An acetylcholine receptor regulatory site in BC3H1 cells: characterized by laser-pulse photolysis in the microsecond-to-millisecond time region.

When a neurotransmitter binds to its specific receptor, the protein forms transmembrane channels through which ions flow, leading to changes in transmembrane voltage that trigger signal transmission between neurons. How do inhibitors affect this process? Interesting and extensive information comes from investigations of the acetylcholine receptor, the best known of these proteins. This receptor is inhibited by cationic inhibitors, including local anesthetics, and acetylcholine at high concentrations. The accepted mechanism, elegant in its simplicity, is that these compounds enter the receptor-channel after it opens and block inorganic ion flux. This mechanism requires that the inhibitors affect only the apparent rate constant for channel closing (k'cl). An alternative mechanism invokes a specific regulatory (inhibitory) site to which inhibitors bind before the channel opens and the signal is transmitted. This mechanism requires that the inhibitors affect the apparent rate constants for both channel opening (k'op) and closing. The effect of inhibitors on k'op has not been determined previously. This report describes the use of a newly developed laser-pulse photolysis technique with a dead time of approximately 120 microseconds to determine the effect of a local anesthetic, procaine, one of the best studied cationic inhibitors of the acetylcholine receptor, on both k'op and k'cl. Both k'op and k'cl were found to decrease with increasing procaine concentration. This effect of the inhibitor of k'op cannot be explained by the open-channel-blocking mechanism but is consistent with the existence of a regulatory (inhibitory) receptor site.