The effects of membrane fatty acid modification of clonal pheochromocytoma cells on depolarization-dependent exocytosis.

We have begun studying the role of membrane lipids in the exocytotic release process using the pheochromocytoma clone, PC12. The phospholipid fatty acid composition of the cells was modified by growth in the presence of specific fatty acids. None of the fatty acid modifications affected K+-stimulated release of [3H] norepinephrine. This observation indicates that the individual steps of the secretion process, including the extent of depolarization produced by K+, the response of the voltage-dependent Ca2+ channels to depolarization, and the subsequent steps in Ca2+-dependent exocytosis were unaffected by the fatty acid changes. In contrast, exocytosis evoked by stimulation of nicotinic cholinergic receptors with carbamylcholine or direct activation of action potential Na+ channels with veratridine was diminished in cells enriched with unsaturated fatty acids. The diminished output of the release systems was observed at all concentrations of carbamylcholine and veratridine tested. Since the events of exocytosis subsequent to Ca2+ influx were unaffected by unsaturated fatty acids, it appears likely that the magnitude of the depolarization produced by carbamylcholine and veratridine was reduced. The loss of carbamylcholine-stimulated release did not correlate with the simple presence of the fatty acids, but paralleled closely the time and concentration-dependent changes in the phospholipid fatty acid composition. However, when oleate and arachidonate were simultaneously added to the culture medium, the inhibitory effects on carbamylcholine-stimulated release were additive, whereas the changes in fatty acid composition were antagonistic. Thus, exposure of PC12 cells to unsaturated fatty acids causes specific, reversible decreases in the activities of at least 2 stimulus/secretion systems. However, the mechanistic explanation for these changes is not readily apparent from a simple analysis of total phospholipid fatty acid composition.