Indirect activation of neuronal noncapacitative Ca2+ entry is the final step involved in the neurotoxic effect of Tityus serrulatus scorpion β-toxin

Authors

  • Françoise Grolleau,

    1. Laboratoire RCIM UPRES EA 2647, Université d'Angers, Faculté des Sciences, 2 bd Lavoisier, F-49045 Angers cedex, France
    Search for more papers by this author
  • Maria Stankiewicz,

    1. Institute of General and Molecular Biology, N. Copernicus University, 87 100 Torun, Poland
    Search for more papers by this author
  • Ewa Kielbasiewicz,

    1. Laboratoire RCIM UPRES EA 2647, Université d'Angers, Faculté des Sciences, 2 bd Lavoisier, F-49045 Angers cedex, France
    2. Institute of General and Molecular Biology, N. Copernicus University, 87 100 Torun, Poland
    Search for more papers by this author
  • Marie-France Martin-Eauclaire,

    1. FRE CNRS 2738, Institut Jean Roche, Université de la Méditerranée, Faculté de Médecine Nord, bd Pierre Dramard, F-13916 Marseille cedex 20, France
    Search for more papers by this author
  • Céline Lavialle,

    1. Laboratoire RCIM UPRES EA 2647, Université d'Angers, Faculté des Sciences, 2 bd Lavoisier, F-49045 Angers cedex, France
    Search for more papers by this author
  • Jan De Vente,

    1. Department of Psychiatry and Neuropsychology, POB 616, European School of Neurosciences (EURON), Universiteit Maastricht 6200 MD Maastricht, the Netherlands
    Search for more papers by this author
  • Bruno Lapied

    1. Laboratoire RCIM UPRES EA 2647, Université d'Angers, Faculté des Sciences, 2 bd Lavoisier, F-49045 Angers cedex, France
    Search for more papers by this author

Professor Françoise Grolleau, as above.
E-mail: francoise.grolleau@univ-angers.fr

Abstract

Interweaving strategies of electrophysiology, calcium imaging and immunocytochemistry bring new insights into the mode of action of the Brazilian scorpion Tityus serrulatusβ-toxin VII. Pacemaker dorsal unpaired median neurons isolated from the cockroach central nervous system were used to study the effects of toxin VII. In current-clamp, 50 nm toxin VII produced a membrane depolarization and reduced spiking. At 200 nm, depolarization associated with multiphasic effects was seen. After artificial hyperpolarization, plateau potentials on which spontaneous electrical activity appeared were observed. In voltage clamp, toxin VII induced a negative shift of the voltage dependence of sodium current activation without significant effect on steady-state inactivation. In addition, toxin VII produced a permanent TTX-sensitive holding inward current, indicating that background sodium channels were targeted by β-toxin. Cell-attached patch recordings indicated that these channels were switched from unclustered single openings to current fluctuating between distinct subconductance levels exhibiting increased open probability and open-time distribution. Toxin VII also produced a TTX-sensitive [Ca2+]i rise. Immunostaining with Cav2.2(α1b) antibodies and calcium imaging data obtained with ω-CgTx GVIA indicated that N-type high-voltage-activated calcium channels initiated calcium influx and were an essential intermediate in the pathway linking toxin VII-modified sodium channels to the activation of an additional route for calcium entry. By using inhibitors of (i) noncapacitative calcium entry (inhibitor LOE-908), (ii) NO-sensitive guanylyl cyclase (ODQ) and (iii) phosphodiesterase 2 (EHNA), together with cGMP antibodies, we demonstrated that noncapacitative calcium entry was the final step in a complex combination of events that was initiated by toxin VII-alteration of sodium channels and then involved successive activation of other membrane ion channels.

Ancillary