Trafficking-Dependent and -Independent Pathways of Neurotransmitter Transporter Regulation Differentially Involving p38 Mitogen-Activated Protein Kinase Revealed in Studies of Insulin Modulation of Norepinephrine Transport in SK-N-SH Cells
Presynaptic, cocaine- and antidepressant-sensitive norepinephrine (NE) transporters (NETs) dictate levels of extracellular NE after vesicular release. Recent studies suggest that G protein-coupled receptors linked to protein kinase C (PKC) down-regulate cell surface NET protein levels and diminish N...
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Published in | The Journal of pharmacology and experimental therapeutics Vol. 299; no. 2; pp. 666 - 677 |
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Main Authors | , , , |
Format | Journal Article |
Language | English |
Published |
United States
American Society for Pharmacology and Experimental Therapeutics
01.11.2001
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Subjects | |
Online Access | Get full text |
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Summary: | Presynaptic, cocaine- and antidepressant-sensitive norepinephrine (NE) transporters (NETs) dictate levels of extracellular
NE after vesicular release. Recent studies suggest that G protein-coupled receptors linked to protein kinase C (PKC) down-regulate
cell surface NET protein levels and diminish NE uptake capacity. We identified distinct phosphatidylinositol 3-OH kinase (PI3K)-linked
pathways supporting basal and insulin-triggered NE transport in the human noradrenergic neuroblastoma, SK-N-SH. Acute (0â60
min) insulin treatments produced a time- and concentration-dependent stimulation of NE transport, resolved in kinetic studies
as an enhancement of NE transport capacity ( V max ) without an alteration in NE K m . Basal and insulin-modulated NET activities were reduced by the tyrosine kinase inhibitor genistein and the PI3K inhibitors
wortmannin and LY-294002, but not by the PKC inhibitor staurosporine. PI3K activation was found to support phosphorylation
of p38 mitogen-activated protein kinase (p38 MAPK). However, basal and insulin-stimulated NET activities were differentiated
by their reliance on p38 MAPK activation. Thus, the p38 MAPK inhibitor SB203580 and SB202190 abolished insulin activation
of NE transport yet failed to impact basal NET activity. Moreover, p38 MAPK activation and insulin activation of NETs were
found to be sensitive to external Ca 2+ depletion, blockade of voltage-sensitive Ca 2+ channels, and inhibition of protein phosphatase 2A. Effects of tyrosine kinase and PI3K inhibitors on basal NET uptake appear
to arise from a loss of cell surface NET protein, whereas the p38 MAPK-dependent enhancement of NE transport occurs without
a detectable enhancement of surface NET. Our findings establish two distinct pathways for regulation of NE uptake involving
PI3K, one linked to transporter trafficking and a second linked to Ca 2+ -dependent, p38 MAPK phosphorylation that promotes activation of cell surface NETs. |
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Bibliography: | ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 23 |
ISSN: | 0022-3565 1521-0103 |