Promotion of Neuritogenesis in Mouse Neuroblastoma Cells by Exogenous Gangliosides. Relationship Between the Effect and the Cell Association of Ganglioside GM1

Abstract: Ganglioside GM 1 promoted neuritogenesis of neuroblastoma cells, neuro‐2a clone, in monolayer culture. GM 1 bound to neuro‐2a cells in three distinct forms, one removable by treatment with serum‐containing solutions, one serum‐resistant and labile to trypsin treatment, and one resistant to serum and trypsin treatments. The proportions among the three forms of cell‐associated GM 1 varied in relation to duration of exposure to ganglioside, ganglioside concentration in the medium, and number of cells in culture. The form removable by serum was predominant at the initial stages of association and at the highest ganglioside concentrations (over 10 −6 M ); the trypsin‐labile and ‐stable forms tended to increase with increasing cell number and decreasing ganglioside concentration. The neuritogenic effect of GM 1 was higher when neuro‐2a cells were incubated for 24 h in the presence of GM 1 and fetal calf serum. Under this condition the percentage of neurite‐bearing cells increased from 11% of control to 62% at the optimal ganglioside concentration of 10 −4 M . The effect was still present, although to a lower extent (from 11% to 28% of neurite‐bearing cells), when cells were first exposed for only 2 h to GM 1 , then washed and incubated for 24 h in the presence of fetal calf serum. The trypsin‐labile and ‐stable forms of cell‐associated GM 1 had a fundamental role in the effect, whereas the form removable by serum was not involved. The preparation of GM 1 used was extremely pure (99%) and, in particular, had a peptide contamination, if any, <1:20,000–1:50,000. Therefore the neuritogenic effect can be attributed to ganglioside itself. The results obtained suggest that under the experimental conditions used the stimulation of neuro‐2a cell differentiation by GM 1 is related to changes of the plasma membrane properties following association of exogenous GM 1 molecules. This would facilitate the spontaneous process of differentiation, or enhance cell responsiveness to differentiating factors present in the serum.