Previous studies with electropermeabilized cells have suggested the occurrence of metabolic compartmentation and Ca2+-dependent channeling of intermediates of
phosphatidylcholine (PC) biosynthesis in C6 rat
glioma cells. With a more accessible permeabilization technique, we investigated whether this is a more general phenomenon also occurring in other cell types and whether channeling is involved in
phosphatidylethanolamine (PE) synthesis as well. C6 rat
glioma cells, C3H10T12 fibroblasts and rat hepatocytes were permeabilized with Staphylococcus aureus alpha-toxin, and the incorporation of the radiolabelled precursors
choline,
phosphocholine (
P-choline),
ethanolamine and
phosphoethanolamine (P-EA) into PC and PE were measured both at high and low Ca2+ concentrations. In
glioma cells, permeabilization at high Ca2+ concentration did not affect [14C]
choline or [14C]
P-choline incorporation into PC. However, reduction of free Ca2+ in the medium from 1.8 mM to <1 nM resulted in a dramatic increase in [14C]
P-choline incorporation into permeabilized cells, whereas [14C]
choline incorporation remained unaffected. Also, in fibroblasts, reduction of extracellular Ca2+ increased [14C]
P-choline and [14C]P-EA incorporation into PC and PE respectively. In hepatocytes, a combination of alpha-toxin and low Ca2+ concentration severely impaired [14C]
choline incorporation into PC. Therefore, alpha-toxin-permeabilized hepatocytes are not a good model in which to study channeling of intermediates in PC biosynthesis. In conclusion, our results indicate that channeling is involved in PC synthesis in
glioma cells and fibroblasts. PE synthesis in fibroblasts is also at least partly dependent on channeling.