Complex channel activity recorded from rat liver GAP junctional membranes incorporated into lipid bilayers

RESUMO

Channels from isolated liver junctional membranes were incorporated into lipid bilayers and studied under voltage clamp conditions. Detergent treatment of junctional membrane fragments greatly increased the incidence of channel incorporation but did not noticeably alter the properties of the incorporated channels. Incorporation resulted in channel activity displaying an approximately symmetric voltage dependence in which conductance was decrease with imposed transmembrane voltage exceeding ñ 20 mV. A residual coltage-independent conductance was also detected in membranes in which liver junctional membranes were incorporated. The magnitude of this voltage-insensitive component varied from less than 20% to more than 75% of the total conductance. These results are generally similar to those described by Young, Chn and Gilula(Cell, 48733-743, 1987) in incorporation experiments following detergent treatment of isolated gap junction membranes. However, we interpret these data as indicating the existence of distint channel populations in the incorporated membrane fractions. Our results suggest that a population of larger conductance channels (* 150 pS) contributes the voltage-dependent component of the membrane conductance, while smailler channels (unitary conductance abouth 50-150 pS) contribute the voltage-independent component. The biophysical proprieties of the larger channel are comparable to those seen in communication-deficient cells transfected with connexin32, confirming a report describing conductance of bilayers in which electroeluted 27-kDa liver gap junction protein was inserted. These findings indicate that connexin32 comprises the larger, voltage-dependent channels seen in the bilayer experiments in which liver junctional membranes are incorporated