| Abstract: | Batrachotoxin-modified, voltage-dependent sodium channels from canine forebrain were incorporated into planar lipid bilayers. Single-channel conductances were studied for Na+] ranging between 0.02 and 3.5 M. Typically, the single-channel currents exhibited a simple two-state behavior, with transitions between closed and fully open states. Two other conductance states were observed: a subconductance state, usually seen at NaCl] greater than or equal to 0.5 M, and a flickery state, usually seen at NaCl] less than or equal to 0.5 M. The flickery state became more frequent as NaCl] was decreased below 0.5 M. The K+/Na+ permeability ratio was approximately 0.16 in 0.5 and 2.5 M salt, independent of the Na+ mole fraction, which indicates that there are no interactions among permeant ions in the channels. Impermeant and permeant blocking ions (tetraethylammonium, Ca++, Zn++, and K+) have different effects when added to the extracellular and intracellular solutions, which indicates that the channel is asymmetrical and has at least two cation-binding sites. The conductance vs. Na+] relation saturated at high concentrations, but could not be described by a Langmuir isotherm, as the conductance at low NaCl] is higher than predicted from the data at NaCl] greater than or equal to 1.0 M. At low NaCl] (less than or equal to 0.1 M), increasing the ionic strength by additions of impermeant monovalent and divalent cations reduced the conductance, as if the magnitude of negative electrostatic potentials at the channel entrances were reduced. The conductances were comparable for channels in bilayers that carry a net negative charge and bilayers that carry no net charge. Together, these results lead to the conclusion that negative charges on the channel protein near the channel entrances increase the conductance, while lipid surface charges are less important. |
