Skip to main content

PIP5K2A-dependent regulation of excitatory amino acid transporter EAAT3

Abstract

Introduction

According to previous observations, the gene encoding the phosphatidylinositol-4-phosphate 5-kinase II alpha (PIP5K2A) is associated with schizophrenia. Specifically, the mutation N251SPIP5K2A has been discovered in schizophrenic patients but not in healthy individuals. A defect of the excitatory amino acid transporter EAAT3 has similarly been implicated in the development of schizophrenia. The present study thus explored whether PIP5K2A is involved in the regulation of EAAT3 activity.

Materials and methods

EAAT3 was expressed in Xenopus oocytes either without or with PIP5K2A, and EAAT3 transporter activity was estimated from the glutamate (2-mM)-induced current (Iglu) in dual electrode voltage clamp experiments. EAAT3 protein abundance in the cell membrane was estimated by Western blotting and confocal microscopy.

Results

In EAAT3-expressing oocytes, Iglu was enhanced by coexpression of wild type PIP5K2A. Coexpression of the schizophrenia-associated mutant N251SPIP5K2A significantly decreased Iglu in oocytes expressing EAAT3 with or without additional expression of wild type PIP5K2A. Thus, N251SPIP5K2A exerts a dominant inhibitory effect.

Discussion

Membrane abundance of EAAT3 was increased by wild type PIP5K2A and decreased by N251SPIP5K2A in both EAAT3-expressing oocytes and human embryonic kidney cells. The present observations disclose a novel mechanism of EAAT3 regulation, which may contribute to the deranged regulation of excitability in schizophrenic patients.

This is a preview of subscription content, access via your institution.

Fig. 1
Fig. 2
Fig. 3
Fig. 4

References

  • Amara SG, Fontana AC (2002) Excitatory amino acid transporters: keeping up with glutamate. Neurochem Int 41:313–318

    PubMed  Article  CAS  Google Scholar 

  • Anderson RA, Boronenkov IV, Doughman SD, Kunz J, Loijens JC (1999) Phosphatidylinositol phosphate kinases, a multifaceted family of signaling enzymes. J Biol Chem 274:9907–9910

    PubMed  Article  CAS  Google Scholar 

  • Bakker SC, Hoogendoorn ML, Hendriks J, Verzijlbergen K, Caron S, Verduijn W, Selten JP, Pearson PL, Kahn RS, Sinke RJ (2007) The PIP5K2A and RGS4 genes are differentially associated with deficit and non-deficit schizophrenia. Genes Brain Behav 6:113–119

    PubMed  Article  CAS  Google Scholar 

  • Boehmer C, Laufer J, Jeyaraj S, Klaus F, Lindner R, Lang F, Palmada M (2008a) Modulation of the voltage-gated potassium channel Kv1.5 by the SGK1 protein kinase involves inhibition of channel ubiquitination. Cell Physiol Biochem 22:591–600

    PubMed  Article  CAS  Google Scholar 

  • Boehmer C, Palmada M, Klaus F, Jeyaraj S, Lindner R, Laufer J, Daniel H, Lang F (2008b) The peptide transporter PEPT2 is targeted by the protein kinase SGK1 and the scaffold protein NHERF2. Cell Physiol Biochem 22:705–714

    PubMed  Article  CAS  Google Scholar 

  • Brown DA, Hughes SA, Marsh SJ, Tinker A (2007) Regulation of M(Kv7.2/7.3) channels in neurons by PIP(2) and products of PIP(2) hydrolysis: significance for receptor-mediated inhibition. J Physiol 582:917–925

    PubMed  Article  CAS  Google Scholar 

  • Chan H, Zwingmann C, Pannunzio M, Butterworth RF (2003) Effects of ammonia on high affinity glutamate uptake and glutamate transporter EAAT3 expression in cultured rat cerebellar granule cells. Neurochem Int 43:137–146

    PubMed  Article  CAS  Google Scholar 

  • Collin M, Backberg M, Ovesjo ML, Fisone G, Edwards RH, Fujiyama F, Meister B (2003) Plasma membrane and vesicular glutamate transporter mRNAs/proteins in hypothalamic neurons that regulate body weight. Eur J NeuroSci 18:1265–1278

    PubMed  Article  Google Scholar 

  • Crino PB, Jin H, Shumate MD, Robinson MB, Coulter DA, Brooks-Kayal AR (2002) Increased expression of the neuronal glutamate transporter (EAAT3/EAAC1) in hippocampal and neocortical epilepsy. Epilepsia 43:211–218

    PubMed  Article  CAS  Google Scholar 

  • Davis KE, Straff DJ, Weinstein EA, Bannerman PG, Correale DM, Rothstein JD, Robinson MB (1998) Multiple signaling pathways regulate cell surface expression and activity of the excitatory amino acid carrier 1 subtype of Glu transporter in C6 glioma. J Neurosci 18:2475–2485

    PubMed  CAS  Google Scholar 

  • Delmas P, Brown DA (2005) Pathways modulating neural KCNQ/M (Kv7) potassium channels. Nat Rev Neurosci 6:850–862

    PubMed  Article  CAS  Google Scholar 

  • Deng X, Shibata H, Takeuchi N, Rachi S, Sakai M, Ninomiya H, Iwata N, Ozaki N, Fukumaki Y (2007) Association study of polymorphisms in the glutamate transporter genes SLC1A1, SLC1A3, and SLC1A6 with schizophrenia. Am J Med Genet B Neuropsychiatr Genet 144:271–278

    Google Scholar 

  • Doughman RL, Firestone AJ, Anderson RA (2003) Phosphatidylinositol phosphate kinases put PI4, 5P(2) in its place. J Membr Biol 194:77–89

    PubMed  Article  CAS  Google Scholar 

  • Dowd LA, Robinson MB (1996) Rapid stimulation of EAAC1-mediated Na+-dependent L-glutamate transport activity in C6 glioma cells by phorbol ester. J Neurochem 67:508–516

    PubMed  CAS  Article  Google Scholar 

  • Dowd LA, Coyle AJ, Rothstein JD, Pritchett DB, Robinson MB (1996) Comparison of Na+-dependent glutamate transport activity in synaptosomes, C6 glioma, and Xenopus oocytes expressing excitatory amino acid carrier 1 (EAAC1). Mol Pharmacol 49:465–473

    PubMed  CAS  Google Scholar 

  • Fedorenko O, Strutz-Seebohm N, Henrion U, Ureche ON, Lang F, Seebohm G, Lang UE (2008) A schizophrenia-linked mutation in PIP5K2A fails to activate neuronal m-channels. Psychopharmacology 199:47–54

    PubMed  Article  CAS  Google Scholar 

  • Furuta A, Martin LJ, Lin CL, Dykes-Hoberg M, Rothstein JD (1997) Cellular and synaptic localization of the neuronal glutamate transporters excitatory amino acid transporter 3 and 4. Neuroscience 81:1031–1042

    PubMed  Article  CAS  Google Scholar 

  • Furuta A, Takashima S, Yokoo H, Rothstein JD, Wada K, Iwaki T (2005) Expression of glutamate transporter subtypes during normal human corticogenesis and type II lissencephaly. Brain Res Dev Brain Res 155:155–164

    PubMed  Article  CAS  Google Scholar 

  • He Z, Li Z, Shi Y, Tang W, Huang K, Ma G, Zhou J, Meng J, Li H, Feng G, He L (2007) The PIP5K2A gene and schizophrenia in the Chinese population—a case-control study. Schizophr Res 94:359–365

    PubMed  Article  Google Scholar 

  • Huang YH, Dykes-Hoberg M, Tanaka K, Rothstein JD, Bergles DE (2004) Climbing fiber activation of EAAT4 transporters and kainate receptors in cerebellar Purkinje cells. J Neurosci 24:103–111

    PubMed  Article  CAS  Google Scholar 

  • Huerta I, McCullumsmith RE, Haroutunian V, Gimenez-Amaya JM, Meador-Woodruff JH (2006) Expression of excitatory amino acid transporter interacting protein transcripts in the thalamus in schizophrenia. Synapse 59:394–402

    PubMed  Article  CAS  Google Scholar 

  • Kim JH, Do SH, Kim YL, Zuo Z (2005) Effects of chronic exposure to ethanol on glutamate transporter EAAT3 expressed in Xenopus oocytes: evidence for protein kinase C involvement. Alcohol Clin Exp Res 29:2046–2052

    PubMed  Article  CAS  Google Scholar 

  • Lang UE, Puls I, Muller DJ, Strutz-Seebohm N, Gallinat J (2007) Molecular mechanisms of schizophrenia. Cell Physiol Biochem 20:687–702

    PubMed  Article  CAS  Google Scholar 

  • Maragakis NJ, Dietrich J, Wong V, Xue H, Mayer-Proschel M, Rao MS, Rothstein JD (2004) Glutamate transporter expression and function in human glial progenitors. Glia 45:133–143

    PubMed  Article  Google Scholar 

  • Mathern GW, Mendoza D, Lozada A, Pretorius JK, Dehnes Y, Danbolt NC, Nelson N, Leite JP, Chimelli L, Born DE, Sakamoto AC, Assirati JA, Fried I, Peacock WJ, Ojemann GA, Adelson PD (1999) Hippocampal GABA and glutamate transporter immunoreactivity in patients with temporal lobe epilepsy. Neurology 52:453–472

    PubMed  CAS  Google Scholar 

  • McCullumsmith RE, Meador-Woodruff JH (2002) Striatal excitatory amino acid transporter transcript expression in schizophrenia, bipolar disorder, and major depressive disorder. Neuropsychopharmacology 26:368–375

    PubMed  Article  CAS  Google Scholar 

  • Miralles VJ, Martinez-Lopez I, Zaragoza R, Borras E, Garcia C, Pallardo FV, Vina JR (2001) Na+ dependent glutamate transporters (EAAT1, EAAT2, and EAAT3) in primary astrocyte cultures: effect of oxidative stress. Brain Res 922:21–29

    PubMed  Article  CAS  Google Scholar 

  • Nieoullon A, Canolle B, Masmejean F, Guillet B, Pisano P, Lortet S (2006) The neuronal excitatory amino acid transporter EAAC1/EAAT3: does it represent a major actor at the brain excitatory synapse? J Neurochem 98:1007–1018

    PubMed  Article  CAS  Google Scholar 

  • Nudmamud-Thanoi S, Piyabhan P, Harte MK, Cahir M, Reynolds GP (2007) Deficits of neuronal glutamatergic markers in the caudate nucleus in schizophrenia. J Neural Transm Suppl 72:281–285

    PubMed  Article  CAS  Google Scholar 

  • O’Kane RL, Martinez-Lopez I, DeJoseph MR, Vina JR, Hawkins RA (1999) Na(+)-dependent glutamate transporters (EAAT1, EAAT2, and EAAT3) of the blood-brain barrier. A mechanism for glutamate removal. J Biol Chem 274:31891–31895

    PubMed  Article  Google Scholar 

  • Proper EA, Hoogland G, Kappen SM, Jansen GH, Rensen MG, Schrama LH, van Veelen CW, van Rijen PC, van Nieuwenhuizen O, Gispen WH, de Graan PN (2002) Distribution of glutamate transporters in the hippocampus of patients with pharmaco-resistant temporal lobe epilepsy. Brain 125:32–43

    PubMed  Article  CAS  Google Scholar 

  • Qian Y, Galli A, Ramamoorthy S, Risso S, DeFelice LJ, Blakely RD (1997) Protein kinase C activation regulates human serotonin transporters in HEK-293 cells via altered cell surface expression. J Neurosci 17:45–57

    PubMed  CAS  Google Scholar 

  • Rakhade SN, Loeb JA (2008) Focal reduction of neuronal glutamate transporters in human neocortical epilepsy. Epilepsia 49:226–236

    PubMed  Article  CAS  Google Scholar 

  • Rameh LE, Tolias KF, Duckworth BC, Cantley LC (1997) A new pathway for synthesis of phosphatidylinositol-4,5-bisphosphate. Nature 390:192–196

    PubMed  Article  CAS  Google Scholar 

  • Schmitt A, Zink M, Petroianu G, May B, Braus DF, Henn FA (2003) Decreased gene expression of glial and neuronal glutamate transporters after chronic antipsychotic treatment in rat brain. Neurosci Lett 347:81–84

    PubMed  Article  CAS  Google Scholar 

  • Schniepp R, Kohler K, Ladewig T, Guenther E, Henke G, Palmada M, Boehmer C, Rothstein JD, Broer S, Lang F (2004) Retinal colocalization and in vitro interaction of the glutamate transporter EAAT3 and the serum- and glucocorticoid-inducible kinase SGK1 [correction]. Invest Ophthalmol Vis Sci 45:1442–1449

    PubMed  Article  Google Scholar 

  • Schwab SG, Knapp M, Sklar P, Eckstein GN, Sewekow C, Borrmann-Hassenbach M, Albus M, Becker T, Hallmayer JF, Lerer B, Maier W, Wildenauer DB (2006) Evidence for association of DNA sequence variants in the phosphatidylinositol-4-phosphate 5-kinase IIalpha gene (PIP5K2A) with schizophrenia. Mol Psychiatry 11:837–846

    PubMed  Article  CAS  Google Scholar 

  • Shashidharan P, Huntley GW, Murray JM, Buku A, Moran T, Walsh MJ, Morrison JH, Plaitakis A (1997) Immunohistochemical localization of the neuron-specific glutamate transporter EAAC1 (EAAT3) in rat brain and spinal cord revealed by a novel monoclonal antibody. Brain Res 773:139–148

    PubMed  Article  CAS  Google Scholar 

  • Shyng SL, Barbieri A, Gumusboga A, Cukras C, Pike L, Davis JN, Stahl PD, Nichols CG (2000) Modulation of nucleotide sensitivity of ATP-sensitive potassium channels by phosphatidylinositol-4-phosphate 5-kinase. Proc Natl Acad Sci U S A 97:937–941

    PubMed  Article  CAS  Google Scholar 

  • Simantov R, Crispino M, Hoe W, Broutman G, Tocco G, Rothstein JD, Baudry M (1999) Changes in expression of neuronal and glial glutamate transporters in rat hippocampus following kainate-induced seizure activity. Brain Res Mol Brain Res 65:112–123

    PubMed  Article  CAS  Google Scholar 

  • Smith RE, Haroutunian V, Davis KL, Meador-Woodruff JH (2001) Expression of excitatory amino acid transporter transcripts in the thalamus of subjects with schizophrenia. Am J Psychiatry 158:1393–1399

    PubMed  Article  CAS  Google Scholar 

  • Stopkova P, Saito T, Fann CS, Papolos DF, Vevera J, Paclt I, Zukov I, Stryjer R, Strous RD, Lachman HM (2003) Polymorphism screening of PIP5K2A: a candidate gene for chromosome 10p-linked psychiatric disorders. Am J Med Genet B Neuropsychiatr Genet 123B:50–58

    PubMed  Article  Google Scholar 

  • Stopkova P, Vevera J, Paclt I, Zukov I, Papolos DF, Saito T, Lachman HM (2005) Screening of PIP5K2A promoter region for mutations in bipolar disorder and schizophrenia. Psychiatr Genet 15:223–227

    PubMed  Article  Google Scholar 

  • Strutz-Seebohm N, Shojaiefard M, Christie D, Tavare J, Seebohm G, Lang F (2007) PIKfyve in the SGK1 mediated regulation of the creatine transporter SLC6A8. Cell Physiol Biochem 20:729–734

    PubMed  Article  CAS  Google Scholar 

  • van Landeghem FK, Weiss T, von Deimling A (2007) Expression of PACAP and glutamate transporter proteins in satellite oligodendrocytes of the human CNS. Regul Pept 142:52–59

    PubMed  Article  CAS  Google Scholar 

  • Wagner CA, Friedrich B, Setiawan I, Lang F, Broer S (2000) The use of Xenopus laevis oocytes for the functional characterization of heterologously expressed membrane proteins. Cell Physiol Biochem 10:1–12

    PubMed  Article  CAS  Google Scholar 

Download references

Acknowledgements

The authors acknowledge the technical support by Darina Iskalieva and the meticulous preparation of the manuscript by Sari Rübe and Lejla Subasic. This study was supported by the Deutsche Forschungsgemeinschaft, Nr. La 315/6-1, La 315/13-1, and GRK1302.

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Florian Lang.

Additional information

Olga Fedorenko and Cai Tang contributed equally and thus share first authorship.

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Fedorenko, O., Tang, C., Sopjani, M. et al. PIP5K2A-dependent regulation of excitatory amino acid transporter EAAT3. Psychopharmacology 206, 429–435 (2009). https://doi.org/10.1007/s00213-009-1621-5

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00213-009-1621-5

Keywords

  • PIP2
  • Schizophrenia
  • Glutamate
  • Mutation
  • Neuroexcitability