Cerebellar ganglioside abnormalities in pcd mutant mice

The distribution of cerebellar gangliosides was studied in Purkinje cell degeneration (pcd/pcd) mutant mice at postnatal days 25, 30, 50, and 150. These mutants lose the majority of Purkinje cells between 18 and 50 days of age. A reactive gliosis accompanies Purkinje cell loss and a partial loss of granule cells occurs in pcd/pcd mice older than p50. Purkinje cell loss is associated with significant reductions in cerebellar weight and ganglioside concentration. This neuronal loss was also developmentally correlated with reductions of gangliosides (GT1a/LD1 and GT1b and with elevations of GD3. These results agree with previous findings in other cerebellar mutants that GT1a/LD1 and GT1b are concentrated in Purkinje cells and that GD3 is enriched in reactive glial cells. A slight, but significant, reduction in GD1a concentration occurred only in older pcd/pcd mice, consistent with previous findings in weaver and staggerer mice that GD1a is enriched in mature granule cells. The findings with pcd/pcd and other neurological mutants indicate that certain gangliosides can serve as cell-surface markers for monitoring changes in cerebellar cytoarchitecture that accompany development or disease.     

Gangliosides of cultured astroglia

Cultured astrocytes prepared from newborn rat brain and 13-day-old chick embryonic brain were analyzed qualitatively and quantitatively for ganglioside content. All preparations contained approximately the same total level: 2.4-3.4 micrograms N-acetylneuraminic acid (NeuAc)/mg protein. In contrast, the value for primary cultures of neurons from chick embryonic brain was 5.9. The non-hexosamine-containing species, GM3 and GD3, comprised 75-85% of the total in astroglial cultures, the remainder consisting mainly of structural types other than the gangliotetraose series; choleragenoid assay revealed the latter to be virtually absent or to comprise at most a few percent. Deficiency of gangliotetraose synthesizing ability was indicated by the very low level of UDP-GalNac:GM3 N-acetylgalactosaminyltransferase detected in the cells. Treatment of cultured astrocytes with astroglial growth factor 2 or dibutyryl cyclic AMP caused little if any change in quantity or pattern of gangliosides. The large majority of cells stained in a manner characteristic of astrocytes: positive for glial fibrillary acidic protein, negative for galactosyl ceramides. Staining with cholera toxin and anti-GM1 antibody was essentially negative, as was that with tetanus toxin, A2B5 monoclonal antibody, and antibody to GD3. All evidence thus points to cultured astrocytes of rat and chick brain containing appreciable gangliosides, most of which are GM3 and GD3 with the majority of the remainder comprising structures other than the gangliotetraose type.     

Developmental changes of ganglioside expressions in postnatal rat cerebellar cortex

We previously described the differential distribution of gangliosides in adult rat brain as detected by specific antibodies. We report here the distribution of gangliosides during the development of postnatal rat cerebellum by an immunofluorescence technique with mouse monoclonal antibodies (mAbs). Eleven mAbs that specifically recognize each ganglioside were used. Our study revealed that the expression of each ganglioside changed dramatically during the development. GD3 and O-Ac-GD3 were expressed intensely in the external granular layer at 1, 5, and 10 days, whereas GD2 was firstly detected in the internal granular layer at 5 days and GD1b was diffusely detected throughout all layers of the cerebellar cortex at early postnatal days. GD2 and GD1b were more intensely expressed in the granular layer at 20, 30, and 80 days, suggesting that premature granule cells express GD3 and its derivative, O-Ac-GD3, whereas mature granule cells express GD2 and GD1b intensely. On the other hand, GM1 was exclusively detected in the external granular layer and the molecular layer at 1 and 5 days. The staining sites spread gradually from these outer layers into the internal granular layer and the white matter after 10 days. The positive cells in the external granular layer and the molecular layer appeared to be Bergmann glial cells and their radially ascending cytoplasmic processes. The intensity of the staining in these specialized astroglial cells decreased gradually during postnatal days. In contrast, the expression of GQ1b was very faint at birth, but gradually increased during the development and was detected intensely in the internal granular layer, particularly in the cerebellar glomeruli in adulthood, suggesting that GQ1b expression may be associated with synapse-related structures. The developmental changes of the expression of other gangliosides were also recognized in the postnatal rat cerebellum. These results suggest that specific gangliosides may play an important role in regulating the early events responsible for the orderly formation of the cerebellar cortex.     

Differentiation of endogenous neural precursors following spinal cord injury in adult rats

BACKGROUND:Studies have shown that cell death can activate proliferation of endogenous neural stem cells and promote newly generated cells to migrate to a lesion site.OBJECTIVE:To observe regeneration and differentiation of neural cells following spinal cord injury in adult rats and to quantitatively analyze the newly differentiated cells.DESIGN,TIME AND SETTING:A cell biology experiment was performed at the Institute of Orthopedics and Medical Experimental Center,Lanzhou University.between August 2005 and October 2007.MATERIALS:Fifty adult,Wistar rats of both sexes;5-bromodeoxyuridine(BrdU,Sigma,USA);antibodies against neuron-specific enolase,glial fibrillary acidic protein,and myelin basic protein(Chemicon,USA).METHODS:Twenty-five rats were assigned to the spinal cord injury group and received a spinal cord contusion injury.Materials were obtained at day 1,3,7,15,and 29 after injury,with 5 rats for each time point.Twenty-five rats were sham-treated by removing the lamina of the vertebral arch without performing a contusion.MAIN OUTCOME MEASURES:The phenotype of BrdU-labeled cells,i.e.,expression and distribution of surface markers for neurons(neuron-specific enolase),astrocytes(glial fibrillary acidic protein),and oligodendrocytes(myelin basic protein),were identified with immunofluorescence double-labeling.Confocal microscopy was used to detect double-labeled cells by immunofluorescence.Quantitative analysis of newly generated cells was performed with stereological counting methods.RESULTS:There was significant cell production and differentiation after adult rat spinal cord injury.The quantity of newly-generated BrdU-labeled cells in the spinal cord lesion was 75-fold greater than in the corresponding area of control animals.Endogenous neural precursor cells differentiated into astrocytes and oligodendrocytes,however spontaneous neuronal difierentiation was not detected.Between 7 and 29 d after spinal cord injury,newly generated cells expressed increasingly more mature oligodendrocyte and astrocyte markers.CONCLUSION:Spinal cord injury is a direct inducer of regeneration and differentiation of neural cells.Endogenous neural precursor cells Can difierentiate into astrocytes and oligodendrocytes following adult rat spinal cord injury.     

GD3 ganglioside is prevalent in fully differentiated neurons from rat retina

Adult mammalian retinas contain unusually high amounts of GD3, a ganglioside of the lactosylceramide series. In this respect, they differ from adult avian retina and other regions of the adult avian and mammalian brain, where GD3 is a minor ganglioside and gangliosides of the gangliotetraosylceramide series (GM1, GD1a, GD1b, GT1b) are the predominant ones. We compare here the ganglioside patterns of rat, human, horse, and guinea pig retinas, which are known to differ in the degree of vascularization and astrocytic cell content. All these retinas showed a prevalence of pathway "b" gangliosides over pathway "a" gangliosides but showed no correlation between GD3 content and the degree of vascularization and astrocytic cell content. Immunostaining of rat retina sections showed the presence of GD3 in the inner and outer plexiform layers and also in the ganglion cell and inner nuclear layers. About 60% of the cells dissociated from rat retina showed immuno-colocalization of GD3 and the neuronal marker class III beta tubulin isotype or cholera toxin binding. All morphologically identifiable glial Muller cells coexpress GD3 and gangliotetraosylgangliosides. GD3 was a minor ganglioside among these axonally transported by ganglion cells in rats and guinea pigs, suggesting that it is either not synthesized by ganglion cells or, if so, it is restricted to the cell soma and/or dendritic tree. Our results demonstrate that, unlike neurons from avian retina and other regions of avian and mammalian brain, neurons from mammalian retina not only contain gangliosides of the gangliotetraosylceramide series but also keep a prevalence of gangliosides of the lactosylceramide series (GD3) when they are fully differentiated.     

Reversibility of ganglioside effects on astrocyte morphology

The B-subunit of cholera toxin (BCT) induces a morphological change in cultured rat cerebral astrocytes from flat (epithelioid) to stellate (process-bearing). This stellation is reversed by the gangliosides GM1 and GD1a at concentrations of 10 microM or higher. Upon changing to a ganglioside-free medium, the flat astrocytes reacquire the stellate morphology within 3 hr, indicating that the antistellation effect of gangliosides is reversible. The possibility that this reversibility was due to a loss of exogenously acquired gangliosides from the cell membrane can be ruled out since pretreatment with GM1, but not GD1a, which does not bind BCT, results in an increased responsiveness to BCT, which was identical whether measured immediately after withdrawal of the ganglioside or 3 hr later. Asialo-GM1, which neither binds BCT nor reverses BCT-induced stellation by itself, prevents the return to stellation after withdrawal of the gangliosides. These data suggest that while gangliosides remain associated with the cell, their effect on astrocytes can change from opposing to permitting the stellate morphology.     

Contributions of immunohistochemistry to the problem of differentiation in medulloblastoma

The problem of differentiation of medulloblastoma is considered. In this regard 43 medulloblastomas, showing cells with glial or neuronal features by routine histologic methods, were studied. The investigation was carried out by means of the immunohistochemical demonstration of the glial fibrillary acidic protein (GFAP) and the neuron-specific enolase (NSE). In most cases, GFAP-positive cells are preexisting astrocytes; in two cases they correspond to the transitional cells of the subependymal layer. NSE was demonstrated in areas filled with cells with neuroblastic features. The relationship between medulloblastoma with neuron-differentiation and cerebellar neuroblastoma is discussed.     

Differentiation of endogenous neural precursors following spinal cord injury in adult rats☆

BACKGROUND： Studies have shown that cell death can activate proliferation of endogenous neural stem cells and promote newly generated cells to migrate to a lesion site.
OBJECTIVE： To observe regeneration and differentiation of neural cells following spinal cord injury in adult rats and to quantitatively analyze the newly differentiated cells.
DESIGN, TIME AND SETTING： A cell biology experiment was performed at the Institute of Orthopedics and Medical Experimental Center, Lanzhou University, between August 2005 and October 2007.
MATERIALS： Fifty adult, Wistar rats of both sexes; 5-bromodeoxyuridine （BrdU, Sigma, USA）; antibodies against neuron-specific enolase, glial fibrillary acidic protein, and myelin basic protein （Chemicon, USA）.
METHODS： Twenty-five rats were assigned to the spinal cord injury group and received a spinal cord contusion injury. Materials were obtained at day 1, 3, 7, 15, and 29 after injury, with 5 rats for each time point. Twenty-five rats were sham-treated by removing the lamina of the vertebral arch without performing a contusion.
MAIN OUTCOME MEASURES： The phenotype of BrdU-labeled cells, i.e., expression and distribution of surface markers for neurons （neuron-specific enolase）, astrocytes （glial fibrillary acidic protein）, and oligodendrocytes （myelin basic protein）, were identified with immunofluorescence double-labeling. Confocal microscopy was used to detect double-labeled cells by immunofluorescence. Quantitative analysis of newly generated cells was performed with stereological counting methods.
RESULTS： There was significant cell production and differentiation after adult rat spinal cord injury. The quantity of newly-generated BrdU-labeled cells in the spinal cord lesion was 75-fold greater than in the corresponding area of control animals. Endogenous neural precursor cells differentiated into astrocytes and oligodendrocytes, however spontaneous neuronal differentiation was not detected. Between 7 and 29 d after spinal cord injury, newl     

Morphological modulation of cultured rat brain astroglial cells: antagonism by ganglioside GM1

Secondary cultures of neonatal rat astroglial cells, maintained in a serum-free, chemically defined medium were treated with several agents thought to activate cyclic AMP-synthesizing systems. Dibutyryl cyclic AMP (dBcAMP), forskolin and cholera toxin promoted, within 2 h, the near-complete conversion of 1-day-old (D1) astroglial cells from a flat, epithelioid morphology to a stellate (star-shaped) morphology. With all 3 agents, cell susceptibility to morphological change declined with culture age, 5-day-old cultures failing to respond altogether. D1 cultures, after 48 h of treatment, had reverted to the flat morphology. Gangliosides reported to stimulate adenylate cyclase were also tested, using purified GM1 X GM1 failed to stimulate the conversion to stellate morphologies. GM1, however, did affect these astroglial cells by causing a block or reversal of their morphological response to dBcAMP, forskolin or cholera toxin. The GM1 response was specific for the intact ganglioside molecule, asialo GM1 and sialic acid having no effect. Gangliosides GD1a, GD1b and GT1b were also active, being effective at ca. 4-fold lower concentrations. The response to GM1 appeared to involve a direct interaction with the astroglial cell, rather than influencing either substratum or medium components.     

富血小板纤维蛋白诱导骨髓间充质干细胞向成骨样细胞和神经细胞分化

将骨髓间充质干细胞置于富血小板纤维蛋白环境下培养14 d,结果发现,富血小板纤维蛋白可以明显促进骨髓间充质干细胞的增殖,Runt相关转录因子2和骨形态发生蛋白2 mRNA表达均呈剂量依赖性升高,神经元特异性烯醇化酶和神经胶质酸性蛋白含量也升高。结果表明,富血小板纤维蛋白呈剂量依赖性促进骨髓间充质干细胞增殖并分化为成骨样细胞及神经细胞。     

海马神经元条件培养液体外诱导大鼠骨髓间质干细胞向神经元样细胞和神经胶质样 细胞的分化：与含b-FGF培养基和无血清培养基的比较*☆

背景：目前关于骨髓间质干细胞能否向神经元方向分化的报道不多,且争论多集中在分化后的神经元是否仅具有神经元形态而不具有神经元功能。 目的：探讨海马神经元条件培养液诱导大鼠骨髓间质干细胞向神经元样细胞和神经胶质样细胞分化的可能性。 方法：将第5代大鼠骨髓间质干细胞分为4组：条件培养基组加入海马神经元和胶质细胞的培养液;b-FGF组加入含b-FGF的DMEM培养基;无血清培养组加入含Neurobasal和B27的无血清培养基;阴性对照组加入含胎牛血清的DMEM。各组诱导12,24 h后,应用免疫细胞化学染色行神经元特异性烯醇化酶、微管相关蛋白2、胶质纤维酸性蛋白的鉴定,Western-blot法检测细胞神经元特异性烯醇化酶、微管相关蛋白2和胶质纤维酸性蛋白的表达。 结果与结论：诱导12,24 h后,条件培养基组、b-FGF组、无血清培养组骨髓间质充干细胞微管相关蛋白2、胶质纤维酸性蛋白、神经元特异性烯醇化酶均呈阳性表达,阴性对照组未见表达。与阴性对照组比较,诱导后24 h,条件培养基组、b-FGF组、无血清培养组微管相关蛋白2表达均明显增强(P < 0.05),且条件培养基组增强幅度显著高于另两组(P < 0.05);条件培养基组、b-FGF组、无血清培养组神经元特异性烯醇化酶及胶质纤维酸性蛋白表达无明显差异。结果证实海马神经元条件培养液可体外诱导大鼠骨髓间质干细胞分化为神经元样细胞和神经胶质样细胞,与含b-FGF的培养基和无血清培养基相比,海马神经元条件培养基诱导的神经元和神经胶质细胞阳性率最高。     

Expression of gangliosides on glial and neuronal cells in normal and pathological adult human brain

Few studies have assessed the glycolipid phenotype of glial cells in the human central nervous system (CNS) in situ. We investigated by immunohistochemistry the expression and cellular distribution of a panel of gangliosides (GM1, GM2, acetyl-GM3, GD1a, GD1b, GD2, GD3, GT1b, GQ1b and the A2B5 antibody) in adult, human normal and pathological brain, namely multiple sclerosis (MS) and other neurological diseases (OND). In normal conditions, we found diffuse expression in the white matter of most gangliosides tested, with the exception of acetyl-GM3, GT1b and GQ1b. By double immunofluorescence with phenotypic markers, GM1 and GD1b were preferentially expressed on GFAP+ astrocytes, GD1a on NG2+ oligodendrocyte precursors, A2B5 immunostained both populations, while GD2 was selectively present on mature oligodendrocytes. In the gray matter, only GM1, GD2 and A2B5 were present on neuronal cells. Interestingly, those gangliosides present on astrocytes in normal conditions were preferentially expressed on NG2+ cells in chronic MS lesions and in OND. Selective expression of GT1b upon astrocytes and NG2+ cells was instead observed in MS lesions, but not in OND. The definition of the glycolipid phenotype of CNS glial cells may be useful to identify distinct biological glial subsets and provide insights on the potential autoantigenic role of gangliosides in CNS autoimmune diseases.     

天然脑活素对骨髓间充质干细胞向神经元定向分化的诱导作用

BACKGROUND： Bone marrow mesenchymal stem cells （MSCs） have been shown to differentiate into neuronal-like cells through the use of several factors, such as 2-mercaptoethanol, dimethyl sulfoxide, or monothioglycero However, these factors are not suitable for human use due to toxicity. Theoretically speaking, traditional Chinese medicine could be used as potential and safe factors. OBJECTIVE： To investigate the effect of natural cerebrolysin on neuronal-like differentiation of MSCs, based on protein and mRNA analyses. DESIGN, TIME AND SETTING： A parallel controlled, in vitro experiment was performed at the Institute of Integrated Chinese and Western Medicine, Shenzhen Hospital, Southern Medical University, between June 2006 and April 2008. MATERIALS： Natural cerebrolysin was provided by Shenzhen Institute of Integrated Chinese and Western Medicine, China. It primarily consisted of Renshen （Radix Ginseng）, Tianma （Rhizoma Gastrodiae）, and Yinxingye （Ginkgo Leaf） at a proportion of 1：2：2. Natural cerebrolysin extract （1：20） was prepared using conventional water extraction methodology. Each gram of extract equaled 20 grams of the crude drug. Twelve adult, male, New Zealand rabbits were included, six of which underwent intragastric administration of natural cerebrolysin extract （0.976 g/kg per day） for 1 month for natural cerebrolysin-containing serum. The remaining six rabbits received intragastric administration of equal volumes of physiological saline for normal blank serum. METHODS： Sprague Dawley male rats, 6-8 weeks old, were used to harvest tibial and femoral bone marrow. Isolation and purification of MSCs were established from the whole bone marrow by removing the non-adherent cells in primary and passage cultures. For cellular identification, MSCs from four to five passages were co-cultured with LG-DMEM media containing 10% natural cerebrolysin. Simultaneously, MSCs cultured in/G-DMEM media containing 10% blank rabbit serum served as the control group. MAIN OUTCOME MEASURES： Morphology of MSCs and neurite outgrowth during differentiation was observed under inverted phase contrast microscope. Neurite-positive cells were classified by neurite length that was longer than 1.5x the cell body diameter. Immunocytochemistry was used to identify purity of MSCs following passage, as well as expression of nidogen, neuron-specific enolase, glial fibrillary acidic protein, and microtubule-associated protein 2 following treatment with natural cerebrolysin, mRNA expression of neuron-specific enolase and glial fibrillary acidic protein was detected using semi-quantitative RT-PCR. RESULTS： After MSCs were treated with natural cerebrolysin for 3-5 hours, the cell bodies were larger, and small neurites - similar to neuronal neurites - were observed. The number of neurite-positive cells significantly increased compared with the control group （P 〈 0.05）. After MSCs were treated with natural cerebrolysin for 12 hours, most expressed nidogen, neuron-specific enolase, and microtubule-associated protein 2 at higher levels than the control group （P 〈 0.01）. No evident expression of glial fibrillary acidic protein was found （P 〉 0.05）. CONCLUSION： Natural cerebrolysin promoted neurite outgrowth and induced neuronal-like differentiation of MSCs.     

血府逐瘀注射液诱导骨髓间充质干细胞分化为神经元样细胞的最佳剂量筛选

背景：目前大量研究证明传统中药可以诱导骨髓间充质干细胞分化为神经元样细胞。 目的：观察中药血府逐瘀注射液对大鼠骨髓间充质干细胞分化为神经元样细胞的影响,并寻找血府逐瘀注射液诱导分化的最佳浓度。 设计,时间及地点：细胞对照观察实验。实验于2009-07/2010-04在 广东医学院附属医院神经病学研究室及中心实验室完成。 材料： 4周龄SPF级SD雄大鼠,体质量约100g;血府逐瘀注射液的主要中药成分为红花、赤芍、川芎、丹参、当归。 方法：SD大鼠麻醉后无菌条件下取出股骨和胫骨,离心后弃上清液,加入含体积分数为15%胎牛血清的L-DMEM培养基重新悬浮细胞并转入培养瓶培养传代,用免疫细胞化学方法检测第5代骨髓间充质干细胞CD44、CD45的表达;取含1.00,3.00,5.00,10.00 g/L 4种剂量血府逐瘀注射液的无血清L-DMEM培养基对体外培养的第5代骨髓间充质干细胞进行诱导。倒置相差显微镜下观察细胞形态变化,免疫细胞化学方法检测已诱导细胞巢蛋白、神经元特异性烯醇化酶和胶质纤维酸性蛋白的表达,比较4种剂量血府逐瘀注射液诱导神经元样细胞抗原表达率。 主要观察指标：骨髓MSCs生长情况和形态观察,骨髓MSCs表面标志物的表达情况及诱导分化后神经细胞特异性标志物的表达。 结果与结论：①原代细胞接种3 d后多数细胞贴壁,传代后细胞贴壁速度和增殖更快,第5代基本纯化为骨髓间充质干细胞,细胞呈放射状或漩涡状排列。②第5代骨髓间充质干细胞(96.18±1.65)% CD44表达阳性,CD45表达阴性。③诱导后细胞出现类似神经元细胞样形态;免疫细胞化学方法检测显示多数细胞巢蛋白、神经元特异性烯醇化酶阳性表达,3.00 g/L浓度组细胞的巢蛋白和神经元特异性烯醇化酶阳性表达率最高。提示血府逐瘀注射液可诱导骨髓间充质干细胞分化为神经元样细胞,3.00 g/L为最适诱导浓度。     

慢病毒转染胶质细胞源性神经营养因子脂肪干细胞脑移植的研究

目的 探讨慢病毒转染胶质细胞源性神经营养因子(GDNF)的脂肪干细胞(ADSCs)脑移植的应用价值.方法 分离、培养SD大鼠ADSCs,用高滴度慢病毒将GDNF转染入细胞,观察细胞形态及用免疫荧光法观察神经标志物的表达.用立体定向仪将转染ADSCs植入大鼠的纹状体,移植1个月和2个月后,用荧光显微镜观察ADSCs脑内存活和分布状况,用Western blot检测GDNF蛋白的表达,并与正常对照组比较.结果 慢病毒转染ADSCs形态类似神经元,胞浆神经元特异性烯醇化酶(NSE)表达阳性.ADSCs移植到大鼠纹状体1个月后,移植部位有大量存活的ADSCs,2个月后,仍有部分存活细胞,并向远处移行,GDNF蛋白水平显著高于正常对照组(均P<0.01).结论 慢病毒转染GDNF的ADSCs可为干细胞移植治疗中枢神经系统疾病提供新的供体.     

Inhibition of neurite outgrowth of neuroblastoma neuro-2a cells by cholera toxin B-subunit and anti-GM1 antibody

The role of cell surface GM1 ganglioside in neurite outgrowth of Neuro-2a neuroblastoma cells was investigated by application of anti-GM1 antibody and the B subunit of cholera toxin (cholera B) to cultured cells stimulated to grow neurites in various ways. When the cells were simultaneously treated with stimulatory agent and cholera B, inhibition, as measured by percent of neurite-bearing cells, was observed with most stimuli: neuraminidase; GD1a ganglioside, retinoic acid, and low serum. However, with dibutyryl cyclic AMP the small reduction observed was not statistically significant. The inhibitory effect of cholera B on neurite outgrowth induced by low serum was dose-dependent, reaching a maximum at 200 ng/mL; 48 h after washout of cholera B the cells were released from inhibition and regrew neurites at nearly the previous rate in the presence of low serum. When the cells were exposed to stimulus for 6 h or more the inhibitory effect of subsequent addition of cholera B was reduced or eliminated; inhibition thus occurs during an early stage of neurite initiation. Anti-GM1 anti-body at dilutions of 1∶100–1∶400 had the same inhibitory effect as cholera B with cells stimulated by GD1a or retinoic acid, whereas anti-GM2 antibody had no effect at 1∶200 or 1∶400; inhibition by the latter antibody at 1∶100 dilution was similar to that attained with control ascites fluid. These results point to a pivotal role for cell surface GM1 in Neuro-2a differentiation induced by many (but not all) neuritogenic agents.     

Cholera Toxin β Subunit Induces the Differentiation of Human Medulloblastoma Cell Line DEV in a Neuronal Pathway

Medulloblastomas are primitive neuroectodermal tumours that are thought to arise from multipotent precursor cells in the cerebellum. Medulloblastoma cells may be undifferentiated or exibit glial, neuronal or ependymal characteristics, suggesting that they may conserve their ability to differentiate in appropriate circumstances. Medulloblastoma cell lines may thus provide models to study the commitment and differentiation of multipotent CNS progenitor cells. A human medulloblastoma cell line, DEV, has previously been shown to differentiate in an astrocytic pathway after infection by the retrovirus HTLV-1. In this study immunofluorescence flow cytometry shows that cholera toxin β subunit (CTβ), which binds to the ganglioside GM1, induces a twofold increase in the number of DEV cells differentiating towards a neuronal pathway, as shown by the increased proportion and labelling intensity of cells stained by an anti-neurofilament antibody. Immunocytochemistry shows that after 3 days in culture with CTβ, DEV cells develop processes which stain positive for neurofilaments and MAP-1. This suggests that CTβ induces DEV cells to express a more neuronal phenotype.