Behaviour of chemically modified sodium channels in frog nerve supports a three-state model of inactivation

Abstract

Voltage clamp experiments were done on single myelinated nerve fibres of the frog,Rana esculenta, with 10 mM TEA⁺ in the external solutions to block potassium channels. Sodium current inactivation was measured in TEA-Ringer solution and after treatment withAnemonia sulcata toxin II (5 μM), internal iodate (20/40 mM), glutaraldehyde (10 mM), chloramine-T (0.6 mM), and 2,4,6-trinitrophenol (1 mM). The diphasic inactivation time course, observed in untreated membranes, is slowed by all these agents in a very similar way. Both time constants are increased and the proportion of inactivation components is changed favouring the slowly inactivating one. Trinitrophenol only slows inactivation, whereas inAnemonia toxin II, internal iodate, glutaraldehyde and chloramine-T inactivation becomes incomplete, so that a persistent current is flowing during depolarizations. None of these agents even at high concentrations however, totally removes inactivation. These modifications of inactivation time course are interpreted as changes of rate constants in a three-state inactivation model with one open and two closed states (o-c-c). After chemical treatment the access to the closed states is impeded and the transitions into the open state are accelerated. If the membrane is depolarized during drug application chloramine-T fails to modify inactivation. The curve relating the steady state inactivation parameter,h _∞, to the conditioning potential,V _pp becomes non-monotonic in chloramine-T, i.e.dh _∞/dV _pp>0 forV _pp>60 mV. Trinitrophenol, which per se fails to produce a persistent current component, increases the persistent current in a fibre pretreated with chloramine-T. This finding is incompatible with the idea of ascribing the persistent current to a non-inactivating channel fraction. It is, however, consistent with a single channel population obeying the o-c-c model of inactivation.