MULTIPLE MECHANISMS OF BLOCK BY THE ANESTHETIC ISOFLURANE OF A GAMMA-AMINOBUTYRIC-ACID ACTIVATED CHLORIDE CHANNEL IN CRAYFISH

Outside-out patches were excised from the membrane of the deep extenso r abdominal muscle (DEAM), containing gamma-aminobutyric acid (GABA)-a ctivated chloride channels, in the crayfish Astacus astacus. GABA and isoflurane (iso) were applied in pulses by a liquid filament switch, a nd their effects on the GABA-elicited chloride currents were investiga ted. Application of iso alone elicited no current responses and pre-ap plication of iso prior to GABA had no effects on the GABA-elicited cur rent. Go-application of GABA and iso resulted in a reduction of the in itial chloride current and subsequent decline of the current to a stea dy state, indicating that iso binds to the receptor after GABA has bou nd. Recovery currents at the end of the co-application pulse, their am plitudes decreasing with pulse duration, confirmed this suggestion. Op en-time distributions of the blocked channel showed a shift of the lon g open-time towards a new time constant, indicating a second block mec hanism via the long open state A(5)O(s) of the channel. Removal of GAB A and iso after reaching the equilibrium state of the block resulted i n recovery currents containing exclusively openings from the long open state A(5)O(s), confirming the suggestion of an open channel block on ly at one of the open states.