Pflügers Archiv

, Volume 415, Issue 3, pp 255–261 | Cite as

Direct modulation of voltage-dependent calcium channels by muscarinic activation of a pertussis toxin-sensitive G-protein in hippocampal neurons

  • M. Toselli
  • J. Lang
  • T. Costa
  • H. D. Lux
Excitable Tissues and Central Nervous Physiology


Acetylcholine (Ach) reversibly reduces the high voltage-activated (HVA) calcium (Ca) current in hippocampal neurons. Pretreatment of the cells with pertussis toxin (PTX) abolishes the Ach effect, suggesting that PTX-sensitive GTP-binding regulatory proteins (G-proteins) are involved in the signal transduction mechanism that links Ach receptor activation to inhibition of Ca channel activity. This effect is mimicked by intracellular application of the nonhydrolyzable GTP analog GTPγS. Intracellular application of purified G-proteins restored the response to Ach in PTX-treated cells. Furthermore, Ach inhibits the Ca current independently of the presence of cyclic AMP and of the protein kinase C inhibitor H-7 and neither does the Ach effect on the Ca current seem to be correlated to a transient increase in intracellular Ca. Our results suggest that activation of the α-subunit of the PTX-sensitive G-protein could directly modulate the HVA Ca channel without involving second messenger systems.

Key words

Ca channels G-proteins Hippocampus Acetylcholine 


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  1. Carbone E, Lux HD (1987) Kinetics and selectivity of a low voltage activated calcium current in chick and rat sensory neurones. J Physiol (Lond) 386:547–570Google Scholar
  2. Codina J, Yatani A, Grenet D, Brown AM, Birnbaumer L (1987) The α subunit of the GTP binding protein Gk opens atrial potassium channels. Science 236:442–445Google Scholar
  3. Davies NW, Lux HD, Morad M (1988) Site and mechanism of activation of proton-induced sodium current in chick dorsal root ganglion neurons. J Physiol (Lond) 400:159–187Google Scholar
  4. Dolphin AC, Scott RH (1987) Calcium channel currents and their inhibition by (-)-baclofen in rat sensory neurones: modulation by guanine nucleotides. J Physiol (Lond) 386:1–17Google Scholar
  5. Dolphin AC, Wootton JF, Scott RH, Trentham DR (1988) Photoactivation of intracellular guanosine triphosphate analogues reduces the amplitude and slows the kinetics of voltageactivated calcium channel currents in sensory neurons. Pflügers Arch 411:628–636Google Scholar
  6. Dunlap K, Holz GG, Rane SG (1987) G proteins as regulators of ion channel function. Trends Neurosci 10:241–244Google Scholar
  7. Evans T, Martin M, Hughes AR, Harden TK (1985) Guanine nucleotide sensitive, high affinity binding of carbachol to muscarinic cholinergic receptors of 1321N1 astrocytoma cells is insensitive to pertussis toxin. Mol Pharmacol 27:32–37Google Scholar
  8. Ewald DA, Sternweis PC, Miller RJ (1988) Guanine nucleotidebinding protein Go-induced coupling of neuropeptide Y receptors to Ca2+ channels in sensory neurons. Proc Natl Acad Sci USA 85:3633–3637Google Scholar
  9. Fisher SK, Bartus RT (1985) Regional differences in the coupling of muscarinic receptors to inositol phospholipid hydrolysis in guinea pig brain. J Neurochem 45:1085–1095Google Scholar
  10. Florio VA, Sternweis PC (1985) Reconstitution of resolved muscarinic cholinergic receptors with purified GTP-binding proteins. J Biol Chem 260:3477–3483Google Scholar
  11. Fox AP, Nowycky MC, Tsien RW (1987) Kinetic and pharmacological properties distinguishing three types of calcium currents in chick sensory neurons. J Physiol (Lond) 394:149–172Google Scholar
  12. Gähwiler BH, Brown DA (1987) Muscarine affects calcium currents in rat hippocampal pyramidal cells in vitro. Neurosci Lett 76:301–306Google Scholar
  13. Gilman AG (1987) G proteins: transducers of receptor-generated signals. Annu Rev Biochem 56:615–649Google Scholar
  14. Goldsmith P, Backlund PS, Rossiter K, Carter A, Milligan G, Unson CG, Spiegel A (1988) Purification of heterotrimeric GTP-binding proteins from brain: identification of a novel form of Go. Biochemistry 27:7085–7090Google Scholar
  15. Hamill OP, Marty A, Neher E, Sakmann B, Sigworth FJ (1981) Improved patch clamp techniques for high-resolution current recording from cells and cell-free membrane patches. Pflügers Arch 391:85–100Google Scholar
  16. Harris-Warrick RM, Hammond C, Paupardin-Tritsch D, Homburger V, Rouot B, Bockaert J, Gerschenfeld HM (1988) An α40 subunit of a GTP-binding protein immunologically related to Go mediates a dopamine-induced decrease of Ca current in snail neurons. Neuron 1:27–32Google Scholar
  17. Hescheler J, Rosenthal W, Trautwein W, Schultz G (1987) The GTP-binding protein, Go, regulates neuronal calcium channels. Nature 325:445–447Google Scholar
  18. Hidaka H, Inagaki M, Kawamoto S, Sasaki Y (1984) Isoquino-linesulfonamides, novel and potent inhibitors of cyclic nucleotide dependent protein kinase and protein kinase C. Biochemistry 23:5036–5041Google Scholar
  19. Hildebrandt JD, Sekura RD, Codina J, Iyengar R, Manclark CR, Birnbaumer L (1983) Stimulation and inhibition of adenylyl cyclases mediated by distinct regulatory proteins. Nature 302:706–709Google Scholar
  20. Huff RM, Neer EJ (1986) Subunit interactions of native and ADP-ribosylated α39 and α41, two guanine nucleotide-binding proteins from bovine cerebral cortex. J Biol Chem 261:1105–1110Google Scholar
  21. Jakobs KH, Gehring U, Gaugler T, Pfeuffer T, Schultz G (1983) Occurrence of an inhibitory guanine nucleotide-binding regulatory component of the adenylate cyclase system in cyc variants of S49 lymphoma cells. Eur J Biochem 130:605–611Google Scholar
  22. Jakobs KH, Aktories K, Schultz G (1984) Mechanism of pertussis toxin action on the adenylate cyclase system. Eur J Biochem 140:177–181Google Scholar
  23. Katada T, Bokoch GM, Smigel MD, Ui M, Gilman AG (1984) The inhibitory guanine nucleotide-binding regulatory component of adenylate cyclase. J Biol Chem 259:3586–3595Google Scholar
  24. Katada T, Oinuma M, Ui M (1986) Two guanine nucleotide-binding proteins in rat brain serving as the specific substrate of isletactivating protein, pertussis toxin. J Biol Chem 261:8182–8191Google Scholar
  25. Katada T, Oinuma M, Kusakabe K, Ui M (1987) A new GTP-binding protein in brain tissues serving as the specific substrate of islet-activating protein, pertussis toxin. FEBS Lett 213:353–358Google Scholar
  26. Kurachi Y, Nakajima T, Sugimoto T (1986) On the mechanism of activation of muscarinic K+ channels by adenosine in isolated atrial cells: involvement of GTP-binding proteins. Pflügers Arch 407:264–274Google Scholar
  27. Kurose H, Katada T, Haga T, Haga K, Ichiyama A, Ui M (1986) Functional interaction of purified muscarinic receptors with purified inhibitory guanine nucleotide regulatory proteins reconstituted in phospholipid vesicles. J Biol Chem 261:6423–6428Google Scholar
  28. Lang J (1989) Purification and characterization of subforms of the guanine nucleotide-binding proteins Gαi and Gαo. Eur J Biochem (in press)Google Scholar
  29. Lang J, Costa T (1987) Antisera against the 3–17 sequence of rat Gαi recognize only a 40-kDa G-protein in brain. Biochem Biophys Res Commun 148:838–848Google Scholar
  30. Lewis PR, Shute CCD (1967) The cholinergic limbic system: projections to hippocampal formation, medial cortex, nuclei of the ascending cholinergic reticular system, and the subformical organ and supra-optic crest. Brain 90:521–540Google Scholar
  31. Lewis DL, Weight FF, Luini A (1986) A guanine nucleotide-binding protein mediates the inhibition of voltage-dependent calcium current by somatostatin in a pituitary cell line. Proc Natl Acad Sci USA 83:9035–9039Google Scholar
  32. Masters SB, Harden TK, Brown JH (1984) Relationships between phosphoinositide and calcium responses to muscarinic agonists in astrocytoma cells. Mol Pharmacol 26:149–155Google Scholar
  33. Masters SB, Martin MW, Harden TK, Brown JH (1985) Pertussis toxin does not inhibit muscarinic receptor-mediated phosphoinositide hydrolysis or calcium mobilization. Biochem J 227:933–937Google Scholar
  34. Misgeld U, Calabresi P, Dodt U (1986) Muscarinic modulation of calcium dependent plateau potentials in rat neostriatal neurons. Pflügers Arch 407:482–487Google Scholar
  35. Nathanson NM (1987) Molecular properties of the muscarinic acetylcholine receptor. Annu Rev Neurosci 10:195–236Google Scholar
  36. Pfeuffer T, Helmreich EJM (1975) Activation of pigeon erythrocyte membrane adenylate cyclase by guanyl nucleotide analogues and separation of a nucleotide binding protein. J Biol Chem 250:867–876Google Scholar
  37. Rodbell M (1980) The role of hormone receptors and GTP-regulatory proteins in membrane transduction. Nature 284:17–22Google Scholar
  38. Segal M (1983) Rat hippocampal neurons in culture: responses to electrical and chemical stimuli. J Neurophysiol 50:1249–1264Google Scholar
  39. Sternweis PC, Robishaw JD (1984) Isolation of two proteins with high affinity for guanine nucleotides from membranes of bovine brain. J Biol Chem 259:838–848Google Scholar
  40. Toselli M, Lux HD (1989a) GTP binding proteins mediate acetylcholine inhibition of voltage dependent calcium channels in hippocampal neurons. Pflügers Arch 413:319–321Google Scholar
  41. Toselli M, Lux HD (1989b) Opposing effects of acetylcholine on the two classes of voltage-dependent calcium channels in hippocampal neurons. In: Frotscher M, Misgeld U (eds) Central cholinergic synaptic transmission. Birkhäuser, Basel (in press)Google Scholar
  42. Trautwein W, Kameyama M, Hescheler J, Hofmann F (1986) Cardiac calcium channels and their transmitter modulation. Fortschr Zool 33:163–182Google Scholar
  43. Ueda H, Harada H, Nazaki M, Katada T, Ui M, Satoh M, Takagi H (1988) Reconstitution of brain μ-opioid receptors with purified guanine nucleotide-binding proteins, Gi and 2 and Go. Proc Natl Acad Sci USA 85:7013–7017Google Scholar
  44. VanDongen AMJ, Codina J, Olate J, Mattera R, Joho R, Birnbaumer L, Brown AM (1988) Newly identified brain potassium channels gated by the guanine nucleotide binding protein Go. Science 242:1433–1437Google Scholar
  45. Wanke E, Ferroni A, Malgaroli A, Ambrosini A, Pozzan T, Meldolesi J (1987) Activation of a muscarinic receptor selectively inhibits a rapidly inactivated Ca current in rat sympathetic neurons. Proc Natl Acad Sci USA 84:4313–4317Google Scholar
  46. Yaari Y, Hamon B, Lux HD (1987) Development of two types of calcium channels in cultured mammalian hippocampal neurons. Science 235:680–682Google Scholar
  47. Yatani A, Codina Y, Brown AM. Birnbaumer L (1987) Direct activation of mammalian atrial muscarinic potassium channels by GTP regulatory protein Gk. Science 235:207–211Google Scholar

Copyright information

© Springer-Verlag 1989

Authors and Affiliations

  • M. Toselli
    • 1
  • J. Lang
    • 2
  • T. Costa
    • 2
  • H. D. Lux
    • 1
  1. 1.Abteilung NeurophysiologieMax-Planck-Institut für PsychiatriePlaneggFederal Republic of Germany
  2. 2.Abteilung NeuropharmakologieMax-Planck-Institut für PsychiatriePlaneggFederal Republic of Germany

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