Distinguishing surface effects of calcium ion from pore-occupancy effects in Na+ channels.
The effects of calcium ion on the Na+ activation gate were studied in squid giant axons. Saxitoxin (STX) was used to block ion entry into Na+ channels without hindering access to the membrane surface, making it possible to distinguish surface effects of calcium from pore-occupancy effects. In the presence of STX, gating kinetics were measured from gating current (Ig). The kinetic effects of external calcium concentration changes were small when STX was present. In the absence of STX, lowering the calcium concentration (from 100 to 10 mM) slowed the closing of Na+ channels (measured from INa tails) by more than a factor of 2. Surprisingly, the voltage sensitivity of closing kinetics changed with calcium concentration, and it was modified by STX. Voltage sensitivity apparently depends in part on the ability of calcium to enter and block the channels as voltage is driven negative. In external medium with no added calcium, INa tail current initially increases in amplitude severalfold with the relief of calcium block, then progressively slows and gets smaller, as calcium diffuses out of the layers investing the axon. INa tails seen just before the current disappears suggest that closing in the absence of channel block is very slow or does not occur. INa amplitude and kinetics are completely restored when calcium is returned. The results strongly suggest that calcium occupancy is a requirement for channel closing and that nonoccupied channels fold reversibly into a nonfunctional conformation.
Armstrong
C M
CM
Department of Physiology, University of Pennsylvania School of Medicine, B701 Richards Building, Philadelphia, PA 19104-6085, USA.
D000818:Animals; D001369:Axons / Q000187:drug effects / Q000502:physiology*; D002118:Calcium / Q000494:pharmacology*; D049832:Decapodiformes; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D018408:Patch-Clamp Techniques; D012530:Saxitoxin / Q000494:pharmacology; D026941:Sodium Channel Blockers; D015222:Sodium Channels / Q000502:physiology*
D026941:Sodium Channel Blockers; D015222:Sodium Channels; D012530:Saxitoxin; D002118:Calcium