1. Recombinant a-amino-3-hydroxy-5-methyl-4-isoxazolepropionate receptor (AMPAR) subunits (GluR-A or GluR-B) and kainate receptor (KAR) subunit (GluR-6) in their unedited (Q)-and edited (R)-forms were expressed in HEK 293 cells. To estimate the dimensions of the narrow portion of these channels, biionic reversal potentials for organic cations of different mean diameters were determined with Cs+ as the internal reference ion. 2. Homomeric channels assembled from Q-form subunits were cation selective. The relation between the relative permeability and the mean size of different organic cations suggests that the diameter of the narrow portion of Q-form channels is approximately 0f78 nm for AMPAR and 0 75 nm for KAR channels. 3. Homomeric channels assembled from R-form subunits were permeant for anions and cations.When probed with CsCl gradients the relative chloride permeability (Pcl/lPc) was estimated as 0-14 for GluR-B(R) and 074 for GluR-6(R)-subunit channels. The permeability versus mean size relation for large cations measured with the weakly permeant F-as anion, indicates that for the R-form KAR channels the apparent pore diameter is close to 0'76 nm. 4. Heteromeric AMPAR and KAR channels co-assembled from Q-and R-form subunits were cation selective. The diameter of the narrow portion of these channels is estimated to be in the range between 0 70 and 0 74 nm. 5. The results indicate that the diameters of the narrow portion of AMPAR and KAR channels of different subunit composition and of widely different ion selectivity are comparable. Therefore, the differences in the anion versus cation selectivity, in Ca2+ permeability and in channel conductance are likely to be determined by the difference in charge density of the channel.Ionotropic glutamate receptors (GluRs) which mediate fast excitatory synaptic transmission in the mammalian central nervous system (CNS) are thought to be cationselective channels (for review see e.g. Dingledine et al. 1988). They are subdivided into three functionally and structurally distinct subtypes, N-methyl-D-aspartate

Glutamate-activated single-channel and ensemble currents were recorded from Xenopus laevis oocytes and HEK 293 cells expressing a recombinant NMDA receptor, assembled from NR1 and NR2A subunits. Cesium was the main charge carrier, and organic cations were used to determine the presence of vestibules of this channel and to estimate its pore diameter. The large organic cations tris-(hydroxymethyl)-aminomethane (Tris), N-methyl-glucamine (NMG), arginine (NMG), arginine (Arg), choline, and tetramethylammonium (TMA), when added in millimolar concentrations to the extracellular or cytoplasmic side, produced a voltage-dependent blockade of single-channel Cs+ currents. These molecules behaved as impermeant ions that only partially traverse the channel from either side. The smaller cations trimethylammonium (TriMA) and dimethylammonium (DMA) produced a small and nearly voltage-independent reduction in current amplitude, suggesting that they are permeant. In biionic experiments with Cs+ as the reference ion, the large blocking cations NMG, Arg, Tris, TMA, choline, hexamethonium (Hme), triethylammonium (TriEA), and tetraethylammonium (TEA) showed no measurable permeability. TriMA and smaller ammonium derivatives were permeant. Both the permeability and single-channel conductance of organic cations, relative to Cs+, decreased as the ion size increased. The results suggest that the NMDA receptor has extracellular and cytoplasmic mouths that can accommodate large cations up to 7.3 A in mean diameter. The narrow portion of the pore is estimated to have a mean diameter of 5.5 A.

A series of quaternary ammonium (QA) ions were used to probe the gross architecture of the ion conduction pathway in a Ca2+-activated K+ channel from rat muscle membrane. The channels were inserted into planar phospholipid membranes and the single channel currents were measured in the presence of the different QA ions. Internally applied monovalent QA ions (e.g. tetramethylammonium and analogues) induced a voltage-dependent blockade with a unique effective valence of the block equal to 0.30, and blocking potency increases as the compound is made more hydrophobic. Blockade is relieved by increasing the K+ concentration of the internal or external side of the channel. The effective valence of block is independent of K+ concentration. These results suggest that, from the internal side, all monovalent QA ions interact with a site located in the channel conduction system. Divalent QA ions of the type n-alkyl-bis-alpha,beta-trimethylammonium (bisQn) applied internally also block the channel in a voltage dependent fashion. For short chains (bisQ2-bisQ5), the effective valence decreases with chain length from 0.41 to 0.27, it remains constant for bisQ5 to bisQ6 and increases up to 0.54 for bisQ10. This dependence of block with chain length implies that 27% of the voltage drop within the channel occurs over a distance of approximately 1 nm. Externally applied monovalent QA ions also block the channel. The site is specific for tetraethylammonium; increasing or decreasing the side chains in one methylene group decrease potency by about 400-fold. It is concluded that the Ca2+-activated K+ channel has wide mouths located at each end and that they are different in molecular nature.

The ¢z4 subunit of the neuronal nicotinic acetylcholine receptor is the first gene shown to be involved in a human idiopathic epileptic disease. A missense mutation, leading to the replacement of serine 248 by phenylalanine in the second transmembrane segment, had been detected in patients with autosomal dominant nocturnal frontal lobe epilepsy. The properties of the wild type receptor composed of ~4 and ~2 subunits and the mutant receptor where ¢z4 subunits carried the mutation at serine 248 were compared by means of cDNA manipulation and expression in Xenopus oocytes. The mutant receptor exhibited faster desensitization upon activation by acetylcholine and recovery from the desensitized state was much slower than in the wild type receptor. We conclude that the reported mutation causes seizures via a diminution of the activity of the ¢x4[~2 neuronal nicotinic acetylcholine receptor.