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 microM or 1 mM 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, tau, decreased beginning at concentrations of TC added to the test solution of > 10 microM, and the peak amplitude of the current decreased markedly at concentrations of TC of > 30 microM 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 microM 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 10(6) M-1 S-1 and 0.8 S-1, respectively, and those of the competitive block were 0.5 x 10(6) M-1 S-1 and 0.1 S-1, respectively (at 20 degrees C).
Bufler
J
J
Physiologisches Institut, Technischen, Universität München, Germany.
Wilhelm
R
R
Parnas
H
H
Franke
C
C
Dudel
J
J
D000109:Acetylcholine / Q000494:pharmacology; D000818:Animals; D004622:Embryo, Mammalian / Q000187:drug effects*; D006321:Heart / Q000187:drug effects*; D007473:Ion Channels / Q000187:drug effects*; D051379:Mice; D008815:Mice, Inbred Strains; D018408:Patch-Clamp Techniques; D011978:Receptors, Nicotinic / Q000187:drug effects*; D014403:Tubocurarine / Q000494:pharmacology*
D007473:Ion Channels; D011978:Receptors, Nicotinic; D000109:Acetylcholine; D014403:Tubocurarine