This Article Full Text Full Text (PDF) All Versions of this Article: biophysj.107.104794v1 93/12/4209    most recent Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via Google Scholar Google Scholar Articles by Szendroedi, J. Articles by Todt, H. Search for Related Content PubMed PubMed Citation Articles by Szendroedi, J. Articles by Todt, H.

Speeding the Recovery from Ultraslow Inactivation of Voltage-Gated Na⁺ Channels by Metal Ion Binding to the Selectivity Filter: A Foot-on-the-Door?

Julia Szendroedi ^*, Walter Sandtner ^*, Touran Zarrabi ^*, Eva Zebedin ^*, Karlheinz Hilber ^*, Samuel C. Dudley, Jr. , Harry A. Fozzard  and Hannes Todt ^*

^* Center for Biomolecular Medicine and Pharmacology, Institute of Pharmacology, Medical University of Vienna, Vienna, Austria; Division of Cardiology, Emory University, Atlanta, Georgia, and the Atlanta Veterans Administration Hospital, Decatur, Georgia; and University of Chicago, Chicago, Illinois

Correspondence: Address reprint requests to Hannes Todt, Center for Biomolecular Medicine and Pharmacology, Institute of Pharmacology, Medical University of Vienna, Waehringerstrasse 13A, A-1090 Vienna, Austria. Tel.: 43-1-4277-64120; E-mail: hannes.todt{at}meduniwien.ac.at.

Slow inactivated states in voltage-gated ion channels can be modulated by binding molecules both to the outside and to the inside of the pore. Thus, external K⁺ inhibits C-type inactivation in Shaker K⁺ channels by a "foot-in-the-door" mechanism. Here, we explore the modulation of a very long-lived inactivated state, ultraslow inactivation (I_US), by ligand binding to the outer vestibule in voltage-gated Na⁺ channels. Blocking the outer vestibule by a mutant µ-conotoxin GIIIA substantially accelerated recovery from I_US. A similar effect was observed if Cd²⁺ was bound to a cysteine engineered to the selectivity filter (K1237C). In K1237C channels, exposed to 30 µM Cd²⁺, the time constant of recovery from I_US was decreased from 145.0 ± 10.2 s to 32.5 ± 3.3 s (P < 0.001). Recovery from I_US was only accelerated if Cd²⁺ was added to the bath solution during recovery (V = –120 mV) from I_US, but not when the channels were selectively exposed to Cd²⁺ during the development of I_US (–20 mV). These data could be explained by a kinetic model in which Cd²⁺ binds with high affinity to a slow inactivated state (I_S), which is transiently occupied during recovery from I_US. A total of 50 µM Cd²⁺ produced an 8 mV hyperpolarizing shift of the steady-state inactivation curve of I_S, supporting this kinetic model. Binding of lidocaine to the internal vestibule significantly reduced the number of channels entering I_US, suggesting that I_US is associated with a conformational change of the internal vestibule of the channel. We propose a molecular model in which slow inactivation (I_S) occurs by a closure of the outer vestibule, whereas I_US arises from a constriction of the internal vestibule produced by a widening of the selectivity filter region. Binding of Cd²⁺ to C1237 promotes the closure of the selectivity filter region, thereby hastening recovery from I_US. Thus, Cd²⁺ ions may act like a foot-on-the-door, kicking the I_S gate to close.