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      A Role for p38 Mitogen-Activated Protein Kinase in the Regulation of the Serotonin Transporter: Evidence for Distinct Cellular Mechanisms Involved in Transporter Surface Expression

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          Abstract

          The serotonin transporter (SERT) is regulated by various signaling mechanisms that may operate to maintain appropriate levels of synaptic serotonin (5-HT). We demonstrate that one of the mitogen-activated protein kinases (MAPKs), p38 MAPK, regulates SERT. Treatment of rat midbrain synaptosomes with p38 MAPK-specific inhibitors, PD169316 [4-(4-fluorophenyl)-2-(4-nitrophenyl)-5-(4-pyridyl)-1H-imidazole] or SB203580 [4-(4-fluorophenyl)-2-(4-methylsulfinylphenyl)-5-(4-pyridyl)-1H-imidazole], reduced 5-HT uptake. An additive SERT inhibition by PD169316 and β-phorbol 12-myristate 13-acetate (β-PMA) indicated the involvement of a protein kinase C (PKC)-independent MAPK pathway. Kinetic studies indicated a significant decrease in the transport capacity ( V max) after PD169316 treatment of synaptosomes. Biotinylation studies showed reduced SERT proteins in the plasma membrane of synaptosomes after p38 MAPK inhibition and PKC activation. Phosphorylation studies using synaptosomes revealed decreased SERT phosphorylation by PD169316 but increased phosphorylation by β-PMA. d-Amphetamine enhanced SERT basal phosphorylation and PD169316 blocked this effect. SERT interaction with protein phosphatase 2A catalytic subunit and syntaxin 1A decreased after PD169316 or β-PMA treatment of synaptosomes. In synaptosomes, PKC activation but not p38 MAPK inhibition resulted in SERT redistribution from cholesterolrich lipid raft fractions to nonlipid raft fractions. The presence of phospho-p38 MAPK in synaptosomes and human embryonic kidney 293 (HEK-293) cells suggested the presence of constitutively active p38 MAPK in these preparations. Cotransfection of HEK-293 cells with SERT and a constitutively active form of MAP kinase kinase 3b(E) [MKK3b(E)] increased 5-HT transport, and RNA interference targeted to p38 MAPK inhibited 5-HT uptake, confirming the involvement of active p38 MAPK in SERT expression. Although PD169316 inhibited SERT insertion to the plasma membrane, β-PMA increased SERT internalization in HEK-293 cells. Together, these results indicate a distinct role of p38 MAPK in SERT regulation.

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          Author and article information

          Journal
          J Neurosci
          J. Neurosci
          jneuro
          The Journal of Neuroscience
          Society for Neuroscience
          0270-6474
          1529-2401
          5 January 2005
          : 25
          : 1
          : 29-41
          Affiliations
          Departments of [1 ]Physiology and Neuroscience and [2 ]Neurology, Medical University of South Carolina, Charleston, South Carolina 29425
          Article
          PMC6725216 PMC6725216 6725216 00250029
          10.1523/JNEUROSCI.3754-04.2005
          6725216
          15634764
          c82473e1-9cce-475e-bc39-6b1b7b356ada
          Copyright © 2005 Society for Neuroscience 0270-6474/05/2529-13.00/0
          History
          : 12 November 2004
          : 26 May 2004
          : 4 November 2004
          Categories
          Cellular/Molecular
          Custom metadata
          29
          ARTICLE

          lipid rafts,trafficking,uptake,presynaptic,phosphorylation,neurotransmitter

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