Steep concentration dependence and fast desensitization of nicotinic channel currents elicited by acetylcholine pulses, studied in adult vertebrate muscle

Abstract

Skeletal muscles of adult mice and frogs were dissociated enzymatically and prepared for patch-clamping within less than 6 h. Outside-out patches were superfused with repetitive pulses of acetylcholine (ACh) with switching times of about 0.2 ms. Peak responses were reached within 1 ms. In mouse muscle the average channel conductance was 65 pS and the average open time 1 ms (20° C). Between 1 and 10 μM ACh, the peak responses increased proportional to the second to third power of the ACh concentration, and less steeply between 10 and 1000 μM ACh. The apparent K _m of the dose-response curve was about 100 μM. After the peak, channel opening probability declined with time constants decreasing from about 1 s with 1 μM ACh to 15–50 ms with 1 mM ACh. After 100 ms desensitization the channel opening had decreased to less than 1/300 peak value. The rate of desensitization increased with rising temperature, with Q ₁₀ values of 1.7–2.5 between 10 and 30° C. The desensitization characteristics of channels from frog muscle were similar to that from mice. With pulses of 100 μM ACh the channels opened with a probability of 0.55, the open probability declining with a time constant of about 60 ms and dropping to less than 0.001 after 300 ms. The results support the view that three binding steps of ACh are necessary for opening of the channel. Desensitization in the presence of high ACh concentrations is slower than the decay of synaptic currents.