Guanine Nucleotide-Sensitive Inhibition of L-Type Ca2+ Current by Lysosphingolipids in RINm5F Insulinoma Cells

  1. Herbert M. Himmel1,2,
  2. Dagmar Meyer zu Heringdorf1,
  3. Bernd Windorfer1,
  4. Chris J. van Koppen1,
  5. Ursula Ravens1,2 and
  6. Karl H. Jakobs1
  1. 1Institut für Pharmakologie, Universitätsklinikum Essen, D-45122 Essen, Germany (H.M.H., D.M. zu H., B.W., C.J. van K., U.R., K.H.J.) and 2Institut für Pharmakologie und Toxikologie, Technische Universität Dresden, D-01109 Dresden, Germany (H.M.H., U.R.)

    Abstract

    The lysosphingolipids sphingosine-1-phosphate (SPP) and sphingosylphosphorylcholine (SPPC) reportedly increase free cytosolic Ca2+ concentration ([Ca2+]i) in a variety of cell types, apparently by activating G protein-coupled plasma membrane receptors. We investigated whether and how sphingolipids modulate Ca2+ homeostasis in the insulinoma cell line RINm5F. The addition of SPPC and glucopsychosine (GPS) did not affect basal [Ca2+]i but inhibited the KCl (30 mm)-induced increase in [Ca2+]i in a pertussis toxin-insensitive and concentration-dependent manner (EC50 ∼ 5 μm). Similar inhibitory effects were observed with dihydro-SPPC and psychosine, whereas SPP and variousN-acylated sphingolipids (at 10 μm each) had little or no effect on the KCl-induced [Ca2+]i increase. Because in RINm5F cells the primary pathway for depolarization-induced [Ca2+]i increase are L-type Ca2+ channels, we studied whether sphingolipids reduceL-type Ca2+ current (ICa.L). When added to the bath, GPS and SPPC, but not SPP (10 μmeach), rapidly reduced maximal ICa.L by ∼35%, similar to the α2-adrenoceptor agonist clonidine (30 μm). However, when applied internally, GPS had no effect on ICa.L. When the electrode solution contained the stable GDP analog guanosine-5′-O-(2-thio)diphosphate (1 and 10 mm), the inhibitory effect of GPS was abolished. In conclusion, a novel cellular action of lysosphingolipids is observed in RINm5F cells (i.e., a guanine nucleotide-sensitive inhibition ofL-type Ca2+ currents). The pharmacological profile of this inhibition is unique and unlike any known lysosphingolipid receptor-mediated action.

    Footnotes

    • Send reprint requests to: Dr. Karl H. Jakobs, Institut für Pharmakologie, Universitätsklinikum Essen, Hufelandstrasse 55, D-45122 Essen, Germany.

    • This work was supported by Grant 0310493A from Bayer AG and the Bundesministerium für Bildung and Wissenschaft, Forschung und Technologie and the Juterne Forschungsförderung Essen program of the Universitätsklinikum Essen.

    • Preliminary results have been published in abstract form: Himmel HM, Meyer zu Heringdorf D, Windorfer B, van Koppen CJ, Ravens U, and Jakobs KH (1997) Sphingolipid receptor-mediated inhibition ofL-type Ca2+ current in the insulinoma cell line RINm5F. Naunyn-Schmiedebergs Arch Pharmacol355:R52.

    • H.M.H. and D.M.z.H. contributed equally to this work.

    • Abbreviations:
      SPP
      sphingosine-1-phosphate
      [Ca2+]i
      cytosolic free Ca2+concentration
      GDPβS
      guanosine-5′-O-(2-thio)diphosphate
      GPS
      glucopsychosine
      GTPγS
      guanosine-5′-O-(3-thio)triphosphate
      HEK
      human embryonic kidney
      HEPES
      4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid
      ICa.L
      L-type Ca2+ current
      PKC
      protein kinase C
      PS
      psychosine
      PTX
      pertussis toxin
      SPPC
      sphingosylphosphorylcholine
      EGTA
      ethylene glycol bis(β-aminoethyl ether)-N,N,N′,N′-tetraacetic acid
      • Received April 30, 1997.
      • Accepted January 21, 1998.
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    1. Molecular Pharmacology May 1, 1998 vol. 53 no. 5 862-869
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