Open channel and competitive block of the embryonic form of the nicotinic receptor of mouse myotubes by (+)-tubocurarine

1. Embryonic-like nicotinic channels were studied in mouse myotubes. Channel currents were measured by patch clamping outside-out excised patches to which pulses of agonists and drugs could be applied by a liquid filament switch. The holding potential of the patches was generally around -10 to -40 mV. 2. Pulses of 100 μM or 1 mM or 1mM acetylcholine (ACh) elicited average channel currents which reached a maximum open probability of 0.93 within 0.5-1.0 ms, decayed with a time constant of desensitization of 20-80 ms, and fell rapidly to zero at the end of the pulse. When such pulses together with increasing concentrations of (+)-tubocurarine (TC) were applied to outside-out patches, the time constant of current decay, τ(B), decreased beginning at concentrations of TC added to the test solution of > 10 μM, and the peak amplitude of the current decreased markedly at concentrations of TC of > 30 μM due to an open channel block of nicotinic channels by TC. 3. When the outside-out patches were pre-incubated with TC, the peak current elicited by pulses of 100 μM ACh or 1 mM ACh + TC decreased markedly, beginning with concentrations of TC > 30 nM due to a competitive block. 4. The results could be quantitatively modelled by computer calculations based on a circular reaction scheme containing desensitization, TC blocked the open state as well as the unliganded closed state of the embryonic-like nicotinic receptors of mouse myotubes. Also the blocked open channel was subject to desensitization. 5. The rates of block and unblock of the open channel were 3 x 10⁶ M^-1 S^-1 and 0.8 s^-1, respectively, and those of the competitive block were 0.5 x 10⁶ M^-1s^-1 and 0.1 s^-1 respectively (at 20°C).