Measurement of the conductance of the sodium channel from current fluctuations at the node of Ranvier.

Abstract

Single myelinated nerve fibers of Rana esculenta were investigated under voltage clamp conditions at 13.degree. C. Fluctuations of steady-state membrane current were measured during the last 152 ms of 190-225 ms pulses depolarizing the membrane by 8-48 mV. Noise power spectral densities were calculated in the frequency range of 6.6-6757 Hz. External application of 150 nM tetrodotoxin (TTX) and/or 10 mM tetraethylammonium (TEA) ion reduced the current fluctuations. The difference of current noise spectra measured in the presence and absence of TTX (TEA) was not changed by the presence of TEA (TTX) during both measurements, and was taken as the spectrum of the Na+ (K+) current fluctuations. Residual current noise during application of both TTX and TEA was, except for some excess noise at the low and high frequency ends of the spectrum, similar to the noise measured from a passive nerve model and could be understood in terms of Nyquist noise of the known resistances and the amplifier noise. Na+ current fluctuation spectra were interpreted as the sum N/f+SNa(f) where SNa(f) represents the spectrum expected for a set of equal, independent Na+ channels with only 2 conductance states (open or closed) which follow Hodgkin-Huxley kinetics. With values of h.infin., .tau.h and m.infin. measured from macroscopic Na+ currents, the measured spectra were fitted well by optimizing N,SNa(0) and .tau.m. Values of .tau.m obtained by this method were in fair agreement with values found from macroscopic currents. The 1/f component of Na+ current noise was roughly proportional to the square of the steady-state Na current, I2. The mean value of N/I2 was (1.1 .+-. 0.3) .times. 10-4. The current carried by a single Na+ channel was calculated from fitted spectra and steady-state Na+ currents measured simultaneously with the current fluctuations. The single channel conductance .gamma. normalized to zero absolute membrane potential was calculated. The average .gamma. from 12 measurements at depolarizations of 8-40 mV was 7.9 .+-. 0.9 pS [siemen] (SE of mean). The apparent value of .gamma. was smallest with small depolarizations. Variations of the assumed kinetic properties of the model did not drastically affect the single channel conductance. External application of 0.1 mM Ni ion lengthened .tau.m in the macroscopic currents and in the fluctuation spectra and enhanced both the steady-state Na current and the current fluctuations. In Ni-treated nodes .gamma. was smaller than in normal nodes.