Single channels activated by acetylcholine in rat superior cervical ganglion

1. The elementary currents flowing through single channels opened by acetylcholine were recorded in rat superior cervical ganglion neurones using patch-clamp methods. Acetylcholine (30 microM) was included in the patch electrode (cell-attached recordings) or applied by ionophoresis (outside-out configuration). All measurements were made at 23-25 degrees C and mostly at -110 mV. 2. Channel openings appeared both as single events and as bursts of events. One population of the currents observed had a conductance of 20.0 +/- 0.2 pS (mean +/- S.E. of mean, n = 4). A second population had a conductance of about 50 pS, occurred more rarely, and was not included in further analysis. 3. Four channel closed time periods and two channel open time periods were found from the distributions of closed and open times. It was found that shorter channel openings (about 0.2 ms) appeared in isolation, whereas longer openings (duration 1.3 +/- 0.2 ms, n = 4) appeared as bursts of openings separated by the shortest channel closed time periods (about 0.15 ms). The next shortest closed time (about 2 ms) apparently corresponds to the lifetime of the channel not activated by acetylcholine. The two longer closed times (about 80 ms and 1 s) may reflect desensitization. The mean burst duration was 8.5 +/- 1.2 ms (n = 4), giving about six openings per burst. 4. Because the time constant of decay of the excitatory post-synaptic current is more similar to the burst duration than to the duration of individual single openings, it is suggested that acetylcholine released from presynaptic nerves may result in a burst of openings rather than a single opening. 5. On the basis of the above assumption, the rate constants were calculated for a sequential model in which acetylcholine binds to the receptor (forward rate k + 1 = 2.3 X 10(7) M-1 s-1; reverse rate k-1 = 1235 s-1) which then undergoes a conformational change to an open state (forward rate beta = 6293 s-1; reverse rate alpha = 894 s-1). 6. When heptamethonium (30 microM) was added to the solution in the patch electrode, the burst duration was markedly shortened, but there was no change in the closed time between two openings within the burst. This effect was voltage-dependent, which suggests that heptamethonium binds to the channel after it is opened by acetylcholine.