Life time and elementary conductance of the channels mediating the excitatory effects of acetylcholine in Aplysia neurones

Abstract

1. The excitatory effects of acetylcholine (ACh) on an identified group of Aplysia neurones have been studied under voltage clamp in an attempt to measure the average life time. tau, of the channels opened by ACh and the elementary current, iel, flowing through these channels. The value of tau was determined both from spectral noise analysis and from current relaxations after voltage steps. Both methods lead to similar values. iel was calculated from the ratio of the variance of the ACh induced noise to the mean ACh induced current. 2. tau is increased by hyperpolarization, or by lowering the temperature. At 12 degrees C, tau = 27 msec AT -80 MV, tau = 17 msec at mV. tau is about 5 times smaller at 21 degrees C than at 12 degrees C. 3. iel increases linearly with hyperpolarization. At -80 mV, in Tris-buffered sea water, the mean value of iel was 0.8 X 10)-12) A at 12 degrees C. At 21 degrees C, this value was multiplied by 1.8. 4. The estimate of the ACh reversal potential Erev obtained by extrapolation of the relation between iel and the membrane potential V was + 30 mV. The estimate obtained from the analysis of the instantaneous current changes produced by voltage steps was + 15 mV. The difference between the two values appears to be due to the development of a K curent activated by the entry of Ca into the cell during the ACh response. This current introduces an error in opposite directions into the two estimates of Erev, which can therefore be assumed to be intermediate between + 15 and + 30 mV. An assumed value of + 20 mV yields an elementary conductance of 8 X 10(-12) omega-1 at 12 degrees C in Tris-buffered sea water. 5. The total ACh induced current measured in steady-state conditions increases more with hyperpolarization than does iel. The difference can be entirely accounted for by the fact that hyperpolarization increases tau. 6. When carbachol or tetramethylammonium is applied instead of ACh, the value of iel is identical to that found with ACh, but tau is slightly shorter (about 75%). 7. Inward ACh induced currents can still be observed in solutions where all Na has been replaced by Cs, Mg, or Ca. 8. iel increases when Na is replaced by Cs; it decreases when Na is replaced by Mg or Ca. In all Na-free solutions, tau is larger than in Na sea water: the lengthening of tau is largest for Ca sea water, smallest for Cs sea water. An interpretation of these changes of gamma is proposed. This interpretation may also account for the voltage sensitivity of gamma in normal sea water. 9. Partial replacement of NaCl by TrisCl strikingly reduces the ACh induced current. gamma is not modified by Tris substitution, and the reduction of the total current is entirely accounted for by a steep decrease of iel. Tris does not seem to affect the pore opening and closing processes, but to block the ACh controlled channel.