Augmentation of recovery from inactivation by site-3 Na channel toxins - Asingle-channel and whole-cell study of persistent currents

Site-3 toxins isolated from several species of scorpion and sea anemone bin d to voltage-gated Na channels and prolong the time course of I-Na by inter fering with inactivation with little or no effect on activation, effects th at have similarities to those produced by genetic diseases in skeletal musc le (myotonias and periodic paralysis) and heart (long QT syndrome). Some pu blished reports have also reported the presence of a noninactivating persis tent current in site-3 toxin-treated cells. We have used the high affinity site-3 toxin Anthopleurin B to study the kinetics of this current and to ev aluate kinetic differences between cardiac (in RT4-B8 cells) and neuronal ( in N1E-115 cells) Na channels. By reverse transcription-PCR from N1E-115 ce ll RNA multiple Na channel transcripts were detected; most often isolated w ere sequences homologous to rBrII, although at low frequency sequences homo logous to rPN1 and rBrIII were also detected. Toxin treatment induced a vol tage-dependent plateau current in both isoforms for which the relative ampl itude (plateau current/peak current) approached a constant value with depol arization, although the magnitude was much greater for neuronal (17%) than cardiac (5%) INa Cell-attached patch recordings revealed distinct quantitat ive differences in open times and burst durations between isoforms, but for both isoforms the plateau current comprised discrete bursts separated by q uiescent periods, consistent with toxin induction of an increase in the rat e of recovery from inactivation rather than a modal failure of inactivation . In accord with this hypothesis, toxin increased the rate of whole-cell re covery at all tested voltages. Moreover, experimental data support a model whereby recovery at negative voltages is augmented through closed states ra ther than through the open state. We conclude that site-3 toxins produce qu alitatively similar effects in cardiac and neuronal channels and discuss im plications for channel kinetics.