Gangliosides in the human brain development and aging.

In this study, brain gangliosides in prenatal and postnatal human life were analyzed. Immunohistochemically, the presence of "c"-pathway of gangliosides (GQ1c) in embryonic brain was only recorded at 5 weeks of gestation. Biochemical results indicated a twofold increase in human cortex ganglioside concentration between 16 and 22 weeks of gestation. The increasing ganglioside concentration was based on an increasing GD1a ganglioside fraction in all regions analyzed except cerebellar cortex, which was characterized by increasing GT1b. In this developmental period, GD3 was found to be localized in the ventricular zone of the cortical wall. After birth, GD1b ganglioside in neuropil of granular cell layer corresponding to growing mossy fibers was expressed in cerebellar cortex. Between birth and 20/30 years of age, a cerebral neocortical difference of ganglioside composition was observed, characterized by lowest GD1a in visual cortex. Analyzing the composition of gangliosides in cortical regions during aging, they were observed to follow region-specific alterations. In frontal cortex, there was a greater decrease in GD1a and GM1 than in GT1b and GD1b, but in occipital (visual) cortex there was no change in individual gangliosides. In hippocampus, GD1a moderately decreased, whereas other fractions were stable. In cerebellar cortex, GD1b and GT1b fractions decreased with aging.     

Human brain gangliosides in development, aging and disease.

In this study, brain gangliosides in prenatal and postnatal human life and Alzheimer's disease were analyzed. Immunohistochemically, the presence of the "c"-series of gangliosides (GQ1c) was only registered in the embryonic brain at 5 weeks of gestation. Biochemical results indicated a two-fold increase in ganglioside concentration in the human cortex between 16 and 22 weeks of gestation. The increasing ganglioside concentration was based on an increasing GD1a ganglioside fraction in all regions analyzed except in the cerebellar cortex, which was characterized by increasing GT1b. During prenatal human development, regional differences in ganglioside composition could only be detected between the cerebrum ("a"-pathway) and the cerebellum ("b"-pathway). Between birth and 20-30 years of age, a cerebral neocortical difference of ganglioside composition occurred, characterized by the lowest GD1a in visual cortex. Analyzing the composition of gangliosides in cortical regions during aging, they were observed to follow region-specific alterations. In the frontal cortex, there was a greater decrease in GD1a and GM1 than in GT1b and GD1b, but in the occipital (visual) cortex there was no change in individual gangliosides. In hippocampus, GD1a moderately decreased, whereas other fractions were stable. In the cerebellar cortex, GD1b and GT1b fractions decreased with aging. In Alzheimer's disease, we found all ganglio-series gangliosides (GM1, GD1a, GD1b, GT1b) to be decreased in regions (temporal and frontal cortex and nucleus basalis of Meynert) involved in pathogenesis of disease. In addition, in Alzheimer's disease we found simple gangliosides (GN2, GM3) to be elevated in the frontal and parietal cortex, which might correlate accelerated lysosomal degradation of gangliosides and/or astrogliosis occurring during neuronal death.     

High-performance thin-layer chromatography and densitometric determination of brain ganglioside compositions of several species.

Improved resolution of complex brain ganglioside mixtures was achieved by high-performance thin-layer chromatography. The percentage distribution of individual gangliosides was then determined by direct densitometric seanning, employing a transmittance mode, of the resorcinol-positive spots on the plate. As little as 90 pmol (29 ng) of lipid-bound sialic acid could be detected with a good signal-to-noise ratio. A linear detector response was observed up to 3.0 μg of lipid-bound sialic acid. The brain white matter ganglioside patterns of eight animal species, including human, chimpanzee, monkey, chicken, bovine, sheep, and pig, were examined in detail. In addition, human brain gray matter, rat cerebral, rat brain gray matter, and rat cerebellar ganglioside patterns were also studied. Ganglioside G_M4 (G₇) was found to be one of the major components in primate and chicken brain white matter, but it represented only a minor ganglioside in other species. Other major gangliosides in all brain samples studied were G_M1, G_D1a, G_D1b, and G_T1b. G_M1 was more abundant in white matter than in gray matter. G_T1a, a recently discovered ganglioside species, was found in all species examined, but was most abundant in the rat cerebellum. The latter source also contained high proportions of G_T1b and G_Q1b.     

Gangliosides,sialoglycoproteins and acetylcholinesterase of the developing mouse brain

Summary The developmental accretion of up to nine individual gangliosides in foetal brains, peri- and postnatal cortices, postnatal cerebelli and olfactory lobes and in the liver and the spleen were investigated in mice and compared with that of glycoprotein-bound sialic acid and the activity of the acetylcholinesterase.In foetal brain and in postnatal liver and spleen more sialic acid was found bound to glycoproteins than to gangliosides. In postnatal brain structures, however, ganglioside-NeuAc predominated and increased between the 7th and 21st d about 2-fold in the olfactory lobes and cerebellum and more than 3-fold in the cortex.During foetal development the relative quantities (mol %) as well as the absolute concentrations (compared with the fresh weight) of GM1, GM2 and GM3 in the brain decreased, whereas those of GD1a, GD1b and GQ increased.This pattern change continued perinatally in the cortex up to the end of the first week. Thereafter the pattern changed little, but the concentration of all gangliosides present increased much more rapidly, especially between the 10th and 13th d.The postnatal cerebellum and olfactory lobes contained higher concentrations of GM1 and GM3 than the cortex, both gangliosides decreasing in favour of their di-, tri- and tetrasialo-homologues during the third postnatal week.In all brains structures the accretion of GD1a and GT1 was proportional to the increase in the activity of the acetylcholinesterase.Unlike the brain structures, the ganglioside pattern in the liver and spleen, characterised by a predominance of monosialogangliosides and of GD3, did not change noticeably during the first three weeks after birth.The coincidence of the changes in ganglioside accretion observed in the different brain structures with successive periods of morphological differentiation further support the suggestion that gangliosides may play an important role in control of the growth and differentiation of developing nerve cells.Abbreviations GM3 II³NeuAc-GgOse₂Cer - GM2 II³NeuAc-GgOse₃Cer - GM1 II³NeuAcGgOse₄Cer - GD1a IV³NeuAc-, II³ NeuAc-GgOse₄Cer - GD3 II³ NeuAc₂-GgOse₂Cer - GD2 II³ NeuAc₂-GgOse₃ Cer - GD1b II³ NeuAc₂-GgOse₄ Cer - GT1 IV³ NeuAc-, II³ NeuAc₂-GgOse₄ Cer - GQ IV³ NeuAc-, II³ NeuAc₃-GgOse₄ Cer - NeuAc N-acetylneuraminic acid (sialic acid) - AChE Acetylcholinesterase     

Glycosphingolipid Glycosyltransferases in Human Fetal Brain

The developmental pattern of gangliosides in human fetal brain should reflect the activities of the respective glycosyltransferases. LA2-synthase activity, along with that of GM3-, GD3-, GM2-, and GM1-synthases, was determined in human fetal brain at 10-22 weeks of gestation. LA2-synthase is the pivotal enzyme in lacto series ganglioside formation. LA2-synthase activity decreased during the study period, mirroring a similar temporal decline in levels of the lacto series gangliosides, particularly 3'-isoLM1. The developmental profiles of the ganglio series glycosyltransferase activities demonstrate distinct changes that correspond to the ganglioside pattern between fetal weeks 10 and 22. In particular, the marked increase in GM2-synthase activity at 20 and 22 weeks of gestation and the decline in GD3-synthase activity after 15 weeks could explain the prominent expression of the a series gangliosides in this period of rapid neuronal outgrowth. However, a similar decline (two- to 2.5-fold) in GM3-synthase activity suggests a more likely conclusion, namely, that the two sialyltransferase activities are derived mainly from astroglial cells, which show a marked proliferation during the 10-15th fetal weeks. The data do not negate the hypothesis that GM3- and GD3-synthase are the critical enzymes in the regulation of ganglioside biosynthesis but do indicate a need to reevaluate the significance of GM2-synthase in expression of the a series gangliosides.     

THE EFFECT OF PHYSOSTIGMINE AND NEOSTIGMINE ON THE CONCENTRATION OF GLYCOGEN IN VARIOUS BRAIN STRUCTURES OF THE RAT

Abstract—

• 1 Hypothalamus, mesencephalon, cerebral cortex, cerebellar cortex and medulla oblongata of the rat brain contain varying amounts of glycogen. The highest concentration was found in the medulla, and the lowest in the hypothalamus.
• 2 Low doses of physostigmine produced a significant decrease in the concentration of glycogen in mesencephalon, cerebral cortex, cerebellar cortex and medulla. Higher doses of physostigmine were necessary to produce glycogenolysis in the hypothalamus. In the first four structures glycogen stores were almost equally sensitive to the action of physostigmine. Neostigmine did not affect brain glycogen. The glycogenolytic effect of physostigmine was dose-dependent.
• 3 Both atropine and propranolol were found to block the glycogenolytic effect of physostigmine in brain.
• 4 It is concluded that probably both cholinergic and adrenergic processes participate in the glycogenolytic effect of physostigmine. It is suggested that physostigmine initiates the cholinergic processes which then trigger adrenergic processes.

     

Human brain gangliosides: developmental changes from early fetal stage to advanced age

The developmental profiles of the four major brain gangliosides, GM1, GD1a, GD1b, and GT1b, were examined in human frontal lobe covering the period from 10 fetal weeks to 80 years of age. The ganglioside concentration increased approx. 3-fold from the 10th gestational week to the age of about 5 years. Gangliosides GM1 and GD1a increased 12-15-fold during the same period. The most rapid increase of GM1 and GD1a occurred around term, during the period for dendrite arborization, outgrowth of axons and synaptogenesis. GT1b showed a quite different developmental curve. It was the major ganglioside during the 3rd to 5th gestational month, whereafter its concentration dropped rapidly to term, from which time the concentration then increased up to 50 years of age. Similar curves were found for the other gangliosides of the b-series, GD3, GD2, GD1b and GQ1b. Ganglioside 3'-isoLM1 was a characteristic early fetal ganglioside which dropped rapidly to the 5th gestational month, reached a small peak around term and then disappeared during adulthood. The concentration of gangliosides of the neolacto series was larger than that of the lacto series during the whole developmental period. In the beginning of the second trimester, 3'-LM1 constituted 2% and LD1 10% of total ganglioside sialic acid. The new findings demonstrate more dynamic changes of the ganglioside patterns during development than noted in previous studies.     

EFFECT OF DEVELOPMENT ON THE GANGLIOSIDES OF HUMAN BRAIN

Abstract— The ganglioside content and composition of brains from twenty-five human fetuses, three new-born babies and ten children, were studied. The ages ranged from 13 weeks gestation to 26 months postpartum. Each brain was divided into forebrain. cerebellum and brain stem. The concentration of total gangliosides rose to a plateau at different stages of development in the different parts, whereas the total amount reached a constant value at 9 months of age in each part. The developmental profile of individual gangliosides differed in the different parts of the brain. Thus, in the forebrain G_D1a. and in the cerebellum G_D1a rose to become the major gangliosides. The brain stem showed little change in its ganglioside pattern during the developmental period studied. The possible significance of these charges in the gangliosides during development is discussed.     

Developmental profiles of gangliosides in trisomy 19 mice

The ganglioside composition of the cerebrum, cerebellum, brainstem, liver, heart, and spleen was analyzed quantitatively in trisomy 19 (Ts19) mice aged 4 to 12 days postpartum. The developmental profiles of cerebral gangliosides were similar in Ts19 mice and control littermates: Total ganglioside-sialic acid as well as the proportions of the individual gangliosides GD1a and GM1 increased with age, while the percentages of GQ1b and GT1b decreased during development. Both the accretion of the total ganglioside content and the development of the individual ganglioside fractions were delayed by 2-3 days in the Ts19 telencephalon. Likewise, the shift from the b- to the a-pathway of ganglioside synthesis was retarded. Ganglioside development was equally delayed in the cerebellum and the brainstem of Ts19 mice. Since in Ts19 mice, morphogenesis of several brain regions is similarly delayed by 2 days, these results confirm the usefulness of gangliosides as biochemical markers for brain maturation. In contrast to brain gangliosides, the ganglioside composition of the Ts19 livers was clearly distinguished from that of control livers. Total ganglioside-bound sialic acid was increased by 35-50% in Ts19 livers. This elevation in ganglioside content not explicable by a simple delay in development was mainly due to an increase in GD3 and fraction 2, which is likely to contain GD1a and GD1b. In contrast, GM2 which increased considerably with age in control mice persisted on a low level in Ts19 livers. Comparable alterations of the ganglioside pattern were neither observed in the spleen nor in the heart of Ts19 mice. The data presented give additional evidence that ganglioside synthesis in the liver is under a different regulation mechanism than that in the brain, heart, and spleen.     

Accumulation of Glucosylceramide and Glucosylsphingosine (Psychosine) in Cerebrum and Cerebellum in Infantile and Juvenile Gaucher Disease

Abstract: Three major clinical variants of Gaucher disease have been defined: Type I, chronic nonneuronopathic; Type II, acute neuronopathic; and Type III, subacute neuronopathic. In a search for the underlying molecular basis of the neurological manifestations, the concentration and composition of cholesterol, phospholipids, neutral glycosphingolipids, and gangliosides were examined in cerebral and cerebellar cortices of five cases of Type II, eight cases of Type III, and one case of presumed Type I/III. In Type II the concentration of glucosylceramide was 140-530 μmol/kg in cerebral cortex and 51-450 μmol/kg in cerebellar cortex, the highest values found in the most fulminant cases. These concentrations were 20-80 times greater than normal in cerebral cortex and 5-40 times normal in cerebellar cortex. In Type III the concentration of glucosylceramide was 37-65 and 59-1750 μmol/kg in cerebral and cerebellar cortex, respectively. The highest concentrations were found in the cerebellum of patients who had survived splenectomy for several years. The ceramide composition of the accumulated glucosylceramide suggested that brain gangliosides were the major precursors of the glucosylceramide in brains of Type II but in cerebellar cortex in Type III was partly of extracerebral origin. The levels of lactosylceramide and oligohexaosylceramides were slightly raised in all brain specimens from the Gaucher cases. The ganglioside concentration was normal, whereas there was a certain increase in the proportion of GM2 and GM3 gangliosides. The brain glycosphingolipid changes in the Type I/III case were similar but slightly less than those in Type III cases of corresponding age. Glucosylsphingosine (psychosine), never detected in normal human brain, was demonstrated in brains from all the Gaucher cases. The psychosine concentration was highest in Type II cases, 3.8-8.8 and 3.9-12.3 μmol/kg in cerebral and cerebellar cortex, respectively, with the highest values found in the most fulminant cases. In Type III the psychosine concentration varied more widely, 0.8-4.6 and 1.4-6.3 μmol/kg in cerebral and cerebellar cortex, respectively. The lowest value, 0.7 μmol/kg, was found in the Type I/III case. Our method detected psychosine down to 0.01 μmol/kg, which means that the concentration of psychosine was increased at least 100- to 1000-fold in Gaucher grey matter. We suggest that the accumulation of the cell-toxic substance psychosine is the basis for the extensive neuronal cell loss in Gaucher disease, which is most striking in Type II disease.     

Characterization of nuclear gangliosides in rat brain: concentration, composition, and developmental changes

Nuclear gangliosides were characterized using two distinct fractions of large (N1) and small (N2) nuclear populations from rat brain. The ganglioside concentration of N1 nuclei from adult rat brain was 0.92 microg sialic acid/mg protein, which was about 3.8 times higher than that of N2 nuclei. N1 and N2 nuclear gangliosides showed similar compositional profiles; they contained major gangliosides of GM1, GD1a, GD1b, and GT1b, with GM3 in lesser amounts. c-Series gangliosides such as GT3, GQ1c, and GP1c were also detected in both nuclear preparations. Nuclear localization of gangliosides was confirmed by immunofluorescence with anti-GM1 antibody, cholera toxin B subunit, and c-series ganglioside-specific monoclonal antibody A2B5. Developmental changes of nuclear gangliosides were examined using rats of different ages ranging from embryonic day 14 (E14) to postnatal 7 weeks. The concentration of N1 nuclear gangliosides changed only slightly during development and did not correlate with that of whole-brain gangliosides. The developmental pattern of ganglioside composition of N1 nuclei was also distinguished from that of microsomal membranes; the ganglioside changes in N1 nuclei included reduced expression of di- and polysialogangliosides at E16 and higher proportions of GM3 at early and late stages of the period. These findings suggest that gangliosides in nuclear membranes are developmentally regulated in a distinct manner in brain cells.     

THE DISTRIBUTION OF LIPIDS IN THE HUMAN NERVOUS SYSTEM-V. GANGLIOSIDES AND ALLIED NEUTRAL GLYCOLIPIDS OF INFANT BRAIN

—Gangliosides and allied neutral glycosylceramides were isolated from human infant (2-24 months of age) cerebral cortex and white matter. The individual glycolipids were separated quantitatively by a combination of column and thin-layer chromatographic methods on silica gel, DEAE-cellulose and Sephadex G-25. In cerebral cortex G_D1a and G_M1 were the major fractions and constituted more than 70 per cent of the total gangliosides. The concentrations of neutral glycolipids, except for galactosylceramides, were very low: lactosylceramide and glucosylceramide comprised 30 and 5 nmol/g wet weight, respectively. In white matter their concentrations were 10 times higher. The ganglioside concentration was only 50 per cent of that in cerebral cortex: the difference was accounted for mainly by the much lower content of the major di- and trisialogangliosides. Stearic acid was the predominant fatty acid of all brain gangliosides. G_M3, and G_D3 had a considerable content of the very long-chain fatty acids, C₂₂-C₂₄, particularly in the white matter. Glucosylceramide and lactosylceramide had almost identical fatty acid patterns between each other in cerebral cortex and white matter. In the cerebral cortex stearic acid and in the white matter the very long-chain acids predominated. d20:1 Sphingosine comprised more than 20 per cent of total sphingosine in all the gangliosides of the G_l- and G₂-series. G_M3, and G_D3 like lactosylceramide contained significantly less of d20:1 sphingosine. The findings suggest the existence of separate compartments for the biosynthesis of the gangliosides. Glucosylceramides and lactosylceramides of white matter have the same ceramide composition as the galactosylceramides with normal fatty acids and are thus unlikely to be intermediates in the metabolism of the major brain gangliosides which have a completely different fatty acid composition.     

THE ACTIVITY OF GANGLIOSIDE SIALIDASE IN THE DEVELOPING HUMAN BRAIN

—The development of ganglioside sialidase and of four different p-nitrophenyl glycosidases in human brain was investigated including both prenatal and postnatal periods. The developmental curve for sialidase was different from that of the other glycosidases. While p-nitrophenyl glycosidases showed high activities at an early stage of development (21 foetal weeks), and remained at about the same level, ganglioside sialidase was not detected before the foetal age of 15-20 weeks. The sialidase activity at term reached about half the adult level and there was a possible decrease in activity during the first year. The sialidase activity then increased, approaching the adult level at about the age of 5 years. The development of the ganglioside sialidase activity in human brain can be related to changes in the concentration of individual brain ganglioside components. The late maturation of the sialidase system in relation to other glycohydrolases is also discussed as a possible protective mechanism for sialic acid-containing compounds during the early period of development.     

Membrane-bound neuraminidase in the developing mouse brain

The specific activity of membrane-bound neuraminidase towards exogenously added gangliosides in the developing mouse brain was investigated. While whole brain was examined in fetuses, studies were carried out comparatively in cortex and cerebellum of postnatal stages. Considerable differences were found among brain areas. There was a rapid increase in forebrain neuraminidase activity before birth. This high level of activity was maintained throughout the first postnatal week, followed by a slow leveling-off to adult values on the 30th day. In the cerebellum a rapid increase in neuraminidase activity occurred between the 10th and 16th postnatal day. After having reached the maximum, enzyme activity declined rapidly, with adult values being reached on the 21st day. Neuraminidase activity in the adult cerebellum exceeds that in the cortex by 65.9 %. The results are discussed in reference to developmental changes in ganglioside metabolism and the possible involvement of neuraminidase in its regulation is pointed out.     

Changes in rabbit cerebrum and cerebellum gangliosides during postnatal life. A study especially referring to alkali labile gangliosides

The present study deals with the developmental behaviour of cytosolic and membrane-bound gangliosides, especially the alkali labile species, in rabbit cerebrum and cerebellum from birth to 6 months of life. At all ages the amount of cytosolic gangliosides was less than 0.5% of total gangliosides. The proportion of alkali labile gangliosides on the total membrane-bound ganglioside content and the profile of the individual alkali labile gangliosides during postnatal life were different in cerebellum and cerebrum. In cerebellum the proportion of alkali labile gangliosides progressively increased from 4.6% at birth to 16.7% at 6 months; in cerebrum the proportion decreased from 8.9% at birth to a minimum of 5.5% at 10 days, then slowly increased up to 8.1% at 6 months.All the major alkali labile gangliosides in cerebellum, especially O-Ac-GT1b and O-Ac-GQ1b, tended to accumulate during postnatal life. In cerebrum only two alkali labile gangliosides, namely O-Ac-GT1b and a species not yet identified, increased starting from the 10th day of life, while the other ones progressively diminished after birth.     

Developmentally regulated O-acetylated sialoglycans in the central nervous system revealed by a new monoclonal antibody 493D4 recognizing a wide range of O-acetylated glycoconjugates

We have previously detected an alkali-labile and developmentally regulated antigen in rat embryonic cerebral cortex, which may be 9-O-acetylsialylated GT3 ganglioside (Hirabayashi Y, Hirota M, Suzuki Y, Matsumoto M, Obata K, Ando S (1989) Neurosci Lett 106:193-98). In this study we established a mouse monoclonal antibody, 493D4, that recognizes 9-O-acetyl GT3 ganglioside, but not non-O-acetyl gangliosides. This antibody also reacted with 9-O-acetyl GD3 to a much lesser extent. By using this antibody, we found that O-acetyl GT3 as well as O-acetyl GD3 were expressed strongly in fetal murine cerebral cortex and decreased to an undetectable level after birth. With the assistance of TLC-immunostaining using 493D4 together with Q-Sepharose column chromatography, O-acetyl gangliosides of bovine brain were purified and the structural analysis showed the presence of O-acetyl GD3, O-acetyl LD1, O-acetyl GD2 and O-acetyl GD1b in the adult brain as extremely minor components. Interestingly, the antibody 493D4 could detect O-acetyl sialoglycoproteins in rat brain tissues. One of the major immunoreactive proteins was shown to be synaptophysin, an integral membrane protein specifically present in synaptic vesicles. This monoclonal antibody was therefore useful for sensitive detection of both O-acetylated gangliosides and glycoproteins with O-acetylated sialic acids. This revised version was published online in November 2006 with corrections to the Cover Date.     

Differential distribution of major gangliosides in rat central nervous system detected by specific monoclonal antibodies

We investigated the localization of major gangliosides in adultrat brain by an immunofluorescence technique with mouse monoclonalantibodies (MAbs). Five MAbs (GMB16, GMR17, GGR12, GMR5 andGMR13) that specifically recognize gangliosides GM1, GD1a, GD1b,GT1b and GQ1b, respectively, were used. We have found that thereis a cell type-specific expression of the ganglioside in therat central nervous system. In cerebellar cortex, GM1 was expressedin myelin and some glial cells. GD1a was detected exclusivelyin the molecular layer. GD1b and GQ1b were present restrictedlyon the granular layer; GD1b was detected on the surface of thegranular cell bodies, whereas GQ1b was present in the cerebellarglomerulus. GT1b was distributed intensely in both the molecularlayer and the granular layer. In cerebral cortex, GM1 was detectedin some glial cells. Dense staining was limited to the whitematter. GD1a was distributed in layers I, II/III and Va, andthe upper part of layer VI, whereas GQ1b was localized in layersIV and Vb, and the lower part of layer VI. GD1b was detectedbeneath layer III. GT1b appeared to be distributed throughoutall layers. In other regions, such as hippocampal formationand spinal cord, the expression of the ganglioside was alsohighly localized to a specific cell type and layer. ganglioside monoclonal antibody rat brain     

Anabolic sialosylation of gangliosides in situ in rat brain cortical slices

Radiolabeling of the sialic acid residues of gangliosides was examined in thin slices of rat brain cerebral cortex incubated under physiologic conditions in the presence of either [14C]N-acetyl-mannosamine (ManNAc) or cytidine 5'-monophosphoryl-[14C]N-acetyl-neuraminic acid (CMP-NeuAc). CMP-NeuAc is the direct donor substrate in the transfer of sialic acid to gangliosides by sialosyl transferases (SATs), including ectosialosyl transferases at the cell surface. ManNAc must be internalized by the neural cells (neuronal or glial) where it serves as an obligate precursor for the biosynthesis of the NeuAc moiety of intracellular CMP-NeuAc, via multiple reactions in the cytosol and nucleus. When exogenous [14C]ManNAc was supplied, there appeared to be a 2-h lag period before label was incorporated measurably into ganglioside sialic acid. That was followed by rapid ganglioside labeling continuing up to 6 h. There was high incorporation into ganglioside GM1. Labeling by ManNAc was inhibited by monensin, a monovalent cationophore that blocks anabolic transport in medial and trans Golgi. Extracellular CMP-NeuAc was not internalized by the cells. CMP-[14C]NeuAc labeling of gangliosides had no lag period, reached a maximum within 2 h, and then began to level. The label distribution among gangliosides was high in GD3, but quite low in GM1. CMP-NeuAc labeling was not inhibited by 10(-7) M monensin. These findings support a model in which ManNAc labels gangliosides by an intracellular route involving monensin-sensitive, Golgi-associated SATs. In this intracellular system, the major labeled products are gangliosides of the gangliotetraosyl series (GM1, GD1a, etc.).(ABSTRACT TRUNCATED AT 250 WORDS)     

Gangliosides associated with microsomal subfractions of brain: Comparison with synaptic plasma membranes

To study ganglioside distribution within subcellular components and test the hypothesis that they are localized at the nerve ending, microsomes and synaptic plasma membranes were isolated from young adult rat brains and compared with respect to ganglioside composition. These were shown to be heterogeneous preparations by fractionation on a discontinuous sucrose gradient into subfractions which had differing ganglioside concentrations. The highest ganglioside concentrations occurred in membranes banding at the 0.8M/1.0M and 1.0M/1.3M interfaces for both microsomes and synaptic plasma membranes. These subfractions had closely similar ganglioside concentrations and pattern distributions. In addition, the kinetics of ganglioside labeling following administration of [³H]-glucosamine were similar for the two preparations. The fact that microsomal subfractions representing heterogeneous mixtures of brain cell membranes showed close similarity to synaptosomal plasma membranes argues against localization of gangliosides at the nerve ending. These results, together with other lines of evidence, support the concept that gangliosides are distributed over large portions of the neuron (and perhaps other brain cells). Data concerning the labeling of gangliosides in different microsomal subfractions indicated a movement of label over time from the more dense to the less dense membranes, as was also noted for the glycoproteins in the same subfractions. Specific radioactivity of the gangliosides increased relative to that of the glycoproteins with time.     

Gangliosides in rat kidney: composition, distribution, and developmental changes

Gangliosides in rat kidney were analyzed for their composition, regional distribution, and developmental changes. Renal tissue from 7-week-old rats showed a GM3-dominant pattern with GD3 and several minor ganglioside components including GM4, GM2, GD1a, and an unknown ganglioside (ganglioside X). The tissue also contained c-series gangliosides that included GT3 as the main component with GT2 in a lesser amount. Ganglioside analysis of cortical and medullary regions of renal tissue suggested the restricted localization of some gangliosides. While GM4 and GD3 were enriched in the cortical region, GM2 was distributed mainly in the medullary area. Renal gangliosides showed unique developmental profiles during a period from Embryonic Day 20 (E20) to 7 weeks postnatal. The content of renal gangliosides increased from E20, reached the highest around Postnatal Day 1, and thereafter, decreased rapidly to the adult level. The ratio of N-glycolylneuraminic acid to total sialic acids in gangliosides tended to change in inverse proportion to the amount of total sialic acids. The composition of major gangliosides in renal tissues shifted from GD3-dominant to GM3-dominant patterns with advancing ages. While GM1 was expressed only at early stages of the development, GM4, GM2, and ganglioside X appeared after Postnatal Day 3. The expression of c-series gangliosides was less affected through the period examined. These results suggest that gangliosides may be implicated with development and function of rat kidney.