Key points Wnt ligands belonging to both canonical and non‐canonical Wnt pathways regulate membrane potential signifying a very early event in the signal transduction. Wnts activate K+ currents by elevating intracellular Ca2+ and trigger Ca2+ release from intracellular stores. Control of potential by Wnt ligands has significant implications for gene transcription and opens up a novel avenue to interfere with this critical pathway. Abstract The Wnt signalling network determines gene transcription with free intracellular Ca2+ (Cai2+) and β‐catenin as major intracellular signal transducers. Despite its critical importance during development and disease, many of the basic mechanisms of Wnt signal activation remain unclear. Here we show by single cell recording and simultaneous Cai2+ imaging in mammalian prostate cancer cells that an early step in the signal cascade is direct action on the cell membrane potential. We show that Wnt ligands 5A, 9B and 10B rapidly hyperpolarized the cells by activating K+ current by Ca2+ release from intracellular stores. Medium‐throughput multi‐well recordings showed responses to Wnts at concentrations of 2 nm. We identify a putative target for early events as a TRPM channel. Wnts thus act as ligands for ion channel activation in mammalian cells and membrane potential is an early indicator of control of transcription.

Class II antiarrhythmics or b-blockers are antisympathetic nervous system agents that act by blocking b-adrenoceptors. Despite their common clinical use, little is known about the effects of b-blockers on free intracellular calcium (Ca 21 i), an important cytosolic second messenger and a key regulator of cell function. We investigated the role of four chemical analogs, commonly prescribed b-blockers (atenolol, metoprolol, propranolol, and sotalol), on Ca 21 i release and whole-cell currents in mammalian cancer cells (PC3 prostate cancer and MCF7 breast cancer cell lines). We discovered that only propranolol activated free Ca 21 i release with distinct kinetics, whereas atenolol, metoprolol, and sotalol did not. The propranololinduced Ca 21 i release was significantly inhibited by the chelation of extracellular calcium with ethylene glycol tetraacetic acid (EGTA) and by dantrolene, an inhibitor of the endoplasmic reticulum (ER) ryanodine receptor channels, and it was completely abolished by 2-aminoethoxydiphenyl borate, an inhibitor of the ER inositol-1,4,5-trisphosphate (IP 3) receptor channels. Exhaustion of ER stores with 4-chloro-m-cresol, a ryanodine receptor activator, or thapsigargin, a sarco/ER Ca 21 ATPase inhibitor, precluded the propranolol-induced Ca 21 i release. Finally, preincubation of cells with sotalol or timolol, nonselective blockers of b-adrenoceptors, also reduced the Ca 21 i release activated by propranolol. Our results show that different b-blockers have differential effects on whole-cell currents and free Ca 21 i release and that propranolol activates store-operated Ca 21 i release via a mechanism that involves calcium-induced calcium release and putative downstream transducers such as IP 3. The differential action of class II antiarrhythmics on Ca 21 i release may have implications on the pharmacology of these drugs.

v 1.5 recordings from cardiomyocyte-cardiomyocyte and cardiomyocyte-fibroblasts junctions. Functional and dominant-negative Cx43 EGFP adenoviruses were used to probe active/inactive junctions. The Na v b1 adhesion inhibitor peptide (badp1) was tested. Whole-cell Na v -currents were recorded to separate non-junctional badp1 effects. To assess transdepolarisation, current-clamp and optical measurements were combined. Plasma membranes were analysed for junctional protein density. Exogenously-delivered Cx43 EGFP trafficked to regions where Na v activity and cluster size were increased relative to native Cx43-junctions (p=0.015). Smaller Na v -clusters (5-10) were abundant at native junctions, larger clusters (20-30) predominated at Cx43 EGFP -junctions. Acute badp1 peptide treatment caused Na v downregulation at native (p<0.01) and Cx43 EGFP -rich junctions (p<0.001). Lack of badp1 effect on the whole-cell current supports a role for b1 as a trans-cellular adhesion molecule. Blockade of cardiomyocyte transactivation using optical/electrical recordings was demonstrated with badp1 pre-treatment. Reduced Na v -current and smaller channel clusters were found at cardiomyocyte-fibroblast versus cardiomyocyte-cardiomyocytes junctions. Membranes from junction-remote regions showed similar currents, but different Na v -cluster size. Strength of intercellular junctions depend on Na v -current, intact Na v b1 and the amount of functional Cx43. Cardiac junction type hetero-vs. homo-cellular governs Na v activity and distribution. Novel badp1 peptide establishes importance of perinexal regions for signal propagation. Our findings and biophysical approach elucidate molecular origin of arrhythmogenic activity within/around GJ, revealing rationale for the anti-arrhythmic effects of Na v b1-mediated adhesion.

TM287/288 ATP binding cassette transporter (pdb-code 4Q4H;~25% sequence identity). These models are currently used to establish the structural basis of known Cantú associated mutants. Two known gain-of function mutations (V65M and C176S) are located in the pore module of the KATP channel. Both amino acids are in close contact in the closed channel state. During channel opening, this region undergoes large conformational changes, including rearrangements of these two amino acids. To further investigate how perturbations in this area influence gating, we are currently performing molecular dynamics simulations. From these studies, a better understanding of Cantú disease and the function of the KATP channel in general (a well-known pharmaceutical target for treatment of diabetes mellitus) are expected.

surface to a large cytoplasmic cavity, and a unique configuration of the photoactive site likely responsible for the delayed Schiff base deprotonation. Using this structural information, we have identified residues that determine absorption wavelength and current-voltage dependence. Our results provide clues for rational engineering of ACR molecules further to increase their optogenetic utility.