CHARACTERIZATION OF A VOLTAGE-DEPENDENT POTASSIUM CHANNEL IN SQUID SCHWANN-CELLS RECONSTITUTED IN PLANAR LIPID BILAYERS

An affinity column prepared with noxiustoxin (NTx), a K+ channel block er from the venom of the Mexican scorpion Centruroides noxius, was use d to purify a functional channel from a detergent extract of Schwann c ell membrane of the giant axon of the squid Loligo vulgaris. The purif ied protein was reconstituted as a functional unit in a planar lipid b ilayer and tested with a sequence of potentials to obtain information about single-channel amplitude and kinetics. The reconstituted channel showed delayed rectifier behavior with a slope conductance of 10 pS u nder 5:1 asymmetric KCl concentrations and a clear tendency to open un der negative potentials. The zero-current potential was +36 mV, which fitted well with the Nernst equation for the CIS/TRANS K+-concentratio n ratio of 5:1. The channel also showed a strong sensitivity to tetrae thylammonium and its activity was inhibited by NTx, as expected from t he purification procedure. The behavior of this protein in the presenc e of 0.5 mM ATP (cis side) was also tested, significantly increasing c urrent fluctuations across the membrane. In order to compare the modul ation of the Schwann cell K+ channel with that of the axonal K+ channe l, a purified protein from the squid axon membrane was also tested in the presence of ATP. This 10-11 pS, delayed rectifier channel from the squid giant axon (Prestipino et al., FEES Lett. 250:570-574, 1989) wa s also tested in the presence of ATP and showed a similar rise in acti vity. (C) 1995 Wiley-Liss, Inc.