Ionic Permeability and the Open Channel Structure of the Nicotinic Acetylcholine Receptor

Abstract

Rat myotubes, clonal BC3H-1 cells, and frog neuromuscular junctions were studied using electrophysiological techniques. The permeation properties of inorganic monovalent, inorganic divalent, and large organic cations were examined in the nicotinic acetylcholine receptor channel. Three results indicated there is one main ion binding site within the pore directly in the permeation pathway. This finding was explained by streaming potential measurements, which indicated that the narrowest cross section of the pore is very short. The narrow region is where the protein can best coordinate around the permeant to form a binding site. Since the narrow region is very short, electrostatic repulsion allows only one cation to bind there at a time. Other results indicated more complicated aspects of permeation. The channel was found to have a high affinity for divalent cations and large organic cations. In addition, some single-channel measurements were not consistent with a transport model simply having one binding site. Even these more complicated results were explained by an ion transport model that accounted for the size, shape, and net negative charge of the pore. In summary, the results were consistent with the channel having wide funnel-shaped vestibules containing net negative charge. The charged residues in the vestibules attract cations and provide binding sites. Although these binding sites and local potentials in the vestibule influence permeation, the binding sites may not be directly in the permeation pathway. The narrowest cross section provides one main ion binding site in the permeation pathway. The tapering region and narrowest region of the pore are lined by transmembrane alpha-helices with a low charge density. Since the narrowest cross section is very short, it is possible that the transmembrane helices are tilted in the open pore. Away from the narrowest region, the helices lining the pore spread out. A second set of helices could then be exposed to the pore at the interstices between the first set of helices.