were examined in order to determine the presence of protein antifreezes. In comparison with eurythermic fishes of temperate climates in both groups the phylogenetical adaptation to cold is correlated with a significantly higher concentration of gangliosides in the brain. The ganglioside concentration of liver in Antarctic fish, but not in Arctic species, is 3 to 5fold higher than in mammals (rat); in muscle the ganglioside content is increased only in red-blooded Antarctic fish as compared with mammals. The concentration of neuronal sialo-glycoproteins generally is lower in Antarctic fish than in other marine teleosts; in muscles the content is 2 to 3fold higher than in mammals. The molecular composition of brain gangliosides is characterized by an extreme high polarity which is due to an equipment with highly sialylated fractions (40 to 50% higher sialylated than tetrasialogangliosides). There are distinct differences between the freezing and melting point of blood serum, especially in the Antarctic species in favour of the existence of protein antifreezes. The results are discussed with regard to the fact that the extremely high polarity of brain gangliosides reflects a very efficient mechanism on molecular level to keep the neuronal membrane functional under low temperature conditions.

Structure related phylogenetic variations in brain gangliosides of vertebrates

1. 1. The concentration and composition of brain gangliosides from five brain structures of vertebrate species belonging to the classes of Chondrichthyes, Osteichthyes, Reptilia, Aves and Mammalia were investigated. 2. 2. The complexity of brain ganglioside composition is strikingly reduced over phyletic lines. 3. 3. In lower vertebrates there is only little variation in the ganglioside pattern between the different brain structures, whereas in higher vertebrates differences distinctly occurred. 4. 4. A similarity over phyletic lines of ganglioside pattern was only noted in phylogenetically old brain structures as for instance in the medulla oblongata and the brain stem.