Single voltage-dependent potassium channels in cultured rat hippocampal neurons

Abstract

Single-channel recordings using the gigohm seal patch-clamp technique were carried out on the somatic membranes of dissociated embryonic rat hippocampal neurons grown in cell culture. The recording medium contained tetrodotoxin to block the voltage-dependent Na+ conductance and Cd2+ to block Ca2+ and Ca2+-activated conductances. In the cell-attached configuration, depolarizing voltage steps activated outward directed single-channel currents with conductance 15-20 pS. The channel openings exhibited a moderate degree of flickering. The mean burst lifetimes ranged from 5 to 13 ms with a tendency to increase slightly at more depolarized potentials (T = 21-25 degrees C). Reversal potential measurements using excised membrane patches indicated that the channels behaved as expected of a K+-selective membrane pore. Channel opening occurred in Ca2+-free EGTA-containing solutions but was never observed in the presence of tetraethylammonium (TEA; 20 mM). The frequency of channel opening increased as the membrane was depolarized by up to 50 mV from resting potential; the fraction of time spent in the open state during the first 300 ms following a step depolarization increased e-fold for a 8-25 mV change in potential. First-latency histograms and simulations of the macroscopic current based on channel data obtained during repeated depolarizing voltage steps indicated that the probability of the channel being in the open state increases gradually with time after a step depolarization. During repeated depolarizing steps the channels appeared to randomly enter and exit a long-lived inactive state. It is concluded that these channels may underly the slowly activating, very slowly inactivating, TEA-sensitive voltage-dependent K+ current (IK) in cultured hippocampal neurons.