Modification of cardiac Na+ channels by anthopleurin-A: effects on gating and kinetics

J. Andrew Wasserstrom*, James E. Kelly, Kristine N. Liberty

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

19 Scopus citations

Abstract

We used the whole cell patch clamp technique to investigate the characteristics of modification of cardiac Na+ channel gating by the sea anemone polypeptide toxin anthopleurin-A (AP-A). Guinea pig ventricular myocytes were isolated enzymatically using a retrograde perfusion apparatus. Holding potential was -140 mV and test potentials ranged from -100 to + 40 mV (pulse duration 100 or 1000 ms). AP-A (50-100 nM) markedly slowed the rate of decay of Na+ current (INa) and increased peak INa conductance (gNa) by 38±5.5% (mean±SEM, P < 0.001, n = 12) with little change in slope factor (n = 12) or voltage midpoint of the gNa/V relationship after correction for spontaneous shifts. The voltage dependence of steady-state INa availability (h) demonstrated an increase in slope factor from 5.9±0.8 mV in control to 8.0±0.7 mV after modification by AP-A (P < 0.01, n = 14) whereas any shift in the voltage midpoint of this relationship could be accounted for by a spontaneous time-dependent shift. AP-A-modified INa showed a use-dependent decrease in peak current amplitude (interpulse interval 500 ms) when pulse duration was 100 ms (-15±2%, P < 0.01, n = 17) but showed no decline when pulse duration was 100 ms (-3±1%). This use-dependent effect was probably the result of a decrease in the rate of recovery from inactivation caused by AP-A which had a small effect on the fast time constant of recovery (from 4.1±0.3 ms in control to 6.0±1.1 ms after AP-A, P < 0.05) but increased the slow time constant from 66.2±6.5 ms in control to 188.9±36.4 ms (P< 0.002, n = 19) after exposure to AP-A. Increasing external divalent cation concentration (either Ca2+ or Mg2+) to 10 mM abolished the effects of AP-A on the rate of INa decay. These results demonstrate that modification of cardiac Na+ channels by AP-A markedly slowed INa inactivation and altered the voltage dependence of activation; these alterations in gating characteristics, in turn, caused an increase in gNa presumably by increasing the number of channels open at peak INa. AP-A slows the rate of recovery of INa from inactivation which is probably the basis for a use-dependent decrease in peak amplitude. Finally, AP-A binding is sensitive to external divalent cation concentrations. Thus, increasing [Mg2+]o or [Ca2+]o displaces AP-A from binding, suggesting that they share related binding sites on the external surface of the Na+ channel.

Original languageEnglish (US)
Pages (from-to)15-24
Number of pages10
JournalPflügers Archiv European Journal of Physiology
Volume424
Issue number1
DOIs
StatePublished - Jun 1993

Keywords

  • Activation
  • Anthopleurin-A
  • Inactivation
  • Na current

ASJC Scopus subject areas

  • Physiology
  • Clinical Biochemistry
  • Physiology (medical)

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