Expression of Plasma Membrane and Cell Surface Phospholipids and Gangliosides of Chick Embryo Neurons Grown in Primary Cultures: Developmental Studies

Abstract

Plasma membranes isolated from chick embryo neurons after 2 days (division phase) and 5 days (maturation phase) in culture showed no differences in the phospholipid (PL) pattern or in the plasma membrane average fluidity. Significant ganglioside (Ggl) variations were only found in GM1 and GD3 which represent 35% of the total plasma membrane N-acetylneuraminic acid (NeuNac) content (1.8-fold GM1 NeuNac increase, 1.4-fold GD3 NeuNac reduction). Lactoperoxidase-catalyzed radioiodination (¹²⁵I) under saturating conditions indicated that the hydrophobic core of cell surface Ggl was more exposed to the hydrophilic extracellular environment than that of PL. Nevertheless, when interneuronal contacts start to form, the ¹²⁵I-labelling of both PL and Ggl dropped progressively throughout the maturation phase. Further analysis revealed that (1) plasma membrane GD3 was not accessible to the enzymatic labelling, and (2) surface phosphatidylethanolamine (PE) and GT1b became more accessible to the external medium during development, whereas the opposite was found for phosphatidylcholine (PC) and GM1. The overall profile of radioiodinated PL and Ggl was unaltered when poly-L-lysine was substituted for polyethyleneimine as culture substratum, except for an increase in GT1b labelling 24 h after plating. These results suggest the existense of a ''buffering'' mechanism protecting the neuronal plasma membrane during cellular growth from marked changes in fluidity, polar lipid composition and relative localization of polar lipids with regard to the extracellular environment. In addition, our findings suggest that: (1) GD3, which is at maximal levels during the division phase, is not exposed to the extracellular environment; (2) PE, PC, GM1 and GT1b undergo subtle topological reorganization as cellular maturation proceeds, and (3) the culture substratum may modulate the exposure of certain cell surface glycolipid components.

Neuronal cells mature faster on polyethyleneimine coated plates than on polylysine coated plates

“…This was not the case for cells grown on PEI which display a pattern quite similar to the pattern found in 5 day-old cultures. This observation is also in agreement with data we reported previously (Lelong et al, 1991). indeed, in this previous paper, we showed a higher level of radiolabeling of GDlb in plasma membranes from 1 day old neurons for cells grown on PEI compared to their counterparts grown on polylysine.…”

Effect of Colchicine on Ganglioside Composition of Rat Primary Culture Neurons

“…Simi larly, cell density-dependent change in ganglioside com position was not so obvious in 1-day-old culture when dissociated cells were seeded on polylysine-coated sur faces at cell densities increased up to 8-fold [unpubl.]. Recently, Lelong et al [40] demonstrated topological changes o f cell surface gangliosides during division and m aturation phases in cultured chick neurons. Based upon lactoperoxidase-catalyzed radioiodination, GD3 was shown not to be exposed to the extracellular environ ment and other gangliosides were to undergo subtle topo logical reorganization during neuronal development.…”

Lack of Cell Density-Dependent Changes in Gangliosides of Rat Primary Culture Neurons

“…The immunocytochemical approach using antiganglioside antibodies revealed the intracellu lar distribution of most gangliosides 24 h after seeding and the subsequent expression of gangliosides on neu ronal membranes with a lag [11]. Inhibition of neurite outgrowth by colchicine during the first 24 h following plating caused neither the compositional change nor syn thetic inhibition of gangliosides [12], Recently, Lelonget al [30] have also reported that some of gangliosides are not exposed to the extracellular environment during divi sion and maturation phases in cultured chick neurons. In the ontogénie development a major ganglioside of fetal neurons, GD3, is reported to be strongly reactive to anti-GD3 antibody in proliferative neuronal cell precursors and to become unreactive in the subsequent stages [25][26][27].…”