Modification of cardiac Na+ channels by anthopleurin-A: effects on gating and kinetics.
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 infinity) 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 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.(ABSTRACT TRUNCATED AT 400 WORDS)
Wasserstrom
J A
JA
Department of Medicine (Cardiology), Northwestern University Medical School, Chicago, IL 60611.
Kelly
J E
JE
Liberty
K N
KN
D000818:Animals; D002118:Calcium / Q000494:pharmacology; D002413:Cations, Divalent; D004594:Electrophysiology; D006168:Guinea Pigs; D006321:Heart / Q000502:physiology*; D006352:Heart Ventricles / Q000187:drug effects; D036341:Intercellular Signaling Peptides and Proteins; D015640:Ion Channel Gating / Q000502:physiology*; D007700:Kinetics; D008274:Magnesium / Q000494:pharmacology; D010455:Peptides / Q000494:pharmacology*; D015222:Sodium Channels / Q000187:drug effects / Q000502:physiology*; D016276:Ventricular Function
D002413:Cations, Divalent; D036341:Intercellular Signaling Peptides and Proteins; D010455:Peptides; D015222:Sodium Channels; C012746:anthopleurin-A; D008274:Magnesium; D002118:Calcium