Voltage-gated ion channels contain a positively charged transmembrane segment termed S4. Recent evidence suggests that depolarisation of the membrane potential causes this segment to undergo conformational changes that, in turn, lead to the opening of the channel pore. In order to define these conformational changes in structural terms, we have introduced single cysteine substitutions into the S4 segment of the prototypical Shaker K+ channel at various positions and expressed the mutants in Xenopus oocytes. The cells were depolarised to induce K+ currents and the effect of application of 100 microM parachloromercuribenzenesulphonate (PCMBS) on these currents was examined by the two-electrode voltage-clamp technique. PCMBS inhibited K+ currents elicited by mutants L358C, L361C, V363C and L366C, but not those by V367C and S376C. Since PCMBS is a membrane-impermeable cysteine-modifying reagent, the data suggest that depolarisation must have caused the S4 segment to move out of the lipid bilayer into the extracellular phase rendering the residues at positions 358, 361, 363 and 366 susceptible to PCMBS attack. The lack of effect of PCMBS on V367C suggests that the exposure of S4 terminates at L366. Detailed analysis of L361C mutant revealed that the S4 movement can occur even below the resting potential of the cell, at which potential voltage-gated K+ channels are normally in a non-conducting closed state.

The 5‐HT3 receptor is a member of the ‘Cys‐loop’ family of ligand‐gated ion channels that mediate fast excitatory and inhibitory transmission in the nervous system. Current evidence points towards native 5‐HT3 receptors originating from homomeric assemblies of 5‐HT3A or heteromeric assembly of 5‐HT3A and 5‐HT3B. Novel genes encoding 5‐HT3C, 5‐HT3D, and 5‐HT3E have recently been described but the functional importance of these proteins is unknown. In the present study, in silico analysis (confirmed by partial cloning) indicated that 5‐HT3C, 5‐HT3D, and 5‐HT3E are not human–specific as previously reported: they are conserved in multiple mammalian species but are absent in rodents. Expression profiles of the novel human genes indicated high levels in the gastrointestinal tract but also in the brain, Dorsal Root Ganglion (DRG) and other tissues. Following the demonstration that these subunits are expressed at the cell membrane, the functional properties of the recombinant human subunits were investigated using patch clamp electrophysiology. 5‐HT3C, 5‐HT3D, and 5‐HT3E were all non‐functional when expressed alone. Co‐transfection studies to determine potential novel heteromeric receptor interactions with 5‐HT3A demonstrated that the expression or function of the receptor was modified by 5‐HT3C and 5‐HT3E, but not 5‐HT3D. The lack of distinct effects on current rectification, kinetics or pharmacology of 5‐HT3A receptors does not however provide unequivocal evidence to support a direct contribution of 5‐HT3C or 5‐HT3E to the lining of the ion channel pore of novel heteromeric receptors. The functional and pharmacological contributions of these novel subunits to human biology and diseases such as irritable bowel syndrome for which 5‐HT3 receptor antagonists have major clinical usage, therefore remains to be fully determined.

A novel series of AMPAR positive modulators is described that were identified by high throughput screening. The molecules of the series have been optimized from a high quality starting point hit to afford excellent developability, tolerability, and efficacy profiles, leading to identification of a clinical candidate. Unusually for an ion channel target, this optimization was integrated with regular generation of ligand-bound crystal structures and uncovered a novel chemotype with a unique and highly conserved mode of interaction via a trifluoromethyl group.

A series of AMPA receptor positive allosteric modulators has been optimized from poorly penetrant leads to identify molecules with excellent preclinical pharmacokinetics and CNS penetration. These discoveries led to 17i, a potent, efficacious CNS penetrant molecule with an excellent pharmacokinetic profile across preclinical species, which is well tolerated and is also orally bioavailable in humans.