title: p38 MAPK activation elevates serotonin transport activity via a trafficking-independent, protein phosphatase 2A-dependent process.
abstract: Presynaptic, plasma membrane serotonin (5-hydroxytryptamine; 5-HT) transporters (SERTs) clear 5-HT following vesicular release and are regulated through trafficking-dependent pathways. Recently, we provided evidence for a trafficking-independent mode of SERT regulation downstream of adenosine receptor (AR) activation that is sensitive to p38 MAPK inhibitors. Here, we probe this pathway in greater detail, demonstrating elevation of 5-HT transport by multiple p38 MAPK activators (anisomycin, H(2)O(2), and UV radiation), in parallel with p38 MAPK phosphorylation, as well as suppression of anisomycin stimulation by p38 MAPK siRNA treatments. Studies with transporter-transfected Chinese hamster ovary cells reveal that SERT stimulation is shared with the human norepinephrine transporter but not the human dopamine transporter. Saturation kinetic analyses of anisomycin-SERT activity reveal a selective reduction in 5-HT K(m) supported by a commensurate increase in 5-HT potency (K(i)) for displacing surface antagonist binding. Anisomycin treatments that stimulate SERT activity do not elevate surface SERT surface density whereas stimulation is lost with preexposure of cells to the surface-SERT inactivating reagent, 2-(trimethylammonium)ethyl methane thiosulfonate. Guanylyl cyclase (1H-(1,2,4)-oxadiazolo[4,3-a]-quinoxalin-1-one) and protein kinase G inhibitors (H8, DT-2) block AR stimulation of SERT yet fail to antagonize SERT stimulation by anisomycin. We thus place p38 MAPK activation downstream of protein kinase G in a SERT-catalytic regulatory pathway, distinct from events controlling SERT surface density. In contrast, the activity of protein phosphatase 2A inhibitors (fostriecin and calyculin A) to attenuate anisomycin stimulation of 5-HT transport suggests that protein phosphatase 2A is a critical component of the pathway responsible for p38 MAPK up-regulation of SERT catalytic activity.
authors.affiliation: Department of Psychiatry, Vanderbilt University School of Medicine, Nashville, Tennessee 37232-8548, USA.
authors.forename: Chong-Bin
authors.initials: CB
authors.lastname: Zhu
authors.forename: Ana M
authors.initials: AM
authors.lastname: Carneiro
authors.forename: Wolfgang R
authors.initials: WR
authors.lastname: Dostmann
authors.forename: William A
authors.initials: WA
authors.lastname: Hewlett
authors.forename: Randy D
authors.initials: RD
authors.lastname: Blakely
mesh_terms: D000818:Animals; D002462:Cell Membrane / Q000378:metabolism; D006224:Cricetinae; D020536:Enzyme Activators / Q000494:pharmacology; D006801:Humans; D008562:Membrane Glycoproteins / Q000378:metabolism*; D026901:Membrane Transport Proteins / Q000378:metabolism*; D009419:Nerve Tissue Proteins / Q000378:metabolism*; D010749:Phosphoprotein Phosphatases / Q000378:metabolism*; D054648:Protein Phosphatase 2; D012701:Serotonin / Q000378:metabolism*; D050486:Serotonin Plasma Membrane Transport Proteins; D048051:p38 Mitogen-Activated Protein Kinases / Q000187:drug effects / Q000378:metabolism*
chemical_list: D020536:Enzyme Activators; D008562:Membrane Glycoproteins; D026901:Membrane Transport Proteins; D009419:Nerve Tissue Proteins; C493230:SLC6A4 protein, human; D050486:Serotonin Plasma Membrane Transport Proteins; D012701:Serotonin; D048051:p38 Mitogen-Activated Protein Kinases; D010749:Phosphoprotein Phosphatases; D054648:Protein Phosphatase 2