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In outside-out patches of mouse-muscle membrane, embryonic-like channels were activated by pulses of acetylcholine (ACh). On increasing the ACh concentration, the rate of desensitization, 1/tau d, increased linearly with the peak open probability, indicating desensitization from the open state. Desensitization had only one time constant tau d at each ACh concentration. Recovery from desensitization was only approximately 10 times slower than desensitization, whereas the probability of steady-state channel opening, declined to < 0.01 with > 10(-6) M ACh. The peak probability of opening in > 10(-4) M ACh pulse was close to 1. A linear reaction scheme was not compatible with these results. The scheme had to be expanded resulting in a circular scheme with two additional ACh binding steps to desensitized channel states. The approximate rate constants of all reaction steps in the circular scheme could be determined using computer simulations. The model predicted that clusters of channel opening had the average duration tau d at the respective ACh concentration. In cell-attached patches on intact muscle fibers, similar average cluster durations were observed at the respective ACh concentration. This indicates that tau d in the intact muscle fibers has similar values as in outside-out patches.

Of all the synapses that have been studied the physiology of neuromuscular transmission in vertebrates is most advanced, mainly as a result of the more recent microphysiological work by del Castillo, Fatt and Katz (Katz, 1958). The chemical nature of the transmission process having been established earlier, they found that transmission occurs by the simultaneous release of many packets or quanta of acetylcholine (ACh) from the motor nerve terminals. These quanta, each of which consists of many molecules of transmitter, cause in the post-synaptic membrane the endplate potential. This sets up conducted impulses in twitch muscle fibres.

Excitatory postsynaptic currents (EPSCs) were recorded extracellularly from large muscle fibers by means of 'patch clamp' electrodes. Compared to usual extracellular recordings, better signal/noise ratio and temporal stability were achieved. In the range of extracellular calcium concentrations [Ca]0 between 2.7 and 13.5 mmol/l (normal), the average amplitude of the EPSC increased more than proportional to [Ca]0. The unit quantum current, C1, and the average release rate, m, were determined from EPSCs and also from spontaneous sEPSCs, using both Poisson and binomial statistics. The main effect of [Ca]0 was on m: at different synaptic sites m depended on the second to fourth power of [Ca]0. In terms of binomial parameters, the release probability p is the [Ca]0-dependent one. In addition, reduction of [Ca]0 from 13.5 to 2.7 mmol/l decreased the unit quantum C1 consistently to 60%; simultaneously the rise and decay of EPSCs and sEPSCs were shortened by 10-20%. [Ca]0 thus has strong presynaptic effects on the release probability, but in addition smaller ones on the postsynaptic channel characteristics.