Plasmalemmal phosphatidylinositol-4,5-bisphosphate level regulates the releasable vesicle pool size in chromaffin cells

被引:167
作者
Milosevic, I
Sorensen, JB [1 ]
Lang, T
Krauss, M
Nagy, G
Haucke, V
Jahn, R
Neher, E
机构
[1] Max Planck Inst Biophys Chem, Dept Membrane Biophys, D-37077 Gottingen, Germany
[2] Max Planck Inst Biophys Chem, Dept Neurobiol, D-37077 Gottingen, Germany
[3] Free Univ Berlin, Inst Chem Biochem, D-14195 Berlin, Germany
关键词
exocytosis; phosphatidylinositol-4,5-bisphospate; chromaffin cell; phosphatidylinositol; 4-phosphate; 5-kinase; plasmalemmal microdomains; LY294002;
D O I
10.1523/JNEUROSCI.3761-04.2005
中图分类号
Q189 [神经科学];
学科分类号
071006 ;
摘要
During exocytosis, certain phospholipids may act as regulators of secretion. Here, we used several independent approaches to perturb the phosphatidylinositol-4,5-bisphosphate[PI(4,5) P-2] level in bovine chromaffin cells to investigatehowchanges of plasmalemmal PI(4,5) P-2 affect secretion. Membrane levels of PI(4,5) P-2 were estimated by analyzing images of lawns of plasma membranes labeled with fluorescent probes specific for PI(4,5) P-2. The specific PI(4,5) P-2 signal was enriched in submicrometer-sized clusters. In parallel patch-clamp experiments on intact cells, we measured the secretion of catecholamines. Overexpression of phosphatidylinositol-4-phosphate-5-kinase Igamma, or infusion of PI(4,5) P-2 through the patch pipette, increased the PI(4,5) P-2 level in the plasma membrane and potentiated secretion. Expression of a membrane-targeted inositol 5-phosphatase domain of synaptojanin 1 eliminated PI(4,5) P-2 from the membrane and abolished secretion. An inhibitor of phosphatidylinositol-3 kinase, 2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one, led to a transient increase in the PI(4,5) P-2 level that was associated with a potentiation of secretion. After prolonged incubation, the level of PI(4,5) P-2 decreased and secretion was inhibited. Kinetic analysis showed that changes in PI(4,5) P-2 levels led to correlated changes in the size of two releasable vesicle pools, whereas their fusion kinetics remained unaffected. We conclude that during both short- and long-term manipulations of PI(4,5) P-2 level secretion scales with plasma membrane PI(4,5) P-2 content and that PI(4,5) P-2 has an early effect on secretion by regulating the number of vesicles ready for release.
引用
收藏
页码:2557 / 2565
页数:9
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