POST-REPOLARIZATION BLOCK OF CLONED SODIUM-CHANNELS BY SAXITOXIN - THE CONTRIBUTION OF PORE-REGION AMINO-ACIDS

Sodium channels expressed in oocytes exhibited isoform differences in phasic block by saxitoxin (STX). Neuronal channels (rat lla co-express ed with beta 1 subunit, Br2a + beta 1) had slower kinetics of phasic b lock for pulse trains than cardiac channels (RHI). After the membrane was repolarized from a single brief depolarizing step, a test pulse at increasing intervals showed first a decrease in current (post-repolar ization block) then eventual recovery in the presence of STX. This blo ck/unblock process for Br2a + beta 1 was 10-fold slower than that for RHI. A model accounting for these results predicts a faster toxin diss ociation rate and a slower association rate for the cardiac isoform, a nd it also predicts a shorter dwell time in a putative high STX affini ty conformation for the cardiac isoform. The RHI mutation (Cys(374)--> Phe), which was previously shown to be neuronal-like with respect to h igh affinity tonic toxin block, was also neuronal-like with respect to the kinetics of post-repolarization block, suggesting that this singl e amino acid is important for conferring isoform-specific transition r ates determining post-repolarization block. Because the same mutation determines both sensitivity for tonic STX block and the kinetics of ph asic STX block, the mechanisms accounting for tonic block and phasic b lock share the same toxin binding site. We conclude that the residue a t position 374, in the putative pore-forming region, confers isoform-s pecific channel kinetics that underlie phasic toxin block.